WorldWideScience

Sample records for alizarin

  1. Marking pike fry otoliths with alizarin complexone and strontium : an evaluation of methods

    DEFF Research Database (Denmark)

    Skov, Christian; Grønkjær, P.; Nielsen, C.

    2001-01-01

    Laboratory experiments demonstrated that both alizarin complexone and strontium are useful in mass marking of pike Esox lucius fry otoliths. Visual detection of alizarin complexone marks was considered more reliable than the quantitative analysis of strontium for differentiating marked and unmarked...

  2. Alizarin Red S as an electrochemical indicator for saccharide recognition

    Energy Technology Data Exchange (ETDEWEB)

    Schumacher, Soeren, E-mail: soeren.schumacher@ibmt.fraunhofer.de [Fraunhofer Institute for Biomedical Engineering, Am Muehlenberg 13, 14476 Potsdam (Germany); Nagel, Thomas; Scheller, Frieder W.; Gajovic-Eichelmann, Nenad [Fraunhofer Institute for Biomedical Engineering, Am Muehlenberg 13, 14476 Potsdam (Germany)

    2011-07-30

    Graphical abstract: Display Omitted Highlights: > Transfer of the established Alizarin Red S (ARS) for saccharide-boronic acid interaction to electrochemistry. > Investigation of electrochemical behaviour of ARS and its interaction with phenylboronic acid at pH 7.4. > Through addition of fructose the ARS was displaced. > Displaced ARS could be monitored electrochemically corresponding to the used fructose concentration. - Abstract: In addition to the well-established spectroscopic Alizarin Red S (ARS) assay for the determination of binding constants between arylboronic acids and different saccharides, we report the use of ARS as a reporter in an electrochemical set-up. The electrochemical properties of ARS, the binding to phenyl boronic acid (PBA) and the competition with fructose in phosphate buffer at pH 7.4 were investigated by cyclic voltammetry (CV). By choosing a negative scan direction (starting at +0.2 V), a quasi-reversible process was detected at E{sup 0}' = -0.59V with {Delta}E{sub p} = 0.1V. An irreversible oxidation peak at +0.42 V could also be detected. These peaks are characterised both as a 2-proton-2-electron transfer and corresponds to the oxidation and reduction of the anthraquinone or the ortho-quinone moiety. After addition of phenylboronic acid a new oxidation peaks occurred at -0.42 V which correlates with the ARS-PBA interaction. The peak current increased with increasing phenylboronic acid concentration according to the release of BA and formation of the ARS-PBA ester. After addition of fructose the peak current decreases again, in proportion to the fructose concentration, enabling the use of ARS as an electrochemical reporter for fructose detection up to 50 mM. Also the interaction with other cis-diol containing compounds such as sorbitol, mannitol, glucose and mannose was investigated and a dependence based on already published binding constants to phenylboronic acid could be shown.

  3. Hydrated alizarin complexes: hydrogen bonding and proton transfer.

    Science.gov (United States)

    Huh, Hyun; Cho, Sung Haeng; Heo, Jiyoung; Kim, Nam Joon; Kim, Seong Keun

    2012-07-07

    We investigated the hydrogen bonding structures and proton transfer for the hydration complexes of alizarin (Az) produced in a supersonic jet using fluorescence excitation (FE), dispersed laser induced fluorescence (LIF), visible-visible hole burning (HB), and fluorescence detected infrared (FDIR) spectroscopy. The FDIR spectrum of bare Az with two O-H groups exhibits two vibrational bands at 3092 and 3579 cm(-1), which, respectively, correspond to the stretching vibration of O1-H1 that forms a strong intramolecular hydrogen bond with the C9=O9 carbonyl group and the stretching vibration of O2-H2 that is weakly hydrogen-bonded to O1-H1. For the 1:1 hydration complex Az(H(2)O)(1), we identified three conformers. In the most stable conformer, the water molecule forms hydrogen bonds with the O1-H1 and O2-H2 groups of Az as a proton donor and proton acceptor, respectively. In the other conformers, the water binds to the C10=O10 group in two nearly isoenergetic configurations. In contrast to the sharp vibronic peaks in the FE spectra of Az and Az(H(2)O)(1), only broad, structureless absorption was observed for Az(H(2)O)(n) (n≥ 2), indicating a facile decay process, possibly due to proton transfer in the electronic excited state. The FDIR spectrum with the wavelength of the probe laser fixed at the broad band exhibited a broad vibrational band near the O2-H2 stretching vibration frequency of the most stable conformer of Az(H(2)O)(1). With the help of theoretical calculations, we suggest that the broad vibrational band may represent the occurrence of proton transfer by tunnelling in the electronic ground state of Az(H(2)O)(n) (n≥ 2) upon excitation of the O2-H2 vibration.

  4. Spectrophotometric determination of piroxicam and tenoxicam in pharmaceutical formulations using alizarin.

    Science.gov (United States)

    Amin, Alaa S

    2002-07-20

    New spectrophotometric procedures have been established for the quantitation of piroxicam and tenoxicam. The procedures are based on the reaction between the examined drug and alizarin (I), alizarin red S (II), alizarin yellow G (III) or quinalizarin (IV) producing ion-pair complexes which can be measured at the optimum wavelength. The optimization of the reaction conditions is investigated. Beer's law is obeyed in the concentration ranges 0.05-2.40 microg ml(-1), whereas optimum concentration as adopted from Ringbom plots was 0.12-2.25 microg ml(-1). The molar absorptivity, Sandell sensitivity, detection and quantification limits are also calculated. The correlation coefficient was >/=0.9990 (n=10) with a relative standard deviation (R.S.D.) of

  5. Nanoparticle-enhanced fluorescence emission for non-separation assays of carbohydrates using a boronic acid-alizarin complex.

    Science.gov (United States)

    Li, Qianjin; Kamra, Tripta; Ye, Lei

    2016-03-01

    Addition of crosslinked polymer nanoparticles into a solution of a 3-nitrophenylboronic acid-alizarin complex leads to significant enhancement of fluorescence emission. Using the nanoparticle-enhanced boronic acid-alizarin system has improved greatly the sensitivity and extended the dynamic range of separation-free fluorescence assays for carbohydrates.

  6. Fast and efficient removal of alizarin yellow dye (Azo dye from water and wastewater samples using modified nanoclay

    Directory of Open Access Journals (Sweden)

    Shahla Elhami

    2014-07-01

    Full Text Available A fast and efficient method has been developed for removal of Alizarin Yellow dye using modified nanoclay. Montmorillonite (MMT was modified by a facile and one-step procedure with diethylenetriamine (DETA and was used as an adsorbent. The effects of pH value of the dye solution, adsorbent dose, adsorption time and the initial dye concentration on the Alizarin Yellow adsorption onto the composite were investigated.  The DETA-MMT had a high uptake capacity in room temperature and could remove Alizarin Yellow dye of about 85 % with 6 g/L of adsorbent, in only 2 min. Langmuir and Freundlich isotherms were employed for the study of the adsorption of Alizarin Yellow dye onto DETA-MMT. The method was applied to the removal of Alizarin Yellow in different tap water, river water and industrial wastewater samples.

  7. Structure and properties of alizarin complex formed with alkali metal hydroxides in methanol solution.

    Science.gov (United States)

    Jeliński, Tomasz; Cysewski, Piotr

    2016-06-01

    Quantum chemical computations were used for prediction of the structure and color of alizarin complex with alkali metal hydroxides in methanolic solutions. The color prediction relying on the single Gaussian-like band once again proved the usefulness of the PBE0 density functional due to the observed smallest color difference between computed and experimentally derived values. It was found that the alkali metal hydroxide molecules can bind to the two oxygen atoms of both hydroxyl groups of alizarin or to one of these atoms and the oxygen atom from the keto group in a complex with three methanol molecules. This means that two electronic transitions need to be taken into account when considering the spectra of the studied complexes. The resulting bond lengths and angles are correlated with the properties of the alkali metal atoms. The molar mass, the atomic radius, and the Pauling electronegativity of studied metals are quite accurate predictors of the geometric properties of hydroxide complexes with alizarin in methanol solution. Graphical abstract The spectra of the neutral and monoanionic form of alizarin together with color changes resulting from addition of different metal hydroxides and represented in CIE color space.

  8. Imaging the Ultrafast Photoelectron Transfer Process in Alizarin-TiO2

    Directory of Open Access Journals (Sweden)

    Tatiana Gomez

    2015-07-01

    Full Text Available In this work, we adopt a quantum mechanical approach based on time-dependent density functional theory (TDDFT to study the optical and electronic properties of alizarin supported on TiO2 nano-crystallites, as a prototypical dye-sensitized solar cell. To ensure proper alignment of the donor (alizarin and acceptor (TiO2 nano-crystallite levels, static optical excitation spectra are simulated using time-dependent density functional theory in response. The ultrafast photoelectron transfer from the dye to the cluster is simulated using an explicitly time-dependent, one-electron TDDFT ansatz. The model considers the δ-pulse excitation of a single active electron localized in the dye to the complete set of energetically accessible, delocalized molecular orbitals of the dye/nano-crystallite complex. A set of quantum mechanical tools derived from the transition electronic flux density is introduced to visualize and analyze the process in real time. The evolution of the created wave packet subject to absorbing boundary conditions at the borders of the cluster reveal that, while the electrons of the aromatic rings of alizarin are heavily involved in an ultrafast charge redistribution between the carbonyl groups of the dye molecule, they do not contribute positively to the electron injection and, overall, they delay the process.

  9. Efficient application of nano-TiO2 thin films in the photocatalytic removal of Alizarin Yellow from aqueous solutions

    Science.gov (United States)

    Tiwari, Diwakar; Lalhriatpuia, C.; Lalhmunsiama; Lee, Seung-Mok; Kong, Sung-Ho

    2015-10-01

    The aim of this investigation is to obtain thin films of nano-TiO2 on a borosilicate glass substrate using sol-gel template method. The thin film was immobilized with and without polyethylene glycol as filler media and annealed at 500 °C. Further, thin films were characterized by the IR, XRD, XRF and XPS analytical methods. The surface morphology of these films was obtained by the FE-SEM images and the BET specific surface area and pore sizes were obtained. The nano-TiO2 was, perhaps, formed a nanopillar onto the substrate. The thin films were successfully employed in the photocatalytic degradation of Alizarin Yellow (AY), an azo dye, from aqueous solutions using the UV-light irradiation under batch reactor operations. Various physico-chemical parametric studies, viz., effect of pH, Alizarin Yellow concentration and interfering ions were studied to deduce the mechanism involved in photocatalytic degradation of this pollutant. The time dependence degradation of Alizarin Yellow was provided to demonstrate the kinetics of degradation of this pollutant from aqueous solutions. It was observed that the degradation of Alizarin Yellow followed pseudo-first-order rate kinetics. Study was further extended with total organic carbon measurement using TOC analyser to demonstrate an apparent mineralization of Alizarin Yellow from aqueous solutions. The presence of several interfering ions or even rad OH scavengers suppressed the photo-catalytic action of thin films in AY degradation from aqueous solutions.

  10. Environmental and complexation effects on the structures and spectroscopic signatures of organic pigments relevant to cultural heritage: the case of alizarin and alizarin-Mg(II)/Al(III) complexes.

    Science.gov (United States)

    Carta, Luciano; Biczysko, Malgorzata; Bloino, Julien; Licari, Daniele; Barone, Vincenzo

    2014-02-21

    An integrated computational approach allowed an unbiased analysis of optical and structural properties of alizarin-based pigments, which can be directly compared with experimental results. Madder lake pigments have been modeled by Mg(II)- and Al(III)-coordinated alizarin taking into account solvation and metal-linkage effects, responsible for colour modifications. Moreover, different environmental conditions have been analyzed for free alizarin, showing in all cases semi-quantitative agreement with experimental spectroscopic data (UV-VIS). Our results point out the ability of in silico approaches to unravel the subtle interplay of stereo-electronic, dynamic, and environmental effects in tuning the physico-chemical properties of pigments relevant to cultural heritage.

  11. Inclusion complex of Alizarin Red S with β-cyclodextrin: Synthesis, spectral, electrochemical and computational studies

    Science.gov (United States)

    Chin, Yuk Ping; Abdul Raof, Siti Farhana; Sinniah, Subathra; Lee, Vannajan Sanghiran; Mohamad, Sharifah; Abdul Manan, Ninie Suhana

    2015-03-01

    Inclusion complex formation of Alizarin Red S (ARS) with β-cyclodextrin was studied by UV-visible, Fourier transform infrared (FTIR), nuclear magnetic resonance (NMR), cyclic voltammetry (CV) and molecular modeling methods. FTIR and NMR results had justified that ARS was partly included into the β-CD cavity. The inclusion complex has 1:1 stoichiometry, where the apparent formation constant achieved was 4.137 × 103 L/mol using Benesi-Hildebrand equation. Cyclic voltammetry results shows the peak current decreased as the ARS molecule entered the hydrophobic cavity of β-CD. Molecular modeling results showed that the aromatic ring of the ARS entered into the secondary hydroxyl rim of the CD cavity was more thermodynamically favorable. The lowest stabilization energy, ΔE was -17.80 kcal/mol, and dipole-dipole interaction is was one of the driving forces for the inclusion complex formation.

  12. Enhanced fluorescence quenching in an acridine orange - alizarin red system through matrine and its analytical application

    Science.gov (United States)

    Wei, Xiaoling; Wang, Xiaojun; Gong, Qi; Wang, Lisheng; Zhou, Shiwu

    2015-01-01

    This study shows that alizarin red (AR) only slightly quenched fluorescence for acridine orange (AO) in an AR/AO mixed solution at pH = 5-6. The reduced fluorescent signal was closely and linearly associated with the level of MT added to the system, which is the basis for a new quantitative MT assay method using the fluorescence quenching reaction in the AO-AR system. The results show that under optimal conditions, this method had a 14.9-43.5 mg L-1 linear detection range with a 1.38 mg L-1 detection limit and 1.24% precision. In addition, this method was used to determine the MT levels in the commercially available MT-containing pesticides and suppositories, which showed a 96.6-103% recovery. Therefore, this method has multiple advantages, including simple and fast operation, high accuracy and low cost. Moreover, herein, we investigated the underlying mechanism in-depth using an ultraviolet (UV) spectroscopic technique.

  13. Osmotic induction marking with Alizarin Red S on juveniles of pejerrey, Odontesthes bonariensis (Atherinopsidae

    Directory of Open Access Journals (Sweden)

    Daniela Campanella

    Full Text Available Juveniles of pejerrey, Odontesthes bonariensis, were exposed to 0.1% Alizarin Red S (ARS alone or with a previous immersion in 2.2% saline solution (Osmotic Induction, OI to enhance the ARS marking method. Fish were marked in the field and immediately released in 1 m3 cages in "La Salada de Monasterio" lagoon, Chascomús, Buenos Aires , Argentina. After 73 days, clear marks were observed in the otoliths, caudal fin rays and scales with both treatments, being the intensity of the signal in the scales of OI+ARS treated fish higher. On the other hand, no marks were observed in the control group on the same structures. Approximately one year post-treatment (385 days, only marks in caudal fin rays were found clearly in OI+ARS treated fish. After this period, no significant differences in total length or weight between marked or control fish were observed and the mortality ranged between 30-40 % in all cages. These results provide strong evidence for the potential applicability of this cost-effective marking technique in differentiation of wild and hatchery-produced pejerrey. The success in the caudal fin rays marking is also important because it is easy to do and does not require the sacrifice of fish.

  14. A Novel Electrochemical Method for Protionamide Determination Based on Its Interaction with Alizarin Red S

    Directory of Open Access Journals (Sweden)

    Weili Zhang

    2015-01-01

    Full Text Available The interaction of protionamide with alizarin red S (ARS and its analytical application were carefully investigated in this contribution. The interaction conditions were carefully studied and optimized by cyclic voltammetry. Under the optimum conditions, the cyclic voltammetry curve of ARS showed an oxidation peak with the peak potential of 0.57 V. After the addition of protionamide to the ARS solution, the peak potential was negatively moved, and meanwhile the oxidation peak current decreased apparently to the concentration of protionamide and then a new method for the protionamide determination was established. The linear equation between the decreasing current (Δip and protionamide concentration was got as Δip (μA = 0.01514C (mg/L −0.01553  (n=9; r=0.991 with the linear range of 10.0~50.0 mg/L, and the detection limit (3σ was got as 8.25 μg/mL. The effects of coexisting substances on the determination were carefully investigated and the protionamide artificial and tablet samples were detected with satisfactory results.

  15. Spectrophotometric determination of molybdenum with Alizarin Red S in the presence of poly(sulfonylpiperidinylmethylene hydroxide).

    Science.gov (United States)

    Alkan, Mahir; Kharun, Myroslava; Chmilenko, Fedor

    2003-03-01

    The present work describes a selective and rapid method for the determination of molybdenum with Alizarin Red S (ARS) in the presence of a water soluble polymer, poly(sulfonylpiperidinylmethylene hydroxide) (PSPMH). The ARS modified by PSPMH reacts with molybdenum(VI) in the solutions of pH 3.4-4.0 to produce a red complex. The composition of the complex is 1:4:1 mol ratio of Mo(VI): ARS:PSPMH. The complex obeys Beer's law from 0.05 to 5.50 mug ml(-1) with an optimum range. The molar absorptivity is 2.1x10(4) l mol(-1) cm(-1) at 500 nm. The interference effects of the foreign cations have been examined and it has been determined that only Cu(II), Al(III) and Fe(III) have to be masked by EDTA and tungsten can be tolerated till 4-fold of molybdenum in case of masking by citrate. The method has been applied to the determination of geological samples without solvent extraction or separation steps.

  16. Preparation of a Modified Nanoalumina Sorbent for the Removal of Alizarin Yellow R and Methylene Blue Dyes from Aqueous Solutions

    Directory of Open Access Journals (Sweden)

    Wasan T. Al-Rubayee

    2016-01-01

    Full Text Available A modified form of γ-alumina nanoparticles prepared by immobilization of 2,4-dinitrophenyl hydrazine on γ-alumina nanoparticles coated with sodium dodecyl sulfate (DNPH-γ-alumina for the removal of the anionic dye (Alizarin yellow R and cationic dye (Methylene blue from aqueous solutions has been investigated. The FTIR, SEM, TEM, XRD, BET, and BJH analysis techniques indicate that the modification reaction has occurred. Batch adsorption study revealed that 0.05 g amount of the modified adsorbent was capable of removing 95.6% and 65.6% of Alizarin yellow (AY and Methylene blue (MB dyes, respectively, in 60 min. The experimental equilibrium data showed that Langmuir isotherm applies well for describing the adsorption behavior, and the maximum adsorption capacity was found to be 47.8 mg/g and 32.8 mg/g for AY and MB on DNPH-γ-alumina, respectively. Kinetic studies showed best applicability of the second-order kinetic model. The DNPH-γ-alumina adsorbent proved capability, effectiveness, and selectivity for the removal of Alizarin yellow R dye. Therefore, it is possible to increase the efficiency of an adsorbent for the removal of pollutants by applying a modification to the surface of the adsorbent, and DNPH as a modifier proved efficient for the removal of a wider range of pollutants including metal ions and dye compounds.

  17. Adsorptive Cathodic Stripping Voltammetric Method with Alizarin for the Simultaneous Determination of Cadmium, and Zinc in Water Samples

    Directory of Open Access Journals (Sweden)

    Deswati

    2016-12-01

    Full Text Available This paper reports on the development of adsorptive cathodic stripping voltammetric (AdCSV method with 1,2-dihydroxyanthraquinone or Alizarin (AZ as a complexing agent used in the simultaneous determination of ultra trace of Cd and Zn because it has a good sensitivity, and selectivity. The influence of several parameters was studied: the effects of 1,2-dihydroxyanthraquinone or Alizarin (AZ concentration, pH, accumulation potential, and accumulation time. The relative standard deviation (RSD, and recovery is determined to get the accuracy and precision method. It also determined the limit of detection (LOD of the method to get the sensitivity. In this case, the optimum conditions were AZ concentration of 0.5 mM, pH 5, step deposition (70 s, -0.5 V. This method has been applied in water samples successfully, was obtained (Cd 23, and Zn 124 µg/L, LOD (Cd 0.006, and Zn 0.004 µg/L, RSD (Cd 0.4, and Zn 1.4 % (n = 10, recovery (Cd 99.36, and Zn 99.28%. The Atomic absorption spectrometric (AAS method is used as a comparison AdCSV optimum, was obtained (Cd 16, and Zn 115 µg/L.

  18. Electrocatalytic oxidation of hydrazine with alizarin red S as a homogenous mediator on the glassy carbon electrode

    Institute of Scientific and Technical Information of China (English)

    Mohammad; Mazloum-Ardakani; Roya; Mazidi; Mohammad; Hossein; Mashhadizadeh; Parvanah; Rahimi; Mohammad; Ali; Karimi

    2010-01-01

    Electro-catalytic oxidation and detection of hydrazine on a glassy carbon electrode,at pH 6.0,was studied by using alizarin red S as a homogeneous mediator.The overall number of electrons involved in the catalytic oxidation of hydrazine and that involved in the rate-determining step were four and one,respectively.The interfering effect of some cations,anions and organic compounds were examined.Peak current for this process varied linearly with the square root of the scan rate.The kinetic parameters,such as the electron transfer coefficient(α) and catalytic rate constant(k) ,were determined using cyclic voltammetry,linear sweep voltammetry and chronoamperometry.The electro-catalytic response was optimized with regards to the pH,scan rate,hydrazine concentration and other variables.

  19. Larvicidal and Pupicidal Activities of Alizarin Isolated from Roots of Rubia cordifolia Against Culex quinquefasciatus Say and Aedes aegypti (L.) (Diptera: Culicidae).

    Science.gov (United States)

    Gandhi, M R; Reegan, A D; Ganesan, P; Sivasankaran, K; Paulraj, M G; Balakrishna, K; Ignacimuthu, S; Al-Dhabi, N A

    2016-08-01

    The mosquitocidal activities of different fractions and a compound alizarin from the methanol extract of Rubia cordifolia roots were evaluated on larvae and pupae of Culex quinquefasciatus Say and Aedes aegypti (L.) (Diptera: Culicidae). Larvae and pupae were exposed to concentrations of 2.5, 5.0, 7.5 and 10 ppm for fractions and 0.5, 1.0, 1.5 and 2.0 ppm for compound. After 24 h, the mortality was assessed and the LC50 and LC90 values were estimated for larvae and pupae. Among the 23 fractions screened, fraction 2 from the methanol extract of R. cordifolia showed good mosquitocidal activity against C. quinquefasciatus and A. aegypti. LC50 and LC90 values of fraction 2 were 3.53 and 7.26 ppm for C. quinquefasciatus and 3.86 and 8.28 ppm for A. aegypti larvae, and 3.76 and 7.50 ppm for C. quinquefasciatus and 3.92 and 8.05 ppm for A. aegypti pupae, respectively. Further, the isolated compound alizarin presented good larvicidal and pupicidal activities. LC50 and LC90 values of alizarin for larvae were 0.81 and 3.86 ppm against C. quinquefasciatus and 1.31 and 6.04 ppm for A. aegypti larvae, respectively. Similarly, the LC50 and LC90 values of alizarin for pupae were 1.97 and 4.79 ppm for C. quinquefasciatus and 2.05 and 5.59 ppm for A. aegypti pupae, respectively. The structure of the isolated compound was identified on the basis of spectroscopic analysis and compared with reported spectral data. The results indicated that alizarin could be used as a potential larvicide and pupicide.

  20. Linear sweep voltammetric studies on the supramolecular complex of alizarin red S with lysozyme and determination of lysozyme

    Indian Academy of Sciences (India)

    Wei Sun; Na Zhao; Xueliang Niu; Yan Wang; Kui Jiao

    2009-03-01

    An electrochemical method for the determination of lysozyme (LYS) based on its interaction with alizarin red S (ARS) was established by linear sweep voltammetry in this paper. The electrochemical behaviour of ARS with LYS was investigated on a dropping mercury working electrode in 0.2 mol/L pH 4.8 Britton-Robinson (B-R) buffer solution. ARS showed a sensitive second order derivative linear sweep voltammetric reductive peak at -0.42 V (vs SCE). After the addition of LYS, the reductive peak current of ARS decreased without the shift of the reductive peak potential and no new waves appeared, which was due to the formation of a supramolecular complex of ARS with LYS in the solution. The stoichiometry of the ARS-LYS complex was further calculated by the electrochemical data with the results of the binding ratio as 3 : 1 and the binding constant as 2.82 × 1014. Under the selected conditions, the decrease of the second order derivative linear sweep voltammetric reductive peak current of ARS was in proportion to the LYS concentration in the range from 0.8 to 35.0 mg/L and the detection limit of LYS was calculated as 0.52 mg/L (3). Different kinds of LYS samples were detected satisfactorily with this method.

  1. Adsorption Capacity of The As-Synthetic Graphene Oxide for The Removal of Alizarin Red S Dye from Aqueous Solution

    Directory of Open Access Journals (Sweden)

    Prawit Nuengmatcha

    2016-06-01

    Full Text Available This research was aimed to study the adsorption of Alizarin Red S (ARS dye using graphene oxide (GO as an adsorbent compared with bare graphite powder (BGP. For optimum conditions, the effects of the initial concentration of ARS, solution pH, adsorbent dosage, and contact time were investigated in detail. The optimum conditions for this work were consisted of 350 mg/L initial concentration of ARS with 0.02 mg adsorbent at pH 2.0. The adsorption equilibrium was completely reached within 30 min. The maximum adsorption capacity of GO was 88.50 mg/g which was higher than that of BGP (34.13 mg/g. The adsorption kinetics well fitted using a pseudo second-order kinetic model. The intraparticle diffusion model described that the intraparticle diffusion was not the only rate-limiting step. In thermodynamics diversion, changes in free energy (DG˚, enthalpy (DH˚ and entropy (DS˚ were also evaluated. The overall adsorption process was exothermic and spontaneous in nature. The adsorption isotherms for GO and BGP fit well with the Langmuir and Freundlich models, respectively. It is, therefore, evident that the as-prepared GO can be used as a high potential adsorbent for the anionic dye and it can be reused for fourth time of adsorption.

  2. Studies on thermo-optic property of chitosan–alizarin yellow GG complex: a direction for devices for biomedical applications

    Indian Academy of Sciences (India)

    Nidhi Nigam; Santosh Kumar; Pradip Kumar Dutta; Tamal Ghosh

    2015-10-01

    The optical parameters including the refractive index () and thermo-optic coefficient, TOC (d/d), the dielectric constant () and its variation with temperature, and the thermal volume expansion coefficient () and its variation with temperature of chitosan–alizarin yellow GG (CS–AY GG) complex were examined. The dn/dT and - values for the polymer derivative were in the range −2.5 × 10−4 to 1.2 × 10−4° C−1 and 2.2 to 2.3, respectively. The dn/dT values were larger than that of inorganic glasses such as zinc silicate glass (5.5 × 10−6° C−1) and borosilicate glass (4.1 × 10−6° C−1) and were larger than that of organic polymers such as polystyrene (−1.23 × 10−4 ° C−1) and PMMA (−1.20 × 10−4 ° C−1). The -values are lower than optically estimated -values of conventional polymer (3.00), aliphatic polyimide (2.5) and semi-aromatic polyamide (2.83). The obtained results of chitosan derivative are expected to be useful for optical switching and optical waveguide areas for devices of biomedical applications.

  3. Degradation of a monoazo dye Alizarin Yellow GG in aqueous solutions by gamma irradiation: Decolorization and biodegradability enhancement

    Science.gov (United States)

    Sun, Weihua; Chen, Lujun; Tian, Jinping; Wang, Jianlong; He, Shijun

    2013-02-01

    The irradiation-induced degradation of an azo dye, Alizarin Yellow GG (AY-GG), was investigated in aqueous solution under gamma irradiation using a 60Cobalt source at a dose rate of 113 Gy/min. The decolorization percentage of AY-GG reached 65% when its initial concentration was 100 mg/l and the absorbed dose was 9 kGy. The decolorization process could be described by first-order kinetic equation. In addition, specific oxygen uptake rate (SOUR, mg O2 (g MLVSS)-1 h-1) of activated sludge using the irradiated azo dye solutions was 8.1 mg O2 (g MLVSS)-1 h-1 after 9 kGy irradiation, indicating that the biodegradability of AY-GG could be enhanced by 30%. However, toxic intermediates including heterocyclic aromatic amines and cyanides were detected during the irradiation process, which inhibited the complete biological degradation of azo dye. Fortunately, the inhibition could be eliminated by further irradiation. The azo dye solution became amenable to biodegradation and can be further treated by biological treatment process.

  4. Use of alizarin red S as a chromogenic agent for the colorimetric determination of dothiepin hydrochloride in pharmaceutical formulations

    Directory of Open Access Journals (Sweden)

    Sameer A.M. Abdulrahman

    2014-04-01

    Full Text Available The present study describes two simple, rapid, selective and cost-effective spectrophotometric methods for the determination of dothiepin hydrochloride (DOTH, an antidepressant drug, in bulk drug and pharmaceutical formulations. The first method (method A is based on the formation of yellow colored ion-pair complex between DOTH and alizarin red S (ARS in acid medium which was extracted into dichloromethane and the absorbance was measured at 445 nm. The second method (method B is based on the breaking of the yellow DOTH–ARS ion-pair complex in alkaline medium followed by the measurement of the violet color free dye at 570 nm. Under the optimized conditions, Beer’s law is obeyed over the concentration ranges of 2.50–55.0 and 1.00–35.0 μg ml−1 DOTH for method A and method B, respectively. The molar absorptivity, Sandell’s sensitivity, detection and quantification limits are also calculated. The methods were validated for intra-day and inter-day accuracy and precision; selectivity and robustness and ruggedness. The proposed methods were applied successfully to the determination of DOTH in pure drug and commercial formulations. The accuracy and reliability of the proposed methods were further established by parallel determination by the official method and also by recovery studies via standard addition technique.

  5. Synthesis and application of Amberlite xad-4 functionalized with alizarin red-s for preconcentration and adsorption of rhodium (III

    Directory of Open Access Journals (Sweden)

    Sid Kalal Hossein

    2012-09-01

    Full Text Available Abstract A new chelating resin was prepared by coupling Amberlite XAD-4 with alizarin red-s through an azo spacer, characterized by infra-red spectroscopy and thermal analysis and studied for Rh(III preconcentration using inductively coupled plasma atomic emission spectroscopy (ICP-AES for rhodium monitoring in the environment. The optimum pH for sorption of the metal ion was 6.5. The sorption capacity was found 2.1 mg/g of resin for Rh(III. A recovery of 88% was obtained for the metal ion with 1.5 M HCl as eluting agent. Kinetic adsorption data were analyzed by adsorption and desorption times of Rh(III on modified resin. Scat chard analysis revealed that the homogeneous binding sites were formed in the polymers. The linear regression equation was Q/C = –1.3169Q + 27.222 (R2 = 0.9239, for Rh were formed in the SPE sorbent,Kd and Qmax for the affinity binding sites were calculated to be 0.76 μmol/mL and 20.67 μmol/g, respectively. The equilibrium data and parameters of Rh(III adsorption on modified resin were analyzed by Langmuir, Freundlich, Temkin and Redlich–Peterson models. The experimental adsorption isotherm was in good concordance with Langmuir and Freundlich models (R2 > 0.998 and based on the Langmuir isotherm the maximum amount of adsorption (qmax was 4.842 mg/g. The method was applied for rhodium ions determination in environmental samples. with high recovery (>80%.

  6. Poly(alizarin red)/Graphene modified glassy carbon electrode for simultaneous determination of purine and pyrimidine

    Energy Technology Data Exchange (ETDEWEB)

    Ba Xi; Luo Liqiang [Department of Chemistry, Shanghai University, Shanghai 200444 (China); Ding Yaping, E-mail: wdingyp@sina.com [Department of Chemistry, Shanghai University, Shanghai 200444 (China); Zhang Zhen [Department of Chemistry, Shanghai University, Shanghai 200444 (China); Chu Yuliang [Instrumental Analysis and Research Center, Shanghai University, Shanghai 200444 (China); Wang Bijun; Ouyang Xiaoqian [Department of Chemistry, Shanghai University, Shanghai 200444 (China)

    2012-11-08

    Graphical abstract: DPVs of PAR/Graphene/GCE (a) and the bare GCE (c) in 0.1 M PBS containing 50.0 {mu}M G, 50.0 {mu}M A, 100.0 {mu}M T and 100.0 {mu}M C, (b) PAR/Graphene/GCE in 0.1 M PBS. Highlights: Black-Right-Pointing-Pointer The sensor exhibited well-separated peaks and low detection limit. Black-Right-Pointing-Pointer The sensor possesses high sensitivity and wide linear range. Black-Right-Pointing-Pointer The sensor was used for simultaneous detection of G, A, T and C successfully. Black-Right-Pointing-Pointer The sensor was applied in a fish sperm DNA sample with satisfactory results. Black-Right-Pointing-Pointer The proposed sensor has good stability and reproducibility. - Abstract: In this work, a poly(alizarin red)/Graphene composite film modified glassy carbon electrode (PAR/Graphene/GCE) was prepared for simultaneous determination of four DNA bases (guanine, adenine, thymine and cytosine) without any pretreatment. The morphology and interface property of PAR/Graphene films were examined by scanning electron microscopy and electrochemical impedance spectroscopy. The PAR/Graphene/GCE exhibited excellent electrocatalytic activity toward purine (guanine and adenine) and pyrimidine (thymine and cytosine) in 0.1 M phosphate buffer solution (pH 7.4). Under optimum conditions, differential pulse voltammetry was used to detect the oxidation of purine and pyrimidine. The results showed that PAR/Graphene/GCE exhibited well-separated peaks, low detection limit, high sensitivity and wide linear range for simultaneous detection of purine and pyrimidine. The proposed sensor also has good stability and reproducibility. Furthermore, the modified electrode was applied for the detection of DNA bases in a fish sperm DNA sample with satisfactory results.

  7. Synthesis and Application of Amberlite Xad-4 Functionalized with Alizarin Red-S for Preconcentration and Adsorption of Rhodium (iii

    Directory of Open Access Journals (Sweden)

    Hossein Sid Kalal

    2012-09-01

    Full Text Available A new chelating resin was prepared by coupling Amberlite XAD-4 with alizarin red-s through an azo spacer, characterized by infra-red spectroscopy and thermal analysis and studied for Rh(III preconcentration using inductively coupled plasma atomic emissionspectroscopy (ICP-AES for rhodium monitoring in the environment. The optimum pH for sorption of the metal ion was 6.5. The sorption capacity was found 2.1 mg/g of resin for Rh(III. A recovery of 88% was obtained for the metal ion with 1.5 M HCl as eluting agent.Kinetic adsorption data were analyzed by adsorption and desorption times of Rh(III on modified resin. Scat chard analysis revealed that the homogeneous binding sites were formed in the polymers. The linear regression equation was Q/C = –1.3169Q + 27.222 (R2 = 0.9239,for Rh were formed in the SPE sorbent,Kd and Qmax for the affinity binding sites were calculated to be 0.76 μmol/mL and 20.67 μmol/g, respectively. The equilibrium data and parameters of Rh(III adsorption on modified resin were analyzed by Langmuir, Freundlich,Temkin and Redlich–Peterson models. The experimental adsorption isotherm was in good concordance with Langmuir and Freundlich models (R2 > 0.998 and based on the Langmuir isotherm the maximum amount of adsorption (qmax was 4.842 mg/g. The method was applied for rhodium ions determination in environmental samples. with high recovery (>80%.

  8. Synthesis and application of Amberlite xad-4 functionalized with alizarin red-s for preconcentration and adsorption of rhodium (III).

    Science.gov (United States)

    Sid Kalal, Hossein; Panahi, Homayon Ahmad; Hoveidi, Hassan; Taghiof, Mohammad; Menderjani, Mahnaz Taheri

    2012-09-18

    A new chelating resin was prepared by coupling Amberlite XAD-4 with alizarin red-s through an azo spacer, characterized by infra-red spectroscopy and thermal analysis and studied for Rh(III) preconcentration using inductively coupled plasma atomic emission spectroscopy (ICP-AES) for rhodium monitoring in the environment. The optimum pH for sorption of the metal ion was 6.5. The sorption capacity was found 2.1 mg/g of resin for Rh(III). A recovery of 88% was obtained for the metal ion with 1.5 M HCl as eluting agent. Kinetic adsorption data were analyzed by adsorption and desorption times of Rh(III) on modified resin. Scat chard analysis revealed that the homogeneous binding sites were formed in the polymers. The linear regression equation was Q/C = -1.3169Q + 27.222 (R2 = 0.9239), for Rh were formed in the SPE sorbent,Kd and Qmax for the affinity binding sites were calculated to be 0.76 μmol/mL and 20.67 μmol/g, respectively. The equilibrium data and parameters of Rh(III) adsorption on modified resin were analyzed by Langmuir, Freundlich, Temkin and Redlich-Peterson models. The experimental adsorption isotherm was in good concordance with Langmuir and Freundlich models (R2 > 0.998) and based on the Langmuir isotherm the maximum amount of adsorption (qmax) was 4.842 mg/g. The method was applied for rhodium ions determination in environmental samples. with high recovery (>80%).

  9. Ag/TiO2光催化降解茜素红%Photocatalytic degradation of alizarin red on Ag/TiO2 catalyst

    Institute of Scientific and Technical Information of China (English)

    付文; 王丽; 黄军左

    2012-01-01

    以钛酸四丁酯为前驱体,冰醋酸为水解抑制剂,用溶胶-凝胶法制备了纳米Ag/TiO2复合催化剂,测定了催化剂对茜素红溶液的光降解活性.结果表明,Ag/TiO2催化剂具有较高的光催化活性.%Using tetrabutyl titanate as the precursor and glacial acetic acid as the hydrolysis inhibitor, Ag/TiO2 catalyst was prepared by means of sol-gel method. The photo-degradation activity of the as-prepared catalyst for alizarin red was investigated. The results showed that Ag/TiO2 catalyst exhibited better catalytic properties.

  10. Kinetic Model for Simultaneous Adsorption/Photodegradation Process of Alizarin Red S in Water Solution by Nano-TiO2 under Visible Light

    Directory of Open Access Journals (Sweden)

    Rita Giovannetti

    2016-06-01

    Full Text Available The simultaneous adsorption and visible light photodegradation of Alizarin Red S in water solutions were studied in real time mode by using nano-TiO2, such as Anatase and Aeroxide P-25, supported on polypropylene strips. Kinetic results of the overall process were compared with those obtained from separated steps of adsorption and photodegradation previously studied; kinetic advantages were evidenced with the simultaneous approach. From the study of different dye concentrations, a kinetic model has been proposed which describes the overall process. This model considered two consecutive processes: The adsorption of dye on TiO2 surface and its photodegradation. The obtained results were in good agreement with experimental data and can predict the profiles of free dye, dye adsorbed on TiO2 and photoproduct concentrations during the total process.

  11. Eco-friendly and green synthesis of BiVO4 nanoparticle using microwave irradiation as photocatalayst for the degradation of Alizarin Red S

    Science.gov (United States)

    Abraham, S. Daniel; David, S. Theodore; Bennie, R. Biju; Joel, C.; Kumar, D. Sanjay

    2016-06-01

    Bismuth vanadate (BiVO4) nanocrystals have been successfully synthesised using microwave-assisted combustion synthesis (MCS), and characterised using Fourier transform infrared (FT-IR) and Raman spectra, surface area analysis (BET), X-ray diffraction (XRD), scanning electron microscopy (SEM), Energy Dispersive X-ray analysis (EDX), diffused reflectance spectroscopy (DRS) and Photoluminescence (PL) spectroscopy. The XRD results confirmed the formation of monoclinic bismuth vanadate. The formations of BiO & VO43-vibrations were ascertained from FT-IR data. The morphology of hallow internal structural micro entities were confirmed by SEM. The optical properties were determined by DRS and PL spectra. Hence, the influence of the preparation methods on the structure, morphology and optical activities of bismuth vanadate was investigated systematically. Photocatalytic degradation (PCD) of Alizarin Red S (ARS), an effective disrupting chemical in aqueous medium was investigated using BiVO4 nanoparticles. The kinetics of PCD was found to follow pseudo first-order.

  12. A Novel Nanofilm Sensor Based on Poly-(Alizarin Red)/Fe3O4 Magnetic Nanoparticles-Multiwalled Carbon Nanotubes Composite Material for Determination of Nitrite.

    Science.gov (United States)

    Qu, Jianying; Dong, Ying; Yong, Wang; Lou, Tongfang; Du, Xueping; Qu, Jianhang

    2016-03-01

    Fe3O4 magnetic nanoparticles were synthesized by chemical co-precipitation with sodium citrate as surfactant and were characterized by FT-IR spectrometer, X-ray diffraction and transmission electron microscopy. A novel nitrite sensor was fabricated by electropolymerization of alizarin red on the surface of glassy carbon electrode modified with Fe3O4-multiwalled carbon nanotubes composite nanofilm. Under the optimal experimental conditions, it was showed that the proposed sensor exhibited good electrocatalytic activity to the oxidation of nitrite, and the peak current increased linearly with the nitrite concentration from 9.64 x 10(-6) mol x L(-1) to 1.30 x 10(-3) mol x L(-1) (R = 0.9976) with a detection limit of 1.19 x 10(-6) mol x L(-1) (S/N = 3). This sensor showed excellent sensitivity, wide linear range, stability and repeatability for nitrite determination with potential applications.

  13. Electrocatalytic oxidation and voltammetric determination of ciprofloxacin employing poly(alizarin red)/graphene composite film in the presence of ascorbic acid, uric acid and dopamine.

    Science.gov (United States)

    Zhang, Xin; Wei, Youli; Ding, Yaping

    2014-07-04

    A glassy carbon electrode modified with poly(alizarin red)/electrodeposited graphene (PAR/EGR) composite film was prepared and applied to detect ciprofloxacin (CPFX) in the presence of ascorbic, uric acid and dopamine. The morphology and interface property of PAR/EGR films were examined by scanning electron microscopy (SEM) and electrochemical impedance spectroscopy (EIS). The electrocatalytic oxidation of CPFX on AR/EGR was investigated by cyclic voltammetry (CV) and differential pulse voltammetry (DPV). The linearity ranged from 4 × 10(-8) to 1.2 × 10(-4) M with a detection limit (S/N=3) of 0.01 μM. The modified electrode could be applied to the individual determination of CPFX as well as the simultaneous determination of CPFX, ascorbic acid, uric acid and dopamine. This method proved to be a simple, selective and rapid way to determine CPFX in pharmaceutical preparation and biological media.

  14. Estudo voltamétrico do complexo de cobre(II com o ligante vermelho de alizarina S, adsorvido na superfície do eletrodo de grafite pirolítico Voltammetric study of complex of copper (II with alizarin red S ligand, absorbed on surface of pyrolytic graphite electrode

    Directory of Open Access Journals (Sweden)

    Victor E. Mouchrek Filho

    1999-06-01

    Full Text Available The alizarin red S (ARS has been used as a spectrophotometric reagent of several metals for a long time. Now this alizarin has been used as modifier agent of electrodes, for voltammetric analyses. In this work cyclic voltammetry experiments was accomplished on closed circuit, with the objective of studying the voltammetric behavior of alizarin red S adsorbed and of its copper complex, on the surface of the pyrolytic graphite electrode. These studies showed that ARS strongly adsorbs on the surface of this electrode. This adsorption was used to immobilize ions copper(II from the solution.

  15. Application of excitation and emission matrix fluorescence (EEM) and UV-vis absorption to monitor the characteristics of Alizarin Red S (ARS) during electro-Fenton degradation process.

    Science.gov (United States)

    Lai, Bo; Zhou, Yuexi; Wang, Juling; Yang, Zhishan; Chen, Zhiqiang

    2013-11-01

    Oxidative degradation of Alizarin Red S (ARS) in aqueous solutions by using electro-Fenton was studied. At first, effect of operating parameters such as current density, aeration rate and initial pH on the degradation of ARS were studied by using UV-vis spectrum, respectively. Then, under the optimal operating conditions (current density: 10.0mAcm(-2), aeration rate: 1000mLmin(-1), initial pH: 2.8), the identification of degradation products of ARS was carried out by using GC-MS and HPLC, meanwhile its degradation pathway was proposed according to the intermediates. Considering the location, intensity and intensity ratio of fluorescence center peak of the ARS in aqueous solution, a convenient and quick monitoring method by using excitation-emission matrix fluorescence spectrum technology was developed to monitor the degradation degree of ARS through electro-Fenton process. Furthermore, it is suggested that the developed method would be promising for the quick analysis and evaluation of the degradation degree of the pollutants with π-conjugated system.

  16. On-line preconcentration system using a microcolumn packed with Alizarin Red S-modified alumina for zinc determination by flame atomic absorption spectrometry

    Directory of Open Access Journals (Sweden)

    A.M. Haji Shabani

    2009-01-01

    Full Text Available A simple and sensitive on-line flow injection system for determination of zinc with FAAS has been described. The method is based on the separation and preconcentration of zinc on a microcolumn of immobilized Alizarin Red S on alumina. The adsorbed analyte is then eluted with 250 µL of nitric acid (1 mol L-1 and is transported to flame atomic absorption spectrometer for quantification. The effect of pH, sample and eluent flow rates and presence of various cations and anions on the retention of zinc was investigated. The sorption of zinc was quantitative in the pH range of 5.5-8.5. For a sample volume of 25 mL an enrichment factor of 144 and a detection limit (3S of 0.2 µg L-1 was obtained. The precision (RSD, n=7 was 3.0% at the 20 µg L-1 level. The developed system was successfully applied to the determination of zinc in water samples, hair, urine and saliva.

  17. Rapid bioremediation of Alizarin Red S and Quinizarine Green SS dyes using Trichoderma lixii F21 mediated by biosorption and enzymatic processes.

    Science.gov (United States)

    Adnan, Liyana Amalina; Sathishkumar, Palanivel; Yusoff, Abdull Rahim Mohd; Hadibarata, Tony; Ameen, Fuad

    2017-01-01

    In this study, a newly isolated ascomycete fungus Trichoderma lixii F21 was explored to bioremediate the polar [Alizarin Red S (ARS)] and non-polar [Quinizarine Green SS (QGSS)] anthraquinone dyes. The bioremediation of ARS and QGSS by T. lixii F21 was found to be 77.78 and 98.31 %, respectively, via biosorption and enzymatic processes within 7 days of incubation. The maximum biosorption (ARS = 33.7 % and QGSS = 74.7 %) and enzymatic biodegradation (ARS = 44.1 % and QGSS = 23.6 %) were observed at pH 4 and 27 °C in the presence of glucose and yeast extract. The laccase and catechol 1,2-dioxygenase produced by T. lixii F21 were involved in the molecular conversions of ARS and QGSS to phenolic and carboxylic acid compounds, without the formation of toxic aromatic amines. This study suggests that T. lixii F21 may be a good candidate for the bioremediation of industrial effluents contaminated with anthraquinone dyes.

  18. Electrocatalytic oxidation and voltammetric determination of ciprofloxacin employing poly(alizarin red)/graphene composite film in the presence of ascorbic acid, uric acid and dopamine

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Xin; Wei, Youli; Ding, Yaping, E-mail: wdingyp@sina.com

    2014-07-04

    Graphical abstract: An electrochemical sensor based on PAR/EGR/GCE via a cooperation of the potentiostatic technique and cyclic voltammetry was first fabricated for the determination of CPFX with satisfied detecting result of real samples. - Highlights: • PAR/EGR composite film was prepared for the first time. • The sensor can be applied to determinate CPFX in the presence of AA, UA and DA. • The sensor indicated the feasibility in drug samples and biological media. - Abstract: A glassy carbon electrode modified with poly(alizarin red)/electrodeposited graphene (PAR/EGR) composite film was prepared and applied to detect ciprofloxacin (CPFX) in the presence of ascorbic, uric acid and dopamine. The morphology and interface property of PAR/EGR films were examined by scanning electron microscopy (SEM) and electrochemical impedance spectroscopy (EIS). The electrocatalytic oxidation of CPFX on AR/EGR was investigated by cyclic voltammetry (CV) and differential pulse voltammetry (DPV). The linearity ranged from 4 × 10{sup −8} to 1.2 × 10{sup −4} M with a detection limit (S/N = 3) of 0.01 μM. The modified electrode could be applied to the individual determination of CPFX as well as the simultaneous determination of CPFX, ascorbic acid, uric acid and dopamine. This method proved to be a simple, selective and rapid way to determine CPFX in pharmaceutical preparation and biological media.

  19. Mathematical analysis of mandibular morphogenesis by micro-CT-based mouse and alizarin red S-stained-based human studies during development.

    Science.gov (United States)

    Rafiq, Ashiq Mahmood; Udagawa, Jun; Lundh, Torbjörn; Jahan, Esrat; Matsumoto, Akihiro; Sekine, Joji; Otani, Hiroki

    2012-02-01

    Prenatal development of the mandible is an important factor in its postnatal function. To examine quantitatively normal and abnormal developmental changes of the mandible, we here evaluated morphological changes in mineralizing mandibles by thin-plate spline (TPS) including bending energy (BE) and Procrustes distance (PD), and by Procrustes analyses including warp analysis, regression analysis, and discriminant function analysis. BE and PD were calculated from lateral views of the mandibles of mice or of human fetuses using scanned micro-computed tomography (CT) images or alizarin red S-stained specimens, respectively. BE and PD were compared (1) between different developmental stages, and further, to detect abnormalities in the data sets and to evaluate the deviation from normal development in mouse fetuses, (2) at embryonic day (E) 18.5 between the normal and deformed mandibles, the latter being caused by suturing the jaw at E15.5, (3) at E15.5 and E18.5 between normal and knockout mutant mice of receptor tyrosine kinase-like orphan receptor (Ror) 2. In mice, BE and PD were large during the prenatal period and small after postnatal day 3, suggesting that the mandibular shape changes rapidly during the prenatal and early postnatal periods. In humans, BE of the mandibles peaked at 16-19 weeks of gestation, suggesting the time-dependent change in the mandibular shape. TPS and Procrustes analyses statistically separated the abnormal mandibles of the sutured or Ror2 mutant mouse fetuses from the normal mandible. These results suggest that TPS and Procrustes analyses are useful for assessing the morphogenesis and deformity of the mandible.

  20. Alizarin Complexone Functionalized Mesoporous Silica Nanoparticles: A Smart System Integrating Glucose-Responsive Double-Drugs Release and Real-Time Monitoring Capabilities.

    Science.gov (United States)

    Zou, Zhen; He, Dinggeng; Cai, Linli; He, Xiaoxiao; Wang, Kemin; Yang, Xue; Li, Liling; Li, Siqi; Su, Xiaoya

    2016-04-06

    The outstanding progress of nanoparticles-based delivery systems capable of releasing hypoglycemic drugs in response to glucose has dramatically changed the outlook of diabetes management. However, the developed glucose-responsive systems have not offered real-time monitoring capabilities for accurate quantifying hypoglycemic drugs released. In this study, we present a multifunctional delivery system that integrates both delivery and monitoring issues using glucose-triggered competitive binding scheme on alizarin complexone (ALC) functionalized mesoporous silica nanoparticles (MSN). In this system, ALC is modified on the surface of MSN as the signal reporter. Gluconated insulin (G-Ins) is then introduced onto MSN-ALC via benzene-1,4-diboronic acid (BA) mediated esterification reaction, where G-Ins not only blocks drugs inside the mesopores but also works as a hypoglycemic drug. In the absence of glucose, the sandwich-type boronate ester structure formed by BA binding to the diols of ALC and G-Ins remains intact, resulting in an fluorescence emission peak at 570 nm and blockage of pores. Following a competitive binding, the presence of glucose cause the dissociation of boronate ester between ALC and BA, which lead to the pores opening and disappearance of fluorescence. As proof of concept, rosiglitazone maleate (RSM), an insulin-sensitizing agent, was doped into the MSN to form a multifunctional MSN (RSM@MSN-ALC-BA-Ins), integrating with double-drugs loading, glucose-responsive performance, and real-time monitoring capability. It has been demonstrated that the glucose-responsive release behaviors of insulin and RSM in buffer or in human serum can be quantified in real-time through evaluating the changes of fluorescence signal. We believe that this developed multifunctional system can shed light on the invention of a new generation of smart nanoformulations for optical diagnosis, individualized treatment, and noninvasive monitoring of diabetes management.

  1. 茜素红S标记分光光度法测定微量人血清白蛋白%Spectrophotometric Determination of Human Serum Albumin Using Alizarin Red

    Institute of Scientific and Technical Information of China (English)

    赵丹华

    2011-01-01

    研究有机染料茜素红S(alizarin red S,ARS)与牛血清白蛋白(bovine serum albumin,BSA)、人血清白蛋白(human serum albumin,HSA)的结合反应,选择实验的最佳条件:PH=5.10的B—R缓冲溶液3.00mL,4.00mL5.0×10-4mol/L茜素红S溶液.反应20min后,体系的吸光度很稳定,在入λ=420nm处有最大吸收峰,并且随着牛血清白蛋白(BSA),人血清白蛋白(HSA)的加入,茜素红S的吸收峰下降.因此以茜素红S为标记物,根据其在波长420nm处吸收峰下降的程度,可用于定量测定BSA和HSA.测量BSA的线性响应范围为0~32.0μg/mL,相关系数为0.9987,测量BSA的线性响应范围为0~28.0μg/mL,相关系数为0.9968.该方法具有较高的灵敏度和选择性,用于实际试样分析,实验结果令人满意.%The protein is a kind of important biological macromolecules in the human body. It is not merely very important and essential for maintaining the life. A acidic dye has been applied to the assay of proteins in the fields of medicine and life science. The reactions of human serum albumen (HSA) and bovine serum albumen (BSA) with alizarin red have been studied. Both BSA and HSA can form orange complexes, which decrease the absorbance. The optimum conditions of complex reactions of a new chromogenic reagent alizarin red with BSA and HSA was studied. The method was used in the photometric determination of BSA and HSA. In a buffer medium of pH 5.10, alizarin red reacts with BSA and HSA to form a complex having its absorption maxima at 420nm and the reactions steady with 80 rain. Beefs law is obeyed in the concentration in the range of 0-32.0 μg/mL(r=0. 998 7) for BSA and 0-28.0μg/mL(r= 0. 996 8) for HSA. The BSA and HSA in human serum samples were determined, and the result is satisfactory.

  2. Uso do violeta de alizarina N (AVN como reagente espectrofotométrico na determinação de alumínio Use of the alizarine violet N (AVN as a spectrophotometric reagent for aluminium determination

    Directory of Open Access Journals (Sweden)

    Alailson Falcão Dantas

    2000-04-01

    Full Text Available The present work proposes the application of the 4-Hidroxy-3-(2-hydroxynaphtylazo-benzenesulphonic acid (C.I. 15670, Alizarine Violet N (AVN, as a reagent for direct aluminium determination using molecular absorption spectrophotometry in the presence of tensoatives. Al(III cation reacts with AVN in pH 9.4, forming a red complex, stable for at least 24 hours, with absorption minimum at 607nm and, against a reagent blank, (epsiloncomplex - epsilonreagent = -2.71x10(4 L.mol-1.cm-1. The reaction occurs in the presence of a Triton-X100 and CTAB tensoatives mixture, in the presence of EDTA. Al(III determination is possible in the linear range of 50 up to 400ng.mL-1, with a detection limit of 41 ng.mL-1.

  3. Degradation of alizarin red solution by TiO2 supported on CNTs%CNTs负载TiO2对茜素红溶液的降解研究

    Institute of Scientific and Technical Information of China (English)

    付文; 王丽

    2014-01-01

    Using tetrabutyl titanate as the precursor and acetic acid as the depressor,TiO2/CNTs composite catalyst was prepared by sol-gel method. The photo-degradation activity of TiO2/CNTs composite catalyst for photocatalytic degradation of alizarin red solution with different concentrations was investigated,and its photocatalytic degradation kinetics was also studied. The results showed that the optimal preparation condition of composite catalyst was as follows:CNTs loading mass fraction 7 . 5%,calcination temperature 450 ℃and calcination time 3. 5 h,and the photocatalytic activity of TiO2/CNTs composite catalyst was improved. Under the same condition,the final degradation rate of alizarin red solution with the concentration of 150 mg·L-1 over TiO2/CNTs catalyst reached 67%,while the final degradation rate of alizarin red solu-tion over TiO2 only was 56%. The reason for photocatalytic activity improvement of composite catalyst was that it could generate more active hydroxyl under UV irradiation when TiO2 was loaded on CNTs. The photocatalytic degradation kinetics of the composite catalyst complied Langmuir-Hinshelwood equation. The catalyst surface adsorption to pollutants was the bottleneck of reaction,and the reaction order of kinetics decreased from one order to zero level with the increase of pollutant concentrations.%以钛酸四丁酯为前驱体,冰醋酸为水解抑制剂,采用溶胶-凝胶法制备纳米TiO2/CNTs复合催化剂。测定复合催化剂对不同浓度茜素红溶液的光降解活性,并对光催化降解动力学进行研究。结果表明,在CNTs负载质量分数7.5%、焙烧温度450℃和焙烧时间3.5 h 条件下制备的TiO2/CNTs复合催化剂光催化活性得到提升。相同条件下,TiO2/CNTs对150 mg·L-1茜素红溶液的最终降解率达67%,而TiO2的最终降解率只有56%。复合催化剂光催化活性提升的原因是CNTs负载后光催化粒子在紫外光照射下生成更多的氢氧根活性自由基

  4. Poly-Alizarin red S/multiwalled carbon nanotube modified glassy carbon electrode for the boost up of electrocatalytic activity towards the investigation of dopamine and simultaneous resolution in the presence of 5-HT: A voltammetric study.

    Science.gov (United States)

    Reddaiah, K; Madhusudana Reddy, T; Venkata Ramana, D K; Subba Rao, Y

    2016-05-01

    Poly-Alizarin red S/multiwalled carbon nanotube film on the surface of glassy carbon electrode (poly-AzrS/MWCNT/GCE) was synthesized by electrochemical process and was used for the sensitive and selective determination of dopamine (DA) by employing voltammetric techniques. The electrocatalytic response of the modified electrode was found to exhibit admirable activity. The simultaneous determination of dopamine in the presence of serotonin (5-HT) was found to exhibit very good response at poly-AzrS/MWCNTs/GCE. The effect of pH, scan rate, accumulation time and concentration of dopamine was studied at the developed poly-AzrS/MWCNTs/GCE. The poly-AzrS/MWCNTs/GCE exhibited an efficient electron mediating behavior together with well resolved peaks for dopamine, in 0.1 mol/dm(3) phosphate buffer (PBS) solution of pH 7.0. The limit of detection (LOD) and limit of quantification (LOQ) were found to be as 1.89 × 10(-7) mol/dm(3) and 6.312 × 10(-7) mol/dm(3) respectively with a dynamic range from 1 × 10(-6) to 1.8 × 10(-5) mol/dm(3). The interfacial electron transfer behavior of DA was studied by electrochemical impedance spectroscopy (EIS); the studies showed that the charge transfer rate was enhanced at poly-AzrS/MWCNTs/GCE when compared with bare GCE and poly-AzrS/GCE.

  5. Poly-Alizarin red S/multiwalled carbon nanotube modified glassy carbon electrode for the boost up of electrocatalytic activity towards the investigation of dopamine and simultaneous resolution in the presence of 5-HT: A voltammetric study

    Energy Technology Data Exchange (ETDEWEB)

    Reddaiah, K. [Electrochemical Research Laboratory, Department of Chemistry, S.V.U. College of Sciences, Sri Venkateswara University, Tirupati 517 502, Andhra Pradesh (India); Madhusudana Reddy, T., E-mail: tmsreddysvu@gmail.com [Electrochemical Research Laboratory, Department of Chemistry, S.V.U. College of Sciences, Sri Venkateswara University, Tirupati 517 502, Andhra Pradesh (India); Department of Chemistry, University of Minnesota, 207 Pleasant Street SE, Minneapolis, MN 55455 (United States); Venkata Ramana, D.K. [Department of Safety Engineering, Dongguk University, 123 Dongdae-ro, Gyeongju, Gyeongbuk 780 714 (Korea, Republic of); Subba Rao, Y. [DST-PURSE Centre, Sri Venkateswara University, Tirupati 517502, Andhra Pradesh (India)

    2016-05-01

    Poly-Alizarin red S/multiwalled carbon nanotube film on the surface of glassy carbon electrode (poly-AzrS/MWCNT/GCE) was synthesized by electrochemical process and was used for the sensitive and selective determination of dopamine (DA) by employing voltammetric techniques. The electrocatalytic response of the modified electrode was found to exhibit admirable activity. The simultaneous determination of dopamine in the presence of serotonin (5-HT) was found to exhibit very good response at poly-AzrS/MWCNTs/GCE. The effect of pH, scan rate, accumulation time and concentration of dopamine was studied at the developed poly-AzrS/MWCNTs/GCE. The poly-AzrS/MWCNTs/GCE exhibited an efficient electron mediating behavior together with well resolved peaks for dopamine, in 0.1 mol/dm{sup 3} phosphate buffer (PBS) solution of pH 7.0. The limit of detection (LOD) and limit of quantification (LOQ) were found to be as 1.89 × 10{sup −7} mol/dm{sup 3} and 6.312 × 10{sup −7} mol/dm{sup 3} respectively with a dynamic range from 1 × 10{sup −6} to 1.8 × 10{sup −5} mol/dm{sup 3}. The interfacial electron transfer behavior of DA was studied by electrochemical impedance spectroscopy (EIS); the studies showed that the charge transfer rate was enhanced at poly-AzrS/MWCNTs/GCE when compared with bare GCE and poly-AzrS/GCE. - Highlights: • The poly-AzrS/MWCNTs/GCE showed good sensitivity towards DA sensing. • The sensor reduced the overoxidation potentials for DA. • This electrode was successfully used for simultaneous sensing of DA and 5-HT. • The electrode was effectively used for the determination of DA in pharmaceutical formulations.

  6. 茜素络合物浸泡标记秦岭细鳞鲑发眼卵及仔鱼耳石%Alizarin marking of otolith at eyed eggs and larvae stages in Brachymystax lenok tsinlingensis

    Institute of Scientific and Technical Information of China (English)

    靳建波; 危起伟; 孙庆亮; 李罗新; 甘芳

    2011-01-01

    采用茜素络合物(ALC)在秦岭细鳞鲑(Brachymystax lenok tsinlingensis)发眼卵和仔鱼阶段进行浸泡标记试验,为确定合适浸泡浓度和持续浸泡时间,试验设置了6个的浓度组和4个时间梯度组.结果显示:在荧光显微镜下观察被浸泡的仔鱼和经浸泡处理的发眼卵而出膜后的仔鱼,其耳石都出现橘红色荧光标记.浸泡发眼卵组最适处理条件是茜素络合物溶液浓度30 mg/L持续浸泡18 h,标记率100%,孵化率91%,出膜后15 d仔鱼存活率91%,标记环"非常明显";仔鱼浸泡组最适处理条件是茜素络合物溶液浓度120 mg/L持续浸泡12 h,标记率83.3%,处理15 d后仔鱼存活率92.3%,相比对照组统计检验无差异(P<0.05),标记环"较明显".相比上述两种方法,浸泡发眼卵试验组相比浸泡30日龄仔鱼试验组所需要的茜素络合物(ALC)溶液浓度更低,而且在标记成功率、标记效果、标记后仔鱼存活率以及标记成本等方面也更为理想.因此,用茜素络合物(ALC)在秦岭细鳞鲑发眼卵阶段进行荧光标记是可行的,这种方法也可在其它鱼类标记中尝试,同时将人工繁育鱼苗与野生鱼苗加以区别,有利于开展人工增殖放流效果及早期生活史等研究.%To study the effects of ALC for marking the eggs and larvae, Brachymystax lenok tsinlingensis eyed eggs and larvae were immersed with twenty-four combinations of six different ALC concentration and four treatment time. The otoliths of larvae were detected by fluorescence microscope. The results showed that the larvae immersed in alizarin complexone solution or which from treated eggs presented distinct scarlet-pink fluorescent marks on otolith under UV light. 30 mg/L for 18 h was found to be the most appropriate for eyed eggs with 100% of marking rate and 91% of hatching and larvae survival rate. The marking effect was “bright mark”. 120 mg/L for 12 h was the most appropriate for 30 days old larvae with 83.3

  7. Two Validated HPLC Methods for the Quantification of Alizarin and other Anthraquinones in Rubia tinctorum Cultivars

    NARCIS (Netherlands)

    Derksen, G.C.H.; Lelyveld, G.P.; Beek, van T.A.; Capelle, A.; Groot, de Æ.

    2004-01-01

    Direct and indirect HPLC-UV methods for the quantitative determination of anthraquinones in dried madder root have been developed, validated and compared. In the direct method, madder root was extracted twice with refluxing ethanol-water. This method allowed the determination of the two major native

  8. Cathodic bacterial community structure applying the different co-substrates for reductive decolorization of Alizarin Yellow R.

    Science.gov (United States)

    Sun, Qian; Li, Zhi-Ling; Wang, You-Zhao; Yang, Chun-Xue; Chung, Jong Shik; Wang, Ai-Jie

    2016-05-01

    Selective enrichment of cathodic bacterial community was investigated during reductive decolorization of AYR fedding with glucose or acetate as co-substrates in biocathode. A clear distinction of phylotype structures were observed between glucose-fed and acetate-fed biocathodes. In glucose-fed biocathode, Citrobacter (29.2%), Enterococcus (14.7%) and Alkaliflexus (9.2%) were predominant, and while, in acetate-fed biocathode, Acinetobacter (17.8%) and Achromobacter (6.4%) were dominant. Some electroactive or reductive decolorization genera, like Pseudomonas, Delftia and Dechloromonas were commonly enriched. Both of the higher AYR decolorization rate (k(AYR)=0.46) and p-phenylenediamine (PPD) generation rate (k(PPD)=0.38) were obtained fed with glucose than acetate (k(AYR)=0.18; k(PPD)=0.16). The electrochemical behavior analysis represented a total resistance in glucose-fed condition was about 73.2% lower than acetate-fed condition. The different co-substrate types, resulted in alteration of structure, richness and composition of bacterial communities, which significantly impacted the performances and electrochemical behaviors during reductive decolorization of azo dyes in biocathode.

  9. Studies of midecamycin and erythromycin with alizarin by spectrophotometry%麦迪霉素、红霉素与茜素的光度分析研究

    Institute of Scientific and Technical Information of China (English)

    江虹; 刘艳; 湛海粼

    2007-01-01

    研究了茜素与麦迪霉素(MID)、红霉素(ERY)的显色反应.在pH 5.5~6.8的BR缓冲溶液中,茜素与麦迪霉素(MID)、红霉素(ERY)反应生成红色络合物.最大吸收波长分别为528 nm (MID)和534 nm (ERY);表观摩尔吸光系数为8.3×103 L·mol-1·cm-1 (MID)和1.01×104 L·mol-1·cm-1(ERY);麦迪霉素质量浓度在0~16.5 mg/L和红霉素质量浓度在0~18.0 mg/L范围内均符合比耳定律,据此建立了测定麦迪霉素和红霉素的分光光度法,并探讨了适宜的反应条件.该法可用于麦迪霉素、红霉素药物的测定.

  10. Study on Reaction of Alizarin Red S-Copper(Ⅱ) Metal Complexes with Lysozyme%茜素红S-铜(Ⅱ)金属配合物与溶菌酶作用研究

    Institute of Scientific and Technical Information of China (English)

    王兴明

    2005-01-01

    采用UV-Vis光谱法研究了pH=4.25的缓冲溶液中茜素红S(ARS)-铜(Ⅱ)金属配合物与溶菌酶(LYS)的结合反应.提出了双波长物质的量比法和平衡透析物质的量比法,与单波长物质的量比法进行对照测定研究,实验结果基本相符.研究发现,ARSCu(Ⅱ)-LYS的最大吸收波长为526 nm,比ARS红移96 nm,比ARS-Cu(Ⅱ)配合物红移10 nm.在526 nm处,测得ARS-Cu(Ⅱ)-LYS三元配合物的结合比为nARS:nCu(Ⅱ):nLYS=6:3:1,摩尔吸光系数ε7.93×104L·mol-1·cm-1,ARS-Cu(Ⅱ)配合物与LYS作用的条件平衡常数K=3.21×1012.ARS-Cu(Ⅱ)与LYS之间的作用力为配位键和电荷力.

  11. Activity and mechanism of photocatalytic degradation of Alizarin Green by cerium ions%铈离子光催化降解茜素绿的性能和机理

    Institute of Scientific and Technical Information of China (English)

    钟恒; 曹文彪; 熊重铎; 夏东升; 曾庆福; 徐爱华

    2014-01-01

    主要研究了简单铈离子(Ce3+)在紫外光(UV)的作用下对蒽醌染料茜素绿(AG)的光催化降解效果和反应机理.结果表明,UV/Ce3+体系能够有效降解AG,初始反应速率随AG浓度的倒数值和Ce3+浓度的增加而线性增加,随初始溶液pH的增加先降低后增加,在酸性条件下有很高的TOC去除率.荧光探针实验表明,反应过程中可以产生·OH自由基.UV/Ce3+体系对其他类型染料和对硝基苯酚都有较好的降解效果.

  12. Determination of Aluminium in Water Samples by Alizarin Red Spectrophotometry after Solid Phase Extraction on XAD-2-Catechol Resin%邻苯二酚螯合树脂的合成及水样中铝的测定

    Institute of Scientific and Technical Information of China (English)

    于涛; 耿伟; 杨枝; 曹维鹏

    2006-01-01

    合成并表征了新螯合树脂--邻苯二酚螯合树脂(XAD-2-Catechol),研究了XAD-2-Catechol吸附铝的特性和茜素红-铝的显色反应,在pH4. 5的HAc-NaAc缓冲介质中,茜素红和铝(Ⅲ)反应生成红色络合物,λmax=500 nm,铝的含量在0-50 μg/25 mL内符合比耳定律.建立了邻苯二酚螯合树脂分离/富集-茜素红分光光度法测定天然水样中铝的新方法,对水样中铝形态进行测定,结果满意.

  13. Red, redder, madder. Analysis and isolation of anthraquinones from madder roots (Rubia tinctorum)

    NARCIS (Netherlands)

    Derksen, G.C.H.

    2001-01-01

    The roots of Rubia tinctorum L. (madder) are the source of a natural dye. The dye components are anthraquinones with alizarin being the main dye component. Alizarin as such is present in madder root in only small quantities, most of the alizarin is present as its glycoside ruberythric acid. The suga

  14. Mass marking of Leuciscus idus larvae using Artemia salina as a vector of fluorescent dyes.

    Science.gov (United States)

    Stańczak, K; Krejszeff, S; Dębowska, M; Mierzejewska, K; Woźniak, M; Hliwa, P

    2015-09-01

    A method for the mass marking of ide Leuciscus idus larvae by feeding them Artemia salina nauplii that were immersed in different solutions of alizarin red S, tetracycline hydrochloride and calcein was tested. The best quality marks were obtained after feeding fish for 4 days with nauplii that had been immersed in 200 mg l(-1) alizarin red S.

  15. Triple bone labeling of canine mandibles

    DEFF Research Database (Denmark)

    Pinholt, E M; Kwon, P H

    1990-01-01

    Fluorescence microscopy was used for evaluation of new bone formation in 16 canine mandibles augmented with hydroxylapatite (HA) granules. Three fluorochromes were injected at different time intervals during therapeutic radiation treatment. Oxytetracycline, DCAF, and alizarin-complexone were give...

  16. Facile colorimetric methods for the quantitative determination of tetramisole hydrochloride

    Science.gov (United States)

    Amin, A. S.; Dessouki, H. A.

    2002-10-01

    A facile, rapid and sensitive methods for the determination of tetramisole hydrochloride in pure and in dosage forms are described. The procedures are based on the formation of coloured products with the chromogenic reagents alizarin blue BB (I), alizarin red S (II), alizarin violet 3R (III) and alizarin yellow G (IV). The coloured products showed absorption maxima at 605, 468, 631 and 388 nm for I-IV, respectively. The colours obtained were stable for 24 h. The colour system obeyed Beer's law in the concentration range 1.0-36, 0.8-32, 1.2-42 and 0.8-30 μg ml -1, respectively. The results obtained showed good recoveries with relative standard deviations of 1.27, 0.96, 1.13 and 1.35%, respectively. The detection and determination limits were found to be 1.0 and 3.8, 1.2 and 4.2, 1.0 and 3.9 and finally 1.4 and 4.8 ng ml -1 for I-IV complexes, respectively. Applications of the method to representative pharmaceutical formulations are represented and the validity assessed by applying the standard addition technique, which is comparable with that obtained using the official method.

  17. Environ: E00286 [KEGG MEDICUS

    Lifescience Database Archive (English)

    Full Text Available E00286 Madder root Rubia root Crude drug Alizarin, Rubierythric acid, Purpurin [CPD...:C10395], Pseudopurpurin [CPD:C10394], Xanthopurpurin, Munjistin Rubia cordifolia [TAX:339321] Rubiaceae (madder family) Rub...ia cordifolia root Crude drugs [BR:br08305] Dicot plants: asterids Rubiaceae (madder family) E00286 Madder root ...

  18. The significance of a differential distribution of phosphomonoesterases on bone surfaces after prolonged demineralization

    DEFF Research Database (Denmark)

    Vilmann, H; Kirkeby, S

    1979-01-01

    with the patterns of apposition and resorption on the periosteal surfaces of this bone, revealed by in vivo staining with alizarin red S. Presence of reaction to acid phosphatase is, as shown before, an indication of resorptive surfaces, while the presence of reaction to alkaline phosphatase is an indication...

  19. Extractive spectrophotometric determination of micro and sub-micro amounts of fluoride

    NARCIS (Netherlands)

    Haarsma, J.P.S.; Agterdenbos, J.

    1971-01-01

    A simple and sensitive extractive spectrophotometric determination of fluoride with the cerium(III)-alizarin complexan chelate has been investigated. The fluoro chelate formed is extracted into n-pentanol containing triethylamine. It is possible to achieve under selected conditions a selective extra

  20. Chemical and enzymatic hydrolysis fo anthraquinone glycosides from madder roots

    NARCIS (Netherlands)

    Derksen, G.C.H.; Naayer, M.; Beek, van T.A.; Capelle, A.; Haaksman, I.K.; Doren, H.A.; Groot, de Æ.

    2003-01-01

    For the production of a commercially useful dye extract from madder, the glycoside ruberythric acid has to be hydrolysed to the aglycone alizarin which is the main dye component. An intrinsic problem is the simultaneous hydrolysis of the glycoside lucidin primeveroside to the unwanted mutagenic agly

  1. Validation of a simple and fast method to quantify in vitro mineralization with fluorescent probes used in molecular imaging of bone.

    Science.gov (United States)

    Moester, Martiene J C; Schoeman, Monique A E; Oudshoorn, Ineke B; van Beusekom, Mara M; Mol, Isabel M; Kaijzel, Eric L; Löwik, Clemens W G M; de Rooij, Karien E

    2014-01-01

    Alizarin Red S staining is the standard method to indicate and quantify matrix mineralization during differentiation of osteoblast cultures. KS483 cells are multipotent mouse mesenchymal progenitor cells that can differentiate into chondrocytes, adipocytes and osteoblasts and are a well-characterized model for the study of bone formation. Matrix mineralization is the last step of differentiation of bone cells and is therefore a very important outcome measure in bone research. Fluorescently labelled calcium chelating agents, e.g. BoneTag and OsteoSense, are currently used for in vivo imaging of bone. The aim of the present study was to validate these probes for fast and simple detection and quantification of in vitro matrix mineralization by KS483 cells and thus enabling high-throughput screening experiments. KS483 cells were cultured under osteogenic conditions in the presence of compounds that either stimulate or inhibit osteoblast differentiation and thereby matrix mineralization. After 21 days of differentiation, fluorescence of stained cultures was quantified with a near-infrared imager and compared to Alizarin Red S quantification. Fluorescence of both probes closely correlated to Alizarin Red S staining in both inhibiting and stimulating conditions. In addition, both compounds displayed specificity for mineralized nodules. We therefore conclude that this method of quantification of bone mineralization using fluorescent compounds is a good alternative for the Alizarin Red S staining.

  2. Beyond Vibrationally Mediated Electron Transfer: Coherent Phenomena Induced by Ultrafast Charge Separation

    CERN Document Server

    Huber, Robert; Moser, Jacques E; Grätzel, Michael; Wachtveitl, Josef

    2016-01-01

    Wave packet propagation succeeding electron transfer (ET) from alizarin dye molecules into the nanocrystalline TiO2 semiconductor has been studied by ultrafast transient absorption spectroscopy. Due to the ultrafast time scale of the ET reaction of about 6 fs the system shows substantial differences to molecular ET systems. We show that the ET process is not mediated by molecular vibrations and therefore classical ET theories lose their applicability. Here the ET reaction itself prepares a vibrational wave packet and not the electromagnetic excitation by the laser pulse. Furthermore, the generation of phonons during polaron formation in the TiO2 lattice is observed in real time for this system. The presented investigations enable an unambiguous assignment of the involved photoinduced mechanisms and can contribute to a corresponding extension of molecular ET theories to ultrafast ET systems like alizarin/TiO2.

  3. Visible light photoactivity of Polypropylene coated Nano-TiO2 for dyes degradation in water

    Science.gov (United States)

    Giovannetti, R.; Amato, C. A. D.'; Zannotti, M.; Rommozzi, E.; Gunnella, R.; Minicucci, M.; di Cicco, A.

    2015-12-01

    The use of Polypropylene as support material for nano-TiO2 photocatalyst in the photodegradation of Alizarin Red S in water solutions under the action of visible light was investigated. The optimization of TiO2 pastes preparation using two commercial TiO2, Aeroxide P-25 and Anatase, was performed and a green low-cost dip-coating procedure was developed. Scanning electron microscopy, Atomic Force Microscopy and X-Ray Diffraction analysis were used in order to obtain morphological and structural information of as-prepared TiO2 on support material. Equilibrium and kinetics aspects in the adsorption and successive photodegradation of Alizarin Red S, as reference dye, are described using polypropylene-TiO2 films in the Visible/TiO2/water reactor showing efficient dyes degradation.

  4. Textile wastewater purification through natural coagulants

    Science.gov (United States)

    Beltrán-Heredia, J.; Sánchez-Martín, J.; Rodríguez-Sánchez, M. T.

    2011-09-01

    A new coagulant obtained through polymerization of Acacia mearnsii de Wild tannin extract has been characterized in the removal of two dangerous dye pollutants: Alizarin Violet 3R and Palatine Fast Black WAN. This coagulant is lab-synthesized according to the etherification of tannins with glycidyltrimethylammonium chloride and formaldehyde and its performance in dye removal in terms of efficiency was high. Reasonably low coagulant dosages (ca. 50 mg L-1) reaches high capacity levels (around 0.8 for Alizarin Violet 3R and 1.6 for Palatine Fast Black WAN mg dye mg-1 of coagulant) and pH and temperature are not extremely affecting variables. The systems coagulant dyes were successfully modeled by applying the Langmuir hypothesis. q max and b parameters were obtained with an adjusted correlation factor ( r 2) above 0.8.

  5. Photoinduced interaction between MPA capped CdTe QDs and certain anthraquinone dyes

    Energy Technology Data Exchange (ETDEWEB)

    Jagadeeswari, S.; Asha Jhonsi, M.; Kathiravan, A. [School of Chemistry, Bharathidasan University, Tiruchirappalli 620 024, Tamil Nadu (India); Renganathan, R., E-mail: rrengas@gmail.co [School of Chemistry, Bharathidasan University, Tiruchirappalli 620 024, Tamil Nadu (India)

    2011-04-15

    Photoinduced interaction of mercapto propionic acid (MPA) capped CdTe quantum dots (QDs) with certain anthraquinone dyes namely alizarin, alizarin red S, acid blue 129 and uniblue has been studied by steady state and time resolved fluorescence measurements. Addition of anthraquinone dyes to CdTe QDs results in the reduction of electron hole recombination has been observed (i.e., fluorescence quenching). The Stern-Volmer constant (K{sub SV}), quenching rate constant (k{sub q}) and association constants (K) were obtained from fluorescence quenching data. The interaction of anthraquinone dyes with QDs occurs through static quenching was confirmed by unaltered fluorescence lifetime. The occurrence of electron transfer quenching mechanism has been proved by the negative free energy change ({Delta}G{sub et}) obtained as per the Rehm-Weller equation.

  6. Isolation and extraction of lucidin primeveroside from Rubia tinctorum L. and crystal structure elucidation.

    Science.gov (United States)

    Henderson, Robert L; Rayner, Christopher M; Blackburn, Richard S

    2013-11-01

    Madder (Rubia tinctorum L.) has been used as a dye for over 2000 years with alizarin and purpurin the major natural dyes analysed from extractions undertaken. The use of ethanol as the solvent in the extraction process produced an extract that yielded four anthraquinone compounds lucidin primeveroside, ruberythric acid, alizarin and lucidin-ω-ethyl ether. Gravitational separation of the extract was used to record the first crystal structure of lucidin primeveroside, which is also the first ever known crystal structure of a glycoside containing anthraquinone moiety. The crystal structure along with (1)H and (13)C NMR helped elucidate and confirm the structure of this overlooked natural dye which has been shown to be a major compound in R. tinctorum L.

  7. Teratogenic Potential of Ethylene Thiourea (ETU), A Positive Control in Sprague-Dawley Rats.

    Science.gov (United States)

    1987-10-01

    skeletal examination. IRTU produced teratogenic and embryotoxic effects in Sprague-Dawley rats. Ile .P DO FORM 1473,.84 MAR di A#114 ’atiaon iTr.y Oae...examination or alizarin red stain for skeletal examination. ETU produced teratogenic and embryotoxic effects in Sprague-Dawley rats. Key Words...frequency in the test groups than in the vehicle control group, this is evidence of embryotoxicity . Other manifestations of embryotoxicity are decreased body

  8. Enhancing Quality of Life for Breast Cancer Patients with Bone Metastases

    Science.gov (United States)

    2007-03-01

    50 µg/ml ascorbic acid and 5 mM sodium phosphate in order to promote mineral formation . Cell Proliferation and Apoptosis: Samples will be assayed...confluent (3-4 days), the medium will be replaced with media containing ascorbic acid (50 µg/ml) and sodium phosphate (4 mM). After 21 days, mineral ... formation will be measured with a quantitative assay for Alizarin Red S (21). Radiation Therapy: Radiation will be administered with a Phillips MGC-30

  9. Alkaloid and other chemical constituents from Psychotria stachyoides Benth

    Energy Technology Data Exchange (ETDEWEB)

    Pimenta, Antonia T.A.; Uchoa, Daniel E.A.; Silveira, Edilberto R.; Lima, Mary Anne S. [Departamento de Quimica Organica e Inorganica, Universidade Federal do Ceara, Fortaleza, CE (Brazil); Braz-Filho, Raimundo, E-mail: mary@dqoi.ufc.br [Centro de Ciencias, Universidade Estadual do Norte Fluminense and Universidade Federal Rural do Rio de Janeiro, Campos dos Goytacazes-RJ (Brazil)

    2011-09-15

    The organic extracts of leaves and roots of Psychotria stachyoides provided the new glucoside monoterpenoid indole alkaloid N-demethylcorreantoside, besides bizantionoside B, a-amyrin, alizarine methyl-ether, rubiadine, scopoletin, barbinevic acid and a mixture of b-sitosterol and stigmasterol glucosides. The structural characterization of the isolates was established based on infrared spectroscopy (IR), mass spectrometry (MS) and, particularly, 1D and 2D nuclear magnetic resonance (NMR). (author)

  10. Electrocoagulation of Quinone Pigments

    Directory of Open Access Journals (Sweden)

    Duang Buddhasukh

    2006-07-01

    Full Text Available Some representative quinones, viz. one naphthoquinone (plumbagin and five anthraquinones (alizarin, purpurin, chrysazin, emodin, and anthrarufin, were subjected to electrocoagulation. It was found that the rate and extent of coagulation of these compounds appears to correlate with the number and relative position of their phenolic substituent groups, and that all of the coagulated quinones could be recovered. Attempts were then made to electrochemically isolate three quinones, namely plumbagin, morindone and erythrolaccin, from natural sources.

  11. Neuronal feedback between brain and inner ear for growth of otoliths in fish

    Science.gov (United States)

    Anken, R. H.; Edelmann, E.; Rahmann, H.

    Previous investigations revealed that fish inner ear otolith growth (concerning otolith size and calcium-incorporation) depends on the amplitude and the direction of gravity, suggesting the existence of a (negative) feedback mechanism. In search for the regulating unit, the vestibular nerve was unilaterally transected in neonate swordtail fish ( Xiphophorus helleri) which were subsequently incubated in the calcium-tracer alizarin-complexone. Calcium incorporation ceased on the transected head sides, indicating that calcium uptake is neurally regulated.

  12. Extraction and Separation of Molybdenum by Using Homogeneous Liquid-Liquid Microextraction via Flotation Assistance

    OpenAIRE

    Rezaee, Mohammad; Mozaffari,Maryam; Haddadi,Hedayat; Pourjavid,Mohammad R.; SEMNANI, Abolfazl

    2015-01-01

    Homogeneous liquid-liquid microextraction via flotation assistance (HLLME-FA) was investigated for the extraction of molybdenum from the water samples. Alizarin Red S and cetyl trimethylammonium bromide (CTAB) were used as a complexing ligand and ion-pairing reagent, respectively. The enriched analyte in the floated organic phase was determined by electrothermal atomic absorption spectrometry (ETAAS). In this work, low density organic solvent was used and no centrifugation was required in thi...

  13. Visual Detection and Determination of Melamine Using Synthetic Dyes

    Directory of Open Access Journals (Sweden)

    Ramesh Thimmasandra Narayan

    2014-01-01

    Full Text Available We have used spectroscopic technique for the detection of melamine. The effect of melamine on the colour as well as the pH of bromophenol, methyl red and alizarin red dye solutions was examined at different mole ratios. It is found that we observe color transition and the absorption maxima for bromophenol were at 598 nm, while for methyl red, and alizarin red-S dye they are at 520 nm and 423 nm, respectively. We observe an increase in the absorption intensities at 598 nm with increase in the concentration of melamine in bromophenol blue dye. The absorption intensities at 520 nm decreases and new peak at 420 nm emerges in methyl red dye-melamine mixture. While the absorption intensities at 420 nm decreases and 520 nm peak emerges in alizarin red S dye-melamine at higher mole ratios. The results indicate that we can choose the appropriate dye of suitable range to detect the concentration of melamine from 3 to 206 mg dm−3. The results demonstrate possible use of the simple method for the qualitative and quantitative detection of melamine in adulterated food samples.

  14. The Application of Electrochemical Impedance Techniques in Analyzing the AC Response of Some Two-electron Transfer Dye Systems

    Directory of Open Access Journals (Sweden)

    Farouk Rashwan

    2005-01-01

    Full Text Available The Electrochemical Impedance Spectroscopic techniques (EIS were used to investigate the behavior of some dye compounds (quinoid systems characterized with 2e-transfer processes. For this purpose, Alizarin Red S (ARS, Alizarin Cyanine (AC, Alizarin Viridin (AV and carminic acid were chosen for the measurements. The EIS experiments were performed using a small AC amplitude (10 mV p-p in addition to a relatively wide frequency range (0.01 Hz ≤ f ≤ 105 Hz. The investigations were carried out at room temperature in aqueous media (HClO4, NaClO4 and KNO3 on the Hanging Mercury Drop Electrode (HMDE and for comparison one experiment only was measured in aprotic solvent (DMF on the Pt-disc electrode. The EIS diagrams of these systems were characterized in the complex plane by two fundamental observations, the first of which is a straight line crossing the real axis at an angle of 45° (or at least nearly so and the second one is two semicircles beside each other corresponding to high-frequency and low-frequency regions, which are implying the presence of well-separated time constants. The EIS characteristic parameters for these dye systems were calculated and discussed.

  15. Oxygen pre-breathing decreases dysbaric diseases in UW sheep undergoing hyperbaric exposure.

    Science.gov (United States)

    Sobakin, A S; Wilson, M A; Lehner, C E; Dueland, R T; Gendron-Fitzpatrick, A P

    2008-01-01

    Prolonged exposure of humans and animals to increased pressure as in a disabled submarine (DISSUB) can saturate the body's tissues with dissolved N2 as compressed air is breathed. Decompression-induced bubble formation in the long bone marrow cavity may lead to a bone compartment syndrome resulting in bone ischemia and necrosis. We tested oxygen pre-breathing prior to decompression in sheep to assess the effect upon dysbaric osteonecrosis (DON) induction in a DISSUB simulation experiment. A total of sixteen adult female sheep were used throughout the experiment. Four sheep were used as controls without oxygen pre-breathing. All sheep (99 +/- 14 kg SD) underwent dry chamber air exposure at 60 fsw (2.79 atm abs) (.2827 MPa) for 24 h followed by oxygen (88-92%) pre-breathing (15-min, 1-h, and 2-h and air for control) before "dropout" decompression at 30 fsw/min (0.91 atm/min). 99mTc-methylene diphosphonate (MDP) bone scans of the distal (radii and tibiae) long bones were used to detect "hot spots" of remodeling suggestive of DON lesions. Alizarin complexone fluorochrome was injected to visualize sites of metabolic activity indicating DON repair of both the proximal and distal long bones (radii, tibiae, femora, and humeri). Our findings showed that the amount of alizarin complexone deposition and bone scan uptake was greater in sheep with shorter oxygen pre-breathing times than those undergoing longer pre-breathing dives (p = 0.0056 and p = 0.001, for one and two hour pre-breathes respectively). Proximal limb bones (femur, humerus) displayed less alizarin complexone deposition than the distal radius and tibia (p < 0.0001).

  16. The Effect of 1α,25(OH)2D3 on Osteogenic Differentiation of Stem Cells from Dental Pulp of Exfoliated Deciduous Teeth

    Science.gov (United States)

    Mojarad, Farzad; Amiri, Iraj; Rafatjou, Rezvan; Janeshin, Atousa; Farhadian, Maryam

    2016-01-01

    Statement of the Problem: Stem cells from human exfoliated deciduous teeth (SHEDs) are a population of highly proliferative cells, being capable of differentiating into osteogenic, odontogenic, adipocytes, and neural cells. Vitamin D3 metabolites such as 1α, 25-dihydroxyvitamin D3 are key factors in the regulation of bone metabolism. Purpose: The aim of this study was to investigate the effect of 1α, 25-dihydroxyvitamin D3 on osteogenic differentiation (alkaline phosphatase activity and alizarin red staining) of stem cells of exfoliated deciduous teeth. Materials and Method: Dental pulp was removed from freshly extracted primary teeth and immersed in a digestive solution. Then, the dental pulp cells were immersed in α-MEM (minimum essential medium) to which 10% fetal bovine serum was added. After the third passage, the cells were isolated from the culture plate and were used for osteogenic differentiation. As a control group, the cells were cultured in osteogenic cell culture medium. As the case group, the cells were cultured in osteogenic culture medium supplemented with 100 nM 1α,25 (OH)2D3. The alkaline phosphatase (ALP) activity and alizarin red staining were analyzed to evaluate the osteogenic differentiation at day 21. The results were analyzed by using t-test. Results: Compared with the control group, significant increase was observed in ALP activity of SHEDs after being treated with 1α,25(OH)2D3 (p= 0.002). Alizarin red staining demonstrated that the cells exposed to 1α,25(OH)2D3 induced higher mineralized nodules (p2D3. It can be concluded that 1α,25(OH)2D3 can improve osteoblastic differentiation. PMID:27942551

  17. Synthesis of benzofuran derivatives as selective inhibitors of tissue-nonspecific alkaline phosphatase: effects on cell toxicity and osteoblast-induced mineralization.

    Science.gov (United States)

    Marquès, Stéphanie; Buchet, René; Popowycz, Florence; Lemaire, Marc; Mebarek, Saïda

    2016-03-01

    Tissue-nonspecific alkaline phosphatase (TNAP) by hydrolyzing pyrophosphate, an inhibitor of apatite formation, promotes extracellular matrix calcification during bone formation and growth, as well as during ectopic calcification under pathological conditions. TNAP is a target for the treatment of soft tissue pathological ossification. We synthesized a series of benzofuran derivatives. Among these, SMA14, displayed TNAP activity better than levamisole. SMA14 was found to be not toxic at doses of up to 40μM in osteoblast-like Saos-2 cells and primary osteoblasts. As probed by Alizarin Red staining, this compound inhibited mineral formation in murine primary osteoblast and in osteoblast-like Saos-2 cells.

  18. Characterization of cultural remains associated to a human skeleton found at the site HMS Swift (1770)

    Science.gov (United States)

    Maier, M. S.; Gómez, B. A.; Parera, S. D.; Elkin, D.; De Rosa, H.; Ciarlo, N. C.; Svoboda, H.

    2010-08-01

    Different types of materials found in association with a human skeleton found in an 18th century shipwreck in Patagonia (Argentina) were analyzed by means of OM, SEM-EDX, HPLC, and chemical analysis. Alizarin and purpurin, the main anthraquinones of the dye plant Rubia tinctorum L. (madder) were identified as the coloring matter of a red fabric attached to the skeleton. Metallographic and chemical analysis of one of the dome-shaped buttons associated to the human bones revealed that it was composed of a Pb-Sn-Cu alloy known as pewter. The results obtained support the hypothesis that the remains originally were part of a private marine uniform.

  19. Colorimetric determination of the fluoride ion - application to uranium metal and to uranous fluoride; Dosage colorimetrique de l'ion fluor - application a l'uranium metal et au fluorure uraneux

    Energy Technology Data Exchange (ETDEWEB)

    Hering, H.; Hure, J.; Legrand, S. [Commissariat a l' Energie Atomique (France)

    1949-12-01

    In the determination described for fluoride in U metal, the U is brought into H{sub 2}SO{sub 4} solution by anodic oxidation, the fluo-silicic acid is distilled by entrainment in water vapor, and the F ion is determined in the distillate by using the fact that it complexes Zr and thus prevents the formation of the Zr-alizarin S lake. For F ion in UF{sub 4}, the compound is dissolved in a Na{sub 2}CO{sub 3}-H{sub 2}O{sub 2} mixture, and F is determined in the solution by the colorimetric method described. (author)

  20. Lumbosacral transitional vertebra and thoracic limb malformations in a Chihuahua puppy.

    Science.gov (United States)

    Schultz, V A; Watson, A G

    1995-01-01

    A three-month-old, male Chihuahua puppy with congenital absence of the distal 40% of the right thoracic limb was examined. The limb ended as a short, rounded, skin-covered stump. Radiography revealed a 40% shortened humerus tapered to a blunt end without its distal extremity. Dissection of the left thoracic limb identified luxation of the elbow joint and absence of the fourth digital pad. Alizarin-red staining and clearing demonstrated syndactylous fourth and fifth digits in the left thoracic limb and an anomalous eighth lumbar vertebra. This additional vertebra was unilaterally sacralized and constituted a lumbosacral transitional vertebra.

  1. Removal of biological stains from aqueous solution using a flow-through decontamination procedure.

    Science.gov (United States)

    Lunn, G; Klausmeyer, P J; Sansone, E B

    1994-01-01

    Chromatography columns filled with Amberlite XAD-16 were used to decontaminate, using a continuous flow-through procedure, aqueous solutions of the following biological stains: acridine orange, alcian blue 8GX, alizarin red S, azure A, azure B, brilliant blue G, brilliant blue R, Congo red, cresyl violet acetate, crystal violet, eosin B, eosin Y, erythrosin B, ethidium bromide, Giemsa stain, Janus green B, methylene blue, neutral red, nigrosin, orcein, propidium iodide, rose Bengal, safranine O, toluidine blue O, and trypan blue. Adsorption was most efficient for stains of lower molecular weight (removing stains from aqueous solution.

  2. Pineal concretions in turkey (Meleagris gallopavo) as a result of collagen-mediated calcification.

    Science.gov (United States)

    Przybylska-Gornowicz, B; Lewczuk, B; Prusik, M; Bulc, M

    2009-04-01

    The intra-pineal calcification is a well-known phenomenon in mammals, however it is almost completely unknown in birds. The aim of the present work was to analyze morphology and genesis of the pineal concretions in the turkey. The studies were performed on the pineals collected from one-year-old turkeys (Meleagris gallopavo). In addition to standard morphological methods, the alizarin red S and potassium pyroantimonate methods were employed for localization of calcium at the light and electron microscopy level. In light microscopy, calcified concretions with diameters from 300 microm to 2 mm and quantities from 3 to 6 per gland were observed in all the examined pineals. They were stained red with alizarin S and showed the presence of collagen in Mallory's staining. Two types of cells were noted inside the concretion: polygonal and elongated ones. Using electron microscopy, three parts were distinguished within the calcification area. The peripheral part contained densely packed collagen fibrils, some elongated cells and numerous pyroantimonate precipitates demonstrating the presence of calcium ions. In the intermediate part, the fibrils were covered by almost continuous sheets of pyroantimonate precipitates and fused side by side. The central part showed an appearance of calcified hard tissue and contained some polygonal (osteocyte-like) cells. The obtained data demonstrated that the formation of the pineal concretions in the turkey is associated with the mineralization of collagen. This process is completely different from the mechanisms responsible for the formation of the concretions in the mammalian pineal.

  3. Effect of silica/titania ratio on enhanced photooxidation of industrial hazardous materials by microwave treated mesoporous SBA-15/TiO2 nanocomposites

    Science.gov (United States)

    Mehta, Akansha; Mishra, Amit; Sharma, Manisha; Singh, Satnam; Basu, Soumen

    2016-07-01

    In this study microwave assisted technique has been adopted for the synthesis of different weight ratios of TiO2 dispersed on Santa barbara amorphous-15 (SBA-15) support. Morphological study revealed TiO2 particles (4-10 nm) uniformly distributed on SBA-15 while increases in SBA-15 content results in higher specific surface area (524-237 m2/g). The diffraction intensity of 101 plane of anatase polymorph was seen increasing with increase in TiO2 ratio. All the photocatalysts were having a mesoporous nature and follow the Langmuir IV isotherm, SBA-15 posses the highest pore volume (0.93 cm3 g-1) which consistently decreased with TiO2 content and was lowest (0.50 cm3 g-1) in case of 5 wt% of TiO2 followed by P25 (0.45 cm3 g-1) while pore diameter increased after TiO2 incorporation due to pore strain. The photocatalytic activity of the nanocomposites were analysed for the photodegradation of alizarin dye and pentachlorophenol under UV light irradiation. The reaction kinetics suggested the highest efficiency (98 % for alizarin and 94 % for PCP) of 5 wt% TiO2 compared to other photocatalysts, these nanocomposites were reused for several cycles, which is most important for heterogeneous photocatalytic degradation reaction.

  4. Investigation of red natural dyes used in historical objects by HPLC-DAD-MS.

    Science.gov (United States)

    Karapanagiotis, Ioannis; Chryssoulakis, Yannis

    2006-01-01

    High performance liquid chromatography (HPLC) with UV-Vis Diode Array Detection (DAD) and electrospray mass spectrometric (ESI-MS) method was utilized for the identification of coloring components of madder, Armenian and Mexican cochineal, lac dye, brazilwood, safflower and dragon blood--probably the most important red natural dyestuffs found in objects of the cultural heritage. UV-Vis detection limits in the range of 0.2-0.6 ng for carminic acid, alizarin and purpurin were achieved using a gradient elution of H2O-0.01% TFA and CH3CN-0.01% TFA. ESI mass spectrometer was also used, as a supportive detection method to the standard DAD, for further analysis of the tested materials, with the ability to analyze dyestuffs as small as one milligram. The presence of madder was revealed in two historical (Hellenistic and Roman period) samples, found in the Mediterranean area, by identifying purpurin in both of them. Munjistin was also identified in one of the samples (Hellenistic period) while alizarin was not detected, raising questions regarding the exact madder type, utilized in the historical samples.

  5. Quantification of in vitro mineralisation using ion chromatography.

    Science.gov (United States)

    Souter, Paul; Horner, Alan; Cunningham, Jim C

    2011-03-15

    Analysis of in vitro mineralisation is an important tool in orthopedic research, allowing assessment of new therapeutic agents and devices; however, access to analytical equipment and accuracy of current methods can be a limiting factor. This current work investigated the use of calcium chelation with citric acid and subsequent analysis by ion chromatography as a method for accurately quantifying the extent of in vitro calcium deposition. Primary human osteoblasts were cultured on tissue culture plastic for 21 days under osteogenic conditions. At 3, 7, 14, and 21 days, alizarin red staining and citric acid calcium chelation of the cultures were performed. The use of alizarin red revealed increased calcium deposition over the culture period but was not sensitive enough to detect mineralisation at early time points after taking in to account background residual staining. The use of ion chromatography gave a limit of detection of 2 μg calcium, sensitive enough to detect mineralisation after 3 days, with no issues relating to background levels. We believe that the use of ion chromatography for quantifying in vitro mineralisation gives researchers an accurate, accessible, and cheap way of assessing novel technologies.

  6. Spectrophotometric determination of some histamine H1-antagonists drugs in their pharmaceutical preparations

    Science.gov (United States)

    Hassan, Wafaa S.; El-Henawee, Magda M.; Gouda, Ayman A.

    2008-01-01

    Two rapid, simple and sensitive extractive specrophotometric methods has been developed for the determination of three histamine H1-antagonists drugs, e.g., chlorphenoxamine hydrochloride (CPX), diphenhydramine hydrochloride (DPH) and clemastine (CMT) in bulk and in their pharmaceutical formulations. The first method depend upon the reaction of molybdenum(V) thiocyanate ions (Method A) with the cited drugs to form stable ion-pair complexes which extractable with methylene chloride, the orange red color complex was determined colorimetrically at λmax 470 nm. The second method is based on the formation of an ion-association complex with alizarin red S as chromogenic reagents in acidic medium (Method B), which is extracted into chloroform. The complexes have a maximum absorbance at 425 and 426 nm for (DPH or CMT) and CPX, respectively. Regression analysis of Beer-Lambert plots showed a good correlation in the concentration ranges of 5.0-40 and 5-70 μg mL -1 for molybdenum(V) thiocyanate (Method A) and alizarin red S (Method B), respectively. For more accurate analysis, Ringbom optimum concentration ranges were calculated. The molar absorptivity, Sandell sensitivity, detection and quantification limits were calculated. Applications of the procedure to the analysis of various pharmaceutical preparations gave reproducible and accurate results. Further, the validity of the procedure was confirmed by applying the standard addition technique and the results obtained in good agreement well with those obtained by the official method.

  7. Raman spectroscopy of organic dyes adsorbed on pulsed laser deposited silver thin films

    Energy Technology Data Exchange (ETDEWEB)

    Fazio, E.; Neri, F. [Dipartimento di Fisica della Materia e Ingegneria Elettronica, Universitá di Messina, V.le F. Stagno d’Alcontres 31, I-98166, Messina, Italy. (Italy); Valenti, A. [Dipartimento di Chimica Inorganica, Chimica Analitica e Chimica Fisica, Universitá di Messina, V.le F. Stagno d’Alcontres 31, I-98166, Messina, Italy. (Italy); Ossi, P.M., E-mail: paolo.ossi@polimi.it [Dipartimento di Energia, Politecnico di Milano, via Ponzio 34-3, 20133 Milano, Italy. (Italy); Trusso, S.; Ponterio, R.C. [CNR-Istituto per i Processi Chimico-Fisici Sede di Messina, V.le F. Stagno d’Alcontres 37, I-98158 Messina, Italy. (Italy)

    2013-08-01

    The results of a surface-enhanced Raman scattering (SERS) study performed on representative organic and inorganic dyes adsorbed on silver nanostructured thin films are presented and discussed. Silver thin films were deposited on glass slides by focusing the beam from a KrF excimer laser (wavelength 248 nm, pulse duration 25 ns) on a silver target and performing the deposition in a controlled Ar atmosphere. Clear Raman spectra were acquired for dyes such as carmine lake, garanza lake and brazilwood overcoming their fluorescence and weak Raman scattering drawbacks. UV–visible absorption spectroscopy measurements were not able to discriminate among the different chromophores usually referred as carmine lake (carminic, kermesic and laccaic acid), as brazilwood (brazilin and brazilein) and as garanza lake (alizarin and purpurin). SERS measurements showed that the analyzed samples are composed of a mixture of different chromophores: brazilin and brazilein in brazilwood, kermesic and carminic acid in carmine lake, alizarin and purpurin in garanza lake. Detection at concentration level as low as 10{sup −7} M in aqueous solutions was achieved. Higher Raman intensities were observed using the excitation line of 632.8 nm wavelength with respect to the 785 nm, probably due to a pre-resonant effect with the molecular electronic transitions of the dyes.

  8. Determinação espectrofotométrica de aspartame em adoçantes por injeção em fluxo usando um reator em fase sólida contendo fosfato de zinco imobilizado

    Directory of Open Access Journals (Sweden)

    Pereira Airton Vicente

    2000-01-01

    Full Text Available A flow injection spectrophotometric method was developed for determining aspartame in sweeteners. Sample was dissolved in water and 250 µL of the solution was injected into a carrier stream of 5.0 x 10-5 mol L-1 sodium borate solution. The sample flowed through a column (14 cm x 2.0 mm packed with Zn3(PO42 immobilized in a polymeric matrix of polyester resin and Zn(II ions were released from the solid-phase reactor by formation of the Zn(II-aspartame complex. The mixture merged with a stream of borate buffer solution (pH 9.0 containing 0.030 % (m/v alizarin red S and the Zn(II-alizarin red complex formed was measured spectrophotometrically at 540 nm. The calibration graph for aspartame was linear in the concentration range from 10 to 80 µg mL-1 with a detection limit of 4 µg mL-1 of aspartame. The RSD was 0.3 % for a solution containing 40 µg mL-1 aspartame (n = 10 and seventy results were obtained per hour. The proposed method was applied for determining aspartame in commercial sweeteners.

  9. Biocompatible thermoresponsive PEGMA nanoparticles crosslinked with cleavable disulfide-based crosslinker for dual drug release.

    Science.gov (United States)

    Ulasan, Mehmet; Yavuz, Emine; Bagriacik, Emin Umit; Cengeloglu, Yunus; Yavuz, Mustafa Selman

    2015-01-01

    Smart materials have been attracting much attention because of their stimuli responsive nature. We have synthesized biocompatible thermoresponsive crosslinked poly(ethylene glycol) methyl ether methacrylate (PEGMA)-co-vinyl pyrrolidone nanoparticles (PEGMA NPs) using disulfide-based crosslinker by surfactant-free emulsion polymerization method. Particle characterization studies were carried out by dynamic light scattering, and scanning electron microscopy. Polymerization kinetics, effect of crosslinker and initiator concentrations on both average hydrodynamic diameter and polydispersity index were investigated. Hydrodynamic diameters of thermoresponsive PEGMA NPs were decreased from 210 nm to 90 nm upon heating over the lowest critical solution temperature (LCST). Disulfide crosslinked PEGMA NPs were demonstrated as a dual delivery system. Rhodamine B, a model of small-sized drug molecule, and poly(ethylene glycol) (PEG)-alizarin yellow, a model of large drug molecule, were loaded into PEGMA NPs where LCST of these NPs was tuned to 37°C, the body temperature. The rhodamine B was released from PEGMA NPs upon heating to 39°C. Then, PEG-alizarin content was released by subsequent degradation of nanoparticles using dithiothreitol (DTT), which reduces disulfide bonds to thiols. Furthermore, cytotoxicity studies of PEGMA NPs were carried out in 3T3 cells, which resulted in no toxic effect on the cells.

  10. Preparation and photocatalyticactivity of CuO/CeO2 nanocomposites%CuO/CeO2复合物的制备及其光催化性质研究

    Institute of Scientific and Technical Information of China (English)

    张晓娟; 张胜义

    2012-01-01

    In the presence of the CTAB soft template, the CuO/CeO2 nanocomposites were prepared by co-precipitation process. The product was characterized by TEM,XRD and UV-vis spectra. The experimental results showed that CuO/CeO2 nanocomposites had a strong adsorptive decolorization activity for organic dye alizarin red. Furthermore, its decolorization activity for alizarin red increased with the increase of the CuO content. Under illumination, this decolorization activity could be further enhanced due to the catalysis of the CuO/ CeO2 nanocomposites.%以CTAB为模板采用共沉淀法合成CuO/CeO2复合物,用透射电镜、X射线粉末衍射和紫外可见光谱对复合物进行表征;研究CuO/CeO2复合物对茜素红的催化脱色效果.实验结果表明,CuO/CeO2复合物既具有吸附作用也具有光催化降解作用,且随着CuO含量和催化剂用量的增加催化脱色效果有明显提高.

  11. Micellar Electrokinetic Chromatography with Laser-Induced Fluorescence Detection for Separation of Red and Yellow Historical Dyes

    Directory of Open Access Journals (Sweden)

    Shokoufeh Ahmadi

    2013-03-01

    Full Text Available In this study, the separation parameters in micellar electrokinetic chromatography (MEKC-laser-induced fluorescence (LIF were optimized for the separation of red and yellow historical dyes in 20 mM borate buffer with 20 mM sodium dodecyl sulfate (SDS. Separation conditions were optimized by changing pH, organic modifier (methanol and acetonitrile concentrations and applied voltage. The mixtures of dyes used in this study included four anthraquinone dyes (alizarin, purpurin, emodin and carmine and six flavonoid dyes (luteolin, apigenin, kaempferol, quercetin, morin and myricetin. For this work, dyes were introduced electro-kinetically (10 kV for 5 s into a 50 cm capillary (10 µm id and separated using a running potential of 18, 20, 22 and 25 kV. Absolute limits of detection for most of these dyes was less than 1 pg. For dyes such as alizarin, improved detection limits were achieved at pH = 9.24; however dyes such as purpurin had significantly improved detection limits at pH = 8.0. The successful extraction and identification of a number of dyes in plants and textiles samples is also described.

  12. Surface-charging behavior of Zn-Cr layered double hydroxide.

    Science.gov (United States)

    Rojas Delgado, R; Arandigoyen Vidaurre, M; De Pauli, C P; Ulibarri, M A; Avena, M J

    2004-12-15

    A Zn-Cr layered double hydroxide (LDH) having the formula Zn(2)Cr(OH)(6)Cl(0.7)(CO(3))(0.15)2.1H(2)O was synthesized and characterized by powder X-ray diffraction, infrared spectroscopy, acid-base potentiometric titration, mass titration, electrophoretic mobility, and modeling of the electrical double layer. Adsorption of alizarin was also performed in order to show some particular features of the HDL. Net hydroxyl adsorption, which increases with increasing pH and decreasing supporting electrolyte concentration, takes place above pH 5. The electrophoretic mobility of the particles was always positive and it decreased when the pH was higher than 9. An isoelectric point of 12 could be estimated by extrapolating the data. The modified MUSIC model was used to estimate deprotonation constants of surface groups and different adsorption models were compared. Good fit of hydroxyl adsorption and electrophoresis could be achieved by considering both OH(-)/Cl(-) exchange at structural sites and proton desorption from surface hydroxyl groups. The modeling, in agreement with alizarin adsorption, indicates that most of the structural positive charge of the LDH is screened at the surface by exchanged anions and negatively charged surface groups. It also suggests that only structural charge sites initially neutralized by chloride ions are active for anion exchange. The remaining sites are blocked by carbonate and do not participate in the exchange.

  13. Prior states: evolution of composition and color in two Barnett Newman paintings

    Science.gov (United States)

    Epley, Bradford A.; Rogge, Corina E.

    2015-11-01

    The color field paintings of Barnett Newman, one of the great American abstract expressionist painters, are seminal works of the modern era. They feature large flat fields of vibrant colors intended to allow the viewer to connect with the paintings in immediate, visceral ways. Despite the apparent simplicity of his compositions, Newman considered himself an intuitive painter and allowed his compositions to evolve during the painting process. Two paintings in the Menil Collection, Untitled 2 (1950) and Unfinished Painting [Blue and Brown 1970— #2] (1970) display visual evidence of former states, but attempts to elucidate earlier compositions by X-radiography were inconclusive due to the lack of contrast in paint densities. We applied limited sampling and used a handheld X-ray fluorescence spectrometer in a `scanning' manner to determine the color and composition of the previous states of these paintings to help us better understand their evolution. Newman altered his initial cadmium red and alizarin composition in Untitled 2 (1950) by overpainting the alizarin region with a wider band of Mars black paint. He then modulated the surface of the black by partially covering it with a carbonaceous black with a different gloss. For Unfinished Painting [Blue and Brown 1970— #2] (1970), Newman not only changed the cadmium red to an umber but simplified the composition, removing multiple zips and refining it to its current monumental state. This evidence of Newman's decision-making processes permits a tantalizing glimpse of the artist consistently looking both ahead and backward, experimenting and revisiting.

  14. Combined effects of proinflammatory cytokines and intermittent cyclic mechanical strain in inhibiting osteogenicity in human periodontal ligament cells.

    Science.gov (United States)

    Sun, Chaofan; Chen, Lijiao; Shi, Xinlian; Cao, Zhensheng; Hu, Bibo; Yu, Wenbin; Ren, Manman; Hu, Rongdang; Deng, Hui

    2016-09-01

    Mechanical strain plays an important role in bone formation and resorption during orthodontic tooth movement. The mechanism has not been fully studied, and the process becomes complex with increased amounts of periodontal patients seeking orthodontic care. Our aims were to elucidate the combined effects of proinflammatory cytokines and intermittent cyclic strain (ICS) on the osteogenic capacity of human periodontal ligament cells. Cultured human periodontal ligament cells were exposed to proinflammatory cytokines (interleukin-1β 5 ng/mL and tumor necrosis factor-α 10 ng/mL) for 1 and 5 days, and ICS (0.5 Hz, 12% elongation) was applied for 4 h per day. The autocrine of inflammatory cytokines was measured by enzyme-linked immunosorbent assay. The expression of osteoblast markers runt-related transcription factor 2 and rabbit collagen type I was determined using real-time polymerase chain reaction and Western blot. The osteogenic capacity was also detected by alkaline phosphatase (ALP) staining, ALP activity, and alizarin red staining. We demonstrated that ICS impaired the osteogenic capacity of human periodontal ligament cells when incubated with proinflammatory cytokines, as evidenced by the low expression of ALP staining, low ALP activity, reduced alizarin red staining, and reduced osteoblast markers. These data, for the first time, suggest that ICS has a negative effect on the inductive inhibition of osteogenicity in human PDL cells mediated by proinflammatory cytokines.

  15. Avaliação de indicadores de uso diverso como inibidores de corrosão

    Directory of Open Access Journals (Sweden)

    Cardoso Sheila Pressentin

    2005-01-01

    Full Text Available Very often hydrochloric acid is employed in acidification operations aiming to dissolve the mineral matrix in petroleum wheel operations, which always require intense use of corrosion inhibitors. This work presents an evaluation of common indicators, phenolfthaleine, fluorescein, methylene blue, alizarine S and methyl orange, as corrosion inhibitors for carbon steel in HCl 15% w/v at temperatures of 26, 40 and 60 ºC. Fluorescein and methyl orange show excelent corrosion inhibition efficiencies at 26 ºC; however at 60 ºC only fluorescein shows good corrosion inhibition when employed with alcohol and/or formaldehyde. For the fluorescein 1% w/v + formaldehyde 0.6% w/v mixture we present polarization and impedance curves and adsorption isotherms.

  16. ADSORPTION AND RELEASING PROPERTIES OF BEAD CELLULOSE

    Institute of Scientific and Technical Information of China (English)

    A. Morales; E. Bordallo; V. Leon; J. Rieumont

    2004-01-01

    The adsorption of some dyes on samples of bead cellulose obtained in the Unit of Research-Production "Cuba 9"was studied. Methylene blue, alizarin red and congo red fitted the adsorption isotherm of Langmuir. Adsorption kinetics at pH = 6 was linear with the square root of time indicating the diffusion is the controlling step. At pH = 12 a non-Fickian trend was observed and adsorption was higher for the first two dyes. Experiments carried out to release the methylene blue occluded in the cellulose beads gave a kinetic behavior of zero order. The study of cytochrome C adsorption was included to test a proteinic material. Crosslinking of bead cellulose was performed with epichlorohydrin decreasing its adsorption capacity in acidic or alkaline solution.

  17. Kinetics of sonophotocatalytic degradation of anionic dyes with Nano-TiO2.

    Science.gov (United States)

    Vinu, R; Madras, Giridhar

    2009-01-15

    The current research work focuses on the combination of photocatalytic and sonocatalytic (sonophotocatalytic) degradation of anionic dyes, viz., Orange G, Remazol Brilliant Blue R, Alizarin Red S, Methyl Blue, and Indigo Carmine, with solution combustion synthesized TiO2 (CS TiO2) and commercial Degussa P-25 TiO2 (DP-25). The rate of sonophotocatalytic degradation of all the dyes and the reduction of total organic carbon was higher compared to the individual photo- and sonocatalytic processes. The effect of dissolved gases and ultrasonic intensity on the sonophotocatalytic degradation of the dyes was evaluated. A dual-pathway network mechanism of sonophotocatalytic degradation was proposed for the first time, and the rate equations were modeled using the network reduction technique. The kinetic rate coefficients of the individual steps were evaluated for all the systems by fitting the model with experimental data.

  18. Sm-Doped Tio2 Nanoparticles with High Photocatalytic Activity for ARS Dye Under Visible Light Synthesized by Ultrasonic Assisted Sol-Gel Method

    Directory of Open Access Journals (Sweden)

    V. Aware Dinkar

    2016-05-01

    Full Text Available In this article series of nano crystalline Sm-doped TiO2 nano particles with various molar concentration of samarium were synthesized by modified ultrasonic assisted sol-gel method and calcined at 500°C for 2 h. The synthesized nanomaterials were characterized in details using XRD, TEM, XPS UV–vis DRS and BET analysis. The detailed photocatalytic activity results revealed that doped samples shows excellent photodegradation efficiency towards model pollutant Alizarin red-S (ARS and almost 93% dye degrades within 120 minutes. The highest photodegradation efficiency was noticed for 1mole % samarium doped sample at 50 mgL-1 of catalyst dose. The photocatalytic activity of synthesized nano particles were also compared with commercially available ZnO and TiO2 (Degussa, P-25 photocatalyst. It was found that synthesized nano materials showed enhanced photocatalytic efficiency than commercially available semiconducting photocatalyst.

  19. Effect of dye-metal complexation on photocatalytic decomposition of the dyes on TiO2 under visible irradiation

    Institute of Scientific and Technical Information of China (English)

    MAHMOOD Tariq; CHEN Chuncheng; LIU Lili; ZHAO Dan; MA Wanghong; LIN Jun; ZHAO Jincai

    2009-01-01

    The photocatalytic degradation of dyes (Acid Chrome Blue K (ACBK) and Alizarin Red (AR)) with strong complexation ability was investigated in the presence of metal ions under visible light irradiation.It was found that, at low dye-metal ratio, the photodegradation of ACBK was markedly inhibited by the addition of high oxidative potential Cu2+.However, at high dye-metal ratio, the presence of Cu2+ enhanced the photodegradation of ACBK.The negtive effect of Cu2+ on the photodegradation of AR was observed for all dye-metal ratios.The relative chemical inert Zn2+ tended to enhance the photodegradation of both anionic dyes.The mechanism underlying the different effect of Cu2+ is discussed from the different roles of surface-adsorbed and dye-coordinated Cu2+ in the photodegradation of dyes.

  20. Morinda citrifolia leaves enhance osteogenic differentiation and mineralization of human periodontal ligament cells.

    Science.gov (United States)

    Boonanantanasarn, Kanitsak; Janebodin, Kajohnkiart; Suppakpatana, Prapan; Arayapisit, Tawepong; Rodsutthi, Jit-aree; Chunhabundit, Panjit; Boonanuntanasarn, Surintorn; Sripairojthikoon, Wanida

    2014-01-01

    This present study investigated the potential of Morinda citrifolia leaf aqueous extract to induce osteogenic differentiation and matrix mineralization of human periodontal ligament (hPDL) cells. Human periodontal ligament cells were cultured in complete medium, ascorbic acid with β-glycerophosphate, or Morinda citrifolia leaf aqueous extract. Morinda citrifolia leaf aqueous extract significantly increased alkaline phosphatase activity compared to culturing in complete medium or ascorbic acid with β-glycerophosphate. Matrixcontaining mineralized nodules were formed only when the cells were cultured in the presence of Morinda citrifolia leaf aqueous extract. These nodules showed positive alizarin red S staining and were rich in calcium and phosphorus according to energy dispersive X-ray analysis. In conclusion, Morinda citrifolia leaf extract promoted osteogenic differentiation and matrix mineralization in human periodontal ligament cells, a clear indication of the therapeutic potential of Morinda citrifolia leaves in bone and periodontal tissue regeneration.

  1. Lithium Chloride Modulates Adipogenesis and Osteogenesis of Human Bone Marrow-Derived Mesenchymal Stem Cells

    Directory of Open Access Journals (Sweden)

    Linjun Tang

    2015-08-01

    Full Text Available Background/Aims: Lithium chloride (LiCl has long been used as a psychiatric medication; however, its role in the differentiation of bone marrow-derived mesenchymal stem cells (BMSCs remains largely unknown. The aim of this study is to explore the effect of LiCl on the differentiation of BMSCs. Methods: The roles of LiCl in osteogenic and adipogenic processes were observed using alizarin red staining and oil red O staining, respectively. The effects of LiCl on the Wnt and Hedgehog (Hh pathways were investigated. Results: Our data showed that LiCl effectively promoted osteogenesis and inhibited adipogenesis by simultaneously affecting the Wnt and Hh pathways. Conclusion: These results suggest that LiCl influences the differentiation of BMSCs directly through the Wnt and Hh pathways and thus may be a candidate drug for the treatment of osteoporosis.

  2. Determination of strontium and simultaneous determination of strontium oxide, magnesium oxide and calcium oxide content of Portland cement by derivative ratio spectrophotometry.

    Science.gov (United States)

    Idriss, K A; Sedaira, H; Ahmed, S S

    2009-04-15

    A derivative spectrophotometric method has been developed for the determination of strontium in Portland cement. The method is applied successfully for the simultaneous determination of SrO, MgO and CaO. It is based on the use of Alizarin Complexone (AC) as a complexing agent and measurement of the derivative ratio spectra of the analytes. Interferences of manganese(II) and zinc(II) were eliminated by precipitation. The validity of the method was examined by analyzing several Standard Reference Material (SRM) Portland cement samples. The strontium complex formed at pH 9.5 allows precise and accurate determination of strontium over the concentration range of 1.5-18 mg L(-1) of strontium. The MDL (at 95% confidence level) was found to be 25 ng mL(-1) for strontium in National Institute of Standards and Technology (NIST) cement samples using the proposed method.

  3. Simultaneous identification of natural dyes in the collection of drawings and maps from The Royal Chancellery Archives in Granada (Spain) by CE.

    Science.gov (United States)

    López-Montes, Ana; Blanc García, Rosario; Espejo, Teresa; Huertas-Perez, José F; Navalón, Alberto; Vílchez, José Luis

    2007-04-01

    A simple and rapid capillary electrophoretic method with UV detection (CE-UV) has been developed for the identification of five natural dyes namely, carmine, indigo, saffron, gamboge and Rubia tinctoria root. The separation was performed in a fused-silica capillary of 64.5 cm length and 50 microm id. The running buffer was 40 mM sodium tetraborate buffer solution (pH 9.25). The applied potential was 30 kV, the temperature was 25 degrees C and detections were performed at 196, 232, 252, 300 and 356 nm. The injections were under pressure of 50 mbar during 13 s. The method was applied to the identification of carminic acid, gambogic acid, crocetin, indigotin, alizarin and purpurin in the collection of drawings and maps at the Royal Chancellery Archives in Granada (Spain). The method was validated by using HPLC as a reference method.

  4. Effects of gadolinium on proliferation, differentiation and calcification of primary mouse osteoblasts in vitro

    Institute of Scientific and Technical Information of China (English)

    ZHANG Jinchao; LI Yaping; ZHANG Qun; HAO Xiaohong; WANG Shuxiang

    2012-01-01

    To evaluate the effects of Gd on proliferation,differentiation and mineralization function of primary osteoblasts (OBs) in vitro,we tested cell viability by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay,cell differentiation by alkaline phosphatase (ALP) activity assay,synthesis of type I collagen,and oil red O and alizarin red S (ARS) stain assays.The results indicated that effects of Gd on the proliferation,osteogenic differentiation,mineralization function and adipocytic transdifferentiation of primary OBs depended on concentration and incubation time,but were not dose-dependent.It was suggested that the effect of Gd on bone metabolism was complicated,and concentration and culture time were key factors for switching the biological effects of Gd from damage to protection.

  5. Strontium Promotes Cementoblasts Differentiation through Inhibiting Sclerostin Expression In Vitro

    Directory of Open Access Journals (Sweden)

    Xingfu Bao

    2014-01-01

    Full Text Available Cementogenesis, performed by cementoblasts, is important for the repair of root resorption caused by orthodontic treatment. Based on recent studies, strontium has been applied for osteoporosis treatment due to its positive effect on osteoblasts. Although promising, the effect of strontium on cementoblasts is still unclear. So the aim of this research was to clarify and investigate the effect of strontium on cementogenesis via employing cementoblasts as model. A series of experiments including MTT, alkaline phosphatase activity, gene analysis, alizarin red staining, and western blot were carried out to evaluate the proliferation and differentiation of cementoblasts. In addition, expression of sclerostin was checked to analyze the possible mechanism. Our results show that strontium inhibits the proliferation of cementoblasts with a dose dependent manner; however, it can promote the differentiation of cementoblasts via downregulating sclerostin expression. Taking together, strontium may facilitate cementogenesis and benefit the treatment of root resorption at a low dose.

  6. Enzymatic Cell Isolation and Explant Cultures of Rat Calvarial Osteoblast Cells

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    Osteoblast cells were isolated from the calvarial bones of newborn Wistar rats and cultured in vitro via both collagenase digestion method and explant technique, and a comparative study was carried out on the two culture methods. The biologic characteristics of tbs osteoblast cells were studied via cell number counting,morphology observation, alkaline phosphatase staining of the cells and alizarine- red staining of the calcified nodules. The results show that osteoblast cells can be cultured in vitro via collagenase digestion method and explant technique, and the obtained cells are of good biologic characteristics. In comparison with the explant techniqne,the operative procedure of the enzymatic digestion method is more complicated. The digestion time must be carefully controlled. However, with this method, one can obtain a lager number of cells in a short time. The operative procedure of the explant technique is simpler, but it usually takes longer time to obtain cells of desirable number.

  7. The potential of human-derived periodontal ligament stem cells to osteogenic differentiation: An In vitro investigation

    Directory of Open Access Journals (Sweden)

    Behzad Houshmand

    2014-10-01

    Full Text Available Background: Periodontal ligament stem cells (PDLSCs are considered as a type of mesenchymal stem cell that is beneficial target for numerous clinical applications in periodontal tissue regeneration therapy. Materials and Methods: This study examined the effects of dexamethasone (Dex on human PDLSCs in vitro. PDLSCs obtained from the roots of patient’s teeth were cultured with Dex (0.01 μM, and their proliferation was measured. The osteogenic differentiation was assessed by alkaline phosphatase (ALP activity and Alizarin Red-S staining for calcium deposition. Results: After the administration of 0.01 μM Dex, the activity of ALP increased significantly. Furthermore, mineralized nodule formation showing the intracellular calcium deposition was significantly higher in the Dex-treated cells than that of the control cells. Conclusion: Collectively, Dex has positive effects on osteogenic differentiation of human PDLSCs in vitro. It is suggested that PDLSCs may serve as a potential material for periodontal tissue regeneration.

  8. Description of skeletal structure and cranial myology of Cobitis keyvani (Cypriniformes: Cobitidae

    Directory of Open Access Journals (Sweden)

    Pariya Jalili

    2014-12-01

    Full Text Available Cobitis keyvani is recently described from the sourhern Caspian Sea basin. Limited information is available about morphological features of C. keyvani, therefore this study was conducted to provide osteological characteristics and cranial myology of this species. For this purpose, nine specimens of C. keyvani were collected from the Talar River. The specimens were cleared and stained with alizarin red S and alcian blue for osteological examinations. The detailed skeletal structure and cranial muscles of C. keyvani were provided. Based on the results, C. keyvani can be distinguished from other members of the genus Cobitis by a contact between sphenotic and supraoccipital and a contact between pterosphenoid, parasphenoid, prootic and sphenotic in terms of osteological features.

  9. Surface-enhanced Raman scattering in art and archaeology

    Science.gov (United States)

    Leona, Marco

    2005-11-01

    The identification of natural dyes found in archaeological objects and in works of art as textile dyes and lake pigments is a demanding analytical task. To address the problems raised by the very low dye content of dyed fibers and lake pigments, and by the requirement to remove only microscopic samples, surface enhanced Raman scattering techniques were investigated for application to museum objects. SERS gives excellent results with the majority of natural dyes, including: alizarin, purpurin, laccaic acid, carminic acid, kermesic acid, shikonin, juglone, lawsone, brazilin and brazilein, haematoxylin and haematein, fisetin, quercitrin, quercetin, rutin, and morin. In this study, limits of detection were determined for representative dyes and different SERS supports such as citrate reduced Ag colloid and silver nanoisland films. SERS was successfully used to identify natural madder in a microscopic fragment from a severely degraded 11th Century Byzantine textile recently excavated in Amorium, Turkey.

  10. BSA-boronic acid conjugate as lectin mimetics.

    Science.gov (United States)

    Narla, Satya Nandana; Pinnamaneni, Poornima; Nie, Huan; Li, Yu; Sun, Xue-Long

    2014-01-10

    We report bovine serum albumin (BSA)-boronic acid (BA) conjugates as lectin mimetics and their glyco-capturing capacity. The BSA-BA conjugates were synthesized by amidation of carboxylic acid groups in BSA with aminophenyl boronic acid in the presence of EDC, and were characterized by Alizarin Red S (ARS) assay and SDS-PAGE gel. The BSA-BA conjugates were immobilized onto maleimide-functionalized silica beads and their sugar capturing capacity and specificity were confirmed by ARS displacement assay. Further, surface plasmon resonance (SPR) analysis of the glyco-capturing activity of the BSA-BA conjugates was conducted by immobilizing BSA-BA onto SPR gold chip. Overall, we demonstrated a BSA-BA-based lectin mimetics for glyco-capturing applications. These lectin mimetics are expected to provide an important tool for glycomics and biosensor research and applications.

  11. Gallic acid inhibits vascular calcification through the blockade of BMP2-Smad1/5/8 signaling pathway.

    Science.gov (United States)

    Kee, Hae Jin; Cho, Soo-Na; Kim, Gwi Ran; Choi, Sin Young; Ryu, Yuhee; Kim, In Kyeom; Hong, Young Joon; Park, Hyung Wook; Ahn, Youngkeun; Cho, Jeong Gwan; Park, Jong Chun; Jeong, Myung Ho

    2014-11-01

    Vascular calcification is associated with increased risk of morbidity and mortality in patients with cardiovascular diseases, chronic kidney diseases, and diabetes. Gallic acid, a natural compound found in gallnut and green tea, is known to be antifungal, antioxidant, and anticancer. Here we investigated the effect of gallic acid on vascular smooth muscle cell (VSMC) calcification and the underlying mechanism. Gallic acid inhibited inorganic phosphate-induced osteoblast differentiation markers as well as calcification phenotypes (as determined by calcium deposition, Alizarin Red, and Von Kossa staining). Knockdown of BMP2 or Noggin blocked phosphate-induced calcification. Gallic acid suppressed phosphorylation of Smad1/5/8 protein induced by inorganic phosphate. Taken together, we suggest that gallic acid acts as a novel therapeutic agent of vascular calcification by mediating BMP2-Smad1/5/8 signaling pathway.

  12. Bio-green synthesis of Ag-GO, Au-GO and Ag-Au-GO nanocomposites using Azadirachta indica: its application in SERS and cell viability

    Science.gov (United States)

    Hareesh, K.; Williams, J. F.; Dhole, N. A.; Kodam, K. M.; Bhoraskar, V. N.; Dhole, S. D.

    2016-07-01

    Silver-graphene oxide, Gold-graphene oxide, Silver-Gold-graphene oxide nanocomposites were synthesized from Neem leaves (Azadirachta indica) extract using a bio-green one-pot method. The synthesized bio-green nanocomposites were characterized by UV-visible spectroscopy, x-ray diffractogram, transmission electron microscopy, x-ray photoelectron spectroscopy and Raman spectroscopy. The results indicated the decoration of ˜10 nm of silver, ˜20 nm of gold and ˜15 nm of silver-gold nanoparticles on a graphene oxide sheet. The synthesized nanocomposites showed enhancement in surface enhanced Raman spectroscopy with alizarin and also enhancement in cell viability of Chang liver cell lines which may be due to a synergetic effect of nanoparticles and graphene oxide.

  13. Contribution à l'étude de la structure et des propriétés des laques de garance

    OpenAIRE

    Sanyova, Jana

    2001-01-01

    Les laques de garance, les pigments artistiques dont les procédés de fabrication étaient souvent des secrets jalousement gardés, ont depuis longtemps éveillé l'intérêt des chimistes. Le premier mode opératoire de laque de garance décrit scientifiquement est dû à Marggrave en 1754, chimiste allemand célèbre surtout pour la découverte du sucre de betterave. L'élucidation de la structure chimique de l'alizarine par Graebe en 1868 est une des étapes fondatrices de la chimie organique. Il a ensuit...

  14. Hybrid pigments resulting from several guest dyes onto γ-alumina host: A spectroscopic analysis

    Science.gov (United States)

    Pérez, Erik; Ibarra, Ilich A.; Guzmán, Ariel; Lima, Enrique

    2017-02-01

    The synthesis of hybrid pigments was made from combination of γ-Al2O3 and some organic chromophores such as carminic acid, alizarin, purpurin, curcumin, fluorescein and betacyanins. The γ-Al2O3 was obtained through sol-gel synthesis with 2-propanol and aluminium tri-sec-butoxide (ATB). This article presents some spectroscopic evidences related to the formation of aluminium complexes between coordinative unsaturated sites (CUS) of aluminium and some organic groups (carboxylic acid, quaternary ammonium and β-keto enol) present in the chromophores structure. The physicochemical properties upcoming from a spectroscopic analysis point out that these materials can be applied in the design of new materials with potential uses in artworks and in the field of cultural heritage.

  15. Characterization of Japanese color sticks by energy dispersive X-ray fluorescence, X-ray diffraction and Fourier transform infrared analysis

    Energy Technology Data Exchange (ETDEWEB)

    Manso, M. [Centro de Fisica Atomica, Universidade de Lisboa, Faculdade de Ciencias, Av. Prof. Gama Pinto 2, 1649-003 Lisboa (Portugal); Valadas, S. [Chemistry Department, Evora Chemistry Centre and HERCULES Centre, University of Evora, Rua Romao Ramalho, 59 Evora (Portugal); Pessanha, S.; Guilherme, A. [Centro de Fisica Atomica, Universidade de Lisboa, Faculdade de Ciencias, Av. Prof. Gama Pinto 2, 1649-003 Lisboa (Portugal); Queralt, I. [Laboratory of X-ray Analytical Applications, Institute of Earth Sciences ' Jaume Almera' , CSIC, Sole Sabaris s/n. 08028 Barcelona (Spain); Candeias, A.E. [Chemistry Department, Evora Chemistry Centre and HERCULES Centre, University of Evora, Rua Romao Ramalho, 59 Evora (Portugal); Carvalho, M.L., E-mail: luisa@cii.fc.ul.p [Centro de Fisica Atomica, Universidade de Lisboa, Faculdade de Ciencias, Av. Prof. Gama Pinto 2, 1649-003 Lisboa (Portugal)

    2010-04-15

    This work comprises the use of energy dispersive X-ray fluorescence (EDXRF), X-ray diffraction (XRD) and Fourier transformed infrared (FTIR) techniques for the study of the composition of twentieth century traditional Japanese color sticks. By using the combination of analytical techniques it was possible to obtain information on inorganic and organic pigments, binders and fillers present in the sticks. The colorant materials identified in the sticks were zinc and titanium white, chrome yellow, yellow and red ochre, vermillion, alizarin, indigo, Prussian and synthetic ultramarine blue. The results also showed that calcite and barite were used as inorganic mineral fillers while Arabic gum was the medium used. EDXRF offered great potential for such investigations since it allowed the identification of the elements present in the sample preserving its integrity. However, this information alone was not enough to clearly identify some of the materials in study and therefore it was necessary to use XRD and FTIR techniques.

  16. In vitro evaluation of three different biomaterials as scaffolds for canine mesenchymal stem cells

    Directory of Open Access Journals (Sweden)

    Oduvaldo Câmara Marques Pereira-Junior

    2013-05-01

    Full Text Available PURPOSE: To evaluate in vitro ability the of three different biomaterials - purified hydroxyapatite, demineralized bone matrix and castor oil-based polyurethane - as biocompatible 3D scaffolds for canine bone marrow mesenchymal stem cell (MSC intending bone tissue engineering. METHODS: MSCs were isolated from canine bone marrow, characterized and cultivated for seven days with the biomaterials. Cell proliferation and adhesion to the biomaterial surface were evaluated by scanning electron microscopy while differentiation into osteogenic lineage was evaluated by Alizarin Red staining and Sp7/Osterix surface antibody marker. RESULTS: The biomaterials allowed cellular growth, attachment and proliferation. Osteogenic differentiation occurred in the presence of hydroxyapatite, and matrix deposition commenced in the presence of the castor oil-based polyurethane. CONCLUSION: All the tested biomaterials may be used as mesenchymal stem cell scaffolds in cell-based orthopedic reconstructive therapy.

  17. Changes in the corneal Na-K ATPase levels in eyes stored in moist chamber at 4°C

    Directory of Open Access Journals (Sweden)

    Devi B

    1996-01-01

    Full Text Available This report deals with a chronological measurement of Na-K ATPase enzyme activity in human and bovine corneas stored in a moist chamber at 4°C. Paired human and bovine eyes were sterilized by the standard eye bank procedure and stored up to 6 days. At the desired time, the corneal endothelium was assayed for Na-K ATPase activity. The protein content of each tissue sample was also determined. In a parallel set of experiments, the viability of identical stored corneas was determined by trypan blue and alizarin red staining technique, and morphometric analysis was done to quantify the extent of the corneal endothelial damage. The human corneas showed that there was a significant progressive decrease in the Na-K ATPase activity as the storage time increased. The decrease was related to morphological endothelial damage.

  18. Ⅲ型前列腺炎纳米细菌快速检测方法研究及临床评价%Rapid detection and clinical evaluation of nanobacteria infection in type Ⅲ prostatitis

    Institute of Scientific and Technical Information of China (English)

    唐金元; 周占松; 沈学成; 袁宇峰

    2011-01-01

    目的 通过对几种Ⅲ型前列腺炎纳米细菌感染检测方法评价比较,找出一种最快速、可靠的纳米细菌检测方法.方法 以培养后间接免疫荧光鉴定结果为基准对照,分别采用间接免疫荧光法、茜素红钙染色法及PCR法对西南医院泌尿外科门诊150例Ⅲ型前列腺炎患者前列腺液标本进行纳米细菌检测,比较其阳性检出率、灵敏度和特异度差异.结果 评价阳性检出率、灵敏度、特异度:间接免疫荧光法依次为23.3%、42.2%、100.0%;茜素红钙染色法依次为21.3%、34.9%、95.5%;PCR法依次为57.3%、95.2%、89.6%.间接免疫荧光法、茜素红钙染色法检出率与培养后间接免疫荧光法比较,均有统计学差异(P0.05).结论 PCR法检出率优于间接免疫荧光法及茜素红钙染色法,灵敏度及特异度均较高,可广泛应用于Ⅲ型前列腺炎患者纳米细菌感染快速检测.%Objective To discover the rapidest and most reliable method for detecting nanobacteria infection in type Ⅲ prostatitis by comparing several available methods. Methods Expressed prostatic secretion (EPS) specimens were obtained from 150 outpatients suffering from type Ⅲ prostatitis from the Department of Urology, Southwest Hospital, Chongqing, China. Nanobacteria infection in the EPS specimens was detected by indirect immunofluorescence staining (IIFS), alizarin red staining of calcium, and polymerase chain reaction (PCR). With the result of indirect immunofluorescence staining after culture (IIFSC) as a benchmark, the differences of the concerning the positive detection rate, sensitivity and specificity were compared. Results The positive detection rate, sensitivity and specificity of IIFS were 23.3%, 42.2% and 100%, respectively.Those of alizarin red staining of calcium were 21.3%, 34.9% and 95.5%, and those of PCR were 57.3%,95.2% and 89.6%, respectively. The positive detection rate of IIFS or alizarin red calcium staining had a

  19. Identification of anthraquinone coloring matters in natural red dyes by electrospray mass spectrometry coupled to capillary electrophoresis.

    Science.gov (United States)

    Puchalska, Maria; Orlińska, Magdalena; Ackacha, Mohamed A; Połeć-Pawlak, Kasia; Jarosz, Maciej

    2003-12-01

    Capillary electrophoresis with UV/visible diode-array detection (DAD) and electrospray mass spectrometric (ESI-MS) detection were used for the identification of anthraquinone color components of cochineal, lac-dye and madder, natural red dyestuffs often used by ancient painters. For the purpose of such analysis, ESI-MS was found to be a much more appropriate detection technique than DAD one owing to its higher sensitivity (detection limits in the range 0.1-0.5 micro g ml(-1)) and selectivity. The method developed made it possible to identify unequivocally carminic acid and laccaic acids A, B and E as coloring matters in the examined preparations of cochineal and lac-dye, respectively. In madder, European Rubia tinctorum, alizarin and purpurin were found. The method allows the rapid, direct and straightforward identification and quantification of components of natural products used in art and could be very helpful in restoration and conservation procedures.

  20. Imaging and morphology in reproductive toxicology - progress to date and future directions.

    Science.gov (United States)

    French, Julian M

    2014-09-01

    This review looks at the recent development and application of imaging techniques for the morphological examination of fetuses from preclinical regulatory reproductive toxicology studies. Full replacement of the examination methods currently used in routine studies (microdissection, Bouin's fluid fixation/sectioning and alizarin red S/alcian blue preparations) by imaging techniques has yet to be achieved. Progress, especially in the application of micro-CT for skeletal examination, has been made but challenges, particularly the financial investment required, remain. Despite this apparent lack of progress the application of imaging techniques to "non-routine" preclinical reproductive toxicology studies has been used to good effect. The ability to acquire multiple images over a time course i.e. longitudinally has enabled the fate, particularly of skeletal features, to be determined. The additional evidence gained from such studies can be used to better inform the prenatal developmental hazard assessment of test compounds.

  1. Determinação de Mn, Cu e Zn em matrizes salinas após separação e pré-concentração usando Amberlite XAD-7 impregnada com Vermelho de Alizarina S

    Directory of Open Access Journals (Sweden)

    Santos Júnior Aníbal de Freitas

    2002-01-01

    Full Text Available The aim of this work was to explore the possibility of the application of a non-ionic resin obtained by impregnation of Alizarin Red S (VAS in Amberlite XAD-7 for manganese, copper and zinc separation and preconcentration in saline matrices. For these system, the metals were quantitatively retained, in the pH range 8.5-10.0, by using 0.50 g of solid phase, stirring time of five minutes and a total mass up to 200 mug of each cation. The sorbed elements were subsequently eluted and a fifty-fold, ten-fold and ten-fold preconcentration factor for to Zn, Cu and Mn were obtained, respectively.

  2. [Application of AOTF in spectral analysis. 2. Application of self-constructed visible AOTF spectrophotometer].

    Science.gov (United States)

    Peng, Rong-fei; He, Jia-yao; Zhang, Zhan-xia

    2002-02-01

    The performances of a self-constructed visible AOTF spectrophotometer are presented. The wavelength calibration of AOTF1 and AOTF2 are performed with a didymium glass using a fourth-order polynomial curve fitting method. The absolute error of the peak position is usually less than 0.7 nm. Compared with the commercial UV1100 spectrophotometer, the scanning speed of the AOTF spectrophotometer is much more faster, but the resolution depends on the quality of AOTF. The absorption spectra and the calibration curves of copper sulfate and alizarin red obtained with AOTF1(Institute for Silicate, Shanghai China) and AOTF2 (Brimrose U.S.A) respectively are presented. Their corresponding correlation coefficients of the calibration curves are 0.9991 and 0.9990 respectively. Preliminary results show that the self-constructed AOTF spectrophotometer is feasible.

  3. Cultured Human Periosteum-Derived Cells Can Differentiate into Osteoblasts in a Perioxisome Proliferator-Activated Receptor Gamma-Mediated Fashion via Bone Morphogenetic Protein signaling.

    Science.gov (United States)

    Chung, Jin-Eun; Park, Jin-Ho; Yun, Jeong-Won; Kang, Young-Hoon; Park, Bong-Wook; Hwang, Sun-Chul; Cho, Yeong-Cheol; Sung, Iel-Yong; Woo, Dong Kyun; Byun, June-Ho

    2016-01-01

    The differentiation of mesenchymal stem cells towards an osteoblastic fate depends on numerous signaling pathways, including activation of bone morphogenetic protein (BMP) signaling components. Commitment to osteogenesis is associated with activation of osteoblast-related signal transduction, whereas inactivation of this signal transduction favors adipogenesis. BMP signaling also has a critical role in the processes by which mesenchymal stem cells undergo commitment to the adipocyte lineage. In our previous study, we demonstrated that an agonist of the perioxisome proliferator-activated receptor γ (PPARγ), a master regulator of adipocyte differentiation, stimulates osteoblastic differentiation of cultured human periosteum-derived cells. In this study, we used dorsomorphin, a selective small molecule inhibitor of BMP signaling, to investigate whether BMP signaling is involved in the positive effects of PPARγ agonists on osteogenic phenotypes of cultured human periosteum-derived cells. Both histochemical detection and bioactivity of ALP were clearly increased in the periosteum-derived cells treated with the PPARγ agonist at day 10 of culture. Treatment with the PPARγ agonist also caused an increase in alizarin red S staining and calcium content in the periosteum-derived osteoblasts at 2 and 3 weeks of culture. In contrast, dorsomorphin markedly decreased ALP activity, alizarin red S staining and calcium content in both the cells treated with PPARγ agonist and the cells cultured in osteogenic induction media without PPARγ agonist during the culture period. In addition, the PPARγ agonist clearly increased osteogenic differentiation medium-induced BMP-2 upregulation in the periosteum-derived osteoblastic cells at 2 weeks of culture as determined by quantitative reverse transcriptase polymerase chain reaction (RT-PCR), immunoblotting, and immunocytochemical analyses. Although further study will be needed to clarify the mechanisms of PPARγ-regulated osteogenesis

  4. Hyaluronic Acid Gel-Based Scaffolds as Potential Carrier for Growth Factors: An In Vitro Bioassay on Its Osteogenic Potential

    Directory of Open Access Journals (Sweden)

    Masako Fujioka-Kobayashi

    2016-11-01

    Full Text Available Hyaluronic acid (HA has been utilized for a variety of regenerative medical procedures due to its widespread presence in connective tissue and perceived biocompatibility. The aim of the present study was to investigate HA in combination with recombinant human bone morphogenetic protein 9 (rhBMP9, one of the most osteogenic growth factors of the BMP family. HA was first combined with rhBMP9 and assessed for the adsorption and release of rhBMP9 over 10 days by ELISA. Thereafter, ST2 pre-osteoblasts were investigated by comparing (1 control tissue culture plastic, (2 HA alone, and (3 HA with rhBMP9 (100 ng/mL. Cellular proliferation was investigated by a MTS assay at one, three and five days and osteoblast differentiation was investigated by alkaline phosphatase (ALP activity at seven days, alizarin red staining at 14 days and real-time PCR for osteoblast differentiation markers. The results demonstrated that rhBMP9 adsorbed within HA scaffolds and was released over a 10-day period in a controlled manner. While HA and rhBMP9 had little effect on cell proliferation, a marked and pronounced effect was observed for cell differentiation. rhBMP9 significantly induced ALP activity, mRNA levels of collagen1α2, and ALP and osteocalcin (OCN at three or 14 days. HA also demonstrated some ability to induce osteoblast differentiation by increasing mRNA levels of OCN and increasing alizarin red staining at 14 days. In conclusion, the results from the present study demonstrate that (1 HA may serve as a potential carrier for various growth factors, and (2 rhBMP9 is a potent and promising inducer of osteoblast differentiation. Future animal studies are now necessary to investigate this combination approach in vivo.

  5. Dose-dependent Effects of Strontium Ranelate on Ovariectomy Rat Bone Marrow Mesenchymal Stem Cells and Human Umbilical Vein Endothelial Cells

    Science.gov (United States)

    Guo, Xiaojing; Wei, Silong; Lu, Mengmeng; Shao, Zhengwei; Lu, Jiayu; Xia, Lunguo; Lin, Kaili; Zou, Derong

    2016-01-01

    In clinic, strontium ranelate (SrR) is a useful drug to treat osteoporosis by orally taken method, but some side effect appeared in recent years. The aim of this study is to evaluate the effectiveness and safety of SrR on cells by direct application, to study the possibility of local application of this drug. Qualitative ALP staining, quantitative ALP activity assay, alizarin red staining, realtime PCR and westernblot assay were used to evaluate the osteogenesis ability of SrR under normal or osteogenic induction environment of ovariectomy bone marrow mesenchymal stem cells (OVX-BMSCs). The angiogenesis ability of SrR was studied by immunofluorescence staining of CD31 and vWF of OVX-BMSCs under angiogenesis induction environment, transwell, tubeformation and realtime PCR assay of HUVECs. Signaling pathway of PI3K/AKT/mTOR was also studied. The result demonstrated that SrR could enhance proliferation and osteogenic differentiation of OVX-BMSCs. The osteogenesis effect of SrR has been proved by the better performed of ALP activity, alizarin red staining and the remarkable up-regulation of ALP, Col-I, Runx2, OCN, BMP-2, BSP, OPG of the OVX-BMSCs, and reduction of RANKL. In addition, SrR promotes angiogenesis differentiation of both OVX-BMSCs and HUVECs. Higher intensity of immunostaining of CD31 and vWF, better result of transwell and tubeformation assay could be observed in SrR treated group, and increasing mRNA levels of VEGF and Ang-1 in the OVX-BMSCs, VEGF in HUVECs were learnt. Signaling pathway assay showed that PI3K/AKT/mTOR signaling pathway was involved in this SrR triggered angiogenesis procedure. The thrombosis marker ET-1, PAI-1 and t-PA were up-regulated, but no significant differences for low concentration (regeneration. PMID:27994515

  6. Influence of quercetin and nanohydroxyapatite modifications of decellularized goat-lung scaffold for bone regeneration.

    Science.gov (United States)

    Gupta, Sweta K; Kumar, Ritesh; Mishra, Narayan C

    2017-02-01

    In the present study, goat-lung scaffold was fabricated by decellularization of lung tissue and verified for complete cell removal by DNA quantification, DAPI and H&E staining. The scaffold was then modified by crosslinking with quercetin and nanohydroxyapatite (nHAp), and characterized to evaluate the suitability of quercetin-crosslinked nHAp-modified scaffold for regeneration of bone tissue. The crosslinking chemistry between quercetin and decellularized scaffold was established theoretically by AutoDock Vina program (in silico docking study), which predicted multiple intermolecular hydrogen bonding interactions between quercetin and decellularized scaffold, and FTIR spectroscopy analysis also proved the same. From MTT assay and SEM studies, it was found that the quercetin-crosslinked nHAp-modified decellularized scaffold encouraged better growth and proliferation of bone-marrow derived mesenchymal stem cells (BMMSCs) in comparison to unmodified decellularized scaffold, quercetin-crosslinked decellularized scaffold and nHAp-modified decellularized scaffold. Alkaline Phosphatase (ALP) assay results showed highest expression of ALP over quercetin-crosslinked nHAp-modified scaffold among all the tested scaffolds (unmodified decellularized scaffold, quercetin-crosslinked decellularized scaffold and nHAp-modified decellularized scaffold) indicating that quercetin and nHAp is very much efficient in stimulating the differentiation of BMMSCs into osteoblast cells. Alizarin red test quantified in vitro mineralization (calcium deposits), and increased expression of alizarin red over quercetin-crosslinked nHAp-modified scaffold indicating better stimulation of osteogenesis in BMMSCs. The above findings suggest that quercetin-crosslinked nHAp-modified decellularized goat-lung scaffold provides biomimetic bone-like microenvironment for BMMSCs to differentiate into osteoblast and could be applied as a potential promising biomaterial for bone regeneration.

  7. Evaluation of natural anthracene-derived compounds as antimitotic agents.

    Science.gov (United States)

    Badria, Farid A; Ibrahim, Ahmed S

    2013-04-01

    Plants that contain anthracene-derived compounds such as anthraquinones have been reported to act as anticancer besides their use for millennia to treat constipation, but the mechanism of action is still unfolding. Therefore we pursue this study to explore a new horizon in the anticancer property of these agents with relevance to mitotic arrest. To achieve this goal, the antimitotic activity of a series of naturally occurring anthracene-derived anthraquinones including anthrone, alizarin (1,2-dihydroxyanthraquinone), quinizarin (1,4-dihydroxyanthraquinone), rhein (4,5-dihydroxyanthraquinone-2-carboxylic acid), emodin (1,6,8-trihydroxy-3-methylanthraquinone), and aloe emodin (1,8-dihydroxy-3-hydroxymethylanthraquinone) were evaluated using Allium cepa root tips. Initial results revealed that the mitosis was inhibited after 3, 6, and 24 h, respectively, of incubation with 500, 250, and 125 ppm of each compound in a dose-dependent manner. Furthermore, alizarin at 500 ppm was proved to be the most active compound to arrest the mitosis after 24 h followed by emodin, aloe emodin, rhein, and finally quinizarin. Interestingly, this inhibition of mitosis was irreversible in root tips incubated with each compound at concentration of 500 ppm but not with 250 ppm or 125 ppm, where the roots regained their normal mitotic activity after 96 h post-incubation in water. This re-evaluation of an old remedy suggests that several bioactive anthraquinones possess promising anti-mitotic activity that may have the potential to be lead compounds for the development of a new class of multifaceted natural anticancer/antimitotic agents.

  8. Hollow mesoporous raspberry-like colloids with removable caps as photoresponsive nanocontainers

    Science.gov (United States)

    Hu, Chi; West, Kevin R.; Scherman, Oren A.

    2016-04-01

    The fabrication, characterisation and controlled cargo release of hollow mesoporous raspberry-like colloids (HMRCs), which are assembled by utilising host-guest complexation of cucurbit[8]uril (CB[8]) are described. CB[8] is employed as a supramolecular linker to `stick' the viologen functionalised paramagnetic iron oxide nanoparticles onto an azobenzene functionalised hollow mesoporous silica core. The formed HMRCs are photoresponsive and can be reversibly disassembled upon light irradiation, endowing them with an ability to release loaded cargo under photocontrol. While the assembled HMRCs retain cargo inside their cavity, disassembled particles with their iron oxide nanoparticle `caps' removed will release the loaded cargo through the mesoporous shell of the hollow silica colloids. A model system using a boronic acid derivative as the cargo in the HMRCs and Alizarin Red salt as a sensor for the released boronic acid is demonstrated.The fabrication, characterisation and controlled cargo release of hollow mesoporous raspberry-like colloids (HMRCs), which are assembled by utilising host-guest complexation of cucurbit[8]uril (CB[8]) are described. CB[8] is employed as a supramolecular linker to `stick' the viologen functionalised paramagnetic iron oxide nanoparticles onto an azobenzene functionalised hollow mesoporous silica core. The formed HMRCs are photoresponsive and can be reversibly disassembled upon light irradiation, endowing them with an ability to release loaded cargo under photocontrol. While the assembled HMRCs retain cargo inside their cavity, disassembled particles with their iron oxide nanoparticle `caps' removed will release the loaded cargo through the mesoporous shell of the hollow silica colloids. A model system using a boronic acid derivative as the cargo in the HMRCs and Alizarin Red salt as a sensor for the released boronic acid is demonstrated. Electronic supplementary information (ESI) available. See DOI: 10.1039/C6NR01016D

  9. Antimony film sensor for sensitive rare earth metal analysis in environmental samples.

    Science.gov (United States)

    Makombe, Martin; van der Horst, Charlton; Silwana, Bongiwe; Iwuoha, Emmanuel; Somerset, Vernon

    2016-07-02

    A sensor for the adsorptive stripping voltammetric determination of rare earth elements has been developed. The electrochemical procedure is based on the oxidation of the rare earth elements complexed with alizarin complexone at a glassy carbon electrode that was in situ modified with an antimony film, during an anodic scan from -0.2 V to 1.1 V (vs. Ag/AgCl) and deposition potential of -0.1 V (vs. Ag/AgCl). The factors influencing the adsorptive stripping capability were optimised, including the complexing agent concentration, plating concentration of antimony and deposition time. The detection of rare earth elements (La, Ce and Pr) were realised in 0.08 M sodium acetate (pH = 5.8) solution as supporting electrolyte, with 2 × 10(-6) M alizarin complexone and 1.0 mg L(-1) antimony solution. Under the optimised conditions, a deposition time of 360 s was obtained and a linear response was observed between 1 and 25 µg L(-1). The reproducibility of the voltammetric measurements was found to be within 5.0% RSD for 12 replicate measurements of cerium(III) concentration of 5 µg L(-1) using the same electrode surface. The detection limits obtained using stripping analysis was 0.06, 0.42 and 0.71 μg L(-1) for Ce(III), La(III) and Pr(III), respectively. The developed sensor has been successfully applied for the determination of cerium, lanthanum and praseodymium in municipal tap water samples.

  10. Azo dye removal in a membrane-free up-flow biocatalyzed electrolysis reactor coupled with an aerobic bio-contact oxidation reactor

    Energy Technology Data Exchange (ETDEWEB)

    Cui, Dan; Guo, Yu-Qi; Cheng, Hao-Yi; Liang, Bin; Kong, Fan-Ying [State Key Laboratory of Urban Water Resource and Environment, Harbin Institute of Technology, No. 202 Haihe Road, Harbin 150090 (China); Lee, Hyung-Sool [Department of Civil and Environmental Engineering, University of Waterloo, 200 University Avenue West Waterloo, Ontario, Canada N2L 3G1 (Canada); Wang, Ai-Jie, E-mail: waj0578@hit.edu.cn [State Key Laboratory of Urban Water Resource and Environment, Harbin Institute of Technology, No. 202 Haihe Road, Harbin 150090 (China)

    2012-11-15

    Highlights: Black-Right-Pointing-Pointer A membrane-free up-flow biocatalyzed electrolysis reactor coupled with an aerobic bio-contact oxidation reactor was developed. Black-Right-Pointing-Pointer Alizarin Yellow R as the mode of azo dyes was efficiently converted to p-phenylenediamine (PPD) and 5-aminosalicylic acid (5-ASA). Black-Right-Pointing-Pointer PPD and 5-ASA were further oxidized in a bio-contact oxidation reactor. Black-Right-Pointing-Pointer The mechanism of UBER for azo dye removal was discussed. - Abstract: Azo dyes that consist of a large quantity of dye wastewater are toxic and persistent to biodegradation, while they should be removed before being discharged to water body. In this study, Alizarin Yellow R (AYR) as a model azo dye was decolorized in a combined bio-system of membrane-free, continuous up-flow bio-catalyzed electrolysis reactor (UBER) and subsequent aerobic bio-contact oxidation reactor (ABOR). With the supply of external power source 0.5 V in the UBER, AYR decolorization efficiency increased up to 94.8 {+-} 1.5%. Products formation efficiencies of p-phenylenediamine (PPD) and 5-aminosalicylic acid (5-ASA) were above 90% and 60%, respectively. Electron recovery efficiency based on AYR removal in cathode zone was nearly 100% at HRTs longer than 6 h. Relatively high concentration of AYR accumulated at higher AYR loading rates (>780 g m{sup -3} d{sup -1}) likely inhibited acetate oxidation of anode-respiring bacteria on the anode, which decreased current density in the UBER; optimal AYR loading rate for the UBER was 680 g m{sup -3} d{sup -1} (HRT 2.5 h). The subsequent ABOR further improved effluent quality. Overall the Chroma decreased from 320 times to 80 times in the combined bio-system to meet the textile wastewater discharge standard II in China.

  11. Hyaluronic Acid Gel-Based Scaffolds as Potential Carrier for Growth Factors: An In Vitro Bioassay on Its Osteogenic Potential

    Science.gov (United States)

    Fujioka-Kobayashi, Masako; Schaller, Benoit; Kobayashi, Eizaburo; Hernandez, Maria; Zhang, Yufeng; Miron, Richard J.

    2016-01-01

    Hyaluronic acid (HA) has been utilized for a variety of regenerative medical procedures due to its widespread presence in connective tissue and perceived biocompatibility. The aim of the present study was to investigate HA in combination with recombinant human bone morphogenetic protein 9 (rhBMP9), one of the most osteogenic growth factors of the BMP family. HA was first combined with rhBMP9 and assessed for the adsorption and release of rhBMP9 over 10 days by ELISA. Thereafter, ST2 pre-osteoblasts were investigated by comparing (1) control tissue culture plastic, (2) HA alone, and (3) HA with rhBMP9 (100 ng/mL). Cellular proliferation was investigated by a MTS assay at one, three and five days and osteoblast differentiation was investigated by alkaline phosphatase (ALP) activity at seven days, alizarin red staining at 14 days and real-time PCR for osteoblast differentiation markers. The results demonstrated that rhBMP9 adsorbed within HA scaffolds and was released over a 10-day period in a controlled manner. While HA and rhBMP9 had little effect on cell proliferation, a marked and pronounced effect was observed for cell differentiation. rhBMP9 significantly induced ALP activity, mRNA levels of collagen1α2, and ALP and osteocalcin (OCN) at three or 14 days. HA also demonstrated some ability to induce osteoblast differentiation by increasing mRNA levels of OCN and increasing alizarin red staining at 14 days. In conclusion, the results from the present study demonstrate that (1) HA may serve as a potential carrier for various growth factors, and (2) rhBMP9 is a potent and promising inducer of osteoblast differentiation. Future animal studies are now necessary to investigate this combination approach in vivo. PMID:27916889

  12. Effect of biomimetic zinc-containing tricalcium phosphate (Zn-TCP) on the growth and osteogenic differentiation of mesenchymal stem cells.

    Science.gov (United States)

    Chou, Joshua; Hao, Jia; Hatoyama, Hirokazu; Ben-Nissan, Besim; Milthorpe, Bruce; Otsuka, Makoto

    2015-07-01

    Several studies have shown the effectiveness of zinc-tricalcium phosphate (Zn-TCP) for bone tissue engineering. In this study, marine calcareous foraminifera possessing uniform pore size distribution were hydrothermally converted to Zn-TCP. The ability of a scaffold to combine effectively with mesenchymal stem cells (MSCs) is a key tissue-engineering aim. In order to demonstrate the osteogenic ability of MSCs with Zn-TCP, the scaffolds were cultured in an osteogenic induction medium to elicit an osteoblastic response. The physicochemical properties of Zn-TCP were characterized by XRD, FT-IR and ICP-MS. MSCs were aspirated from rat femurs and cultured for 3 days before indirectly placing four samples into each respective well. After culture for 7, 10 and 14 days, osteoblastic differentiation was evaluated using alizarin red S stain, measurement of alkaline phosphatase (ALP) levels, cell numbers and cell viability. XRD and FT-IR patterns both showed the replacement of CO(3)(2-) with PO(4)(3-). Chemical analysis showed zinc incorporation of 5 mol%. Significant increases in cell numbers were observed at 10 and 14 days in the Zn-TCP group, while maintaining high levels of cell viability (> 90%). ALP activity in the Zn-TCP group was statistically higher at 10 days. Alizarin red S staining also showed significantly higher levels of calcium mineralization in Zn-TCP compared with the control groups. This study showed that MSCs in the presence of biomimetically derived Zn-TCP can accelerate their differentiation to osteoblasts and could potentially be useful as a scaffold for bone tissue engineering.

  13. Human dental pulp mesenchymal stem cells isolation and osteoblast differentiation

    Directory of Open Access Journals (Sweden)

    Moustafa Alkhalil

    2015-02-01

    Full Text Available Aim This study was focused on the isolation and characterization of mesenchymal stem cells (MSCs from human dental pulp (DPSC. Methods The study was performed in the Department for Oral and Cranio-Maxillo- Facial Surgey Hamad Medical Corporation, Doha, Qatar and Weill Cornell Medical Colleague Doha, Qatar, in period 2010-2011. Dental pulp was extracted from premolars and third molars of 19 healthy patients. The pulp was digested in a solution of 3 mg/mL collagenase type I and 4 mg/mL dispase for 1 hour at 37C. After filtration, cells were cultured in Dulbecco’s Modified Eagle Medium (DMEM Low Glucoses with 20% Fetal Bovine Serum (FBS, 2mM L-glutamine and antibiotics (100 U/mL penicillin, 100 ug/mL streptomycin at 37 °C under 5% CO2. Cultures were treated with osteoinductive medium for differentiation MSC in to the osteoblast cell line. Staining with Alizarin red were used for the detection of the osteoblast production and calcification new formed tissue. Results On the total of three out of 19 patients it was possible to isolate DPMSCs after 2 to 3 weeks: in one patient it was not possible to expand MSCs because of infection, and in other two patients positive Alizarin red staining reaction showed osteogenic differentiation capability and strong mineralization in vitro. Conclusion The main advantage of using DPSC is absence of morbidity. MSCs could be isolated noninvasively from teeth, routinely extracted in the clinic and discarded as medical waste. Standardization of clinical and laboratory protocols for DPMSCs isolation and team work coordination could lead to significantly improved result.

  14. Spatiotemporal Progression of Microcalcification in the Hippocampal CA1 Region following Transient Forebrain Ischemia in Rats: An Ultrastructural Study.

    Directory of Open Access Journals (Sweden)

    Tae-Ryong Riew

    Full Text Available Calcification in areas of neuronal degeneration is a common finding in several neuropathological disorders including ischemic insults. Here, we performed a detailed examination of the onset and spatiotemporal profile of calcification in the CA1 region of the hippocampus, where neuronal death has been observed after transient forebrain ischemia. Histopathological examinations showed very little alizarin red staining in the CA1 pyramidal cell layer until day 28 after reperfusion, while prominent alizarin red staining was detected in CA1 dendritic subfields, particularly in the stratum radiatum, by 14 days after reperfusion. Electron microscopy using the osmium/potassium dichromate method and electron probe microanalysis revealed selective calcium deposits within the mitochondria of degenerating dendrites at as early as 7 days after reperfusion, with subsequent complete mineralization occurring throughout the dendrites, which then coalesced to form larger mineral conglomerates with the adjacent calcifying neurites by 14 days after reperfusion. Large calcifying deposits were frequently observed at 28 days after reperfusion, when they were closely associated with or completely engulfed by astrocytes. In contrast, no prominent calcification was observed in the somata of CA1 pyramidal neurons showing the characteristic features of necrotic cell death after ischemia, although what appeared to be calcified mitochondria were noted in some degenerated neurons that became dark and condensed. Thus, our data indicate that intrahippocampal calcification after ischemic insults initially occurs within the mitochondria of degenerating dendrites, which leads to the extensive calcification that is associated with ischemic injuries. These findings suggest that in degenerating neurons, the calcified mitochondria in the dendrites, rather than in the somata, may serve as the nidus for further calcium precipitation in the ischemic hippocampus.

  15. 人绒毛膜来源的间充质干细胞成骨成脂分化潜能%Osteogenic and Adipogenic Differentiation Potential of Human Chorion-derived Mesenchymal Stem Cells

    Institute of Scientific and Technical Information of China (English)

    陈霞; 尹晓娟

    2011-01-01

    目的:探讨人绒毛膜来源的间充质干细胞(hCDMSC)体外生长特性和成骨成脂分化潜能,证实人绒毛膜来源的间充质干细胞作为组织工程种子细胞的可行性.方法:取胎盘组织基蜕膜面用胶原酶和胰蛋白酶法分离培养,通过传代扩增观察细胞形态,MTT法检测细胞增殖曲线,体外向成骨、成脂诱导分化,茜素红和油红O染色鉴定细胞分化能力,RT-PCR测定骨细胞和脂肪细胞特异性基因表达.结果:hCDMSC细胞具有间充质干细胞特性,茜素红和油红O染色呈阳性反应,成骨和成脂标志性基因表达阳性.结论:人绒毛膜来源的间充质干细胞在体外诱导条件下可以向成骨细胞和脂肪细胞诱导分化,可以作为组织工程的种子细胞.%Objective: To study the growth characteristics and the potency of osteogenic and adipogenic differentiation of human chorion-derived mesenchymal stem cells ( hCDMS) in vitro, to validate whether hCDMS could be used in tissue engineering. Methods: The hCDMSC were isolated from placental basal deciduas by collagens and trypsinase digestion methods. After serial sub cultivation in vitro, the stem cells were introduced. Morphologic appearance of hCDMS was observed and the proliferation rate was measured by MTT assay. The osteogenic potential was evaluated by alizarin red staining, otherwise the adipogenic potential by oil red 0 staining. Then the adipogenic and osteogenic specific markers of differentiated cells assayed by RT-PCR method. Results: hCDMSC possed characteristics of mesenchymal stem cells. The alizarin red staining and oil red 0 staining results were positive. Under induction, cells expressed osteogenic and adipogenic marker genes. Conclusions: hCDMSC can be induced to differentiate into osteoblastes and lipoblastes in appropriate condition in vitro, so it can be used as a cell source in tissue engineering.

  16. Platelet-rich concentrate in serum free medium enhances osteogenic differentiation of bone marrow-derived human mesenchymal stromal cells

    Science.gov (United States)

    Ramasamy, Thamil Selvee; Karunanithi, Puvanan; Naveen, Sangeetha Vasudevaraj; Murali, Malliga Raman; Abbas, Azlina A.; Kamarul, Tunku

    2016-01-01

    Previous studies have shown that platelet concentrates used in conjunction with appropriate growth media enhance osteogenic differentiation of human mesenchymal stromal cells (hMSCs). However, their potential in inducing osteogenesis of hMSCs when cultured in serum free medium has not been explored. Furthermore, the resulting osteogenic molecular signatures of the hMSCs have not been compared to standard osteogenic medium. We studied the effect of infrequent supplementation (8-day interval) of 15% non-activated platelet-rich concentrate (PRC) in serum free medium on hMSCs proliferation and differentiation throughout a course of 24 days, and compared the effect with those cultured in a standard osteogenic medium (OM). Cell proliferation was analyzed by alamar blue assay. Gene expression of osteogenic markers (Runx2, Collagen1, Alkaline Phosphatase, Bone morphogenetic protein 2, Osteopontin, Osteocalcin, Osteonectin) were analyzed using Q-PCR. Immunocytochemical staining for osteocalcin, osteopontin and transcription factor Runx2 were done at 8, 16 and 24 days. Biochemical assays for the expression of ALP and osteocalcin were also performed at these time-points. Osteogenic differentiation was further confirmed qualitatively by Alizarin Red S staining that was quantified using cetylpyridinium chloride. Results showed that PRC supplemented in serum free medium enhanced hMSC proliferation, which peaked at day 16. The temporal pattern of gene expression of hMSCs under the influence of PRC was comparable to that of the osteogenic media, but at a greater extent at specific time points. Immunocytochemical staining revealed stronger staining for Runx2 in the PRC-treated group compared to OM, while the staining for Osteocalcin and Osteopontin were comparable in both groups. ALP activity and Osteocalcin/DNA level were higher in the PRC group. Cells in the PRC group had similar level of bone mineralization as those cultured in OM, as reflected by the intensity of Alizarin red

  17. 新生大鼠下颌骨成骨细胞的分离、培养与鉴定%The isolation, culture and identification of osteoblasts from neonatal rat mandible

    Institute of Scientific and Technical Information of China (English)

    张巍; 史册; 倪世磊; 李琛; 乔春燕; 孙宏晨; 李德超

    2011-01-01

    Objective To explore an efficient, economical and convenient method for isolating neonatal rat mandibular osteoblasts. Methods Under sterile conditions,the mandibles were taken from neonatal rats which were bom within 24 hours. Tissue block adherent method was used. ALP staining and alizarin red staining were used for identification. The mandibular osteoblasts were compared to the osteoblasts of skull. Results The cultured cells exhibited the typical morphological characteristics of osteoblasts. ALP staining was positive and alizarin red staining showed the formation of mineralized nudules. These cells had the similar shape and function with the osteoblasts of skulL Conclusion Osteoblasts of neonatal rat mandible were successfully isolated. This laid the foundation for in vitro study of osteoblasts of mandible.%目的 探求高效、经济、方便的新生大鼠下颌骨成骨细胞的原代培养方法,为研究下颌骨成骨细胞的相关特性奠定基础.方法 无菌条件下取新生24h大鼠的下颌骨,采用组织块贴壁法培养成骨细胞,通过细胞形态学观察、碱性磷酸酶染色(alkaline phosphatase,ALP)、茜素红染色对细胞进行鉴定,并与颅骨分离的成骨细胞进行对比.结果 所分离培养的细胞通过倒置相差显微镜观察呈梭型,碱性磷酸酶染色阳性,能形成钙结节,与颅骨分离的成骨细胞有相似的形态和相同的功能.结论 采用组织块贴壁法成功分离出大量纯化的新生大鼠下颌骨成骨细胞,为体外研究下颌骨成骨细胞的特性奠定了实验基础.

  18. 蒙古马脂肪来源间充质干细胞体外成脂和成骨诱导分化%Differentiation of Mongolia Horse Adipose Tissue-Derived Mesenchymal Stem Cells into Adipocytes and Osteoblasts in Vitro

    Institute of Scientific and Technical Information of China (English)

    刘宗正; 韦林盖; 苏小虎; 张焱如; 芒来

    2011-01-01

    To investigate the multilineage differentiation capacity of mesenchymal stem cells isolated and cultured from equine adipose tissue, adipose tissue-derived mesenchymal stem cells ( ADSCs) were obtained from adipose tissue of Mongolia horse. The cells appeared like fibroblast in the culture medium. Adipose tissue was minced and digested with collagenase type I. The obtained cells were plated and expanded in DMEM /F12 medium. Whereas the passage cells were cultured in adipogenisis medium and stained with Oil Red 0 for identification. The cells were cultivated in osteoblast-inducing culture medium , and osteoblast phenotype was assayed with Alizarin Red staining. The cells were daily observed under inverted microscope. Results indicated that ADSCs grew as adherent cells, appeared like fibroblast in vitro, stably proliferate and passed. Under the inverted microscope, significant lipid drops were found a-round the cell nucleus after adipogenisis-inducing cultivation. Alizarin Red staining resulted in the formation of mineralized nods in extracellular matrix. It proved that ADSCs isolated and cultured from equine adipose tissue can be induced to adipogenisis and osteo-inducing, suggesting that the cells have multilineage differentiation.%取蒙古马背臀部皮下脂肪组织,通过Ⅰ型胶原酶消化、离心等步骤分离培养脂肪组织来源的间充质干细胞(Adipose tissue-derived mesenchymal stem cells,ADSCs),经过原代培养和传代培养,分别加入成脂诱导剂和成骨诱导剂培养,采用倒置显微镜观察诱导后的细胞形态变化,并通过油红O染色和茜素红染色法对其脂肪细胞和成骨细胞表型进行鉴定.结果显示:ADSCs呈成纤维细胞样贴壁生长,其经成脂、成骨诱导培养2周后形态、体积发生明显改变.经油红O染色,细胞质内出现橙红色脂滴;茜素红染色表明聚集的细胞团中央能形成钙化结节.说明马ADSCs经体外诱导培养后可向脂肪细胞和成骨细胞

  19. Cytotoxicity and DNA cleavage with core-shell nanocomposites functionalized by a KH domain DNA binding peptide

    Science.gov (United States)

    Bazak, Remon; Ressl, Jan; Raha, Sumita; Doty, Caroline; Liu, William; Wanzer, Beau; Salam, Seddik Abdel; Elwany, Samy; Paunesku, Tatjana; Woloschak, Gayle E.

    2013-11-01

    A nanoconjugate was composed of metal oxide nanoparticles decorated with peptides and fluorescent dye and tested for DNA cleavage following UV light activation. The peptide design was based on a DNA binding domain, the so called KH domain of the hnRNPK protein. This ``KH peptide'' enabled cellular uptake of nanoconjugates and their entry into cell nuclei. The control nanoconjugate carried no peptide; it consisted only of the metal oxide nanoparticle prepared as Fe3O4@TiO2 nanocomposite and the fluorescent dye alizarin red S. These components of either construct are responsible for nanoconjugate activation by UV light and the resultant production of reactive oxygen species (ROS). Production of ROS at different subcellular locations causes damage to different components of cells: only nanoconjugates inside cell nuclei can be expected to cause DNA cleavage. Degradation of cellular DNA with KH peptide decorated nanoconjugates exceeded the DNA damage obtained from control, no-peptide nanoconjugate counterparts. Moreover, caspase activation and cell death were more extensive in the same cells.A nanoconjugate was composed of metal oxide nanoparticles decorated with peptides and fluorescent dye and tested for DNA cleavage following UV light activation. The peptide design was based on a DNA binding domain, the so called KH domain of the hnRNPK protein. This ``KH peptide'' enabled cellular uptake of nanoconjugates and their entry into cell nuclei. The control nanoconjugate carried no peptide; it consisted only of the metal oxide nanoparticle prepared as Fe3O4@TiO2 nanocomposite and the fluorescent dye alizarin red S. These components of either construct are responsible for nanoconjugate activation by UV light and the resultant production of reactive oxygen species (ROS). Production of ROS at different subcellular locations causes damage to different components of cells: only nanoconjugates inside cell nuclei can be expected to cause DNA cleavage. Degradation of cellular DNA

  20. Mild extraction methods using aqueous glucose solution for the analysis of natural dyes in textile artefacts dyed with Dyer's madder (Rubia tinctorum L.).

    Science.gov (United States)

    Ford, Lauren; Henderson, Robert L; Rayner, Christopher M; Blackburn, Richard S

    2017-03-03

    Madder (Rubia tinctorum L.) has been widely used as a red dye throughout history. Acid-sensitive colorants present in madder, such as glycosides (lucidin primeveroside, ruberythric acid, galiosin) and sensitive aglycons (lucidin), are degraded in the textile back extraction process; in previous literature these sensitive molecules are either absent or present in only low concentrations due to the use of acid in typical textile back extraction processes. Anthraquinone aglycons alizarin and purpurin are usually identified in analysis following harsh back extraction methods, such those using solvent mixtures with concentrated hydrochloric acid at high temperatures. Use of softer extraction techniques potentially allows for dye components present in madder to be extracted without degradation, which can potentially provide more information about the original dye profile, which varies significantly between madder varieties, species and dyeing technique. Herein, a softer extraction method involving aqueous glucose solution was developed and compared to other back extraction techniques on wool dyed with root extract from different varieties of Rubia tinctorum. Efficiencies of the extraction methods were analysed by HPLC coupled with diode array detection. Acidic literature methods were evaluated and they generally caused hydrolysis and degradation of the dye components, with alizarin, lucidin, and purpurin being the main compounds extracted. In contrast, extraction in aqueous glucose solution provides a highly effective method for extraction of madder dyed wool and is shown to efficiently extract lucidin primeveroside and ruberythric acid without causing hydrolysis and also extract aglycons that are present due to hydrolysis during processing of the plant material. Glucose solution is a favourable extraction medium due to its ability to form extensive hydrogen bonding with glycosides present in madder, and displace them from the fibre. This new glucose method offers an

  1. The impact of glucocorticoid on proliferation and differentiation of bone marrow mesenchymal stem cells by regulating DKK1

    Directory of Open Access Journals (Sweden)

    Jie YANG

    2012-04-01

    Full Text Available Objective To observe the influence of different concentrations of glucocorticoid on the expression of DKK1 gene and the effect of DKK1 on the proliferation and differentiation of bone marrow mesenchymal stem cells (BMSCs. Methods The experiment consisted three groups, namely, control group (no Dex, Dex8 group (10-8mol/L Dex, and Dex6 group (10-6mol/L Dex. Culture fluids with different concentrations of Dex, including 0, 10-8, and 10-6mol/L, were used to stimulate BMSCs. Realtime PCR was used to detect the expression of the DKK1 gene, and clone formation experiment was used to detect the influence on the proliferation of BMSCs. Osteogenic- and adipogenic-induced media were used which contained 0, 10-8, and 10-6mol/L Dex, respectively, to stimulate BMSCs, and to extract RNA after 3d. Real-time PCR was used to detect the expression of DKK1 genes, osteogenic genes (Runx2 and OCN, and adipogenic genes (C/EBP and PPARγ. After three weeks, Alizarin red staining and Oil red O staining were adopted to detect the influence on osteogenic and adipogenic differentiations. Results The expression of DKK1 was the highest, whereas the corresponding cloning formation ability was the lowest, after the stimulation of glucocorticoid with high concentration. The expressions of Runx2 and OCN under osteogenic induction in Dex8 group were the highest, whereas the expression of DKK1 was the lowest. Based on Alizarin red staining, calcified cortical tubers were only found in Dex8 group. The expressions of C/EBP and PPARγ under adipogenic induction in Dex6 group were the highest, so was the expression of DKK1. With Oil red O staining, there were abundant lipid drops in Dex6 group. Conclusion The expression of DKK1 was upregulated under high concentration of glucocorticoid, but it inhibited the proliferation of BMSCs. DKK1 inhibited osteogenic differentiation but promoted adipogenic differentiation of BMSCs. Osteoporosis after glucocorticoid

  2. Diferenciação in vitro de células-tronco mesenquimais da medula óssea de cães em precursores osteogênicos

    Directory of Open Access Journals (Sweden)

    Sílvia A.F. Lima

    2012-05-01

    Full Text Available O objetivo principal da nossa pesquisa foi avaliar o potencial de diferenciação osteogênica de células-tronco mesenquimais (MSC obtidas da medula óssea do cão. As MSC foram separadas pelo método Ficoll e cultivadas sob duas condições distintas: DMEM baixa glicose ou DMEM/F12, ambos contendo L-glutamina, 20% de SFB e antibióticos. Marcadores de MSC foram testados, confirmando células CD44+ e CD34- através da citometria de fluxo. Para a diferenciação osteogênica, as células foram submetidas a quatro diferentes condições: Grupo 1, as mesmas condições utilizadas para a cultura de células primárias com os meios DMEM baixa glicose suplementado; Grupo 2, as mesmas condições do Grupo 1, mais os indutores de diferenciação dexametasona, ácido ascórbico e b-glicerolfosfato; Grupo 3, células cultivadas com meios DMEM/F12 suplementado; e Grupo 4, nas mesmas condições que no Grupo 3, mais indutores de diferenciação de dexametasona, ácido ascórbico e b-glicerolfosfato. A diferenciação celular foi confirmada através da coloração com alizarin red e da imunomarcação com o anticorpo SP7/Osterix. Nós observamos através da coloração com alizarin red que o depósito de cálcio foi mais evidente nas células cultivadas em DMEM/F12. Além disso, usando a imunomarcação com o anticorpo SP/7Osterix obtivemos positividade em 1:6 células para o Meio DMEM/F12 comparada com 1:12 para o meio DMEM-baixa glicose. Com base nos nossos resultados concluímos que o meio DMEM/F12 é mais eficiente para a indução da diferenciação de células-tronco mesenquimais caninas em promotores osteogênicos. Este efeito provavelmente ocorre em decorrência da maior quantidade de glicose neste meio, bem como da presença de diversos aminoácidos.

  3. Sesamin stimulates osteoblast differentiation through p38 and ERK1/2 MAPK signaling pathways

    Directory of Open Access Journals (Sweden)

    Wanachewin Orawan

    2012-05-01

    Full Text Available Abstract Background Osteoporosis is a worldwide health problem predominantly affecting post-menopausal women. Therapies aimed at increasing bone mass in osteoporetic patients lag behind comparable investigation of therapeutic strategies focusing on the bone resorption process. Sesamin, a major lignan compound found in Sesamun indicum Linn., has a variety of pharmacological effects, though its activity on bone cell function is unclear. Herein we examine the effect of this lignan on osteoblast differentiation and function. Method Cell cytotoxicity and proliferative in hFOB1.19 were examined by MTT and alamar blue assay up to 96 h of treatment. Gene expression of COL1, ALP, BMP-2, Runx2, OC, RANKL and OPG were detected after 24 h of sesamin treatment. ALP activity was measured at day 7, 14 and 21 of cultured. For mineralized assay, ADSCs were cultured in the presence of osteogenic media supplement with or without sesamin for 21 days and then stained with Alizarin Red S. MAPK signaling pathway activation was observed by using western blotting. Results Sesamin promoted the gene expression of COL1, ALP, OCN, BMP-2 and Runx2 in hFOB1.19. On the other hand, sesamin was able to up-regulate OPG and down-regulate RANKL gene expression. ALP activity also significantly increased after sesamin treatment. Interestingly, sesamin induced formation of mineralized nodules in adipose derived stem cells (ADSCs as observed by Alizarin Red S staining; this implies that sesamin has anabolic effects both on progenitor and committed cell stages of osteoblasts. Western blotting data showed that sesamin activated phosphorylation of p38 and ERK1/2 in hFOB1.19. Conclusions The data suggest that sesamin has the ability to trigger osteoblast differentiation by activation of the p38 and ERK MAPK signaling pathway and possibly indirectly regulate osteoclast development via the expression of OPG and RANKL in osteoblasts. Therefore, sesamin may be a promising phytochemical

  4. Platelet-rich concentrate in serum free medium enhances osteogenic differentiation of bone marrow-derived human mesenchymal stromal cells

    Directory of Open Access Journals (Sweden)

    Shani Samuel

    2016-09-01

    Full Text Available Previous studies have shown that platelet concentrates used in conjunction with appropriate growth media enhance osteogenic differentiation of human mesenchymal stromal cells (hMSCs. However, their potential in inducing osteogenesis of hMSCs when cultured in serum free medium has not been explored. Furthermore, the resulting osteogenic molecular signatures of the hMSCs have not been compared to standard osteogenic medium. We studied the effect of infrequent supplementation (8-day interval of 15% non-activated platelet-rich concentrate (PRC in serum free medium on hMSCs proliferation and differentiation throughout a course of 24 days, and compared the effect with those cultured in a standard osteogenic medium (OM. Cell proliferation was analyzed by alamar blue assay. Gene expression of osteogenic markers (Runx2, Collagen1, Alkaline Phosphatase, Bone morphogenetic protein 2, Osteopontin, Osteocalcin, Osteonectin were analyzed using Q-PCR. Immunocytochemical staining for osteocalcin, osteopontin and transcription factor Runx2 were done at 8, 16 and 24 days. Biochemical assays for the expression of ALP and osteocalcin were also performed at these time-points. Osteogenic differentiation was further confirmed qualitatively by Alizarin Red S staining that was quantified using cetylpyridinium chloride. Results showed that PRC supplemented in serum free medium enhanced hMSC proliferation, which peaked at day 16. The temporal pattern of gene expression of hMSCs under the influence of PRC was comparable to that of the osteogenic media, but at a greater extent at specific time points. Immunocytochemical staining revealed stronger staining for Runx2 in the PRC-treated group compared to OM, while the staining for Osteocalcin and Osteopontin were comparable in both groups. ALP activity and Osteocalcin/DNA level were higher in the PRC group. Cells in the PRC group had similar level of bone mineralization as those cultured in OM, as reflected by the intensity of

  5. Azanitrile Cathepsin K Inhibitors: Effects on Cell Toxicity, Osteoblast-Induced Mineralization and Osteoclast-Mediated Bone Resorption.

    Directory of Open Access Journals (Sweden)

    Zhong-Yuan Ren

    Full Text Available The cysteine protease cathepsin K (CatK, abundantly expressed in osteoclasts, is responsible for the degradation of bone matrix proteins, including collagen type 1. Thus, CatK is an attractive target for new anti-resorptive osteoporosis therapies, but the wider effects of CatK inhibitors on bone cells also need to be evaluated to assess their effects on bone. Therefore, we selected, among a series of synthetized isothiosemicarbazides, two molecules which are highly selective CatK inhibitors (CKIs to test their effects on osteoblasts and osteoclasts.Cell viability upon treatment of CKIs were was assayed on human osteoblast-like Saos-2, mouse monocyte cell line RAW 264.7 and mature mouse osteoclasts differentiated from bone marrow. Osteoblast-induced mineralization in Saos-2 cells and in mouse primary osteoblasts from calvaria, with or without CKIs,; were was monitored by Alizarin Red staining and alkaline phosphatase activity, while osteoclast-induced bone resorption was performed on bovine slices.Treatments with two CKIs, CKI-8 and CKI-13 in human osteoblast-like Saos-2, murine RAW 264.7 macrophages stimulated with RANKL and mouse osteoclasts differentiated from bone marrow stimulated with RANKL and MCSF were found not to be toxic at doses of up to 100 nM. As probed by Alizarin Red staining, CKI-8 did not inhibit osteoblast-induced mineralization in mouse primary osteoblasts as well as in osteoblast-like Saos-2 cells. However, CKI-13 led to a reduction in mineralization of around 40% at 10-100 nM concentrations in osteoblast-like Saos-2 cells while it did not in primary cells. After a 48-hour incubation, both CKI-8 and CKI-13 decreased bone resorption on bovine bone slices. CKI-13 was more efficient than the commercial inhibitor E-64 in inhibiting bone resorption induced by osteoclasts on bovine bone slices. Both CKI-8 and CKI-13 created smaller bone resorption pits on bovine bone slices, suggesting that the mobility of osteoclasts was slowed

  6. In vitro evaluation of osteoblast responses to carbon nanotube-coated titanium surfaces

    Directory of Open Access Journals (Sweden)

    K. Subramani

    2016-07-01

    Full Text Available Abstract Background The effects of surface roughness and carboxyl functionalization of multi-walled carbon nanotubes (MWCNTs mixed with collagen coated onto titanium (Ti substrates on MC3T3-E1 osteoblasts were evaluated. Methods The proliferation, differentiation, and matrix mineralization were investigated using (1 smooth-surfaced Ti discs, (2 Ti discs coated with collagen and MWCNT (Ti-MWCNT, and (3 Ti discs coated with collagen and MWCNT-COOH (Ti-MWCNT-COOH for applications in orthodontic mini screw implants (MSIs. The coatings were uniform when analyzed using scanning electron microscopy (SEM, and surface roughness was evaluated by surface profilometry that demonstrated similar surface roughness (R a , mean ± SD in the MWCNT (0.83 ± 0.02 μm and MWCNT-COOH (0.84 ± 0.01 μm groups. MTT (3-(4,5-dimethylthiazol-2-Yl-2,5-diphenyltetrazolium bromide assay was performed after days 1, 3, and 7 to assess proliferation. Alkaline phosphatase (ALP-specific activity was assessed after day 7 to quantify differentiation. Alizarin red staining was measured after day 28 to quantify matrix mineralization. All data were analyzed with JMP Pro11 software (SAS, USA with a statistical significance of p  < 0.05. Results Surface profilometry demonstrated similar surface roughness (R a , mean ± SD in the MWCNT (0.83 ± 0.02 μm and MWCNT-COOH (0.84 ± 0.01 μm groups. On day 7, ALP assay showed that MWCNT-COOH (mean ± SD 0.98 ± 0.26 U/μg of protein enhanced cell differentiation when compared to the uncoated group (p = 0.05. Alizarin red staining after 28 days of cell culture revealed that MWCNT-COOH (mean ± SD 1.5 ± 0.2 OD405 increased (p = 0.03 matrix mineralization when compared to the uncoated group (0.9 ± 0.09 OD405. Conclusions This study showed that coatings containing MWCNT-COOH (increased hydrophilic surface chemistry influence osteoblast proliferation, differentiation, and matrix

  7. Establishment and Characterization of New Canine and Feline Osteosarcoma Primary Cell Lines

    Directory of Open Access Journals (Sweden)

    Florian R. L. Meyer

    2016-06-01

    Full Text Available Osteosarcomas are the most abundant form of bone malignancies in multiple species. Canine osteosarcomas are considered a valuable model for human osteosarcomas because of their similar features. Feline osteosarcomas, on the other hand, are rarely studied but have interesting characteristics, such as a better survival prognosis than dogs or humans, and less likelihood of metastasis. To enable experimental approaches to study these differences we have established five new canine osteosarcoma cell lines out of three tumors, COS_1186h, COS_1186w, COS_1189, and COS_1220, one osteosarcoma-derived lung metastasis, COS_1033, and two new feline osteosarcoma cell lines, FOS_1077 and FOS_1140. Their osteogenic and neoplastic origin, as well as their potential to produce calcified structures, was determined by the markers osteocalcin, osteonectin, tissue unspecific alkaline phosphatase, p53, cytokeratin, vimentin, and alizarin red. The newly developed cell lines retained most of their markers in vitro but only spontaneously formed spheroids produced by COS_1189 showed calcification in vitro.

  8. Translationally controlled tumor protein supplemented chitosan modified glass ionomer cement promotes osteoblast proliferation and function.

    Science.gov (United States)

    Sangsuwan, Jiraporn; Wanichpakorn, Supreya; Kedjarune-Leggat, Ureporn

    2015-09-01

    The objective of this study was to evaluate the effect of translationally controlled tumor protein (TCTP) supplemented in a novel glass ionomer cement (BIO-GIC) on normal human osteoblasts (NHost cells). BIO-GIC was a glass ionomer cement (GIC) modified by adding chitosan and albumin to promote the release of TCTP. NHost cells were seeded on specimens of GIC, GIC+TCTP, BIO-GIC and BIO-GIC+TCTP. Cell proliferation was determined by BrdU assay. It was found that BIO-GIC+TCTP had significantly higher proliferation of cells than other specimens. Bone morphogenetic protein-2 (BMP-2) and osteopontin (OPN) gene expressions assessed by quantitative real time PCR and alkaline phosphatase (ALP) activity were used to determine cell differentiation. Bone cell function was investigated by calcium deposition using alizarin assay. Both BMP-2 and OPN gene expressions of cells cultured on specimens with added TCTP increased gradually up-regulation after day 1 and reached the highest on day 3 then down-regulation on day 7. The ALP activity of cells cultured on BIO-GIC+TCTP for 7 days and calcium content after 14 days were significantly higher than other groups. BIO-GIC+TCTP can promote osteoblast cells proliferation, differentiation and function.

  9. Biomolecule-assisted synthesis of In(OH)3 nanocubes and In2O3 nanoparticles: photocatalytic degradation of organic contaminants and CO oxidation

    Science.gov (United States)

    Nayak, Arpan Kumar; Lee, Seungwon; Sohn, Youngku; Pradhan, Debabrata

    2015-12-01

    The synthesis of nanostructured materials without any hazardous organic chemicals and expensive capping reagents is one of the challenges in nanotechnology. Here we report on the L-arginine (a biomolecule)-assisted synthesis of single crystalline cubic In(OH)3 nanocubes of a size in the range of 30-60 nm along the diagonal using hydrothermal methods. Upon calcining at 750 °C for 1 h in air, In(OH)3 nanocubes are transformed into In2O3 nanoparticles (NPs) with voids. The morphology transformation and formation of voids with the increase of the calcination temperature is studied in detail. The possible mechanism of the voids’ formation is discussed on the basis of the Kirkendall effect. The photocatalytic properties of In(OH)3 nanocubes and In2O3 NPs are studied for the degradation of rhodamin B and alizarin red S. Furthermore, the CO oxidation activity of In(OH)3 nanocubes and In2O3 NPs is examined. The photocatalytic and CO oxidation activity are measured to be higher for In2O3 NPs than for In(OH)3 nanocubes. This is attributed to the lower energy gap and higher specific surface area of the former. The present green synthesis has potential for the synthesis of other inorganic nanomaterials.

  10. Effect of Emdogain enamel matrix derivative and BMP-2 on the gene expression and mineralized nodule formation of alveolar bone proper-derived stem/progenitor cells.

    Science.gov (United States)

    Fawzy El-Sayed, Karim M; Dörfer, Christof; Ungefroren, Hendrick; Kassem, Neemat; Wiltfang, Jörg; Paris, Sebastian

    2014-07-01

    The objective of this study was to evaluate the effect of Emdogain (Enamel Matrix Derivative, EMD) and Bone Morphogenetic Protein-2 (BMP-2), either solely or in combination, on the gene expression and mineralized nodule formation of alveolar bone proper-derived stem/progenitor cells. Stem/progenitor cells were isolated from human alveolar bone proper, magnetically sorted using STRO-1 antibodies, characterized flowcytometrically for their surface markers' expression, and examined for colony formation and multilineage differentiation potential. Subsequently, cells were treated over three weeks with 100 μg/ml Emdogain (EMD-Group), or 100 ng/ml BMP-2 (BMP-Group), or a combination of 100 ng/ml BMP-2 and 100 μg/ml Emdogain (BMP/EMD-Group). Unstimulated stem/progenitor cells (MACS(+)-Group) and osteoblasts (OB-Group) served as controls. Osteogenic gene expression was analyzed using RTq-PCR after 1, 2 and 3 weeks (N = 3/group). Mineralized nodule formation was evaluated by Alizarin-Red staining. BMP and EMD up-regulated the osteogenic gene expression. The BMP Group showed significantly higher expression of Collagen-I, III, and V, Alkaline phosphatase and Osteonectin compared to MACS(+)- and OB-Group (p Emdogain and BMP-2 up-regulate the osteogenic gene expression of stem/progenitor cells. The combination of BMP-2 and Emdogain showed no additive effect and would not be recommended for a combined clinical stimulation.

  11. Greek plant extracts exhibit selective estrogen receptor modulator (SERM)-like properties.

    Science.gov (United States)

    Kassi, Eva; Papoutsi, Zoi; Fokialakis, Nikolaos; Messari, Ioanna; Mitakou, Sophia; Moutsatsou, Paraskevi

    2004-11-17

    To prevent bone loss that occurs with increasing age, nutritional and pharmacological factors are needed. Traditional therapeutic agents (selective estrogen receptor modulators or SERMs, biphosphonates, calcitonin) may have serious side effects or contraindications. In an attempt to find food components potentially acting as SERMs, we submitted four plant aqueous extracts derived from Greek flora (Sideritis euboea, Sideritis clandestina, Marticaria chamomilla, and Pimpinella anisum) in a series of in vitro biological assays reflective of SERM profile. We examined their ability (a) to stimulate the differentiation and mineralization of osteoblastic cell culture by histochemical staining for alkaline phosphatase and Alizarin Red-S staining, (b) to induce, like antiestrogens, the insulin growth factor binding protein 3 (IGFBP3) in MCF-7 breast cancer cells, and (c) to proliferate cervical adenocarcinoma (HeLa) cells by use of MTT assay. Our data reveal that all the plant extracts studied at a concentration range 10-100 microg/mL stimulate osteoblastic cell differentiation and exhibit antiestrogenic effect on breast cancer cells without proliferative effects on cervical adenocarcinoma cells. The presence of estradiol inhibited the antiestrogenic effect induced by the extracts on MCF-7 cells, suggesting an estrogen receptor-related mechanism. In conclusion, the aqueous extracts derived from Sideritis euboea, Sideritis clandestina, Marticaria chamomilla, and Pimpinella anisum may form the basis to design "functional foods" for the prevention of osteoporosis.

  12. Effect of Increasing Doses of γ-Radiation on Bone Marrow Stromal Cells Grown on Smooth and Rough Titanium Surfaces

    Directory of Open Access Journals (Sweden)

    Bo Huang

    2015-01-01

    Full Text Available Radiation therapy for oral and maxillofacial tumors could damage bone marrow stromal cells (BMSCs in jaw, which caused dental implant failure. However, how radiation affects BMSCs on SLA (sandblasted with large-grits, acid-etched surfaces is still unknown. The aim of this study was to investigate effect of different dose of γ-radiation on BMSCs on SLA and PT (polished titanium surfaces. Rat BMSCs were radiated with 2, 4, and 8 Gy γ-radiation and then seeded on both surfaces. Cell adhesion, spreading, and proliferation were tested. The osteogenesis and the adipogenesis ability were examined by Alizarin-Red and Oil-Red staining, respectively. Real-time PCR was performed to detect osteogenic (osteocalcin, OCN; runt-related transcription factor 2, Runx2 and adipogenic (peroxisome proliferator-activated receptor gamma, PPARγ gene expression at days 7 and 14 postirradiation. Results showed that γ-radiation reduced cell proliferation, adhesion, spreading, and osteogenic differentiation. 2 Gy radiation promoted adipogenic differentiation, but it was significantly decreased when dosage reached 4 Gy. In conclusion, results suggest that γ-radiation influenced BMSCs behaviors in a dosage-dependent manner except adipogenic differentiation, low dose promoted it, and high dose inhibited it. This effect was influenced by surface characteristics, which may explain the different failure rate of various implants in patients after radiation.

  13. Characterization of mesenchymal stem cells derived from equine adipose tissue

    Directory of Open Access Journals (Sweden)

    A.M. Carvalho

    2013-08-01

    Full Text Available Stem cell therapy has shown promising results in tendinitis and osteoarthritis in equine medicine. The purpose of this work was to characterize the adipose-derived mesenchymal stem cells (AdMSCs in horses through (1 the assessment of the capacity of progenitor cells to perform adipogenic, osteogenic and chondrogenic differentiation; and (2 flow cytometry analysis using the stemness related markers: CD44, CD90, CD105 and MHC Class II. Five mixed-breed horses, aged 2-4 years-old were used to collect adipose tissue from the base of the tail. After isolation and culture of AdMSCs, immunophenotypic characterization was performed through flow cytometry. There was a high expression of CD44, CD90 and CD105, and no expression of MHC Class II markers. The tri-lineage differentiation was confirmed by specific staining: adipogenic (Oil Red O, osteogenic (Alizarin Red, and chondrogenic (Alcian Blue. The equine AdMSCs are a promising type of adult progenitor cell for tissue engineering in veterinary medicine.

  14. Peroxisome proliferator-activated receptor delta agonist attenuates nicotine suppression effect on human mesenchymal stem cell-derived osteogenesis and involves increased expression of heme oxygenase-1.

    Science.gov (United States)

    Kim, Dong Hyun; Liu, Jiayong; Bhat, Samerna; Benedict, Gregory; Lecka-Czernik, Beata; Peterson, Stephen J; Ebraheim, Nabil A; Heck, Bruce E

    2013-01-01

    Smoking has long been associated with osteoporosis, decreased bone mineral density, increased risk of bone fracture, and increased health costs. Nicotine, the main component of cigarette smoke, has major negative effects on bone metabolism and skeletal remodeling in vivo. Although osteoblasts and osteoblast-like cells have been used extensively to study the impact of nicotine, few studies have been performed on human mesenchymal stem cells (hMSCs). In this context, we examined the impact of nicotine on (a) hMSCs proliferation, (b) osteoblastic differentiation, (c) alkaline phosphatase (ALP) activity, and (d) expression of canonical genes during differentiation of hMSCs. MSCs isolated from human bone marrow were treated with different concentrations (0, 0.1, 1 and 10 μM) of nicotine for 7 days. Nicotine caused a dose-dependent decrease in cell proliferation, decreased heme oxygenase-1 (HO-1) expression (p nicotine caused a dose-dependent decrease in alizarin red staining for calcium and staining for ALP. Induction of HO-1 by peroxisome proliferator-activated receptor delta agonist (GW0742) prevented the effect of nicotine. Nicotine caused a dose-dependent reduction in the expression of BMP-2, a well-known marker for bone formation; however, this was prevented by GW0742 treatment. Therefore, induction of HO-1 prevents the deleterious effects of nicotine on osteogenesis in hMSC. This offers insight into both how nicotine affects bone remodeling and a therapeutic approach to prevent fracture and osteoporosis in smokers.

  15. HIF-1α transgenic bone marrow cells can promote tissue repair in cases of corticosteroid-induced osteonecrosis of the femoral head in rabbits.

    Directory of Open Access Journals (Sweden)

    Hao Ding

    Full Text Available Although corticosteroid-induced osteonecrosis of the femoral head (ONFH is common, the treatment for it remains limited and largely ineffective. We examined whether implantation of hypoxia inducible factor-1α (HIF-1α transgenic bone marrow cells (BMCs can promote the repair of the necrotic area of corticosteroid-induced ONFH. In this study, we confirmed that HIF-1α gene transfection could enhance mRNA expression of osteogenic genes in BMCs in vitro. Alkaline phosphatase activity assay and alizarin red-S staining indicated HIF-1α transgenic BMCs had enhanced osteogenic differentiation capacity in vitro. Furthermore, enzyme linked immunosorbent assay (ELISA for VEGF revealed HIF-1α transgenic BMCs secreted more VEGF as compared to normal BMCs. An experimental rabbit model of early-stage corticosteroid-induced ONFH was established and used for an evaluation of cytotherapy. Transplantation of HIF-1α transgenic BMCs dramatically improved the bone regeneration of the necrotic area of the femoral head. The number and volume of blood vessel were significantly increased in the necrotic area of the femoral head compared to the control groups. These results support HIF-1α transgenic BMCs have enhanced osteogenic and angiogenic activity in vitro and in vivo. Transplantation of HIF-1α transgenic BMCs can potentially promote the repair of the necrotic area of corticosteroid-induced ONFH.

  16. Simultaneous preconcentration of cadmium and chromium(III) in water samples by cloud point extraction and their determination by flame atomic absorption spectrometry.

    Science.gov (United States)

    Meng, Lifen; Ning, Jinyan; Yang, Yaling

    2014-01-01

    A sensitive and simple method for flame atomic absorption spectrometry determination of traces of cadmium and chromium(III) species in water samples after preconcentration by cloud point extraction has been developed. A novel complex agent of alizarin complexone with cadmium (Cd) and chromium (Cr(III)) was quantitatively extracted in surface primary alcohol ethoxylate-rich phase at 33 °C. The effects of experimental conditions including pH of sample solution, concentration of chelating agent and salt, equilibration temperature and time, and foreign ions were evaluated in order to enhance sensitivity of the method. Under optimal conditions, the low limit detections were 6.7 and 3.2 μg/L, and the enrichment factors were 24 and 20 for Cd and Cr(III), respectively. The relative standard deviations were 3.8 and 2.5% for Cd and Cr(II), respectively (n = 11). The high recoveries of the spiked Cd and Cr(III) ions were obtained in the range of 90-116%. The proposed method has been successfully applied for the determination of Cd and Cr(III) in water samples.

  17. Osteogenic Cells Derived From Embryonic Stem Cells Produced Bone Nodules in Three-Dimensional Scaffolds

    Directory of Open Access Journals (Sweden)

    Chaudhry G. R.

    2004-01-01

    Full Text Available An approach for 3D bone tissue generation from embryonic stem (ES cells was investigated. The ES cells were induced to differentiate into osteogenic precursors, capable of proliferating and subsequently differentiating into bone-forming cells. The differentiated cells and the seeded scaffolds were characterized using von Kossa and Alizarin Red staining, electron microscopy, and RT-PCR analysis. The results demonstrated that ES-derived bone-forming cells attached to and colonized the biocompatible and biodegradable scaffolds. Furthermore, these cells produced bone nodules when grown for 3–4 weeks in mineralization medium containing ascorbic acid and beta-glycerophosphate both in tissue culture plates and in scaffolds. The differentiated cells also expressed osteospecific markers when grown both in the culture plates and in 3D scaffolds. Osteogenic cells expressed alkaline phosphatase, osteocalcin, and osteopontin, but not an ES cell-specific marker, oct-4. These findings suggest that ES cell can be used for in vitro tissue engineering and cultivation of graftable skeletal structures.

  18. Degradation of textile dyes using immobilized lignin peroxidase-like metalloporphines under mild experimental conditions

    Directory of Open Access Journals (Sweden)

    Zucca Paolo

    2012-12-01

    Full Text Available Abstract Background Synthetic dyes represent a broad and heterogeneous class of durable pollutants, that are released in large amounts by the textile industry. The ability of two immobilized metalloporphines (structurally emulating the ligninolytic peroxidases to bleach six chosen dyes (alizarin red S, phenosafranine, xylenol orange, methylene blue, methyl green, and methyl orange was compared to enzymatic catalysts. To achieve a green and sustainable process, very mild conditions were chosen. Results IPS/MnTSPP was the most promising biomimetic catalyst as it was able to effectively and quickly bleach all tested dyes. Biomimetic catalysis was fully characterized: maximum activity was centered at neutral pH, in the absence of any organic solvent, using hydrogen peroxide as the oxidant. The immobilized metalloporphine kept a large part of its activity during multi-cycle use; however, well-known redox mediators were not able to increase its catalytic activity. IPS/MnTSPP was also more promising for use in industrial applications than its enzymatic counterparts (lignin peroxidase, laccase, manganese peroxidase, and horseradish peroxidase. Conclusions On the whole, the conditions were very mild (standard pressure, room temperature and neutral pH, using no organic solvents, and the most environmental-friendly oxidant and a significant bleaching and partial mineralization of the dyes was achieved in approximately 1 h. Therefore, the process was consistent with large-scale applications. The biomimetic catalyst also had more promising features than the enzymatic catalysts.

  19. Adhesion, vitality and osteogenic differentiation capacity of adipose derived stem cells seeded on nitinol nanoparticle coatings.

    Directory of Open Access Journals (Sweden)

    Sarah Strauss

    Full Text Available Autologous cells can be used for a bioactivation of osteoimplants to enhance osseointegration. In this regard, adipose derived stem cells (ASCs offer interesting perspectives in implantology because they are fast and easy to isolate. However, not all materials licensed for bone implants are equally suited for cell adhesion. Surface modifications are under investigation to promote cytocompatibility and cell growth. The presented study focused on influences of a Nitinol-nanoparticle coating on ASCs. Possible toxic effects as well as influences on the osteogenic differentiation potential of ASCs were evaluated by viability assays, scanning electron microscopy, immunofluorescence and alizarin red staining. It was previously shown that Nitinol-nanoparticles exert no cell toxic effects to ASCs either in soluble form or as surface coating. Here we could demonstrate that a Nitinol-nanoparticle surface coating enhances cell adherence and growth on Nitinol-surfaces. No negative influence on the osteogenic differentiation was observed. Nitinol-nanoparticle coatings offer new possibilities in implantology research regarding bioactivation by autologous ASCs, respectively enhancement of surface attraction to cells.

  20. Adhesion, vitality and osteogenic differentiation capacity of adipose derived stem cells seeded on nitinol nanoparticle coatings.

    Science.gov (United States)

    Strauss, Sarah; Neumeister, Anne; Barcikowski, Stephan; Kracht, Dietmar; Kuhbier, Jörn W; Radtke, Christine; Reimers, Kerstin; Vogt, Peter M

    2013-01-01

    Autologous cells can be used for a bioactivation of osteoimplants to enhance osseointegration. In this regard, adipose derived stem cells (ASCs) offer interesting perspectives in implantology because they are fast and easy to isolate. However, not all materials licensed for bone implants are equally suited for cell adhesion. Surface modifications are under investigation to promote cytocompatibility and cell growth. The presented study focused on influences of a Nitinol-nanoparticle coating on ASCs. Possible toxic effects as well as influences on the osteogenic differentiation potential of ASCs were evaluated by viability assays, scanning electron microscopy, immunofluorescence and alizarin red staining. It was previously shown that Nitinol-nanoparticles exert no cell toxic effects to ASCs either in soluble form or as surface coating. Here we could demonstrate that a Nitinol-nanoparticle surface coating enhances cell adherence and growth on Nitinol-surfaces. No negative influence on the osteogenic differentiation was observed. Nitinol-nanoparticle coatings offer new possibilities in implantology research regarding bioactivation by autologous ASCs, respectively enhancement of surface attraction to cells.

  1. Mesenchymal stem cell proliferation and differentiation on load-bearing trabecular Nitinol scaffolds.

    Science.gov (United States)

    Gotman, Irena; Ben-David, Dror; Unger, Ronald E; Böse, Thomas; Gutmanas, Elazar Y; Kirkpatrick, C James

    2013-09-01

    Bone tissue regeneration in load-bearing regions of the body requires high-strength porous scaffolds capable of supporting angiogenesis and osteogenesis. 70% porous Nitinol (NiTi) scaffolds with a regular 3-D architecture resembling trabecular bone were produced from Ni foams using an original reactive vapor infiltration technique. The "trabecular Nitinol" scaffolds possessed a high compressive strength of 79 MPa and high permeability of 6.9×10(-6) cm2. The scaffolds were further modified to produce a near Ni-free surface layer and evaluated in terms of Ni ion release and human mesenchymal stem cell (hMSC) proliferation (AlamarBlue), differentiation (alkaline phosphatase activity, ALP) and mineralization (Alizarin Red S staining). Scanning electron microscopy was employed to qualitatively corroborate the results. hMSCs were able to adhere and proliferate on both as-produced and surface-modified trabecular NiTi scaffolds, to acquire an osteoblastic phenotype and produce a mineralized extracellular matrix. Both ALP activity and mineralization were increased on porous scaffolds compared to control polystyrene plates. Experiments in a model coculture system of microvascular endothelial cells and hMSCs demonstrated the formation of prevascular structures in trabecular NiTi scaffolds. These data suggest that load-bearing trabecular Nitinol scaffolds could be effective in regenerating damaged or lost bone tissue.

  2. Increases in discontinuous rib cartilage and fused carpal bone in rat fetuses exposed to the teratogens, busulfan, acetazolamide, vitamin A, and ketoconazole.

    Science.gov (United States)

    Dodo, T; Uchida, K; Hirose, T; Fukuta, T; Kojima, C; Shiraishi, I; Kato, E; Horiba, T; Mineshima, H; Okuda, Y; Maeda, M; Katsutani, N; Hirano, K; Aoki, T

    2010-06-01

    Skeletal changes induced by treatment of pregnant rats with four potent teratogens, busulfan, acetazolamide, vitamin A palmitate, and ketoconazole, were evaluated using Alizarin Red S and Alcian Blue double-staining to investigate the relationship between drug-induced skeletal malformations and cartilaginous changes in the fetuses. Pregnant rats (N = 8/group) were treated once or twice between gestation days (GDs) 10 to 13 with busulfan at doses of 3, 10, or 30 mg/kg; acetazolamide at 200, 400, or 800 mg/kg; vitamin A palmitate at 100,000, 300,000, or 1,000,000 IU/kg; or ketoconazole at doses of 10, 30, or 100 mg/kg. Uterine evaluations and fetal external and skeletal examinations were conducted on GD 20. Marked skeletal abnormalities in ribs and hand/forelimb bones such as absent/ short/bent ribs, fused rib cartilage, absent/fused forepaw phalanx, and misshapen carpal bones were induced at the mid- and high-doses of busulfan and acetazolamide and at the high-dose of vitamin A palmitate and ketoconazole. Increased incidences of discontinuous rib cartilage (DRC) and fused carpal bone (FCB) were observed from the low- or mid-dose in the busulfan and acetazolamide groups, and incidences of FCB were increased from the mid-dose in the vitamin A palmitate and ketoconazole groups. Therefore, DRC and FCB were detected at lower doses than those at which ribs and hand/forelimb malformations were observed in the four potent teratogens.

  3. Teratogenic effects of silymarin on mouse fetuses

    Science.gov (United States)

    Gholami, Mahbobe; Moallem, Seyed Adel; Afshar, Mohammad; Amoueian, Sakineh; Etemad, Leila; Karimi, Gholamreza

    2016-01-01

    Objective: Silybum marianum has been used for centuries in herbal medicine for treatment of liver diseases. Currently, there is no data available on the possible effects of silymarin on fetal development. This study aimed to investigate the teratogenic effect of silymarin on BALB/c mice fetuses. Materials and Methods: A total of 40 pregnant mice were divided into 4 groups of 10 mice each. Three groups received silymarin at three different doses of 50, 100 and 200 mg/kg/day during gestational days (GDs). The control group received normal saline and tween (solvent). Dams were sacrificed on GD 18 and all fetuses were examined for gross malformations, size and body weight. Malformed fetuses were double stained with alizarin red and alcian blue. Results: Silymarin administration at all doses resulted in reduction of the mean fetal body weights. The abnormalities included limb, vertebral column and craniofacial malformations. Craniofacial malformations were the most common abnormalities, but they were not observed in a dose-dependent manner. The percentage of fetal resorption significantly increased (up to 15%) in all treatment groups. Conclusion: Based on our results, silymarin, especially at high doses can lead to fetal resorption, intrauterine growth retardation and limb, vertebral column and craniofacial abnormalities. More precise studies should be conducted about the teratogenic effects of herbal medicine investigating the underlying mechanisms. Thus, caution should be taken when administering S. marianum to pregnant woman. PMID:27761424

  4. Label-free nonlinear optical microscopy detects early markers for osteogenic differentiation of human stem cells

    Science.gov (United States)

    Hofemeier, Arne D.; Hachmeister, Henning; Pilger, Christian; Schürmann, Matthias; Greiner, Johannes F. W.; Nolte, Lena; Sudhoff, Holger; Kaltschmidt, Christian; Huser, Thomas; Kaltschmidt, Barbara

    2016-05-01

    Tissue engineering by stem cell differentiation is a novel treatment option for bone regeneration. Most approaches for the detection of osteogenic differentiation are invasive or destructive and not compatible with live cell analysis. Here, non-destructive and label-free approaches of Raman spectroscopy, coherent anti-Stokes Raman scattering (CARS) and second harmonic generation (SHG) microscopy were used to detect and image osteogenic differentiation of human neural crest-derived inferior turbinate stem cells (ITSCs). Combined CARS and SHG microscopy was able to detect markers of osteogenesis within 14 days after osteogenic induction. This process increased during continued differentiation. Furthermore, Raman spectroscopy showed significant increases of the PO43‑ symmetric stretch vibrations at 959 cm‑1 assigned to calcium hydroxyapatite between days 14 and 21. Additionally, CARS microscopy was able to image calcium hydroxyapatite deposits within 14 days following osteogenic induction, which was confirmed by Alizarin Red-Staining and RT- PCR. Taken together, the multimodal label-free analysis methods Raman spectroscopy, CARS and SHG microscopy can monitor osteogenic differentiation of adult human stem cells into osteoblasts with high sensitivity and spatial resolution in three dimensions. Our findings suggest a great potential of these optical detection methods for clinical applications including in vivo observation of bone tissue–implant-interfaces or disease diagnosis.

  5. Compression of Multilayered Composite Electrospun Scaffolds: A Novel Strategy to Rapidly Enhance Mechanical Properties and Three Dimensionality of Bone Scaffolds

    Directory of Open Access Journals (Sweden)

    Parthasarathy A. Madurantakam

    2013-01-01

    Full Text Available One major limitation of electrospun scaffolds intended for bone tissue engineering is their inferior mechanical properties. The present study introduces a novel strategy to engineer stiffer scaffolds by stacking multiple layers and cold welding them under high pressure. Electrospun polydioxanone (PDO and PDO:nanohydroxyapatite (PDO:nHA scaffolds (1, 2, or 4 layered stacks were compressed either before or after mineralizing treatment with simulated body fluid (SBF. After two weeks in SBF, scaffolds were analyzed for total mineral content and stiffness by Alizarin red S and uniaxial tensile testing, respectively. Scaffolds were also analyzed for permeability, pore size, and fiber diameter. Results indicated that compression of multiple layers significantly increased the stiffness of scaffolds while reducing mineralization and permeability. This phenomenon was attributed to increased density of fibers and loss of surface area due to fiber welding. Statistics revealed, the 4-layered PDO:nHA scaffold compressed first followed by mineralization in revised SBF had maximal stiffness, low permeability and pore size, and mineralization second only to noncompressed scaffolds. Within the limitations of permeability and pore size, this scaffold configuration represents an optimal midway for desired stiffness and mineral content for bone tissue engineering.

  6. Sequential Fluorescent Labeling Observation of Maxillary Sinus Augmentation by a Tissue-engineered Bone Complex in Canine Model

    Institute of Scientific and Technical Information of China (English)

    Xin-quan Jiang; Shao-yi Wang; Jun Zhao; Xiu-li Zhang; Zhi-yuan Zhang

    2009-01-01

    Aim To evaluate the effects of maxillary sinus floor elevation by a tissue-engineered bone complex of β-tricalcium phosphate (β-TCP) and autologous osteoblasts in dogs. Methodology Autologous osteoblasts from adult Beagle dogs were cultured in vitro. They were further combined with β-TCP to construct the tissue-engineered bone complex. 12 cases of maxillary sinus floor elevation surgery were made bilaterally in 6 animals and randomly repaired with the following 3 groups of materials: Group A (osteoblasts/β-TCP); Group B (β-TCP); Group C (autogenous bone) (n-4 per group). A polychrome sequential fluorescent labeling was performed post-operatively and the animals were sacrificed 24 weeks after operation for histological observation.Results Our results showed that autologous osteoblasts were successfully expanded and the osteoblastic phenoltypes were confirmed by ALP and Alizarin red staining. The cells could attach and proliferate well on the surface of the β-TCP scaffold. The fluorescent and histological observation showed that the tissue-engineered bone complex had an earlier mineralization and more bone formation inside the scaffold than β-TCP along or even autologous bone. It had also maximally maintained the elevated sinus height than both control groups. Conclusion Porous β-TCP has served as a good scaffold for autologous osteoblasts seeding. The tissue-engineered bone complex with β-TCP and autologous osteoblasts might be a better alternative to autologous bone for the clinical edentulous maxillary sinus augmentation.

  7. Beyond vibrationally mediated electron transfer: interfacial charge injection on a sub-10-fs time scale

    Science.gov (United States)

    Huber, Robert; Moser, Jacques E.; Gratzel, Michael; Wachtveitl, Josef L.

    2003-12-01

    The electron transfer (ET) from organic dye molecules to semiconductor-colloidal systems is characterized by a special energetic situation with a charge transfer reaction from a system of discrete donor levels to a continuum of acceptor states. If these systems show a strong electronic coupling they are amongst the fastest known ET systems with transfer times of less than 10 fs. In the first part a detailed discussion of the direct observation of an ET reaction with a time constant of about 6 fs will be given, with an accompanying argumentation concerning possible artifacts or other interfering signal contributions. In a second part we will try to give a simple picture for the scenario of such superfast ET reactions and one main focus will be the discussion of electronic dephasing and its consequences for the ET reaction. The actual ET process can be understood as a kind of dispersion process of the initially located electron into the colloid representing a real motion of charge density from the alizarin to the colloid.

  8. Cyclophosphamide-induced temporomandibular synostosis.

    Science.gov (United States)

    Bacon, W

    1983-06-01

    The study of malformations helps toward a better understanding of normal development, which is of significance to the orthodontist. Experiments in teratology have induced an extensive variety of facial abnormalities, but temporomandibular joint (TMJ) synostosis has never been previously reported. Ten pregnant female rabbits were treated with a daily injection of 50 mg. cyclophosphamide (DNA synthesis inhibitor), from day 11 to day 14, which is the period that precedes formation of the face. The control sample comprised five female rabbits. The fetuses were obtained by cesarean section on day 28 and stained with alizarin. Six of the ten treated female animals produced offspring that had TMJ synostosis. The skull with TMJ synostosis showed a retrognathic mandibular pattern in relation to the maxilla, and the bony trabeculae in the mandibular angle showed a downward orientation instead of the horizontal orientation seen in animals without synostosis. The length of the heads was significantly smaller in the treatment group than in the control group; within the treatment group, the heads with synostosis were significantly smaller than those without synostosis. It could be hypothesized that the cyclophosphamide might have affected intrinsic factors in the temporomandibular mesenchyma; an impairment in the development and function of the mandibular musculature, which is a vital factor in joint development and maintenance, might also have contributed to the genesis of the malformation. The association of immobilization and mandibular hypodevelopment seems to be in agreement with today's theories on maxillofacial growth.

  9. One-pot synthesis of biocompatible boronic acid-functionalized poly(methyl methacrylate) nanoparticles at sub-100 nm scale for glucose sensing

    Energy Technology Data Exchange (ETDEWEB)

    Sakalak, Huseyin [Selcuk University, Metallurgy and Materials Engineering (Turkey); Ulasan, Mehmet; Yavuz, Emine [Selcuk University, Advanced Technology Research and Application Center (Turkey); Camli, Sevket Tolga, E-mail: tolgacamli@gmail.com [Biyotez Machinery Chemistry R& D Co. Ltd. (Turkey); Yavuz, Mustafa Selman, E-mail: selmanyavuz@selcuk.edu.tr [Selcuk University, Metallurgy and Materials Engineering (Turkey)

    2014-12-15

    Poly(methyl methacrylate) nanoparticles containing 4-vinylphenyl boronic acid were synthesized in one pot by surfactant-free emulsion polymerization. The nanoparticles were characterized by scanning electron microscopy and dynamic light scattering. Boron content in the nanoparticles was confirmed by electron-dispersive X-ray spectroscopy. In polymerization process, several co-monomer ratios were studied in order to obtain optimum nanoparticle size. Average hydrodynamic diameter and polydispersity index of nanoparticles versus variation of acetone percentage in the solvent mixture and total monomer concentration were investigated. The effect of boronic acid concentration in the monomer mixture on nanoparticle size and size distribution was also reported. Without further functionalization to the nanoparticles, the catechol dye, alizarin red S, was bound to boronic acid-containing nanoparticles. These nanoparticles behave as a nanosensor by which glucose or fructose can be easily detected. Dye-containing nanoparticles were undertaken displacement reaction by glucose or fructose. The glucose or fructose content was also monitored by UV–Visible spectrophotometer. Furthermore, cytotoxicity studies of boronic acid-carrying poly(methyl methacrylate) nanoparticles were carried out in 3T3 cells, which showed no toxicity effect on the cells.

  10. Anthraquinone Content in Noni (Morinda citrifolia L.).

    Science.gov (United States)

    Bussmann, Rainer W; Hennig, Lothar; Giannis, Athanassios; Ortwein, Jutta; Kutchan, Toni M; Feng, Xi

    2013-01-01

    Noni has been used in traditional medicine and as food for thousands of years. While the fruits serve as food and internal medicine, leaves were traditionally used only topically. In recent years, concern regarding the possible content of anthraquinones in noni has led to scrutiny by the European Food Safety Authority. Little research existed on the content of anthraquinones in different noni preparations, with no information about the potential effect of harvest and preparation methods. Our research focused on lucidin, alizarin, and rubiadin, the most important anthraquinones from a health perspective. We found that the production process (fermentation/juice production versus drying/lyophilization) has no effect on the anthraquinone content. The source product, however, does have implications: noni fruit puree from which seeds had been removed as well as consumer products produced from such puree had no detectable amounts of any anthraquinones. Products that did contain seed or leaf material in all cases did contain partly significant amounts of anthraquinones. To alleviate safety concerns, we suggest that noni products, whether fermented or unfermented juice or powder, should be derived only from fully ripe noni fruits, and that any seed material needs to be removed during the production process.

  11. Anthraquinone Content in Noni (Morinda citrifolia L.

    Directory of Open Access Journals (Sweden)

    Rainer W. Bussmann

    2013-01-01

    Full Text Available Noni has been used in traditional medicine and as food for thousands of years. While the fruits serve as food and internal medicine, leaves were traditionally used only topically. In recent years, concern regarding the possible content of anthraquinones in noni has led to scrutiny by the European Food Safety Authority. Little research existed on the content of anthraquinones in different noni preparations, with no information about the potential effect of harvest and preparation methods. Our research focused on lucidin, alizarin, and rubiadin, the most important anthraquinones from a health perspective. We found that the production process (fermentation/juice production versus drying/lyophilization has no effect on the anthraquinone content. The source product, however, does have implications: noni fruit puree from which seeds had been removed as well as consumer products produced from such puree had no detectable amounts of any anthraquinones. Products that did contain seed or leaf material in all cases did contain partly significant amounts of anthraquinones. To alleviate safety concerns, we suggest that noni products, whether fermented or unfermented juice or powder, should be derived only from fully ripe noni fruits, and that any seed material needs to be removed during the production process.

  12. Petroleum ether extract of Cissus quadrangularis (LINN stimulates the growth of fetal bone during intra uterine developmental period: a morphometric analysis

    Directory of Open Access Journals (Sweden)

    Bhagath Kumar Potu

    2008-01-01

    Full Text Available OBJECTIVE: The aim of the present study was to analyze the effect Cissus quadrangularis plant petroleum ether extract on the development of long bones during the intra-uterine developmental stage in rats. METHODS: Pregnant rats (n=12 were randomly assigned into either a control group (n=6 or a Cissus quadrangularis treatment (n=6 group. Pregnant rats in the Cissus quadrangularis group were treated with Cissus quadrangularis petroleum ether extract at a dose of 500 mg/kg body weight from gestation day 9 until delivery. The animals in the control group received an equal volume of saline. Newborn pups were collected from both groups for alizarin red S - alcian blue staining to differentiate ossified and unossified cartilage. The ossified cartilage (bone was morphometrically analyzed using Scion image software. RESULTS: Morphometric analysis revealed that the percentage of the total length of ossified cartilage (bone in pups born to treated dams was significantly higher (P<0.001- -0.0001 than that of the control group. CONCLUSION: The results of the present study suggest that maternal administration of Cissus quadrangularis petroleum ether extract during pregnancy can stimulate the development of fetal bone growth during the intra-uterine developmental period.

  13. Imaging the zebrafish dentition: from traditional approaches to emerging technologies.

    Science.gov (United States)

    Bruneel, Bart; Mathä, Markus; Paesen, Rik; Ameloot, Marcel; Weninger, Wolfgang J; Huysseune, Ann

    2015-02-01

    The zebrafish, a model organism for which a plethora of molecular and genetic techniques exists, has a lifelong replacing dentition of 22 pharyngeal teeth. This is in contrast to the mouse, which is the key organism in dental research but whose teeth are never replaced. Employing the zebrafish as the main organism to elucidate the mechanisms of continuous tooth replacement, however, poses at least one major problem, related to the fact that all teeth are located deep inside the body. Investigating tooth replacement thus relies on conventional histological methods, which are often laborious, time-consuming and can cause tissue deformations. In this review, we investigate the advantages and limitations of adapting current visualization techniques to dental research in zebrafish. We discuss techniques for fast sectioning, such as vibratome sectioning and high-resolution episcopic microscopy, and methods for in toto visualization, such as Alizarin red staining, micro-computed tomography, and optical projection tomography. Techniques for in vivo imaging, such as two-photon excitation fluorescence and second harmonic generation microscopy, are also covered. Finally, the possibilities of light sheet microscopy are addressed.

  14. Osteoblast and monocyte responses to 444 ferritic stainless steel intended for a magneto-mechanically actuated fibrous scaffold.

    Science.gov (United States)

    Malheiro, Vera N; Spear, Rose L; Brooks, Roger A; Markaki, Athina E

    2011-10-01

    The rationale behind this work is to design an implant device, based on a ferromagnetic material, with the potential to deform in vivo promoting osseointegration through the growth of a healthy periprosthetic bone structure. One of the primary requirements for such a device is that the material should be non-inflammatory and non-cytotoxic. In the study described here, we assessed the short-term cellular response to 444 ferritic stainless steel; a steel, with a very low interstitial content and a small amount of strong carbide-forming elements to enhance intergranular corrosion resistance. Two different human cell types were used: (i) foetal osteoblasts and (ii) monocytes. Austenitic stainless steel 316L, currently utilised in many commercially available implant designs, and tissue culture plastic were used as the control surfaces. Cell viability, proliferation and alkaline phosphatase activity were measured. In addition, cells were stained with alizarin red and fluorescently-labelled phalloidin and examined using light, fluorescence and scanning electron microscopy. Results showed that the osteoblast cells exhibited a very similar degree of attachment, growth and osteogenic differentiation on all surfaces. Measurement of lactate dehydrogenase activity and tumour necrosis factor alpha protein released from human monocytes indicated that 444 stainless steel did not cause cytotoxic effects or any significant inflammatory response. Collectively, the results suggest that 444 ferritic stainless steel has the potential to be used in advanced bone implant designs.

  15. Raloxifene microsphere-embedded collagen/chitosan/β-tricalcium phosphate scaffold for effective bone tissue engineering.

    Science.gov (United States)

    Zhang, Ming-Lei; Cheng, Ji; Xiao, Ye-Chen; Yin, Ruo-Feng; Feng, Xu

    2017-02-25

    Engineering novel scaffolds that can mimic the functional extracellular matrix (ECM) would be a great achievement in bone tissue engineering. This paper reports the fabrication of novel collagen/chitosan/β-tricalcium phosphate (CCTP) based tissue engineering scaffold. In order to improve the regeneration ability of scaffold, we have embedded raloxifene (RLX)-loaded PLGA microsphere in the CCTP scaffold. The average pore of scaffold was in the range of 150-200μm with ideal mechanical strength and swelling/degradation characteristics. The release rate of RLX from the microsphere (MS) embedded scaffold was gradual and controlled. Also a significantly enhanced cell proliferation was observed in RLX-MS exposed cell group suggesting that microsphere/scaffold could be an ideal biomaterial for bone tissue engineering. Specifically, RLX-MS showed a significantly higher Alizarin red staining indicating the higher mineralization capacity of this group. Furthermore, a high alkaline phosphatase (ALP) activity for RLX-MS exposed group after 15days incubation indicates the bone regeneration capacity of MC3T3-E1 cells. Overall, present study showed that RLX-loaded microsphere embedded scaffold has the promising potential for bone tissue engineering applications.

  16. Carboxymethyl cellulose enables silk fibroin nanofibrous scaffold with enhanced biomimetic potential for bone tissue engineering application.

    Science.gov (United States)

    Singh, B N; Panda, N N; Mund, R; Pramanik, K

    2016-10-20

    Novel silk fibroin (SF) and carboxymethyl cellulose (CMC) composite nanofibrous scaffold (SFC) were developed to investigate their ability to nucleate bioactive nanosized calcium phosphate (Ca/P) by biomineralization for bone tissue engineering application. The composite nanofibrous scaffold was prepared by free liquid surface electrospinning method. The developed composite nanofibrous scaffold was observed to control the size of Ca/P particle (≤100nm) as well as uniform nucleation of Ca/P over the surface. The obtained nanofibrous scaffolds were fully characterized for their functional, structural and mechanical property. The XRD and EDX analysis depicted the development of apatite like crystals over SFC scaffolds of nanospherical in morphology and distributed uniformly throughout the surface of scaffold. Additionally, hydrophilicity as a measure of contact angle and water uptake capacity is higher than pure SF scaffold representing the superior cell supporting property of the SF/CMC scaffold. The effect of biomimetic Ca/P on osteogenic differentiation of umbilical cord blood derived human mesenchymal stem cells (hMSCs) studied in early and late stage of differentiation shows the improved osteoblastic differentiation capability as compared to pure silk fibroin. The obtained result confirms the positive correlation of alkaline phosphatase activity, alizarin staining and expression of runt-related transcription factor 2, osteocalcin and type1 collagen representing the biomimetic property of the scaffolds. Thus, the developed composite has been demonstrated to be a potential scaffold for bone tissue engineering application.

  17. Atomic Oxygen Treatment for Non-Contact Removal of Organic Protective Coatings from Painting Surfaces

    Science.gov (United States)

    Rutledge, Sharon K.; Banks, Bruce A.; Cales, Michael

    1994-01-01

    Current techniques for removal of varnish (lacquer) and other organic protective coatings from paintings involve contact with the surface. This contact can remove pigment, or alter the shape and location of paint on the canvas surface. A thermal energy atomic oxygen plasma, developed to simulate the space environment in low Earth orbit, easily removes these organic materials. Uniform removal of organic protective coatings from the surfaces of paintings is accomplished through chemical reaction. Atomic oxygen will not react with oxides so that most paint pigments will not be affected by the reaction. For paintings containing organic pigments, the exposure can be carefully timed so that the removal stops just short of the pigment. Color samples of Alizarin Crimson, Sap Green, and Zinc White coated with Damar lacquer were exposed to atomic oxygen. The lacquer was easily removed from all of the samples. Additionally, no noticeable change in appearance was observed after the lacquer was reapplied. The same observations were made on a painted canvas test sample obtained from the Cleveland Museum of Art. Scanning electron microscope photographs showed a slight microscopic texturing of the vehicle after exposure. However, there was no removal or disturbance of the paint pigment on the surface. It appears that noncontact cleaning using atomic oxygen may provide a viable alternative to other cleaning techniques. It is especially attractive in cases where the organic protective surface cannot be acceptably or safely removed by conventional techniques.

  18. Influence of the ortho-methoxyalkyl substituent on the properties of phenylboronic acids

    Science.gov (United States)

    Adamczyk-Woźniak, Agnieszka; Brzózka, Zbigniew; Dąbrowski, Marek; Madura, Izabela D.; Scheidsbach, Roy; Tomecka, Ewelina; Żukowski, Kamil; Sporzyński, Andrzej

    2013-03-01

    Novel phenylboronic acids with methoxyalkyl groups at ortho position were synthesized. Molecular and crystal structures for two compounds were determined by single crystal X-ray diffraction. In both cases the O-H⋯O hydrogen-bonded dimers are the primary supramolecular motives in which the relatively short intramolecular B-O-H⋯O hydrogen bonds are observed between boronic group and oxygen atom of the ortho-substituent. Based on the CSD data for ortho-substitued boronic acids, the relation between the twist of the boronic moiety towards phenyl ring and the intramolecular H-bond angle is discussed. The intermolecular interactions between dimeric motives were investigated with the aid of Hirshfeld surface analysis. The weak C-H⋯O and C-H⋯π interactions were detected together with the agostic B⋯H ones. Sugar-binding ability of the methoxyalkyl compounds was evaluated for D-glucose, D-fructose and D-galactose by the competition assay with Alizarin Red S.

  19. Effect of electrode position on azo dye removal in an up-flow hybrid anaerobic digestion reactor with built-in bioelectrochemical system

    Science.gov (United States)

    Cui, Min-Hua; Cui, Dan; Lee, Hyung-Sool; Liang, Bin; Wang, Ai-Jie; Cheng, Hao-Yi

    2016-04-01

    In this study, two modes of hybrid anaerobic digestion (AD) bioreactor with built-in BESs (electrodes installed in liquid phase (R1) and sludge phase (R2)) were tested for identifying the effect of electrodes position on azo dye wastewater treatment. Alizarin yellow R (AYR) was used as a model dye. Decolorization efficiency of R1 was 90.41 ± 6.20% at influent loading rate of 800 g-AYR/ m3·d, which was 39% higher than that of R2. The contribution of bioelectrochemical reduction to AYR decolorization (16.23 ± 1.86% for R1 versus 22.24 ± 2.14% for R2) implied that although azo dye was mainly removed in sludge zone, BES further improved the effluent quality, especially for R1 where electrodes were installed in liquid phase. The microbial communities in the electrode biofilms (dominant by Enterobacter) and sludge (dominant by Enterococcus) were well distinguished in R1, but they were similar in R2. These results suggest that electrodes installed in liquid phase in the anaerobic hybrid system are more efficient than that in sludge phase for azo dye removal, which give great inspirations for the application of AD-BES hybrid process for various refractory wastewaters treatment.

  20. Utility of Charge Transfer and Ion-Pair Complexation for Spectrophotometric Determination of Eletriptan Hydrobromide in Pure and Dosage Forms

    Directory of Open Access Journals (Sweden)

    Ayman A. Gouda

    2013-01-01

    Full Text Available Three simple, sensitive, and accurate spectrophotometric methods have been developed for the determination of eletriptan hydrobromide (ELT in pure and dosage forms. The first two methods are based on charge transfer complex formation between ELT and chromogenic reagents quinalizarin (Quinz and alizarin red S (ARS producing charge transfer complexes which showed an absorption maximum at 569 and 533 nm for Quinz and ARS, respectively. The third method is based on the formation of ion-pair complex between ELT with molybdenum(V-thiocyanate inorganic complex in hydrochloric acid medium followed by extraction of the colored ion-pair with dichloromethane and measured at 470 nm. Different variables affecting the reactions were studied and optimized. Beer's law is obeyed in the concentration ranges 2.0–18, 1.0–8.0, and 2.0–32 μg mL−1 for Quinz, ARS, and Mo(V-thiocyanate, respectively. The molar absorptivity, Sandell sensitivity, detection, and quantification limits are also calculated. The correlation coefficients were ≥0.9994 with a relative standard deviation (R.S.D%. of ≤0.925. The proposed methods were successfully applied for simultaneous determination of ELT in tablets with good accuracy and precision and without interferences from common additives, and the validity is assessed by applying the standard addition technique, which is compared with those obtained using the reported method.

  1. The marine-derived, multi-mineral formula, Aquamin, enhances mineralisation of osteoblast cells in vitro.

    Science.gov (United States)

    O'Gorman, Denise M; Tierney, Claire M; Brennan, Orlaith; O'Brien, Fergal J

    2012-03-01

    Osteoporosis is a global health problem characterized by low bone mass and an increase in bone fragility. It is now well accepted that dietary factors play a central role in bone development and health. Diet that lacks adequate minerals is considered to be a risk factor for osteoporosis. The food supplement, Aquamin, is a natural, multi-mineral derived from the red algae Lithothamnion corallioides, rich in calcium, magnesium and 72 other trace minerals. The aim of this study was to evaluate the effect of Aquamin on osteoblastic behaviour and mineralisation in a pre-osteoblastic cell line. Cell number and metabolic activity were assessed using Hoescht DNA and AlamarBlue assays respectively. Osteogenic differentiation was measured using an alkaline phosphatase assay while mineralisation was determined using von Kossa and alizarin red staining. It is reported here that Aquamin promotes increased mineralisation in osteoblast cell culture. These data suggest that the nutritional supplement Aquamin plays an important role in promoting bone formation and may be useful in treating bone diseases such as osteoporosis.

  2. Effects of hypoxia on pluripotency in murine iPS cells.

    Science.gov (United States)

    Sugimoto, Kouji; Yoshizawa, Yuu; Yamada, Shizuka; Igawa, Kazunari; Hayashi, Yoshihiko; Ishizaki, Hidetaka

    2013-10-01

    Retroviral transduction of four transcription factors (Oct4, Sox2, Klf4 and c-Myc) or three factors, excluding c-Myc, has been shown to initiate a reprogramming process that results in the transformation of murine fibroblasts to induced pluripotent stem (iPS) cells, and there has been a rapid increase in the number of iPS cell-based preclinical trials. In this study, the effects of these transcription factors were evaluated regarding the growth and differentiation of murine iPS cells under hypoxia. Based on the results of RT-PCR and alizarin red S staining, there were no statistical differences in the growth and differentiation of iPS cells or the induction of iPS cells to osteoblasts under hypoxia between the transcription factor groups. Furthermore, the function of hypoxia inducible factors (HIFs) in murine iPS cells under hypoxia was investigated in relation to the morphology and expression of transcription factors using RT-PCR and Western blotting. The HIF-2α knockdown group exhibited a decrease in the colony size of the iPS cells. The HIF-2α or -3α knockdown group demonstrated a statistically significant decrease in the transcription factor expression compared to that observed in the control group. These results demonstrate that HIF-2α among HIFs is the most influential candidate for the maintenance of the pluripotency of murine iPS cells.

  3. OBSERVATION ON SKELETAL DEFORMITY IN HATCHERY-REARED RED SPOTTED GROUPER, Epinephelus akaara (Temmick et Schlegel FROM LARVAL TO JUVENILE STAGE

    Directory of Open Access Journals (Sweden)

    Eri Setiadi

    2007-06-01

    Full Text Available Skeletal deformity is a significant problem in fish culture. The skeletal deformities in red spotted grouper from yolk-sac to juvenile stages were examined through clearing and staining of the cartilage and bone using Alcian Blue and Alizarin Red S. The overall results showed that the pattern of incidence of deformities showed an increase from preflexion to juvenile stages. The rate of deformities based on ten elements of bone from preflexion to juvenile stages were as follows: vertebral (42.6%—9.0%, dorsal proximal radials (4.8%—25.2%, neural spine (0%—8.4%, haemal spine (0%—6.8%, hypural (1.3%—5.4%, anal proximal radials (0%—5.4%, epural (1.3%—4.9%, arypural (2.0%—4.5%, lower jaw (1.3%—2.5%, and upper jaw (0%. Vertebral and dorsal proximal radials were recognized as the most susceptible parts to deformation. The main types of bone deformity were lordosis, scoliosis, fusion, shortening, branching, supernumerary elements, and saddleback syndrome. Development of saddleback syndrome was detected initially in preflexion stage, which was accompanied by deformity of the neural spines, dorsal proximal radials, and disposition of the distal radials and dorsal spines in later life stages. The skeletal deformity encountered during the larval rearing period could be caused by water surface tension.

  4. Influence of diabetes mellitus on the mineralization ability of two endodontic materials

    Directory of Open Access Journals (Sweden)

    João Eduardo GOMES FILHO

    2016-01-01

    Full Text Available Abstract The aim of this study was to evaluate the influence of diabetes mellituson tissue response and mineralization ability of Sealapex®and MTA Fillapex® sealers. Twenty-four Wistar rats were divided into two groups: diabetic and non-diabetic. The materials were placed in polyethylene tubes and implanted into dorsal connective tissue of rats for 7 and 30 days. Six animals from each group received injection of calcein, alizarin, and oxytetracycline on days 7, 14, and 21, respectively. The animals were killed after 7 and 30 days and specimens were prepared for histologic analysis by staining with hematoxylin and eosin or Von Kossa or left unstained for polarized light or fluorescence microscopy. On day 7, inflammatory reactions were characterized. Moderate inflammatory responses were observed for all groups and on day 30, a mild inflammatory response against MTA Fillapex® and a moderate inflammatory response against Sealapex® were observed. Von Kossa-positive structures were observed in response to both materials and birefringent structures were observed upon polarized light analysis; these had no relation to the diabetic condition (p > 0.05. The fluorescence intensity was unaffected in diabetic rats (p > 0.05. In conclusion, diabetes mellitus did not influence the tissue response or mineralization stimulated by Sealapex® or MTA Fillapex®.

  5. Sampling and identification of natural dyes in historical maps and drawings by liquid chromatography with diode-array detection.

    Science.gov (United States)

    Blanc, Rosario; Espejo, Teresa; López-Montes, Ana; Torres, David; Crovetto, Guillermo; Navalón, Alberto; Vílchez, José Luis

    2006-07-28

    A simple and rapid liquid chromatographic with diode-array UV-vis spectrophotometric detection (HPLC-DAD) method for identification of natural dyes has been developed. Chromatographic retention of carminic acid, indigotin, crocetin, gambogic acid, alizarin and purpurin has been studied. The mobile phase consisted of 40 mM SDS-10 mM phosphate buffer solution (pH 2.3)-0.1% TFA (eluent A) and acetonitrile (eluent B) using a programmed gradient (5% B to 95% B). Analyses were carried out on a Phenomenex, Luna 5u NH2 100(a) column (250 mm x 4.60 mm i.d., 5 microm particle) and the operating conditions were: 0.6 ml min(-1) flow rate, 20 microl volume injection and 35 degrees C column temperature. Extracts of samples of natural dyes taken from historical maps belonging to The Royal Chancellery Archives in Granada were successfully analyzed using the proposed method including a new technique for sampling.

  6. HPLC-MS of anthraquinoids, flavonoids, and their degradation products in analysis of natural dyes in archeological objects.

    Science.gov (United States)

    Surowiec, Izabella; Szostek, Bogdan; Trojanowicz, Marek

    2007-08-01

    LC with MS detection was optimized for sensitive and selective analysis of main classes of natural dyes used in ancient times for dyeing textiles -- red anthraquinoids, yellow flavonoids, and known degradation products of flavonols -- hydroxybenzoic acids. Fragmentation patterns of both negative and positive molecular ions for the above mentioned compounds were investigated. Three acquisition modes of MS analysis: scanning, SIM, and multiple reaction monitoring (MRM) in both positive and negative ion modes were optimized and compared with each other and with the UV-Vis diode-array detection. Even though in the applied chromatographic system formic acid was used in the mobile phase, SIM in the negative ion mode was the most selective and sensitive detection for all the investigated compounds when both mixtures of standards and analysis of extracts from archeological samples were concerned, with one exception -- alizarin, for which MS detection in positive ion mode was more sensitive. Detection limits obtained with MS detection for all investigated compounds except quinizarin were lower than the ones obtained with the diode-array UV-Vis detection, making MS detection the most suitable tool for the analysis of natural dyes and their degradation products in extracts from archeological samples.

  7. Gold nanoparticles promote osteogenic differentiation in human adipose-derived mesenchymal stem cells through the Wnt/β-catenin signaling pathway

    Directory of Open Access Journals (Sweden)

    Choi SY

    2015-07-01

    Full Text Available Seon Young Choi,1 Min Seok Song,1 Pan Dong Ryu,1 Anh Thu Ngoc Lam,2 Sang-Woo Joo,2 So Yeong Lee1 1Laboratory of Veterinary Pharmacology, Research Institute for Veterinary Science, College of Veterinary Medicine, Seoul National University, 2Department of Chemistry, Soongsil University, Seoul, South Korea Abstract: Gold nanoparticles (AuNPs are attractive materials for use in biomedicine due to their physical properties. Increasing evidence suggests that several nanoparticles induce the differentiation of human mesenchymal stem cells into osteoblasts and adipocytes. In this study, we hypothesized that chitosan-conjugated AuNPs promote the osteogenic differentiation of human adipose-derived mesenchymal stem cells. For the evaluation of osteogenic differentiation, alizarin red staining, an alamarBlue® assay, and a quantitative real-time polymerase chain reaction analysis were performed. In order to examine specific signaling pathways, immunofluorescence and a western blotting assay were performed. Our results demonstrate that chitosan-conjugated AuNPs increase the deposition of calcium content and the expression of marker genes related to osteogenic differentiation in human adipose-derived mesenchymal stem cells at nontoxic concentrations. These results indicate that chitosan-conjugated AuNPs promote osteogenesis through the Wnt/β-catenin signaling pathway. Therefore, chitosan-conjugated AuNPs can be used as a reagent for promoting bone formation. Keywords: chitosan-conjugated gold nanoparticle, mineralization, nonphosphorylated beta-catenin

  8. Fidelity of the Sr/Ca proxy in recording ocean temperature in the western Atlantic coral Siderastrea siderea

    Science.gov (United States)

    Kuffner, Ilsa B.; Roberts, Kelsey E.; Flannery, Jennifer A.; Morrison, Jennifer M.; Richey, Julie

    2017-01-01

    Massive corals provide a useful archive of environmental variability, but careful testing of geochemical proxies in corals is necessary to validate the relationship between each proxy and environmental parameter throughout the full range of conditions experienced by the recording organisms. Here we use samples from a coral-growth study to test the hypothesis that Sr/Ca in the coral Siderastrea siderea accurately records sea-surface temperature (SST) in the subtropics (Florida, USA) along 350 km of reef tract. We test calcification rate, measured via buoyant weight, and linear extension (LE) rate, estimated with Alizarin Red-S staining, as predictors of variance in the Sr/Ca records of 39 individual S. siderea corals grown at four outer-reef locations next to in-situ temperature loggers during two, year-long periods. We found that corals with calcification rates sample and drill-path selection when using long cores for SST paleoreconstruction. For our corals that passed this quality control step, the Sr/Ca-SST proxy performed well in estimating mean annual temperature across three sites spanning 350 km of the Florida reef tract. However, there was some evidence that extreme temperature stress in 2010 (cold snap) and 2011 (SST above coral-bleaching threshold) may have caused the corals not to record the temperature extremes. Known stress events could be avoided during modern calibrations of paleoproxies.

  9. Osteoporosis Recovery by Antrodia camphorata Alcohol Extracts through Bone Regeneration in SAMP8 Mice

    Directory of Open Access Journals (Sweden)

    Hen-Yu Liu

    2016-01-01

    Full Text Available Antrodia camphorata has previously demonstrated the efficacy in treating cancer and anti-inflammation. In this study, we are the first to evaluate Antrodia camphorata alcohol extract (ACAE for osteoporosis recovery in vitro with preosteoblast cells (MC3T3-E1 and in vivo with an osteoporosis mouse model established in our previous studies, ovariectomized senescence accelerated mice (OVX-SAMP8. Our results demonstrated that ACAE treatment was slightly cytotoxic to preosteoblast at 25 μg/mL, by which the osteogenic gene expression (RUNX2, OPN, and OCN was significantly upregulated with an increased ratio of OPG to RANKL, indicating maintenance of the bone matrix through inhibition of osteoclastic pathway. Additionally, evaluation by Alizarin Red S staining showed increased mineralization in ACAE-treated preosteoblasts. For in vivo study, our results indicated that ACAE inhibits bone loss and significantly increases percentage bone volume, trabecular bone number, and bone mineral density in OVX-SAMP8 mice treated with ACAE. Collectively, in vitro and in vivo results showed that ACAE could promote osteogenesis and prevent bone loss and should be considered an evidence-based complementary and alternative medicine for osteoporosis therapy through the maintenance of bone health.

  10. Acoustic-Frequency Vibratory Stimulation Regulates the Balance between Osteogenesis and Adipogenesis of Human Bone Marrow-Derived Mesenchymal Stem Cells

    Directory of Open Access Journals (Sweden)

    Xi Chen

    2015-01-01

    Full Text Available Osteoporosis can be associated with the disordered balance between osteogenesis and adipogenesis of bone marrow-derived mesenchymal stem cells (BM-MSCs. Although low-frequency mechanical vibration has been demonstrated to promote osteogenesis, little is known about the influence of acoustic-frequency vibratory stimulation (AFVS. BM-MSCs were subjected to AFVS at frequencies of 0, 30, 400, and 800 Hz and induced toward osteogenic or adipogenic-specific lineage. Extracellular matrix mineralization was determined by Alizarin Red S staining and lipid accumulation was assessed by Oil Red O staining. Transcript levels of osteogenic and adipogenic marker genes were evaluated by real-time reverse transcription-polymerase chain reaction. Cell proliferation of BM-MSCs was promoted following exposure to AFVS at 800 Hz. Vibration at 800 Hz induced the highest level of calcium deposition and significantly increased mRNA expression of COL1A1, ALP, RUNX2, and SPP1. The 800 Hz group downregulated lipid accumulation and levels of adipogenic genes, including FABP4, CEBPA, PPARG, and LEP, while vibration at 30 Hz supported adipogenesis. BM-MSCs showed a frequency-dependent response to acoustic vibration. AFVS at 800 Hz was the most favorable for osteogenic differentiation and simultaneously suppressed adipogenesis. Thus, acoustic vibration could potentially become a novel means to prevent and treat osteoporosis.

  11. Effects of {gamma}-secretase inhibition on the proliferation and vitamin D{sub 3} induced osteogenesis in adipose derived stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Jing, Wei [State Key Laboratory of Oral Diseases, West China College of Stomatology, Sichuan University, 14, 3rd Section of Renmin South Road, Chengdu 610041 (China); Xiong, Zhonghua [Department of Intensive Care Unit, Sichuan Cancer Hospital and Research Institute, Chengdu (China); Cai, Xiaoxiao; Huang, Yuanding; Li, Xiaoyu; Yang, Xingmei; Liu, Lei; Tang, Wei [State Key Laboratory of Oral Diseases, West China College of Stomatology, Sichuan University, 14, 3rd Section of Renmin South Road, Chengdu 610041 (China); Lin, Yunfeng, E-mail: yunfenglin@scu.edu.cn [State Key Laboratory of Oral Diseases, West China College of Stomatology, Sichuan University, 14, 3rd Section of Renmin South Road, Chengdu 610041 (China); Tian, Weidong, E-mail: drtianwd@hotmail.com [State Key Laboratory of Oral Diseases, West China College of Stomatology, Sichuan University, 14, 3rd Section of Renmin South Road, Chengdu 610041 (China)

    2010-02-12

    As a {gamma}-secretase inhibitor, DAPT has been widely used to evaluate the biological behaviors and Notch signaling pathway in various cells. This study was aimed to examine the effects of DAPT on the growth and vitamin D{sub 3} induced osteogenesis in adipose derived stem cells (ASCs). The cells were treated with or without DAPT and induced to osteoblastic lineage in the presence of vitamin D{sub 3}. Alizarin red staining and real-time PCR results indicated that the addition of DAPT to vitamin D{sub 3} treatments enhanced osteogenesis in ASCs. According to the fold increase and colony-forming unit assay results, the cells cultured in DAPT exhibited lower proliferation rate than those cultured in control medium. Hey1, expressed in the nucleus of ASCs to act as a transcriptional repressor, was downregulated when Notch signaling was inhibited by DAPT. Whereas the expression of Runx2 increased in the nucleus of osteogenic induced ASCs after DAPT treatment. This study demonstrated that DAPT reduced the proliferation and enhanced the osteogenesis in ASCs via regulation of Notch and Runx2 expression.

  12. Possible contribution of rubiadin, a metabolite of madder color, to renal carcinogenesis in rats.

    Science.gov (United States)

    Inoue, Kaoru; Yoshida, Midori; Takahashi, Miwa; Fujimoto, Hitoshi; Ohnishi, Kuniyoshi; Nakashima, Koichi; Shibutani, Makoto; Hirose, Masao; Nishikawa, Akiyoshi

    2009-04-01

    Madder color (MC) has been shown to exert carcinogenic potential in the rat kidney in association with degeneration, karyomegaly, increased cell proliferation of renal tubule cells and increased renal 8-OHdG levels. To clarify the causal relationship of components and metabolites of MC to renal carcinogenesis, male F344 rats were fed lucidin-3-O-primeveroside (LuP) or alizarin (Alz), and the genotoxic LuP metabolites lucidin (Luc) or rubiadin (Rub) for up to 26 weeks. After one week and four weeks, Luc did not induce any renal changes. In contrast, after one week, cortical tubule degeneration was apparent in the Alz and LuP groups, and cytoplasmic swelling with basophilic change and karyomegaly in the outer medulla was observed only in the Rub group. LuP and Rub increased the proliferative activity of tubule cells in the outer medulla, and Alz and LuP increased renal 8-OHdG levels. After 26 weeks, Rub but not Alz induced atypical tubules, a putative preneoplastic lesion, and karyomegaly in the outer medulla. These results indicate that Rub may be a potent carcinogenic metabolite of MC, targeting proximal tubule cells in the outer medulla, although oxidative stress increased by Alz or LuP might also be involved in renal carcinogenesis by MC.

  13. Effect of crosslinker on the swelling and adsorption properties of cationic superabsorbent

    Indian Academy of Sciences (India)

    TARUN SHARMA; GIRIDHAR MADRAS

    2016-06-01

    In the present study, superabsorbents (SAPs) of cationicmonomer [2-(methacryloyloxy) ethyl] trimethylammonium chloride have been prepared by free radical solution polymerization with different crosslinkers. They were subjected to repeated cycles of swelling and de-swelling in deionized water and NaCl solution. The conductivity of the swelling medium was measured and related to the swelling/de-swelling characteristics of the SAPs. The swelling capacity was also determined in saline solution. The swelling and de-swelling processes were described by first-order kinetics. The SAPs exhibited varied swelling capacity for crosslinkers of the same functionality as well as different functionality. The SAPs were used to adsorb the dye Orange G at different initial concentrations of the dye. The equilibrium adsorption data followed the Langmuir adsorption isotherms. The SAPs were also used to adsorb three other dyes, namely, Congo red, Amido black and Alizarin cyanine green. They exhibited different adsorption capacities for different dyes. The adsorption phenomenon was found to follow first-order kinetics.

  14. Growth in the area of the inferior dental foramen of rats.

    Science.gov (United States)

    Engel, G; West, V C

    1983-01-01

    The object of the present investigation was to see if the bone around the inferior dental nerve remodelled during mandibular growth and development. The investigation was carried out by injecting 27 albino Lewis rats with three fluorescent bone seeking dyes--oxytetracycline HCl (OTC), alizarin red S (ARS), and 2,4 bis-[N,N'-di' (carbomethyl-aminomethyl)] fluorescein (DCAF)--and then studying the bone around the inferior dental foramen. The mandibles of the animals were studied both macroscopically and microscopically under ultraviolet light to investigate the growth processes occurring and to see if the inferior dental foramen was relocated during growth. A quantitative analysis utilizing two specimens was also carried out for the same purpose. The results of both the qualitative and the quantitative analyses showed that the bone around the inferior dental nerve remodeled during mandibular growth. The mandible grew in an upward and backward direction, and the inferior dental foramen was correspondingly relocated in an upward and backward direction to maintain exactly the same position relative to the condyle and the posterior border of the ramus. This study, then, supports Moss's concept of the "unloaded" nerve, and is in keeping with his view of mandibular growth based on the functional matrix theory.

  15. Micellar electrokinetic chromatography method for the determination of several natural red dyestuff and lake pigments used in art work.

    Science.gov (United States)

    Maguregui, M I; Alonso, R M; Barandiaran, M; Jimenez, R M; García, N

    2007-06-22

    The identification of organic colorants used in artistic paintings is an important information source for reconstructing the working techniques found in a particular work and for defining a programme for the restoration and conservation of the painting. In this work, sodium dodecyl sulfate (SDS) was used as a surfactant in micellar electrokinetic chromatography (MEKC) for separating a broad range of red organic pigments, based on their colouring matters: madder (colouring matters: alizarin, quinizarin and purpurin), cochineal (colouring matter: carminic acid), red sandalwood (colouring matter: santalin), brazilwood (colouring matter: brazilin), lac dye (colouring matter: laccaic acid) and dragon's blood (colouring matter: dracorhodin). The running electrolyte used was 20 mM borax (pH 9), containing 20 mM SDS and 10% acetonitrile as organic modifier. Separation was carried out by applying a +20 kV voltage at the injection end, 25 degrees C and 214 nm/254 nm as detection wavelengths. All colorants were separated within less than 13 min with a good baseline resolution. The method was applied to the analysis of paint samples obtained from the Diocesan Museum of Holy Art of Bilbao.

  16. Sorption of hydrophilic dyes on anodic aluminium oxide films and application to pH sensing.

    Science.gov (United States)

    Silina, Yuliya E; Kuchmenko, Tatyana A; Volmer, Dietrich A

    2015-02-07

    The sorption of selected hydrophilic pH-sensitive dyes (bromophenol blue, bromothymol blue, bromocresol purple, alizarin red, methyl orange, congo red, rhodamine 6G) on films of anodized aluminium oxide (AAO) was investigated in this study. Depth and pore structure of the AAO channels were adjusted by changing electrolysis time and current density during treatment of aluminium foil in oxalic acid, sulfosalycilic acid and sulfuric acid at concentration levels between 0.2 and 0.6 M. The dyes were immobilized on the AAO surface by direct saturation of the films in dye solutions. It was shown by scanning electron microscopy and X-ray spectral analysis that the dyes penetrated into the AAO channels by more than 1.5 μm, even at static saturation conditions. The anionic dyes linked to the porous AAO surface exhibited differential shifts of the UV absorption bands in their acidic/basic forms. By combining several dyes, the films have an application range between pH = 0.5-9 in aqueous media. The dye-modified AAO film was a simple, portable, inexpensive and reusable pH sensor with very fast response time and clear colour transitions.

  17. Enhanced decolorization of azo dye in a small pilot-scale anaerobic baffled reactor coupled with biocatalyzed electrolysis system (ABR-BES): a design suitable for scaling-up.

    Science.gov (United States)

    Cui, Dan; Guo, Yu-Qi; Lee, Hyung-Sool; Wu, Wei-Min; Liang, Bin; Wang, Ai-Jie; Cheng, Hao-Yi

    2014-07-01

    A four-compartment anaerobic baffled reactor (ABR) incorporated with membrane-less biocatalyzed electrolysis system (BES) was tested for the treatment of azo dye (alizarin yellow R, AYR) wastewater (AYR, 200 mg L(-1); glucose, 1000 mg L(-1)). The ABR-BES was operated without and with external power supply to examine AYR reduction process and reductive intermediates with different external voltages (0.3, 0.5 and 0.7 V) and hydraulic retention times (HRT: 8, 6 and 4h). The decolorization efficiency in the ABR-BES (8h HRT, 0.5 V) was higher than that in ABR-BES without electrolysis, i.e. 95.1 ± 1.5% versus 86.9 ± 6.3%. Incorporation of BES with ABR accelerated the consumption of VFAs (mainly acetate) and attenuated biogas (methane) production. Higher power supply (0.7 V) enhanced AYR decolorization efficiency (96.4 ± 1.8%), VFAs removal, and current density (24.1 Am(-3) TCV). Shorter HRT increased volumetric AYR decolorization rates, but decreased AYR decolorization efficiency.

  18. Vascular Effects of Advanced Glycation End-Products: Content of Immunohistochemically Detected AGEs in Radial Artery Samples as a Predictor for Arterial Calcification and Cardiovascular Risk in Asymptomatic Patients with Chronic Kidney Disease

    Directory of Open Access Journals (Sweden)

    Katarzyna Janda

    2015-01-01

    Full Text Available Objectives. Our aim was to determine whether vascular deposition of advanced glycation end-products (AGEs is associated with arterial calcification and cardiovascular mortality in chronic kidney disease (CKD patients and to assess the relationships between vascular content of AGEs and selected clinical and biochemical parameters. Materials and Methods. The study comprised 54 CKD patients (33 hemodialyzed, 21 predialyzed. Examined parameters included BMI, incidence of diabetes, plasma fasting glucose, AGEs, soluble receptor for AGEs and 2,2-diphenyl-1-picrylhydrazyl (DPPH scavenging, serum C-reactive protein (hsCRP, plasminogen activator inhibitor-1 (PAI-1, and fetuin-A. Fragments of radial artery obtained during creation of hemodialysis access were stained for calcifications using alizarin red. AGEs deposits were identified immunohistochemically and their relative content was quantified. Results. Vascular content of AGEs was positively correlated with BMI, hsCRP, fetuin-A, PAI-1, and DPPH scavenging in simple regression; only fetuin-A was an independent predictor in multiple regression. There was a significant positive trend in the intensity of AGEs immunostaining among patients with grades 1, 2, and 3 calcifications. AGEs immunostaining intensity predicted 3-year cardiovascular mortality irrespective of patient’s age. Conclusions. The present study demonstrates an involvement of AGEs in the development of medial arterial calcification and the impact of arterial AGE deposition on cardiovascular mortality in CKD patients.

  19. Impaired Fasting Glucose and Diabetes as Predictors for Radial Artery Calcification in End Stage Renal Disease Patients

    Directory of Open Access Journals (Sweden)

    Katarzyna Janda

    2013-01-01

    Full Text Available Objective. The objective of the study was to assess the relationship between selected clinical and biochemical parameters of end stage renal disease (ESRD patients and arterial calcification. Materials and Methods. The study comprised 59 stage 5 chronic kidney disease patients (36 hemodialyzed and 23 predialysis. The examined parameters included common carotid artery intima-media thickness (CCA-IMT, BMI, incidence of diabetes and impaired fasting glucose (IFG, dyslipidemia, hypertension, and 3-year mortality. Plasma levels asymmetric dimethylarginine (ADMA, osteopontin (OPN, osteoprotegerin (OPG, and osteocalcin (OC were also measured. Fragments of radial artery obtained during creation of hemodialysis access were stained for calcifications using von Kossa method and alizarin red. Results. Calcification of radial artery was significantly associated with higher prevalence of IFG and diabetes (P=0.0004 and older age (P=0.003, as well as higher OPG (P=0.014 and ADMA concentrations (P=0.022. Fasting glucose >5.6 mmol/l (IFG and diabetes significantly predicted vascular calcification in multiple logistic regression. The calcification was also associated with higher CCA-IMT (P=0.006 and mortality (P=0.004; OR for death 5.39 [1.20–24.1] after adjustment for dialysis status and age. Conclusion. Combination of renal insufficiency and hyperglycemic conditions exerts a synergistic effect on vascular calcification and increases the risk of death.

  20. Targeted ablation of the abcc6 gene results in ectopic mineralization of connective tissues.

    Science.gov (United States)

    Klement, John F; Matsuzaki, Yasushi; Jiang, Qiu-Jie; Terlizzi, Joseph; Choi, Hae Young; Fujimoto, Norihiro; Li, Kehua; Pulkkinen, Leena; Birk, David E; Sundberg, John P; Uitto, Jouni

    2005-09-01

    Pseudoxanthoma elasticum (PXE), characterized by connective tissue mineralization of the skin, eyes, and cardiovascular system, is caused by mutations in the ABCC6 gene. ABCC6 encodes multidrug resistance-associated protein 6 (MRP6), which is expressed primarily in the liver and kidneys. Mechanisms producing ectopic mineralization as a result of these mutations remain unclear. To elucidate this complex disease, a transgenic mouse was generated by targeted ablation of the mouse Abcc6 gene. Abcc6 null mice were negative for Mrp6 expression in the liver, and complete necropsies revealed profound mineralization of several tissues, including skin, arterial blood vessels, and retina, while heterozygous animals were indistinguishable from the wild-type mice. Particularly striking was the mineralization of vibrissae, as confirmed by von Kossa and alizarin red stains. Electron microscopy revealed mineralization affecting both elastic structures and collagen fibers. Mineralization of vibrissae was noted as early as 5 weeks of age and was progressive with age in Abcc6(-/-) mice but was not observed in Abcc6(+/-) or Abcc6(+/+) mice up to 2 years of age. A total body computerized tomography scan of Abcc6(-/-) mice revealed mineralization in skin and subcutaneous tissue as well as in the kidneys. These data demonstrate aberrant mineralization of soft tissues in PXE-affected organs, and, consequently, these mice recapitulate features of this complex disease.

  1. Effect of acemannan, an extracted polysaccharide from Aloe vera, on BMSCs proliferation, differentiation, extracellular matrix synthesis, mineralization, and bone formation in a tooth extraction model.

    Science.gov (United States)

    Boonyagul, Sani; Banlunara, Wijit; Sangvanich, Polkit; Thunyakitpisal, Pasutha

    2014-07-01

    Aloe vera is a traditional wound healing medicine. We hypothesized acemannan, a polysaccharide extracted from Aloe vera gel, could affect bone formation. Primary rat bone marrow stromal cells (BMSCs) were treated with various concentrations of acemannan. New DNA synthesis, VEGF, BMP-2, alkaline phosphatase activity, bone sialoprotein, osteopontin expression, and mineralization were determined by [(3)H] thymidine incorporation assay, ELISA, biochemical assay, western blotting, and Alizarin Red staining, respectively. In an animal study, mandibular right incisors of male Sprague-Dawley rats were extracted and an acemannan treated sponge was placed in the socket. After 1, 2, and 4 weeks, the mandibles were dissected. Bone formation was evaluated by dual-energy X-ray absorptiometry and histopathological examination. The in vitro results revealed acemannan significantly increased BMSC proliferation, VEGF, BMP-2, alkaline phosphatase activity, bone sialoprotein and osteopontin expression, and mineralization. In-vivo results showed acemannan-treated groups had higher bone mineral density and faster bone healing compared with untreated controls. A substantial ingrowth of bone trabeculae was observed in acemannan-treated groups. These data suggest acemannan could function as a bioactive molecule inducing bone formation by stimulating BMSCs proliferation, differentiation into osteoblasts, and extracellular matrix synthesis. Acemannan could be a candidate natural biomaterial for bone regeneration.

  2. Acemannan, an extracted product from Aloe vera, stimulates dental pulp cell proliferation, differentiation, mineralization, and dentin formation.

    Science.gov (United States)

    Jittapiromsak, Nawaporn; Sahawat, Dusida; Banlunara, Wijit; Sangvanich, Polkit; Thunyakitpisal, Pasutha

    2010-06-01

    This study investigated the effect of acemannan (Aloe vera gel polysaccharide) on dentin formation. Primary human dental pulp cells were treated with acemannan. New DNA synthesis, bone morphogenetic protein-2, alkaline phosphatase activity, dentin sialoprotein expression, and mineralization were determined by [(3)H]-thymidine incorporation, enzyme-linked immunosorbent assay, biochemical assay, western blotting, and Alizarin Red staining, respectively. Then the upper first molars of 24 male Sprague Dawley rats were intentionally exposed and capped with either acemannan or calcium hydroxide. At day 28, the teeth were histopathologically examined and evaluated for the degree of inflammation, dentin bridge formation, and pulp tissue organization. The results revealed that acemannan significantly increased pulp cell proliferation, bone morphogenetic protein-2, alkaline phosphatase activity, dentin sialoprotein expression, and mineralization, compared with the untreated group. The acemannan-treated group also exhibited a complete homogeneous calcified dentin bridge and good pulp tissue organization, whereas neither was detected in the calcium hydroxide-treated and sham groups. In the acemannan-treated group, either mild or no inflammation was found, whereas the other groups had various degrees of inflammation. The data suggest that acemannan promotes dentin formation by stimulating primary human dental pulp cell proliferation, differentiation, extracellular matrix formation, and mineralization. Acemannan also has pulpal biocompatibility and promotes soft tissue organization.

  3. An Injectable Hydrogel as Bone Graft Material with Added Antimicrobial Properties

    Science.gov (United States)

    Tommasi, Giacomo; Perni, Stefano

    2016-01-01

    Currently, the technique which provides the best chances for a successful bone graft, is the use of bone tissue from the same patient receiving it (autograft); the main limitations are the limited availability and the risks involved in removing living bone tissue, for example, explant site pain and morbidity. Allografts and xenografts may overcome these limitations; however, they increase the risk of rejection. For all these reasons the development of an artificial bone graft material is particularly important and hydrogels are a promising alternative for bone regeneration. Gels were prepared using 1,4-butanediol diacrylate as crosslinker and alpha tricalciumphosphate; ZnCl2 and SrCl2 were added to the aqueous phase. MTT results demonstrated that the addition of strontium had a beneficial effect on the osteoblast cells density on hydrogels, and zinc instead did not increase osteoblast proliferation. The amount of calcium produced by the osteoblast cells quantified through the Alizarin Red protocol revealed that both strontium and zinc positively influenced the formation of calcium; furthermore, their effect was synergistic. Rheology properties were used to mechanically characterize the hydrogels and especially the influence of crosslinker's concentration on them, showing the hydrogels presented had extremely good mechanical properties. Furthermore, the antimicrobial activity of strontium and zinc in the hydrogels against methicillin-resistant Staphylococcus aureus and Staphylococcus epidermidis was determined. PMID:27174392

  4. Antioxidant impregnated ultra-high molecular weight polyethylene wear debris particles display increased bone remodeling and a superior osteogenic:osteolytic profile vs. conventional UHMWPE particles in a murine calvaria model.

    Science.gov (United States)

    Chen, Yu; Hallab, Nadim J; Liao, Yen-Shuo; Narayan, Venkat; Schwarz, Edward M; Xie, Chao

    2016-05-01

    Periprosthetic osteolysis remains a major limitation of long-term successful total hip replacements with ultra-high molecular weight polyethylene (UHMWPE) bearings. As intra and extracellular reactive oxygen species are know to contribute to wear debris-induced osteoclastic bone resorption and decreased osteoblastic bone formation, antioxidant doped UHMWPE has emerged as an approach to reduce the osteolytic potential of wear debris and maintain coupled bone remodeling. To test this hypothesis in vivo, we evaluated the effects of crosslinked UHMWPE wear debris particles (AltrX(™) ), versus similar wear particles made from COVERNOX(™) containing UHMWPE (AOX(™) ), in an established murine calvaria model. Eight-week-old female C57B/6 mice (n = 10/Group) received a pre-op micro-CT scan prior to surgical implantation of the UHMWPE particles (2mg), or surgery without particles (sham). Dynamic labeling was performed by intraperitoneal injection of calcein on day 7 and alizarin on day 9, and the calvaria were harvested for micro-CT and histology on day 10. Surprisingly, we found that AOX particles induced significantly more bone resorption (1.72-fold) and osteoclast numbers (1.99-fold) vs. AltrX (p UHMWPE particles have decreased osteolytic potential due to their increased osteogenic properties that support coupled bone remodeling. © 2015 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:845-851, 2016.

  5. Identification of Natural Dyes in Ancient Textiles by Time-of-Flight Secondary Ion Mass Spectrometry and Surface-Enhanced Raman Spectroscopy.

    Science.gov (United States)

    Lee, Jihye; Kim, Min Jung; van Elslande, Elsa; Walter, Philippe; Lee, Yeonhee

    2015-11-01

    The identification of dyes in archaeological remains is a long standing challenge. Major problems include contamination by environmental conditions over long periods of time, small amounts and limited availability of excavated samples, and low concentrations of dyestuff in the obtained samples. To address these issues, highly sensitive and non-destructive techniques are required. In response, in this work, two non-destructive analytical techniques, Time-of-Flight Secondary Ion Mass Spectrometry (TOF-SIMS) and Surface-Enhanced Raman Spectroscopy (SERS), were used for dye detection and the analysis results are compared. TOF-SIMS provides high detection efficiency for the analysis of organic materials whereas SERS is a useful technique for the detection of dyes in ancient textiles. An Ag colloid was employed to surmount the limitations of normal Raman measurement such as background fluorescence and weak Raman signals in small amounts of components. To identify the dyes used in ancient textiles, standard samples prepared using various dyestuffs and historical samples were analyzed with TOF-SIMS and Raman techniques. From the TOF-SIMS and the SERS spectra, dyestuffs such as alizarin, berberine, an indigo were identified in ancient textiles. The results suggest that TOF-SIMS and SERS are efficient non-destructive techniques for the characterization of archaeological textiles.

  6. Microscopic analysis of an opacified OFT CRYL® hydrophilic acrylic intraocular lens.

    Science.gov (United States)

    Ventura, Bruna Vieira; MacLean, Kyle Douglas; Lira, Wagner; Oliveira, Daniele Mendes de; Ventura, Camila Vieira; Werner, Liliana

    2016-01-01

    A 51-year-old patient underwent posterior vitrectomy with perfluoropropane gas injection, phacoemulsification, and implantation of an Oft Cryl® hydrophilic acrylic intraocular lens (IOL) because of traumatic retinal detachment and cataract in the right eye. On the first postoperative day, gas was filling the anterior chamber because of patient's non-compliance in terms of head positioning, and was reabsorbed within one week. Eight months later, the patient returned complaining of a significant decrease in vision. IOL opacification was noticed by slit-lamp examination. The lens was explanted to undergo gross and light microscopic analysis. The lens was also stained with the alizarin red method for calcium identification. Light microscopic analysis confirmed the presence of granular deposits, densely distributed in an overall circular pattern in the central part of the lens optic. The granules stained positive for calcium. This is the first case of the opacification of this type of hydrophilic lens. Surgeons should be aware of this potential postoperative complication, and the use of hydrophilic IOLs should be avoided in procedures involving intracameral gas because of the risk of IOL opacification.

  7. Activation of Cannabinoid Receptor 2 Enhances Osteogenic Differentiation of Bone Marrow Derived Mesenchymal Stem Cells

    Directory of Open Access Journals (Sweden)

    Yong-Xin Sun

    2015-01-01

    Full Text Available Bone marrow derived mesenchymal stem cells (BM-MSCs are considered as the most promising cells source for bone engineering. Cannabinoid (CB receptors play important roles in bone mass turnover. The aim of this study is to test if activation of CB2 receptor by chemical agonist could enhance the osteogenic differentiation and mineralization in bone BM-MSCs. Alkaline phosphatase (ALP activity staining and real time PCR were performed to test the osteogenic differentiation. Alizarin red staining was carried out to examine the mineralization. Small interference RNA (siRNA was used to study the role of CB2 receptor in osteogenic differentiation. Results showed activation of CB2 receptor increased ALP activity, promoted expression of osteogenic genes, and enhanced deposition of calcium in extracellular matrix. Knockdown of CB2 receptor by siRNA inhibited ALP activity and mineralization. Results of immunofluorescent staining showed that phosphorylation of p38 MAP kinase is reduced by knocking down of CB2 receptor. Finally, bone marrow samples demonstrated that expression of CB2 receptor is much lower in osteoporotic patients than in healthy donors. Taken together, data from this study suggested that activation of CB2 receptor plays important role in osteogenic differentiation of BM-MSCs. Lack of CB2 receptor may be related to osteoporosis.

  8. In vitro analysis and mechanical properties of twin screw extruded single-layered and coextruded multilayered poly(caprolactone) scaffolds seeded with human fetal osteoblasts for bone tissue engineering.

    Science.gov (United States)

    Ergun, Asli; Yu, Xiaojun; Valdevit, Antonio; Ritter, Arthur; Kalyon, Dilhan M

    2011-12-01

    In vitro culturing and mechanical properties of three types of three-dimensional poly(caprolactone) scaffolds with interconnecting open-foam networks are reported. The scaffolds targeted bone tissue regeneration and were fabricated using twin screw extrusion and coextrusion techniques, for continuous mixing/shaping and formation of single or multilayers with distinct and tailorable porosities and pore sizes. Human fetal preosteoblastic cells, hFOB, were cultured on the extruded and coextruded scaffolds under osteogenic supplements and the samples of the resulting tissue constructs were removed and characterized for cell viability and proliferation using the MTS assay, differentiation, and mineralized matrix synthesis via the alkaline phosphatase, ALP, activity and Alizarin Red staining and cell migration using confocal microscopy and scanning electron microscopy. The hFOB cells formed a confluent lining on scaffold surfaces, migrated to the interior and generated abundant extracellular matrix after 2 weeks of culturing, indicative of the promise of such scaffolds for utilization in tissue engineering. The scaffolds and tissue constructs exhibited compressive fatigue behavior that was similar to that of cancellous bone, suggesting the suitability of their use as bone graft substitutes especially for repair of critical-sized defects or nonunion fractures.

  9. Sulphated glycosaminoglycans and proteoglycans in the developing vertebral column of juvenile Atlantic salmon (Salmo salar).

    Science.gov (United States)

    Hannesson, Kirsten O; Ytteborg, Elisabeth; Takle, Harald; Enersen, Grethe; Bæverfjord, Grete; Pedersen, Mona E

    2015-08-01

    In the present study, the distribution of sulphated glycosaminoglycans (GAGs) in the developing vertebral column of Atlantic salmon (Salmo salar) at 700, 900, 1100 and 1400 d° was examined by light microscopy. The mineralization pattern was outlined by Alizarin red S and soft structures by Alcian blue. The temporal and spatial distribution patterns of different types of GAGs: chondroitin-4-sulphate/dermatan sulphate, chondroitin-6-sulphate, chondroitin-0-sulphate and keratan sulphate were addressed by immunohistochemistry using monoclonal antibodies against the different GAGs. The specific pattern obtained with the different antibodies suggests a unique role of the different GAG types in pattern formation and mineralization. In addition, the distribution of the different GAG types in normal and malformed vertebral columns from 15 g salmon was compared. A changed expression pattern of GAGs was found in the malformed vertebrae, indicating the involvement of these molecules during the pathogenesis. The molecular size of proteoglycans (PGs) in the vertebrae carrying GAGs was analysed with western blotting, and mRNA transcription of the PGs aggrecan, decorin, biglycan, fibromodulin and lumican by real-time qPCR. Our study reveals the importance of GAGs in development of vertebral column also in Atlantic salmon and indicates that a more comprehensive approach is necessary to completely understand the processes involved.

  10. A Comparison Study of the Effects of Echinacea purpurea Ethanolic Extract and Mesna on Cyclophosphamide-Induced Macroscopic Fetal Defects in Rats

    Directory of Open Access Journals (Sweden)

    Hossein Najafzadeh Varzi

    2009-03-01

    Full Text Available Objective(s There are some reports that the teratogenic effects of cyclophosphamide (CPA can be prevented by application of antioxidant drugs and stimulation of the maternal immune system. Echinacea purpurea extract is antioxidative and immunomodulator drug. Mesna (Sodium 2-mercaptoethane sulfonate is used for decreasing side effects of CPA, especially hemorrhagic cystitis. In this study, we compared the prophylactic effects of mesna and Echinacea extract on teratogenic effects of CPA. Materials and Methods This study was performed on 32 pregnant rats that were divided into 4 groups. The first group (control group received normal saline and the other groups received CPA (15 mg/kg intraperitoneally on 13th day of gestation. Mesna and E. purpurea extracts were administrated at doses of 100 and 400 mg/kg by IP injection, respectively, along with it and 12 hr later, after CPA injection. Rats were dissected on day 20 of gestation, embryos harvested and after determination of gross malformations they were stained by Alizarin red-Alcian blue method. ResultsCleft palate incidence was 38.46, 30.77 and 14.28% in fetuses of rats that received only CPA, CPA with mesna and CPA with Echinacea extract, respectively. In addition, skeletal anomalies incidence including limbs, vertebra, sternum, and scapula defects were decreased by Echinacea extract.ConclusionE. purpurea has significant effect on preventing CPA-induced malformations and better prophylactic effect than mesna on cases like CPA-induced cleft palate.

  11. Induction of osteoblast differentiation in human adipose derived stem cells by lanthanum ions

    Institute of Scientific and Technical Information of China (English)

    Harini D; Indra R; Rajaram A; Rama Rajaram

    2014-01-01

    Adipose derived stem cells represent a readily available source of adult stem cells for various biomedical applications. In this study, the proliferation and osteogenic differentiation potential of lanthanum nitrate (La3+) on human adipose derived mesenchy-mal stem cells (hADSCs) were investigated for the first time and compared with that of dexamethasone (Dex). Our results provided evidence that La3+at 50 µmol/L concentration promoted proliferation of hADSCs upto 2.4 fold when treated for 21 d in DMEM me-dium. Treatment of hADSCs with La3+containing osteogenic induction medium (α-MEM with ascorbic acid andβ-glycerophosphate) for 7 d resulted in higher calcium deposition than that in the presence of Dex (0.1 µmol/L) as shown by Alizarin red S and von Kossa staining. Scanning electron micrographs also showed more extracellular matrix mineralization in the presence of La3+. After 7 d of treatment with La3+(10 µmol/L) the expression of RunX2, osteopontin (OP) and osteocalcin (OC) increased 3.4, 5.5 and 2.7 fold re-spectively. Our results provided evidence that in the presence of La3+osteogenic differentiation occurred earlier than that in the pres-ence of Dex.

  12. Induction of dental epithelial cell differentiation marker gene expression in non-odontogenic human keratinocytes by transfection with thymosin beta 4

    Directory of Open Access Journals (Sweden)

    Tamotsu Kiyoshima

    2014-01-01

    Full Text Available Previous studies have shown that the recombination of cells liberated from developing tooth germs develop into teeth. However, it is difficult to use human developing tooth germ as a source of cells because of ethical issues. Previous studies have reported that thymosin beta 4 (Tmsb4x is closely related to the initiation and development of the tooth germ. We herein attempted to establish odontogenic epithelial cells from non-odontogenic HaCaT cells by transfection with TMSB4X. TMSB4X-transfected cells formed nodules that were positive for Alizarin-red S (ALZ and von Kossa staining (calcium phosphate deposits when cultured in calcification-inducing medium. Three selected clones showing larger amounts of calcium deposits than the other clones, expressed PITX2, Cytokeratin 14, and Sonic Hedgehog. The upregulation of odontogenesis-related genes, such as runt-related transcription factor 2 (RUNX2, Amelogenin (AMELX, Ameloblastin (AMBN and Enamelin (ENAM was also detected. These proteins were immunohistochemically observed in nodules positive for the ALZ and von Kossa staining. RUNX2-positive selected TMSB4X-transfected cells implanted into the dorsal subcutaneous tissue of nude mice formed matrix deposits. Immunohistochemically, AMELX, AMBN and ENAM were observed in the matrix deposits. This study demonstrated the possibility of induction of dental epithelial cell differentiation marker gene expression in non-odontogenic HaCaT cells by TMSB4X.

  13. Calcium-induced alteration of mitochondrial morphology and mitochondrial-endoplasmic reticulum contacts in rat brown adipocytes.

    Science.gov (United States)

    Golic, I; Velickovic, K; Markelic, M; Stancic, A; Jankovic, A; Vucetic, M; Otasevic, V; Buzadzic, B; Korac, B; Korac, A

    2014-09-09

    Mitochondria are key organelles maintaining cellular bioenergetics and integrity, and their regulation of [Ca2+]i homeostasis has been investigated in many cell types. We investigated the short-term Ca-SANDOZ® treatment on brown adipocyte mitochondria, using imaging and molecular biology techniques. Two-month-old male Wistar rats were divided into two groups: Ca-SANDOZ® drinking or tap water (control) drinking for three days. Alizarin Red S staining showed increased Ca2+ level in the brown adipocytes of treated rats, and potassium pyroantimonate staining localized electron-dense regions in the cytoplasm, mitochondria and around lipid droplets. Ca-SANDOZ® decreased mitochondrial number, but increased their size and mitochondrial cristae volume. Transmission electron microscopy revealed numerous enlarged and fusioned-like mitochondria in the Ca-SANDOZ® treated group compared to the control, and megamitochondria in some brown adipocytes. The Ca2+ diet affected mitochondrial fusion as mitofusin 1 (MFN1) and mitofusin 2 (MFN2) were increased, and mitochondrial fission as dynamin related protein 1 (DRP1) was decreased. Confocal microscopy showed a higher colocalization rate between functional mitochondria and endoplasmic reticulum (ER). The level of uncoupling protein-1 (UCP1) was elevated, which was confirmed by immunohistochemistry and Western blot analysis. These results suggest that Ca-SANDOZ® stimulates mitochondrial fusion, increases mitochondrial-ER contacts and the thermogenic capacity of brown adipocytes.

  14. Multi-Walled Carbon Nanotubes Promote Cementoblast Differentiation and Mineralization through the TGF-β/Smad Signaling Pathway

    Directory of Open Access Journals (Sweden)

    Lu Li

    2015-02-01

    Full Text Available Excretion of cementum by cementoblasts on the root surface is a process indispensable for the formation of a functional periodontal ligament. This study investigated whether carboxyl group-functionalized multi-walled carbon nanotubes (MWCNT-COOH could enhance differentiation and mineralization of mammalian cementoblasts (OCCM-30 and the possible signaling pathway involved in this process. Cementoblasts were incubated with various doses of MWCNT-COOH suspension. Cell viability was detected, and a scanning electron microscopy (SEM observed both the nanomaterials and the growth of cells cultured with the materials. Alizarin red staining was used to investigate the formation of calcium deposits. Real-time PCR and western blot were used to detect cementoblast differentiation and the underlying mechanisms through the expression of the osteogenic genes and the downstream effectors of the TGF-β/Smad signaling. The results showed that 5 µg/mL MWCNT-COOH had the most obvious effects on promoting differentiation without significant toxicity. Alp, Ocn, Bsp, Opn, Col1 and Runx2 gene expression was up-regulated. Smad2 and Smad3 mRNA was up-regulated, while Smad7 was first down-regulated on Day 3 and later up-regulated on Day 7. The elevated levels of phospho-Smad2/3 were also confirmed by western blot. In sum, the MWCNT-COOH promoted cementoblast differentiation and mineralization, at least partially, through interactions with the TGF-β/Smad pathway.

  15. The effect of bisphosphonates on the endothelial differentiation of mesenchymal stem cells.

    Science.gov (United States)

    Sharma, Dileep; Hamlet, Stephen Mark; Petcu, Eugen Bogdan; Ivanovski, Saso

    2016-02-09

    The contribution of the local stem cell niche to providing an adequate vascular framework during healing cannot be overemphasized. Bisphosphonates (BPs) are known to have a direct effect on the local vasculature, but their effect on progenitor cell differentiation is unknown. This in vitro study evaluated the effect(s) of various BPs on the differentiation of human placental mesenchymal stem cells (pMSCs) along the endothelial lineage and their subsequent functional and morphogenic capabilities. pMSC multipotency was confirmed by successful differentiation into cells of both the osteogenic and endothelial lineages, as demonstrated by positive Alizarin Red S staining and Ac-LDL uptake. pMSC differentiation in the presence of non-cytotoxic BP concentrations showed that nitrogen containing BPs had a significant inhibitory effect on cell migration and endothelial marker gene expression, as well as compromised endothelial differentiation as demonstrated using von Willebrand factor immunofluorescence staining and tube formation assay. This in vitro study demonstrated that at non-cytotoxic levels, nitrogen-containing BPs inhibit differentiation of pMSCs into cells of an endothelial lineage and affect the downstream functional capability of these cells supporting a multi-modal effect of BPs on angiogenesis as pathogenic mechanism contributing to bone healing disorders such as bisphosphonate related osteonecrosis of the jaws (BRONJ).

  16. Recovery of molybdenum using alumina microspheres and precipitation with selective organic reagents

    Energy Technology Data Exchange (ETDEWEB)

    Carvalho, Fatima Maria Sequeira de; Abrao, Alcidio [Instituto de Pesquisas Energeticas e Nucleares (IPEN), Sao Paulo, SP (Brazil). Dept. de Engenharia Quimica e Ambiental]. E-mail: fatimamc@net.ipen.br; aabrao@net.ipen.br

    1998-07-01

    In this paper is presented a study for the optimization of dissolution of the UAL{sub x} plates used for irradiation and production of radiomolybdenum. The alloy is dissolved in nitric acid with mercury as catalyst. The separation and concentration of the molybdenum was achieved using a chromatographic grade alumina microspheres column. the purified eluted molybdenum is finally precipitated using one of the selective reagents: alizarine blue, {alpha},{alpha}'- bipyridine and 1,10-phenanthroline. Any one of the obtained precipitate can be fired to the molybdenum trioxide. The interference of the following elements was studied: Re(VII), U(VI), Cr(VI), W(VI), V(V), Te(IV), Ti(IV), Zr(IV), Th(IV), Fe(III), Au(III), Ru(III), Al(III), Bi(III), Sb(III), Ce(IV), Pr(III), Sc(III), Y(III), Sm(III), Ba(II), Sr(II), Ni(II), Co(II), Cs(I). The molybdenum precipitates were characterized by gravimetric, CHN, TG, DTG, IR and X-ray diffraction analyses. (author)

  17. Cementum attachment protein manifestation is restricted to the mineralized tissue forming cells of the periodontium

    Energy Technology Data Exchange (ETDEWEB)

    Bar-Kana, I.; Pitaru, S. [Tel Aviv Univ., Dept. of Oral Biology, Goldschleger School of dental Medicine (Israel); Savion, N. [Tel Aviv Univ., Goldschleger Eye Research Inst. (Israel); Narayanan, A.S. [Univ. of Washington, Dept. of Pathology, Faculty of Medicine (United States)

    1998-08-01

    The mechanisms that regulate cementogenesis are mainly unknown. A specific cementum attachment protein (CAP) has been recently partially characterized and found to be more efficient in supporting the attachment of alveolar bone cells (ABC) and periodontal ligament cells (PLC) than that of gingival fibroblasts (GF). The purpose of this study was to determine the capacity of human periodontal-derived cells to bind an express CAP and to relate these properties to their capacity to express alkaline phosphatase (AlP) and form mineralized tissue (MTF). ABC, PLC and GF were tested. Human stromal bone marrow cells (SBMC) and a cementoma-derived cell line (CC) served as controls. CAP binding was determined using {sup 125}I-CAP. The amount of MTF was assessed by alizarin red staining and image analysis determination of the amount of red-stained material. AlP and CAP expression were examined by histochemistry and immuno-chemistry, respectively. The highest expression of CAP was observed in CC, followed by PLC and ABC in decreasing order, whereas SBMC and GF did not express CAP, SBMC manifested the highest CAP binding capacity followed by CC, ABC, PLC and GF. MTF and AlP manifestation were greatest in SBMC, followed by ABC, PLC and CC. Collectively, the results indicate that CAP binding and secretion are not linked and that CAP manifestation is restricted to periodontal derived cell lineages with the potential of forming mineralized tissues. (au) 39 refs.

  18. In vitro reparative dentin: a biochemical and morphological study

    Directory of Open Access Journals (Sweden)

    G. Teti

    2013-08-01

    Full Text Available In this study, starting from human dental pulp cells cultured in vitro, we simulated reparative dentinogenesis using a medium supplemented with different odontogenic inductors. The differentiation of dental pulp cells in odontoblast-like cells was evaluated by means of staining, and ultramorphological, biochemical and biomolecular methods. Alizarin red staining showed mineral deposition while transmission electron microscopy revealed a  synthesis of extracellular matrix fibers during the differentiation process. Biochemical assays demonstrated that the differentiated phenotype expressed odontoblast markers, such as Dentin Matrix Protein 1 (DMP1 and Dentin Sialoprotein (DSP, as well as type I collagen. Quantitative data regarding the mRNA expression of DMP1, DSP and type I  collagen were obtained by Real Time PCR. Immunofluorescence data demonstrated the various localizations of DSP and DMP1 during odontoblast differentiation. Based on our results, we obtained odontoblast-like cells which simulated the reparative dentin processes in order to better investigate the mechanism of odontoblast differentiation, and dentin extracellular matrix deposition and mineralization. 

  19. A novel basalt fiber-reinforced polylactic acid composite for hard tissue repair.

    Science.gov (United States)

    Chen, Xi; Li, Yan; Gu, Ning

    2010-08-01

    A basalt fiber (BF) was, for the first time, introduced into a poly(l-lactic acid) (PLLA) matrix as innovative reinforcement to fabricate composite materials for hard tissue repair. Firstly, BF/PLLA composites and pure PLLA were produced by the methods of solution blending and freeze drying. The results showed that basalt fibers can be uniformly dispersed in the PLLA matrix and significantly improve the mechanical properties and hydrophilicity of the PLLA matrix. The presence of basalt fibers may retard the polymer degradation rate and neutralize the acid degradation from PLLA. Osteoblasts were cultured in vitro to evaluate the cytocompatibility of the composite. An MTT assay revealed that osteoblasts proliferated well for 7 days and there was little difference found in their viability on both PLLA and BF/PLLA films, which was consistent with the alkaline phosphatase (ALP) activity results. A fluorescent staining observation showed that osteoblasts grew well on the composites. SEM images displayed that osteoblasts tended to grow along the fiber axis. The formation of mineralized nodules was observed on the films by Alizarin red S staining. These results suggest that the presence of basalt fibers does not noticeably affect osteoblastic behavior and the designed composites are osteoblast compatible. It is concluded that basalt fibers, as reinforcing fibers, may have promising applications in hard tissue repair.

  20. Trace Determination of Zirconium(Ⅳ) by Anodic Adsorptive Voltammetry at a Carbon Paste Electrode

    Institute of Scientific and Technical Information of China (English)

    Xun MAO; Huan Xin LAI; Ju Nan LI; Peng GAO; Zhi Hong YAN

    2004-01-01

    A new sensitive adsorptive voltammetric method was described for the determination of zirconium at a carbon paste electrode (CPE) in the presence of alizarin complexone (ALC). Optimal analytical conditions are: 1.0×10-6 or 5.0×10-7 mol/L ALC, 0. 20 mol/L HAC-NaAc (pH 4.3), accumulation for 60 s at 0 V (vs. SCE), and linear scanning from 0 V to 1.0 V at 250 mV/s. The peak potential of the complex is at 0.81 V. By using a model JP-303 polarographic analyzer, 2.0×10-10 mol/L (S/N=3) zirconium can be detected with a 90 s accumulation, when the 2nd-order derivative linear sweep technique is used, and the linear range is 6.0×10-10-2.0×10-8 mol/L (5.0×10-7 mol/L ALC) and 2.0×10-8-2.0×10-7 mol/L (1.0×10-6 mol/L ALC), respectively. The developed method was applied to the determination of trace zirconium in the ore samples with satisfactory results.

  1. The significance of azo-reduction in the mutagenesis and carcinogenesis of azo dyes.

    Science.gov (United States)

    Chung, K T

    1983-04-01

    Azo dyes are widely used in textile, printing, cosmetic, drug and food-processing industries. They are also used extensively in laboratories as either biological stains or pH indicators. The extent of such use is related to the degree of industrialization. Since intestinal cancer is more common in highly industrialized countries, a possible connection may exist between the increase in the number of cancer cases and the use of azo dyes. Azo dyes can be reduced to aromatic amines by the intestinal microflora. The mutagenicity of a number of azo dyes is reviewed in this paper. They include Trypan Blue, Ponceau 3R, Pinceau 2R, Methyl Red, Methyl Yellow, Methyl Orange, Lithol Red, Orange I, Orange II, 4-Phenylazo-Naphthylamine, Sudan I, Sudan IV, Acid Alizarin Violet N, Fast Garnet GBC, Allura Red, Ponceau SX, Sunset Yellow, Tartrazine, Citrus Red No. 2, Orange B, Yellow AB, Carmoisine, Mercury Orange, Ponceau S, Versatint Blue, Phenylazophenol, Evan's Blue and their degraded aromatic amines. The significance of azo reduction in the mutagenesis and carcinogenesis of azo dyes is discussed.

  2. Biomolecule-assisted synthesis of In(OH)₃ nanocubes and In₂O₃ nanoparticles: photocatalytic degradation of organic contaminants and CO oxidation.

    Science.gov (United States)

    Nayak, Arpan Kumar; Lee, Seungwon; Sohn, Youngku; Pradhan, Debabrata

    2015-12-04

    The synthesis of nanostructured materials without any hazardous organic chemicals and expensive capping reagents is one of the challenges in nanotechnology. Here we report on the L-arginine (a biomolecule)-assisted synthesis of single crystalline cubic In(OH)3 nanocubes of a size in the range of 30-60 nm along the diagonal using hydrothermal methods. Upon calcining at 750 °C for 1 h in air, In(OH)3 nanocubes are transformed into In2O3 nanoparticles (NPs) with voids. The morphology transformation and formation of voids with the increase of the calcination temperature is studied in detail. The possible mechanism of the voids' formation is discussed on the basis of the Kirkendall effect. The photocatalytic properties of In(OH)3 nanocubes and In2O3 NPs are studied for the degradation of rhodamin B and alizarin red S. Furthermore, the CO oxidation activity of In(OH)3 nanocubes and In2O3 NPs is examined. The photocatalytic and CO oxidation activity are measured to be higher for In2O3 NPs than for In(OH)3 nanocubes. This is attributed to the lower energy gap and higher specific surface area of the former. The present green synthesis has potential for the synthesis of other inorganic nanomaterials.

  3. Tissue transglutaminase is involved in mechanical load-induced osteogenic differentiation of human ligamentum flavum cells.

    Science.gov (United States)

    Chao, Yuan-Hung; Huang, Shih-Yung; Yang, Ruei-Cheng; Sun, Jui-Sheng

    2016-07-01

    Mechanical load-induced osteogenic differentiation might be the key cellular event in the calcification and ossification of ligamentum flavum. The aim of this study was to investigate the influence of tissue transglutaminase (TGM2) on mechanical load-induced osteogenesis of ligamentum flavum cells. Human ligamentum flavum cells were obtained from 12 patients undergoing lumbar spine surgery. Osteogenic phenotypes of ligamentum flavum cells, such as alkaline phosphatase (ALP), Alizarin red-S stain, and gene expression of osteogenic makers were evaluated following the administration of mechanical load and BMP-2 treatment. The expression of TGM2 was evaluated by real-time PCR, Western blotting, and enzyme-linked immunosorbent assay (ELISA) analysis. Our results showed that mechanical load in combination with BMP-2 enhanced calcium deposition and ALP activity. Mechanical load significantly increased ALP and OC gene expression on day 3, whereas BMP-2 significantly increased ALP, OPN, and Runx2 on day 7. Mechanical load significantly induced TGM2 gene expression and enzyme activity in human ligamentum flavum cells. Exogenous TGM2 increased ALP and OC gene expression; while, inhibited TG activity significantly attenuated mechanical load-induced and TGM2-induced ALP activity. In summary, mechanical load-induced TGM2 expression and enzyme activity is involved in the progression of the calcification of ligamentum flavum.

  4. Staphylococcal lipoteichoic acid promotes osteogenic differentiation of mouse mesenchymal stem cells by increasing autophagic activity.

    Science.gov (United States)

    Liu, Xin; Wang, Yuan; Cao, Zhen; Dou, Ce; Bai, Yun; Liu, Chuan; Dong, Shiwu; Fei, Jun

    2017-02-16

    This study sought to explore the effect of staphylococcal lipoteichoic acid (LTA) on autophagy in mouse mesenchymal stem cells (MSCs), and then influence osteogenesis through the change of autophagy. C3H10T1/2 cells were induced by osteogenic medium with the treatment of LTA at different concentrations (1, 5, 10 μg/mL); 3-methyladenine (3-MA) were used as the autophagy inhibitor, and rapamycin (rapamycin, Rap) were used to activate autophagy; the effects on osteogenesis were detected by alkaline phosphatase staining, alizarin red staining, real-time quantitative PCR, and western blotting; autophagic activity was investigated by the expression of LC3-Ⅱand p62 proteins. Compared with control group, the expression of osteogenesis markers was significantly up-regulated with the LTA treatment on the mRNA and protein level; the positive rate of alkaline phosphatase was enhanced in the LTA groups; and the formation of calcium nodules was increased simultaneously. The expression of LC3-Ⅱ protein was increased in LTA groups, while the expression of p62 protein was decreased. Inhibition of autophagy significantly reduced the effect of LTA on osteogenesis of MSCs; the promotion of LTA on osteogenic differentiation was further enhanced when adding rapamycin to activate autophagic activity. It provides new insight of prevention and treatment for bone infection.

  5. Osteogenic differentiation of human adipose-derived mesenchymal stem cells on gum tragacanth hydrogel.

    Science.gov (United States)

    Haeri, Seyed Mohammad Jafar; Sadeghi, Yousef; Salehi, Mohammad; Farahani, Reza Masteri; Mohsen, Nourozian

    2016-05-01

    Currently, natural polymer based hydrogels has attracted great attention of orthopedic surgeons for application in bone tissue engineering. With this aim, osteoinductive capacity of Gum Tragacanth (GT) based hydrogel was compared to collagen hydrogel and tissue culture plate (TCPS). For this purpose, adipose-derived mesenchymal stem cells (AT-MSCs) was cultured on the hydrogels and TCPS and after investigating the biocompatibility of hydrogels using MTT assay, osteoinductivity of hydrogels were evaluated using pan osteogenic markers such as Alizarin red staining, alkaline phosphatase (ALP) activity, calcium content and osteo-related genes. Increasing proliferation trend of AT-MSCs on GT hydrogel demonstrated that TG has no-cytotoxicity and can even be better than the other groups i.e., highest proliferation at day 5. GT hydrogel displayed highest ALP activity and mineralization when compared to the collagen hydrogel and TCPS. Relative gene expression levels have demonstrated that highest expression of Runx2, osteonectin and osteocalcin in the cells cultured GT hydrogel but the expression of collagen type-1 remains constant in hydrogels. Above results demonstrate that GT hydrogel could be an appropriate scaffold for accelerating and supporting the adhesion, proliferation and osteogenic differentiation of stem cells which further can be used for orthopedic applications.

  6. Synthesis of ZnS/CQDs nanocomposite and its application as a photocatalyst for the degradation of an anionic dye, ARS

    Science.gov (United States)

    Kaur, Sharanjit; Sharma, Shelja; Kansal, Sushil Kumar

    2016-10-01

    Novel carbon quantum dots (CQDs)-modified ZnS nanocomposite was prepared via a fast and facile chemical precipitation technique and was employed for the first time as a photocatalyst for the degradation of Alizarin red S (ARS) dye under visible light irradiation. The structural, morphological and optical properties of the prepared ZnS/CQDs nanocomposite were characterized by multiple analytical techniques such as X-ray diffraction, X-ray photoelectron spectroscopy, Fourier-transform infrared spectroscopy, thermogravimetric analysis, photoluminescence spectroscopy, transmission electron microscopy, field emission scanning electron microscope equipped with energy dispersive spectroscopy and UV-vis diffuse reflectance spectroscopy. Impact of affecting parameters on the photocatalytic activity of the ZnS/CQDs was studied and optimized. The results showed that the ZnS/CQDs exhibited excellent photocatalytic activity for the degradation of ARS dye i.e. 89% within 250 min, higher than that of the bare ZnS (63%). This enhancement in photocatalytic activity of ZnS/CQDs was attributed to the introduction of CQDs, which could absorb visible light efficiently, suppressing the recombination of electron-hole pairs and improving charge separation. Moreover, various scavengers have been used to study the role of reactive species in the photocatalytic degradation process.

  7. In-situ synthesis of high stable CdS quantum dots and their application for photocatalytic degradation of dyes

    Science.gov (United States)

    Samadi-Maybodi, Abdolraouf; Sadeghi-Maleki, Mohammad-Rasool

    2016-01-01

    Photocatalysis based on semiconductor quantum dots, which utilize the solar energy can be used for elimination of pollutants from aqueous media and applied for water purification. In this paper, high stable CdS quantum dots (QDs) with good optical properties were successfully synthesized in a facile in-situ method, using Na2S2O3 as precursor and thioglycolic acid (TGA) as a catalyst, as well as capping agent in aqueous media. The synthesis process was optimized with a 2IV7-3 fractional factorial design method. Then, we studied the degradation of some industrial dyes including: alizarin, acid violet, mordant red and thymol blue as a tool to check the photocatalytic activity of synthesized CdS QDs. Results specified that the synthesized CdS QDs are capable for degradation of organic dyes under visible light irradiation with good recycling stability during photocatalytic experiments. Structural and spectroscopic properties of the synthesized CdS QDs were studied by TEM, XRD and absorption and fluorescence spectroscopy techniques. The synthesized TGA-capped CdS QDs have sizes in the range of 2.65-2.93 nm with cubic crystalline structures.

  8. In vitro direct osteogenesis of murine embryonic stem cells without embryoid body formation.

    Science.gov (United States)

    Hwang, Yu-Shik; Polak, Julia M; Mantalaris, Athanasios

    2008-10-01

    Embryonic stem cells (ESCs) posses the ability to self-renew and differentiate into a multitude of lineages, including the osteogenic lineage in vitro. Currently, most approaches have focused on embryonic body (EB)-mediated osteogenic differentiation, which relies on formation of all three germ layers resulting in limited yields and labour-intensive culture processes. Our study aimed at developing an efficient culture strategy resulting in the upregulated in vitro osteogenic differentiation of murine ESCs (mESCs), which completely avoided EB formation. Specifically, mESCs were cultured in HepG2 conditioned medium for 3 days and then directed into osteogenic differentiation for 21 days without prior EB formation. The mineralised bone nodules generated were characterized by Alizarin red S-staining, phenotypic alkaline phosphatase expression, time-course analysis of ALPase activity, the presence of type I collagen and osteopontin, and osteocalcin, cbfa-1/runx-2, and osterix gene expression. Our method of direct osteogenic differentiation of mESCs represents a novel and efficient approach that results in enhanced yields and could have significant applications in bone tissue engineering.

  9. Photothermal excitation setup for a modified commercial atomic force microscope

    Energy Technology Data Exchange (ETDEWEB)

    Adam, Holger; Rode, Sebastian; Schreiber, Martin; Kühnle, Angelika, E-mail: kuehnle@uni-mainz.de [Institute of Physical Chemistry, Johannes Gutenberg University Mainz, Duesbergweg 10-14, 55099 Mainz (Germany); Kobayashi, Kei; Yamada, Hirofumi [Department of Electronic Science and Engineering, Kyoto University, Katsura, Nishikyo, Kyoto 615-8510 (Japan)

    2014-02-15

    High-resolution imaging in liquids using frequency modulation atomic force microscopy is known to suffer from additional peaks in the resonance spectrum that are unrelated to the cantilever resonance. These unwanted peaks are caused by acoustic modes of the liquid and the setup arising from the indirect oscillation excitation by a piezoelectric transducer. Photothermal excitation has been identified as a suitable method for exciting the cantilever in a direct manner. Here, we present a simple design for implementing photothermal excitation in a modified Multimode scan head from Bruker. Our approach is based on adding a few components only to keep the modifications as simple as possible and to maintain the low noise level of the original setup with a typical deflection noise density of about 15 fm/√(Hz) measured in aqueous solution. The success of the modification is illustrated by a comparison of the resonance spectra obtained with piezoelectric and photothermal excitation. The performance of the systems is demonstrated by presenting high-resolution images on bare calcite in liquid as well as organic adsorbates (Alizarin Red S) on calcite with simultaneous atomic resolution of the underlying calcite substrate.

  10. Human Adipose-Derived Mesenchymal Stem Cells Cryopreservation and Thawing Decrease α4-Integrin Expression

    Directory of Open Access Journals (Sweden)

    Ana Carolina Irioda

    2016-01-01

    Full Text Available Aim. The effects of cryopreservation on adipose tissue-derived mesenchymal stem cells are not clearly documented, as there is a growing body of evidence about the importance of adipose-derived mesenchymal stem cells for regenerative therapies. The aim of this study was to analyze human adipose tissue-derived mesenchymal stem cells phenotypic expression (CD34, CD45, CD73, CD90, CD105, and CD49d, colony forming unit ability, viability, and differentiation potential before and after cryopreservation. Materials and Methods. 12 samples of the adipose tissue were collected from a healthy donor using the liposuction technique. The cell isolation was performed by enzymatic digestion and then the cells were cultured up to passage 2. Before and after cryopreservation the immunophenotype, cellular viability analysis by flow cytometer, colony forming units ability, differentiation potential into adipocytes and osteoblasts as demonstrated by Oil Red O and Alizarin Red staining, respectively. Results. The immunophenotypic markers expression was largely preserved, and their multipotency was maintained. However, after cryopreservation, the cells decreased α4-integrin expression (CD49d, cell viability, and number of colony forming units. Conclusions. These findings suggest that ADMSC transplanted after cryopreservation might compromise the retention of transplanted cells in the host tissue. Therefore, further studies are warranted to standardize protocols related to cryopreservation to attain full benefits of stem cell therapy.

  11. Calcium-induced alteration of mitochondrial morphology and mitochondrial-endoplasmic reticulum contacts in rat brown adipocytes

    Directory of Open Access Journals (Sweden)

    I. Golic

    2014-09-01

    Full Text Available Mitochondria are key organelles maintaining cellular bioenergetics and integrity, and their regulation of [Ca2+]i homeostasis has been investigated in many cell types. We investigated the short-term Ca-SANDOZ® treatment on brown adipocyte mitochondria, using imaging and molecular biology techniques. Two-month-old male Wistar rats were divided into two groups: Ca-SANDOZ® drinking or tap water (control drinking for three days. Alizarin Red S staining showed increased Ca2+ level in the brown adipocytes of treated rats, and potassium pyroantimonate staining localized electron-dense regions in the cytoplasm, mitochondria and around lipid droplets. Ca-SANDOZ® decreased mitochondrial number, but increased their size and mitochondrial cristae volume. Transmission electron microscopy revealed numerous enlarged and fusioned-like mitochondria in the Ca-SANDOZ® treated group compared to the control, and megamitochondria in some brown adipocytes. The Ca2+ diet affected mitochondrial fusion as mitofusin 1 (MFN1 and mitofusin 2 (MFN2 were increased, and mitochondrial fission as dynamin related protein 1 (DRP1 was decreased. Confocal microscopy showed a higher colocalization rate between functional mitochondria and endoplasmic reticulum (ER. The level of uncoupling protein-1 (UCP1 was elevated, which was confirmed by immunohistochemistry and Western blot analysis. These results suggest that Ca-SANDOZ® stimulates mitochondrial fusion, increases mitochondrial-ER contacts and the thermogenic capacity of brown adipocytes

  12. Generation of bovine (Bos indicus) and buffalo (Bubalus bubalis) adipose tissue derived stem cells: isolation, characterization, and multipotentiality.

    Science.gov (United States)

    Sampaio, R V; Chiaratti, M R; Santos, D C N; Bressan, F F; Sangalli, J R; Sá, A L A; Silva, T V G; Costa, N N; Cordeiro, M S; Santos, S S D; Ambrosio, C E; Adona, P R; Meirelles, F V; Miranda, M S; Ohashi, O M

    2015-01-15

    Adult stem cells are known for their plasticity and their potential to differentiate into several different cell types; these characteristics have implications for cell therapy and reproductive biotechnologies. In this study, we report on the isolation and characterization of mesenchymal stem cells (MSC) derived from bovine and buffalo adipose tissue. Cells isolated using enzymatic digestion of bovine and buffalo adipose-tissue biopsy samples were grown in vitro for at least 15 passages, verifying their capacity to proliferate. These cells were also subjected to immunophenotypic characterization for the presence of CD90, CD105, and CD79, and the absence of CD45, CD34, and CD73, which are positive and negative markers of MSC, respectively. To prove their multipotency, the cells were induced to differentiate into three different cell types, chondrocytes, osteoblasts, and adipocytes, which were stained with tissue-specific dyes (Chondrogenic-Alcian Blue, Osteogenic-Alizarin Red, and Adipogenic-Oil-Red O, respectively) to confirm differentiation. Gene expression analysis of pluripotency-related genes was also conducted. Our results suggest that adipose tissue from bovines and buffalos can be used as a source of MSC, making adipose tissue-derived cells an interesting option for cell therapy and regenerative medicine. Additionally, these findings have implications for reproductive biotechnology because the use of MSC as nuclear donors has been linked to an increase in the efficiency of nuclear transfer.

  13. Protective effect of aqueous jujube extract in Carbamazepine induced teratogenicity on Balb/c mice fetuses

    Directory of Open Access Journals (Sweden)

    Doostabadi Mohammadreza

    2016-06-01

    Full Text Available Aim: Carbamazepine (CBZ is an anticonvulsant medication that can produce congenital anomalies. This study aimed to assess protective role of aqueous jujube extract (JE on CBZ induced congenital anomalies in mice fetuses. Methods:One hundred pregnant Balb/c mice were divided into 8 experimental (E and 2 control (C groups equally. The groups (E1, E5, E6 and (E2, E7, E8 received 50 and 100 mg/kg of CBZ, respectively IP, from GD 0 to GD15. Besides, groups (E5, E7 and (E6, E8 in addition to CBZ, were treated with 200 and 400 mg/kg JE, respectively from ten days prior to gestation, till GD15. The groups E3 and E4 received only 200 and 400 mg/kg of JE respectively. The control groups (C1, C2 received normal saline and tween-20 in turn. On GD18 dams cesarianed and their fetuses assessed for skeletal anomalies by using Alizarin red-alcian blue staining. Results:CBZ induced various anomalies such as; limb defects, craniofacial malformations and etc in mice fetuses. However, these anomalies significantly decreased in groups which were co-administered with CBZ and JE. Conclusion: Co-administration of JE and CBZ significantly decrease teratogenicity of CBZ. Therefore, JE may play a protective role against those properties of CBZ inducing teratogenicity

  14. Optimization of dual effects of Mg-1Ca alloys on the behavior of chondrocytes and osteoblasts in vitro

    Institute of Scientific and Technical Information of China (English)

    Yana Dou; Ayeesha Mujeeb; Yufeng Zheng; Zigang Ge

    2014-01-01

    Mg ions can enhance the proliferation and redifferentiation of chondrocytes and the osteogenic differentiation of osteoblasts at specific concentrations, respectively. However, degradation of Mg alloys at varying degradation rates could lead to complex changes in the surrounding tissue environment, such as changes in the dynamic concentration of Mg ions and subsequent pH value. Considering the above mentioned factors, the comprehensive effects of Mg alloys on chondrocytes and osteoblasts behaviors have not yet been optimized. In this study, we evaluated the effects of Mg–1Ca microspheres on cell behavior with an aim to optimize conditions favorable for both cell types. Cells were cultured with Mg–1Ca microspheres prepared using the following concentrations:250μg/ml, 500μg/ml and 1000μg/ml. At specific time points, cytotoxicity, expression of specific genes and extracellular matrix deposition by cells (Alizarin Red Staining of osteoblasts and Alcian blue staining for chondrocytes) were evaluated. The experimental results revealed that Mg–1Ca microspheres prepared at a concentration of 250μg/ml were optimum for both cell types, where chondrocytes were found to be in hypertrophy state while osteoblasts in close proximity to the microspheres showed osteogenetic differentiation. Interestingly, a slight change in osteoblasts behavior was observed nearer to and at a relative distance away from Mg–1Ca microspheres, an important observation for administering the application of microspheres as potential scaffolds.

  15. Bioprinting Organotypic Hydrogels with Improved Mesenchymal Stem Cell Remodeling and Mineralization Properties for Bone Tissue Engineering.

    Science.gov (United States)

    Duarte Campos, Daniela Filipa; Blaeser, Andreas; Buellesbach, Kate; Sen, Kshama Shree; Xun, Weiwei; Tillmann, Walter; Fischer, Horst

    2016-06-01

    3D-manufactured hydrogels with precise contours and biological adhesion motifs are interesting candidates in the regenerative medicine field for the culture and differentiation of human bone-marrow-derived mesenchymal stem cells (MSCs). 3D-bioprinting is a powerful technique to approach one step closer the native organization of cells. This study investigates the effect of the incorporation of collagen type I in 3D-bioprinted polysaccharide-based hydrogels to the modulation of cell morphology, osteogenic remodeling potential, and mineralization. By combining thermo-responsive agarose hydrogels with collagen type I, the mechanical stiffness and printing contours of printed constructs can be improved compared to pure collagen hydrogels which are typically used as standard materials for MSC osteogenic differentiation. The results presented here show that MSC not only survive the 3D-bioprinting process but also maintain the mesenchymal phenotype, as proved by live/dead staining and immunocytochemistry (vimentin positive, CD34 negative). Increased solids concentrations of collagen in the hydrogel blend induce changes in cell morphology, namely, by enhancing cell spreading, that ultimately contribute to enhanced and directed MSC osteogenic differentiation. 3D-bioprinted agarose-collagen hydrogels with high-collagen ratio are therefore feasible for MSC osteogenic differentiation, contrarily to low-collagen blends, as proved by two-photon microscopy, Alizarin Red staining, and real-time polymerase chain reaction.

  16. Microscopic analysis of an opacified OFT CRYL® hydrophilic acrylic intraocular lens

    Directory of Open Access Journals (Sweden)

    Bruna Vieira Ventura

    Full Text Available ABSTRACT A 51-year-old patient underwent posterior vitrectomy with perfluoropropane gas injection, phacoemulsification, and implantation of an Oft Cryl® hydrophilic acrylic intraocular lens (IOL because of traumatic retinal detachment and cataract in the right eye. On the first postoperative day, gas was filling the anterior chamber because of patient's non-compliance in terms of head positioning, and was reabsorbed within one week. Eight months later, the patient returned complaining of a significant decrease in vision. IOL opacification was noticed by slit-lamp examination. The lens was explanted to undergo gross and light microscopic analysis. The lens was also stained with the alizarin red method for calcium identification. Light microscopic analysis confirmed the presence of granular deposits, densely distributed in an overall circular pattern in the central part of the lens optic. The granules stained positive for calcium. This is the first case of the opacification of this type of hydrophilic lens. Surgeons should be aware of this potential postoperative complication, and the use of hydrophilic IOLs should be avoided in procedures involving intracameral gas because of the risk of IOL opacification.

  17. Extractive determination of ephedrine hydrochloride and bromhexine hydrochloride in pure solutions, pharmaceutical dosage form and urine samples.

    Science.gov (United States)

    Abdel-Ghani, N T; Rizk, M S; Mostafa, M

    2013-07-01

    Simple, rapid, sensitive, precise and accurate spectrophotometeric methods for the determination of ephedrine hydrochloride (E-HCl) and bromhexine hydrochloride (Br-HCl) in bulk samples, dosage form and in spiked urine samples were investigated. The methods are based on the formation of a yellow colored ion-associates due to the interaction between the examined drugs with picric acid (PA), chlorophyllin coppered trisodium salt (CLPH), alizarin red (AR) and ammonium reineckate (Rk) reagents. A buffer solution had been used and the extraction was carried out using organic solvent, the ion associates exhibit absorption maxima at 410, 410, 430 and 530 nm of (Br-HCl)with PA, CLPH, AR and Rk respectively; 410, 410, 435 and 530 of (E-HCl) with PA, CLPH, AR and Rk respectively. (E-HCl) and (Br-HCl) could be determined up to 13, 121, 120 and 160; 25, 200, 92 and 206 μg mL(-1), using PA, CLPH, AR and Rk respectively. The optimum reaction conditions for quantitative analysis were investigated. In addition, the molar absorptivity, Sandell sensitivity were determined for the investigated drug. The correlation coefficient was ≥0.995 (n=6) with a relative standard deviation (RSD) ≤1.15 for five selected concentrations of the reagents. Therefore the concentration of Br-HCl and E-HCl drugs in their pharmaceutical formulations and spiked urine samples had been determined successfully.

  18. Hydrothermal synthesis and characterization of hydroxyapatite and fluorhydroxyapatite nano-size powders

    Energy Technology Data Exchange (ETDEWEB)

    Montazeri, Leila; Javadpour, Jafar [School of Metallurgy and Materials Engineering, Iran University of Science and Technology, Tehran (Iran, Islamic Republic of); Shokrgozar, Mohammad Ali; Bonakdar, Shahin [National Cell Bank of Iran, Pasteur Institute of Iran, Tehran (Iran, Islamic Republic of); Javadian, Sayfoddin, E-mail: javadpourj@iust.ac.i, E-mail: mashokrgozar@pasteur.ac.i [Department of Biochemistry, Pasteur Institute of Iran, Tehran (Iran, Islamic Republic of)

    2010-08-01

    Pure hydroxyapatite (HAp) and fluoride-containing apatite powders (FHAp) were synthesized using a hydrothermal method. The powders were assessed by x-ray diffraction (XRD), Fourier transform infrared (FTIR), scanning electron microscope (SEM) and F-selective electrode. X-ray diffraction results revealed the formation of single phase apatite structure for all the compositions synthesized in this work. However, the addition of a fluoride ion led to a systematic shift in the (3 0 0) peak of the XRD pattern as well as modifications in the FTIR spectra. It was found that the efficiency of fluoride ion incorporation decreased with the increase in the fluoride ion content. Fluorine incorporation efficiency was around 60% for most of the FHAp samples prepared in the current study. Smaller and less agglomerated particles were obtained by fluorine substitution. The bioactivity of the powder samples with different fluoride contents was compared by performing cell proliferation, alkaline phosphatase (ALP) and Alizarin red staining assays. Human osteoblast cells were used to assess the cellular responses to the powder samples in this study. Results demonstrated a strong dependence of different cell activities on the level of fluoridation.

  19. Three-dimensional Expansion: In Suspension Culture of SD Rat's Osteoblasts in a Rotating Wall Vessel Bioreactor

    Institute of Scientific and Technical Information of China (English)

    KE-DONG SONG; TIAN-QING LIU; XIANG-QIN LI; ZHAN-FENG CUI; XIANG-YU SUN; XUE-HU MA

    2007-01-01

    Objective To study large-scale expansion of SD (Sprague-Dawley) rat's osteoblasts in suspension culture in a rotating wall vessel bioreactor (RWVB). Methods The bioreactor rotation speeds were adjusted in the range of 0 to 20 rpm,which could provide low shear on the microcarriers around 1 dyn/cm2. The cells were isolated via sequential digestions of neonatal (less than 3 days old) SD rat calvaria. After the primary culture and several passages, the cells were seeded onto the microcarriers and cultivated in T-flask, spinner flask and RWVB respectively. During the culture period, the cells were counted and observed under the inverted microscope for morphology every 12 h. After 7 days, the cells were evaluated with scanning electron microscope (SEM) for histological examination of the aggregates. Also, the hematoxylin-eosin (HE) staining and alkaline phosphatase (ALP) staining were performed. Moreover, von-Kossa staining and Alizarin Red S staining were carried out for mineralized nodule formation. Results The results showed that in RWVB, the cells could be expanded by more than ten times and they presented better morphology and vitality and stronger ability to form bones. Conclusions The developed RWVB can provide the culture environment with a relatively low shear force and necessary three-dimensional (3D)interactions among cells and is suitable for osteopath expansion in vitro.

  20. Ultrafast photoelectron migration in dye-sensitized solar cells: Influence of the binding mode and many-body interactions

    Science.gov (United States)

    Hermann, G.; Tremblay, J. C.

    2016-11-01

    In the present contribution, the ultrafast photoinduced electron migration dynamics at the interface between an alizarin dye and an anatase TiO2 thin film is investigated from first principles. Comparison between a time-dependent many-electron configuration interaction ansatz and a single active electron approach sheds light on the importance of many-body effects, stemming from uniquely defined initial conditions prior to photoexcitation. Particular emphasis is put on understanding the influence of the binding mode on the migration process. The dynamics is analyzed on the basis of a recently introduced toolset in the form of electron yields, electronic fluxes, and flux densities, to reveal microscopic details of the electron migration mechanism. From the many-body perspective, insight into the nature of electron-electron and hole-hole interactions during the charge transfer process is obtained. The present results reveal that the single active electron approach yields quantitatively and phenomenologically similar results as the many-electron ansatz. Furthermore, the charge migration processes in the dye-TiO2 model clusters with different binding modes exhibit similar mechanistic pathways but on largely different time scales.

  1. Surface modification by allylamine plasma polymerization promotes osteogenic differentiation of human adipose-derived stem cells.

    Science.gov (United States)

    Liu, Xujie; Feng, Qingling; Bachhuka, Akash; Vasilev, Krasimir

    2014-06-25

    Tuning the material properties in order to control the cellular behavior is an important issue in tissue engineering. It is now well-established that the surface chemistry can affect cell adhesion, proliferation, and differentiation. In this study, plasma polymerization, which is an appealing method for surface modification, was employed to generate surfaces with different chemical compositions. Allylamine (AAm), acrylic acid (AAc), 1,7-octadiene (OD), and ethanol (ET) were used as precursors for plasma polymerization in order to generate thin films rich in amine (-NH2), carboxyl (-COOH), methyl (-CH3), and hydroxyl (-OH) functional groups, respectively. The surface chemistry was characterized by X-ray photoelectron spectroscopy (XPS), the wettability was determined by measuring the water contact angles (WCA) and the surface topography was imaged by atomic force microscopy (AFM). The effects of surface chemical compositions on the behavior of human adipose-derive stem cells (hASCs) were evaluated in vitro: Cell Count Kit-8 (CCK-8) analysis for cell proliferation, F-actin staining for cell morphology, alkaline phosphatase (ALP) activity analysis, and Alizarin Red S staining for osteogenic differentiation. The results show that AAm-based plasma-polymerized coatings can promote the attachment, spreading, and, in turn, proliferation of hASCs, as well as promote the osteogenic differentiation of hASCs, suggesting that plasma polymerization is an appealing method for the surface modification of scaffolds used in bone tissue engineering.

  2. Osteogenic differentiation of osteoblasts induced by calcium silicate and calcium silicate/β-tricalcium phosphate composite bioceramics.

    Science.gov (United States)

    Fei, Lisha; Wang, Chen; Xue, Yang; Lin, Kaili; Chang, Jiang; Sun, Jiao

    2012-07-01

    In this study, calcium silicate (CS) and CS/β-tricalcium phosphate (CS/β-TCP) composites were investigated on their mechanism of osteogenic proliferation and differentiation through regulating osteogenic-related gene and proteins. Osteoblast-like cells were cultured in the extracts of these CS-based bioceramics and pure β-TCP, respectively. The main ionic content in extracts was analyzed by inductively coupled plasma-atomic emission spectroscopy. The cell viability, mineralization, and differentiation were evaluated by MTT assay, Alizarin Red-S staining and alkaline phosphatase (ALP) activity assay. The expressions of BMP-2, transforming growth factor-β (TGF-β), Runx2, ALP, and osteocalcin (OCN) at both gene and protein level were detected by real-time polymerase chain reaction analysis and Western blot. The result showed that the extracts of CS-based bioceramics promoted cells proliferation, differentiation, and mineralization when compared with pure β-TCP. Accordingly, pure CS and CS/β-TCP composites stimulated osteoblast-like cells to express BMP-2/TGF-β gene and proteins, and further regulate the expression of Runx2 gene and protein, and ultimately affect the ALP activity and OCN deposition. This study suggested that the CS-based bioceramics could not only promote the expression of osteogenic-related genes but also enhance the genes to encode the corresponding proteins, which could finally control osteoblast-like cells proliferation and differentiation.

  3. Extractive determination of ephedrine hydrochloride and bromhexine hydrochloride in pure solutions, pharmaceutical dosage form and urine samples

    Science.gov (United States)

    Abdel-Ghani, N. T.; Rizk, M. S.; Mostafa, M.

    2013-07-01

    Simple, rapid, sensitive, precise and accurate spectrophotometeric methods for the determination of ephedrine hydrochloride (E-HCl) and bromhexine hydrochloride (Br-HCl) in bulk samples, dosage form and in spiked urine samples were investigated. The methods are based on the formation of a yellow colored ion-associates due to the interaction between the examined drugs with picric acid (PA), chlorophyllin coppered trisodium salt (CLPH), alizarin red (AR) and ammonium reineckate (Rk) reagents. A buffer solution had been used and the extraction was carried out using organic solvent, the ion associates exhibit absorption maxima at 410, 410, 430 and 530 nm of (Br-HCl)with PA, CLPH, AR and Rk respectively; 410, 410, 435 and 530 of (E-HCl) with PA, CLPH, AR and Rk respectively. (E-HCl) and (Br-HCl) could be determined up to 13, 121, 120 and 160; 25, 200, 92 and 206 μg mL-1, using PA, CLPH, AR and Rk respectively. The optimum reaction conditions for quantitative analysis were investigated. In addition, the molar absorptivity, Sandell sensitivity were determined for the investigated drug. The correlation coefficient was ⩾0.995 (n = 6) with a relative standard deviation (RSD) ⩽1.15 for five selected concentrations of the reagents. Therefore the concentration of Br-HCl and E-HCl drugs in their pharmaceutical formulations and spiked urine samples had been determined successfully.

  4. Effect of BMP2-Platelet-rich Plasma-Biphasic Calcium Phosphate Scaffold on Accelerated Osteogenesis in Mastoid Obliteration.

    Science.gov (United States)

    Jang, Chul Ho; Choi, Cheol Hee; Cho, Yong Beom

    The aim of this study was to evaluate the synergistic effect of platelet-rich plasma (PRP) and recombinant human bone morphogenic protein (BMP)-2 on accelerated osteogenesis of hydroxyapatite/β-tricalcium phosphate mixture and biphasic calcium phosphate (BCP) in mastoid obliteration. To the best of our knowledge, there have been no studies reporting the enhancing effects of BCP, combined with BMP2 and PRP, on osteogenesis in mastoid obliteration. Mastoid obliteration was performed in a control group (BCP only, n=7), a group treated with BMP2 and BCP (experimental group I, n=7), and a group treated with BMP2, PRP and BCP (experimental group II, n=7). The animals were administered fluorescent bone labels for a qualitative evaluation of bone formation; oxytetracycline hydrochloride was administered at 2 weeks, calcein at 4 weeks, and alizarin red at 8 weeks. The animals were sacrificed 12 weeks post-surgery and osteogenesis was evaluated by micro-computed tomography, histological investigation, and histomorphometry. Both experimental groups showed accelerated osteogenesis compared to the control group. However, there were no statistically significant differences between experimental groups I and II. From these results, it can be concluded that BMP2 activated BCP for the enhancement of bone regeneration. However, no synergistic effect of BMP2 and PRP on the osteogenesis of BCP was observed.

  5. Enhanced healing of rat calvarial defects with MSCs loaded on BMP-2 releasing chitosan/alginate/hydroxyapatite scaffolds.

    Directory of Open Access Journals (Sweden)

    Xiaoning He

    Full Text Available In this study, we designed a chitosan/alginate/hydroxyapatite scaffold as a carrier for recombinant BMP-2 (CAH/B2, and evaluated the release kinetics of BMP-2. We evaluated the effect of the CAH/B2 scaffold on the viability and differentiation of bone marrow mesenchymal stem cells (MSCs by scanning electron microscopy, MTS, ALP assay, alizarin-red staining and qRT-PCR. Moreover, MSCs were seeded on scaffolds and used in a 8 mm rat calvarial defect model. New bone formation was assessed by radiology, hematoxylin and eosin staining 12 weeks postoperatively. We found the release kinetics of BMP-2 from the CAH/B2 scaffold were delayed compared with those from collagen gel, which is widely used for BMP-2 delivery. The BMP-2 released from the scaffold increased MSC differentiation and did not show any cytotoxicity. MSCs exhibited greater ALP activity as well as stronger calcium mineral deposition, and the bone-related markers Col1α, osteopontin, and osteocalcin were upregulated. Analysis of in vivo bone formation showed that the CAH/B2 scaffold induced more bone formation than other groups. This study demonstrates that CAH/B2 scaffolds might be useful for delivering osteogenic BMP-2 protein and present a promising bone regeneration strategy.

  6. Alveolar bone dynamics in osteoporotic rats treated with raloxifene or alendronate: confocal microscopy analysis

    Science.gov (United States)

    Ramalho-Ferreira, Gabriel; Faverani, Leonardo Perez; Grossi-Oliveira, Gustavo Augusto; Okamoto, Tetuo; Okamoto, Roberta

    2015-03-01

    In this study, the characteristics of the alveolar bone of rats with induced osteoporosis were examined. Thirty-two rats were divided into four groups according to the induction of osteoporosis and drugs administered: OG, osteoporotic rats without treatment (negative control); SG, rats which underwent sham surgery ovariectomy (SHAM); alendronate (AG), osteoporotic rats treated with alendronate; and RG, osteoporotic rats treated with raloxifene (RG). On the 8th day after ovariectomy and SHAM surgeries, drug therapy was started with AG or RG. On the 52nd day, 20 mg/kg calcein was administered to all of the rats, and on the 80th day, 20 mg/kg alizarin red was administered. Euthanasia was performed on the 98th day. The bone area marked by fluorochromes was calculated and data were subjected to two-way ANOVA test and Tukey's post-hoc test (pbone turnover only between RG and SG (p=0.074) and AG and OG (p=0.138). All other comparisons showed significant differences (pbone turnover was observed in RG and SG groups. RG was the medication that improved the dynamics of the alveolar bone of rats with induced osteoporosis, resembling that of healthy rats.

  7. In vitro culture and characterization of alveolar bone osteoblasts isolated from type 2 diabetics

    Energy Technology Data Exchange (ETDEWEB)

    Sun, Dao-Cai [Department of Implant Dentistry, School of Stomatology, Fourth Military Medical University, Xi' an (China); Department of Stomatology, The 291st Hospital of P.L.A, Baotou (China); Li, De-Hua [Department of Implant Dentistry, School of Stomatology, Fourth Military Medical University, Xi' an (China); Ji, Hui-Cang [Military Sanatorium of Retired Cadres, Baotou (China); Rao, Guo-Zhou [Center of Laboratory, School of Stomatology, Xi' an Jiaotong University, Xi' an (China); Liang, Li-Hua [Department of Implant Dentistry, School of Stomatology, Fourth Military Medical University, Xi' an (China); Ma, Ai-Jie [Xi' an Technology University, Xi' an (China); Xie, Chao; Zou, Gui-Ke; Song, Ying-Liang [Department of Implant Dentistry, School of Stomatology, Fourth Military Medical University, Xi' an (China)

    2012-04-05

    In order to understand the mechanisms of poor osseointegration following dental implants in type 2 diabetics, it is important to study the biological properties of alveolar bone osteoblasts isolated from these patients. We collected alveolar bone chips under aseptic conditions and cultured them in vitro using the tissue explants adherent method. The biological properties of these cells were characterized using the following methods: alkaline phosphatase (ALP) chemical staining for cell viability, Alizarin red staining for osteogenic characteristics, MTT test for cell proliferation, enzyme dynamics for ALP contents, radio-immunoassay for bone gla protein (BGP) concentration, and ELISA for the concentration of type I collagen (COL-I) in the supernatant. Furthermore, we detected the adhesion ability of two types of cells from titanium slices using non-specific immunofluorescence staining and cell count. The two cell forms showed no significant difference in morphology under the same culture conditions. However, the alveolar bone osteoblasts received from type 2 diabetic patients had slower growth, lower cell activity and calcium nodule formation than the normal ones. The concentration of ALP, BGP and COL-I was lower in the supernatant of alveolar bone osteoblasts received from type 2 diabetic patients than in that received from normal subjects (P < 0.05). The alveolar bone osteoblasts obtained from type 2 diabetic patients can be successfully cultured in vitro with the same morphology and biological characteristics as those from normal patients, but with slower growth and lower concentration of specific secretion and lower combining ability with titanium than normal ones.

  8. Osteogenic potential of human umbilical cord-derived mesenchymal stromal cells cultured with umbilical cord blood-derived fibrin: a preliminary study.

    Science.gov (United States)

    Baba, Kyoko; Yamazaki, Yasuharu; Ishiguro, Masashi; Kumazawa, Kenichi; Aoyagi, Kazuya; Ikemoto, Shigehiro; Takeda, Akira; Uchinuma, Eiju

    2013-12-01

    This study examined the potential for osteogenesis via regenerative medicine using autologous tissues (umbilical cord (UC) and umbilical cord blood (UCB)) in nude mice. The study was designed to provide the three elements required for regenerative medicine (cell, scaffold, and growth factor) and autoserum for culture by means of autologous tissues. Mesenchymal stromal cells were obtained from UC (UC-MSCs). Fibrin, platelet-rich-plasma, and autoserum were obtained from UCB as scaffold, growth factor and serum for culture respectively. UC-MSCs were obtained from Wharton jelly and cultured with UCB-derived fibrin (UCB-fibrin) for 3-4 weeks to induce their differentiation into osteoblasts. They were implanted subcutaneously into the dorsum of male nude mice for 6 weeks prior to undergoing assessment. The assessments performed were haematoxylin and eosin, and alizarin red staining, immunohistochemical staining of human mitochondria, scanning electron microscopy, scanning electron microscopy with energy dispersive X-ray spectrometry and real-time reverse transcriptase-polymerase chain reaction to assess the expressions of osteoblast markers. Consequently, the differentiation of UC-MSCs into osteoblasts and the production of hydroxyapatite were verified. This study suggested the possible formation of bone tissue using biomedical materials obtained from UC and UCB.

  9. Induction of differentiation by down-regulation of Nanog and Rex-1 in cord blood derived unrestricted somatic stem cells.

    Science.gov (United States)

    Langroudi, Lida; Forouzandeh, Mehdi; Soleimani, Masoud; Atashi, Amir; Golestaneh, Azadeh Fahim

    2013-07-01

    Stem cells with high self-renewal and tissue regeneration potentials are the core components of regenerative medicine. Adult stem cells with many available sources, high repairing ability, and also possessing no ethical issues are popular candidates in the clinical field. In this study we looked upon the effects of two transcription factors Nanog and Rex-1 in self-renewal and differentiation abilities of a subpopulation of cord blood stem cells known as unrestricted somatic stem cells (USSCs). USSCs were expanded and transfected in vitro with siRNAs targeting either Nanog, Rex-1, and in combination. Gene suppressions were achieved at both transcript and proteome level. Differentiations were evaluated by specific Real time PCR and differentiating staining. Nanog knock down revealed a significant increase in osteogenic markers, Osteocalcin and Osteopontin expression as well as a positive Alizarin Red staining, which proposes Osteogenesis. This treatment also became positive for Oil Red staining, implying adipogenic differentiation as well. In contrast, Rex-1 knock down showed an increase in MAP II and Nestin expression, which is a hall mark of neural differentiation. Surprisingly, treatment with both siRNAs did not express any changes in any of the assessed markers. Therefore, our results indicated a bilateral mesenchymal differentiation for Nanog and a neural lineage fate for Rex-1 suppression. Considering that both transcription factors are core activators of self-renewal and also are orchestrating with other factors, our results imply a positive feedback in response to changes in the regulatory network of self-renewal.

  10. Involvement of Toll-Like Receptor 2 and Pro-Apoptotic Signaling Pathways in Bone Remodeling in Osteomyelitis

    Directory of Open Access Journals (Sweden)

    Qianbo Chen

    2014-11-01

    Full Text Available Background and Aims: Osteomyelitis is a common manifestation of invasive Staphylococcus aureus infection characterized by bone loss and destruction. We investigated the role of toll-like receptor 2 (TLR2 in bacterial recognition and clearance in response to infection with an osteomyelitis isolate of S. aureus. Methods: Apoptosis was assessed in the osteoblastic cell line MC3T3-E1 by Annexin V-FITC/PI staining and flow cytometry. The expression of TLR2 and apoptosis-related and mitogen-activated protein kinase pathway proteins was assessed by qRT-PCR and western blotting. Alkaline phosphatase (ALP activity and calcium deposition were assessed by ALP activity assay and Alizarin red staining. Results: S. aureus induced apoptosis, upregulated TLR2 expression, and activated mitogen-activated protein kinase pathways in a time dependent manner. Inhibition of the c-Jun N-terminal kinase (JNK pathway downregulated TLR2 and suppressed the S. aureus induced activation of pro-apoptotic pathways. Short-hairpin RNA mediated silencing of TLR2 reversed S. aureus induced apoptosis and decrease in ALP activity and calcium deposition, and inhibition of JNK had a similar effect. Conclusion: We showed that osteoblast apoptosis and osteogenic differentiation in response to bacterial invasion are dependent on TLR2 expression and JNK activation, suggesting novel potential therapeutic targets for the treatment of osteomyelitis.

  11. Combination of Bioactive Polymeric Membranes and Stem Cells for Periodontal Regeneration: In Vitro and In Vivo Analyses

    Science.gov (United States)

    Gonçalves, Flávia; de Moraes, Míriam Santos; Ferreira, Lorraine Braga; Carreira, Ana Cláudia Oliveira; Kossugue, Patrícia Mayumi; Boaro, Letícia Cristina Cidreira; Bentini, Ricardo; Garcia, Célia Regina da Silva; Sogayar, Mari Cleide; Arana-Chavez, Victor Elias; Catalani, Luiz Henrique

    2016-01-01

    Regeneration of periodontal tissues requires a concerted effort to obtain consistent and predictable results in vivo. The aim of the present study was to test a new family of bioactive polymeric membranes in combination with stem cell therapy for periodontal regeneration. In particular, the novel polyester poly(isosorbide succinate-co-L-lactide) (PisPLLA) was compared with poly(L-lactide) (PLLA). Both polymers were combined with collagen (COL), hydroxyapatite (HA) and the growth factor bone morphogenetic protein-7 (BMP7), and their osteoinductive capacity was evaluated via in vitro and in vivo experiments. Membranes composed of PLLA/COL/HA or PisPLLA/COL/HA were able to promote periodontal regeneration and new bone formation in fenestration defects in rat jaws. According to quantitative real-time polymerase chain reaction (qRT-PCR) and Alizarin Red assays, better osteoconductive capacity and increased extracellular mineralization were observed for PLLA/COL/HA, whereas better osteoinductive properties were associated with PisPLLA/COL/HA. We concluded that membranes composed of either PisPLLA/COL/HA or PLLA/COL/HA present promising results in vitro as well as in vivo and that these materials could be potentially applied in periodontal regeneration. PMID:27031990

  12. Low-power laser irradiation promotes the proliferation and osteogenic differentiation of human periodontal ligament cells via cyclic adenosine monophosphate

    Institute of Scientific and Technical Information of China (English)

    Jyun-Yi Wu; Chia-Hsin Chen; Li-Yin Yeh; Ming-Long Yeh; Chun-Chan Ting; Yan-Hsiung Wang

    2013-01-01

    Retaining or improving periodontal ligament (PDL) function is crucial for restoring periodontal defects. The aim of this study was to evaluate the physiological effects of low-power laser irradiation (LPLI) on the proliferation and osteogenic differentiation of human PDL (hPDL) cells. Cultured hPDL cells were irradiated (660 nm) daily with doses of 0, 1, 2 or 4 J?cm22. Cell proliferation was evaluated by the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay, and the effect of LPLI on osteogenic differentiation was assessed by Alizarin Red S staining and alkaline phosphatase (ALP) activity. Additionally, osteogenic marker gene expression was confirmed by real-time reverse transcription-polymerase chain reaction (RT-PCR). Our data showed that LPLI at a dose of 2 J?cm22 significantly promoted hPDL cell proliferation at days 3 and 5. In addition, LPLI at energy doses of 2 and 4 J?cm22 showed potential osteogenic capacity, as it stimulated ALP activity, calcium deposition, and osteogenic gene expression. We also showed that cyclic adenosine monophosphate (cAMP) is a critical regulator of the LPLI-mediated effects on hPDL cells. This study shows that LPLI can promote the proliferation and osteogenic differentiation of hPDL cells. These results suggest the potential use of LPLI in clinical applications for periodontal tissue regeneration.

  13. Efficient azo dye decolorization in a continuous stirred tank reactor (CSTR) with built-in bioelectrochemical system.

    Science.gov (United States)

    Cui, Min-Hua; Cui, Dan; Gao, Lei; Cheng, Hao-Yi; Wang, Ai-Jie

    2016-10-01

    A continuous stirred tank reactor with built-in bioelectrochemical system (CSTR-BES) was developed for azo dye Alizarin Yellow R (AYR) containing wastewater treatment. The decolorization efficiency (DE) of the CSTR-BES was 97.04±0.06% for 7h with sludge concentration of 3000mg/L and initial AYR concentration of 100mg/L, which was superior to that of the sole CSTR mode (open circuit: 54.87±4.34%) and the sole BES mode (without sludge addition: 91.37±0.44%). The effects of sludge concentration and sodium acetate (NaAc) concentration on azo dye decolorization were investigated. The highest DE of CSTR-BES for 4h was 87.66±2.93% with sludge concentration of 12,000mg/L, NaAc concentration of 2000mg/L and initial AYR concentration of 100mg/L. The results in this study indicated that CSTR-BES could be a practical strategy for upgrading conventional anaerobic facilities against refractory wastewater treatment.

  14. Effects of novel hydroxyapatite-based 3D biomaterials on proliferation and osteoblastic differentiation of mesenchymal stem cells.

    Science.gov (United States)

    Karadzic, Ivana; Vucic, Vesna; Jokanovic, Vukoman; Debeljak-Martacic, Jasmina; Markovic, Dejan; Petrovic, Snjezana; Glibetic, Marija

    2015-01-01

    The aim of this study was to examine the differential capacity of isolated dental pulp stem cells (SHED) cultured onto four different scaffold materials. The differential potential of isolated SHED was examined on the following scaffolds: porous hydroxyapatite (pHAP) alone or combined with three polymers [polylactic-co-glycolic acid (PLGA), alginate, and ethylene vinylacetate / ethylene vinylversatate (EVA/EVV)]. SHED were isolated by "outgrowth" method and characterized by the flow cytometry. Viability of cells grown with scaffolds was assessed by MTT and LDH assays. No significant cytotoxic effect of any of the tested materials was shown. Staining with alizarin red and estimated alkaline phosphatase activity to identify differentiation, demonstrated osteoblastic phenotype of SHED and newly deposited and mineralized extra cellular matrix (ECM) in presence of all tested scaffolds. The developed ECM seen at scanning electronic micrographs additionally confirmed the osteogenic differentiation and biocompatibility between cells and materials. In summary, all studied biomaterials are suitable carriers for proliferation and osteoblastic differentiation of dental pulp mesenchymal stem cells in vitro.

  15. Calcium phosphate nanoparticles carrying BMP-7 plasmid DNA induce an osteogenic response in MC3T3-E1 pre-osteoblasts.

    Science.gov (United States)

    Hadjicharalambous, Chrystalleni; Kozlova, Diana; Sokolova, Viktoriya; Epple, Matthias; Chatzinikolaidou, Maria

    2015-12-01

    Functionalized calcium phosphate nanoparticles with osteogenic activity were prepared. Polyethyleneimine-stabilized calcium phosphate nanoparticles were coated with a shell of silica and covalently functionalized by silanization with thiol groups. Between the calcium phosphate surface and the outer silica shell, plasmid DNA which encoded either for bone morphogenetic protein 7 (BMP-7) or for enhanced green fluorescent protein was incorporated as cargo. The plasmid DNA-loaded calcium phosphate nanoparticles were used for the transfection of the pre-osteoblastic MC3T3-E1 cells. The cationic nanoparticles showed high transfection efficiency together with a low cytotoxicity. Their potential to induce an osteogenic response by transfection was demonstrated by measuring the alkaline phosphatase (ALP) activity and calcium deposition with alizarin red staining. The expression of the osteogenic markers Alp, Runx2, ColIa1 and Bsp was investigated by means of real-time quantitative polymerase chain reaction. It was shown that phBMP-7-loaded nanoparticles can provide a means of transient transfection and localized production of BMP-7 in MC3T3-E1 cells, with a subsequent increase of two osteogenic markers, specifically ALP activity and calcium accumulation in the extracellular matrix. Future strategies to stimulate bone regeneration focus into enhancing transfection efficiency and achieving higher levels of BMP-7 produced by the transfected cells.

  16. EFFECT OF RADIX SALVIAE MILTIORRHIZAE ON GROWTH OF ISOLATED CELLS FROM EMBRYONIC CHICKEN FRONTAL BONE CULTURED IN VITRO (A HISTOCHEMICAL STUDY) Ⅱ.THE DEVELOPMENT AND MATURATION OF OSTEOBLAST-LIKE CELLS

    Institute of Scientific and Technical Information of China (English)

    徐荣辉; 柴本甫; 朱雅萍

    1993-01-01

    The isolated osteoblast-like cells from embryonic chicken frontal bone werecultured in vitro and histochemical methods adopted to observe the effect of RadixSalviac Miltiorrhizae (RSM) on proliferation, differentiation, and osteogenic capacity ofthese cells. It was found that: 1. The mitosis and proliferation of the osteoblast-like cellscould be accelerated by RSM, resulting in increased density of the cells in RSM groupas compared with the control. 2. After 48 h, the pseudopodia stretched out and drew backactively in osteoblast-like cells in RSM group. Small particles produced in the cells weresecreted through exocytosis to the extracellular medium. However, in the control group,the capacity to form and secrete these particles was limited. These particles showed posi-tive Alcian blue staining in Alcian blue-Sirius red reaction, so they were acidmucopolysaccharide particles. 3. The osteoblast-like cells could secrete vesicular particles 3micra in diameter. These vesicular particles could be stained with Alcian blue in earlystage, then they could be stained with Sirius red, and finally by Alizarin red S. Thesevesicular particles could aggregate and fuse around the cell colonies, forming bonenodules and bone flakes. The quantity and volume of the bone nodules and flakes inRSM group were larger than in the control group. 4. The bone nodules and flakes couldbe labeled vitally with tetracycline, and show strong yellow fluorescence under thefluorescence microscope. Therefore, these substances were the newly formed bone sub-stances.

  17. The Biomineralization of a Bioactive Glass-Incorporated Light-Curable Pulp Capping Material Using Human Dental Pulp Stem Cells

    Science.gov (United States)

    Jun, Soo-Kyung; Lee, Hae-Hyoung

    2017-01-01

    The aim of this study was to investigate the biomineralization of a newly introduced bioactive glass-incorporated light-curable pulp capping material using human dental pulp stem cells (hDPSCs). The product (Bioactive® [BA]) was compared with a conventional calcium hydroxide-incorporated (Dycal [DC]) and a light-curable (Theracal® [TC]) counterpart. Eluates from set specimens were used for investigating the cytotoxicity and biomineralization ability, determined by alkaline phosphatase (ALP) activity and alizarin red staining (ARS). Cations and hydroxide ions in the extracts were measured. An hDPSC viability of less than 70% was observed with 50% diluted extract in all groups and with 25% diluted extract in the DC. Culturing with 12.5% diluted BA extract statistically lowered ALP activity and biomineralization compared to DC (p 0.05). Ca (~110 ppm) and hydroxide ions (pH 11) were only detected in DC and TC. Ionic supplement-added BA, which contained similar ion concentrations as TC, showed similar ARS mineralization compared to TC. In conclusion, the BA was similar to, yet more cytotoxic to hDPSCs than, its DC and TC. The BA was considered to stimulate biomineralization similar to DC and TC only when it released a similar amount of Ca and hydroxide ions. PMID:28232937

  18. The Biomineralization of a Bioactive Glass-Incorporated Light-Curable Pulp Capping Material Using Human Dental Pulp Stem Cells

    Directory of Open Access Journals (Sweden)

    Soo-Kyung Jun

    2017-01-01

    Full Text Available The aim of this study was to investigate the biomineralization of a newly introduced bioactive glass-incorporated light-curable pulp capping material using human dental pulp stem cells (hDPSCs. The product (Bioactive® [BA] was compared with a conventional calcium hydroxide-incorporated (Dycal [DC] and a light-curable (Theracal® [TC] counterpart. Eluates from set specimens were used for investigating the cytotoxicity and biomineralization ability, determined by alkaline phosphatase (ALP activity and alizarin red staining (ARS. Cations and hydroxide ions in the extracts were measured. An hDPSC viability of less than 70% was observed with 50% diluted extract in all groups and with 25% diluted extract in the DC. Culturing with 12.5% diluted BA extract statistically lowered ALP activity and biomineralization compared to DC (p0.05. Ca (~110 ppm and hydroxide ions (pH 11 were only detected in DC and TC. Ionic supplement-added BA, which contained similar ion concentrations as TC, showed similar ARS mineralization compared to TC. In conclusion, the BA was similar to, yet more cytotoxic to hDPSCs than, its DC and TC. The BA was considered to stimulate biomineralization similar to DC and TC only when it released a similar amount of Ca and hydroxide ions.

  19. Effect of Erythromycin on Albendazole-Induced Teratogenicity in Pregnant Rats

    Directory of Open Access Journals (Sweden)

    Reza Ranjbar

    2013-05-01

    Full Text Available Background: Albendazole is utilized as an anthelmentic agent. One its side effect is teratogenicity. This effect apparently is related to its metabolites especially albendazole sulfoxid. The aim of present study was evaluation effect of erythromycin (as enzyme inhibitor in biotransformation on albendazole biotransformation and consequently fetal malformation. Materials and Methods: Four groups of female pregnant wistar rats (8 rats each group were used. First group received normal saline (as control group. A single oral dose 30 mg/kg of albendazole was administered to rats on day 10 of gestation in group 2. Rats in group 3 received albendazole similar group 2 and erythromycin at dose 60 mg/kg. Rats in group 4 received only erythromycin on day 10 of gestation. The rats were euthanatized on day 20 of gestation. The skeletal malformation of fetus was studied by stereomicroscope after staining by Alizarin red-Alcian blue.Results: The length and weight of fetuses were significantly decreased by albendazole but erythromycin did not prevent this effect. In group that received only erythromycin, the length and weight of fetuses was similar to control group. Erythromycin decreased albendazole effect on weight of placenta. There was an increase in resorption by erythromycin when co-administrated with albendazole. The incidence of skeletal malformations (mostly of the limbs, vertebrae and palate decreased significantly by erythromycin when co-administrated with albendazole.Conclusion: Thus, erythromycin may inhibit albendazole biotransformation and decrease teratogenicity of it metabolites; but this subject needs more detailed evaluation.

  20. The effects of ultraviolet radiation on growth and bleaching in three species of Hawaiian coral

    Energy Technology Data Exchange (ETDEWEB)

    Goodman, G.D. (California State Univ., Long Beach (United States))

    1990-01-09

    Long term exposure to ultraviolet radiation is harmful to many organisms, including hermatypic corals, which obtain much of their nutrition from photosynthetic zooxanthellae. Therefore, increased UV radiation from atmospheric ozone depletion could inhibit growth of such corals. Moreover, coral bleaching, which has been attributed to loss of pigment and/or expulsion of zooxanthellae, may be a specific response to UV light. Does UV-A reduce skeletal growth or influence population density and pigment content of zooxanthellae In addition, do zooxanthellae migrate to shaded areas of the colony to avoid ultraviolet light Using alizarin red stain and suitable filters, I compared the stain and suitable filters, I compared the effects of UV-A (320-400nm) and full-spectrum UV (280-400nm) on the skeletal growth of two Hawaiian corals, Montipora verrucosa, Pocillopora damicornis, in situ. In the perforate corals, M. Verrucosa and Porites compressa, I measured concentration of zooxanthellae and their chlorophyll content to quantify bleaching in response to UV light. Reduction in skeletal growth by the two corals in response to different ranges of UV light appears to be species specific. Bleaching by UV appears to be characterized by an initial loss of pigment followed by the expulsion and migration of the zooxanthellae to shaded areas of the colony. Differences in tolerance and adaptation to decreasing ozone levels and increasing UV light should confer a competitive advantage on various species and morphologies of reef-building corals.

  1. Vascular Adventitia Calcification and Its Underlying Mechanism.

    Directory of Open Access Journals (Sweden)

    Na Li

    Full Text Available Previous research on vascular calcification has mainly focused on the vascular intima and media. However, we show here that vascular calcification may also occur in the adventitia. The purpose of this work is to help elucidate the pathogenic mechanisms underlying vascular calcification. The calcified lesions were examined by Von Kossa staining in ApoE-/- mice which were fed high fat diets (HFD for 48 weeks and human subjects aged 60 years and older that had died of coronary heart disease, heart failure or acute renal failure. Explant cultured fibroblasts and smooth muscle cells (SMCswere obtained from rat adventitia and media, respectively. After calcification induction, cells were collected for Alizarin Red S staining. Calcified lesions were observed in the aorta adventitia and coronary artery adventitia of ApoE-/-mice, as well as in the aorta adventitia of human subjects examined. Explant culture of fibroblasts, the primary cell type comprising the adventitia, was successfully induced for calcification after incubation with TGF-β1 (20 ng/ml + mineralization media for 4 days, and the phenotype conversion vascular adventitia fibroblasts into myofibroblasts was identified. Culture of SMCs, which comprise only a small percentage of all cells in the adventitia, in calcifying medium for 14 days resulted in significant calcification.Vascular calcification can occur in the adventitia. Adventitia calcification may arise from the fibroblasts which were transformed into myofibroblasts or smooth muscle cells.

  2. Human Amnion-Derived Mesenchymal Stem Cells Promote Osteogenic Differentiation in Human Bone Marrow Mesenchymal Stem Cells by Influencing the ERK1/2 Signaling Pathway

    Directory of Open Access Journals (Sweden)

    Yuli Wang

    2016-01-01

    Full Text Available Human amnion-derived mesenchymal stem cells (HAMSCs are considered to be an important resource in the field of tissue engineering because of their anti-inflammatory properties and fewer ethical issues associated with their use compared with other sources of stem cells. HAMSCs can be obtained from human amniotic membranes, a readily available and abundant tissue. However, the potential of HAMSCs as seed cells for treating bone deficiency is unknown. In this study, HAMSCs were used to promote proliferation and osteoblastic differentiation in human bone marrow mesenchymal stem cells (HBMSCs in a Transwell coculture system. Proliferation levels were investigated by flow cytometry and immunofluorescence staining of 5-ethynyl-2′-deoxyuridine (EdU. Osteoblastic differentiation and mineralization were evaluated in chromogenic alkaline phosphatase (ALP activity substrate assays, Alizarin red S staining, and RT-PCR analysis of early HBMSCs osteogenic marker expression. We demonstrated that HAMSCs stimulated increased alkaline phosphatase (ALP activity, mRNA expression of osteogenic marker genes, and mineralized matrix deposition. Moreover, the effect of HAMSCs was significantly inhibited by U0126, a highly selective inhibitor of extracellular signaling-regulated kinase 1/2 (ERK1/2 signaling. We demonstrate that HAMSCs promote osteogenic differentiation in HBMSCs by influencing the ERK1/2 signaling pathway. These observations confirm the potential of HAMSCs as a seed cell for the treatment of bone deficiency.

  3. 犬牙囊干细胞膜片的构建及生物学特性的研究%Construction of Dental Follicle Stem Cell Sheet and its Biological Characteristics in Beagle Dogs

    Institute of Scientific and Technical Information of China (English)

    黄闯; 宋镜明; 宋扬; 刘佳; 王丽颖; 金作林

    2012-01-01

    Objective: To study the construction of dental follicle stem cell sheet and its biological characteristics in Beagle dogs. Methods: After identification.DFSCs were sub-cultured to construct DFSCs sheet. Cell sheet was investaged by inverted microscope, HE staining and scanning electron microscope (SEM). DFSCs sheets were induced by adipogenesis inducing medium and osteogenic medium for 14 days separately. Oil red staining and Alizarin red staining was applied to examine adipogenic induction and osteogenic induction. Results: DFSCs showed typical spindle shape.. Colony-forming assay results showed about 5.1% DFSCs colony formation. DFSCs were positive for CD29 and CD44, but negative for CD34. MTT manifested the growth and proliferation was good. Cell cycle testing showed: G1=87.1%,G2=5.54%. DFSCs sheets were constructed successfully and its growth in multilayer. It found that DFSCs expanded adequately and extracellular matrix(ECM) was clear and numerous in scanning electron micrescopy.Oil red staining and alizarin red staining both demonstrated positive reactions in DFSCs sheet after induction. Conclusion: It suggested that DFSCs cell sheet may be constructed and has a strong bone-forming ability.%目的:利用犬牙囊干细胞(Dental Follicle Stem Cells,DFSCs)构建细胞膜片并研究其生物学特性.方法:取4至6月龄犬尖牙牙胚,分离培养DFSCs,鉴定.用含抗坏血酸的培养基诱导2周构建细胞膜片,并通过倒置显微镜、HE染色、茜素红染色、油红染色、扫描电镜(SEM)对膜片进行形态学检测,检测成骨、成脂能力.结果:DFSCs于体外被成功分离、纯化、培养,细胞克隆形成率约为5.1%.流式鉴定为CD29+CD44+CD34-,增殖能力及克隆形成能力较强,并能成功构建成细胞膜片.光镜和电镜显示膜片细胞排列紧密,细胞基质分泌多,油红O染色后可见细胞内有大量脂滴形成.(B)茜素红染色后可见大量清晰的钙结节形成.结论:成功构建犬DFSCs膜

  4. Experimental study of human BMP-2 on osteogenic induction in BMSCs of dogs in vitro%人BMP-2体外定向诱导犬BMSCs向成骨方向分化的实验研究

    Institute of Scientific and Technical Information of China (English)

    许蕾; 韩建国; 李家锋

    2015-01-01

    Objective:To provide seed cells for bone tissue engineering in the late establishment by establishing the cul-ture system of bone marrow mesenchymal stem cells( BMSCs)of dogs in vitro,and using human BMP-2 to make them in-duced to differentiate into osteoblasts. Methods:The extraction of BMSCs of adult beagle dogs was made,then the whole marrow adherence method and density gradient centrifugation were used to isolate and culture BMSCs in vitro,and observe the cell growth morphology everyday. The third generation BMSCs with good growth form was divided into two groups. The experimental group were cultured with adding 200ng/ml human BMP-2 containing fetal bovine serum(FBS)while the control group were cultured only with complete medium containing FBS. Then we used the detection of alkaline phosphatase staining after 3 weeks′induction,alizarin red staining and Von-Kossa staining after 4 weeks′induction to identify the differentiation of osteoblasts. Results:After 3 weeks of induction of experimental group with alkaline phosphatase,staining showed the cyto-plasm of positive expression of black particles,and it was negative in the control group;After 4 weeks of induction of experi-mental group with alizarin red staining and Von-Kossa staining showed positive expression of calcium nodules,and it was negative in the control group. All the staining results in the experimental group showed the characteristics of osteoblasts. Conclusion:BMSCs of dogs,which are extracted and cultivated in vitro,can directionally differentiate into osteoblasts under the action of human BMP-2.%目的:通过将犬骨髓间充质干细胞( bone marrow mesenchymal stem cells,BMSCs)建立体外培养体系,运用人骨形态发生蛋白-2(bone morphogenetic protein-2,BMP-2)体外定向诱导分化为成骨细胞,为后期建立骨组织工程提供种子细胞。方法提取比格犬BMSCs,全骨髓贴壁法结合密度梯度离心法行体外分离培养,每日观察细

  5. Differentiation of endometrial stromal stem cells into osteoblasts and adipocytes in vitro%子宫内膜基质干细胞的体外成骨诱导和成脂诱导分化

    Institute of Scientific and Technical Information of China (English)

    杨新园; 李旭; 陈葳

    2012-01-01

    目的 探讨子宫内膜基质干细胞的多向分化潜能.方法 取因子宫肌瘤行全子宫切除术患者的子宫内膜组织,磁珠分选子宫内膜基质干细胞,进行传代培养.传代细胞分别加入成骨诱导剂和成脂诱导剂培养,并通过茜素红和油红O染色对成骨细胞和脂肪细胞形态进行鉴定.结果 子宫内膜基质干细胞呈成纤维细胞样贴壁生长,其经成骨、成脂诱导培养3周后形态、体积发生明显改变.茜素红染色显示细胞团中央能形成钙化结节;成脂诱导后油红O染色可见细胞质内出现橙红色脂滴.结论 子宫内膜基质干细胞经体外诱导培养后可向成骨细胞和脂肪细胞分化,并具有明显的成骨和成脂细胞形态,表明子宫内膜基质干细胞具有多向分化潜能.%Objective To investigate the multilineage differentiation capacity of stromal stem cells isolated and cultured from human endometrial tissues. Methods Single-cell suspensions of endometrial stromal stem cells were obtained from hysterectomy tissues of women experiencing normal menstrual cycles. Purified stromal stem cell suspensions were then obtained by selecting cells with a further round of magnetic bead sorting using anti-EpCAM-coated Dynabeads. Rare human endometrial EpCAM-stromal stem cells were screened by inverted microscope each day and identified by flow cytometer. Then the cells were cultured in osteoblast-inducing culture medium, and osteoblast phenotype was assayed with alizarin red staining. The passage cells were cultured in adipogenesis-medium and stained with oil red O for identification. Results Endometrial stromal stem cells grew as adherent cells and presented fibroblast-like in vitro, and could stably proliferate and be passed. The cells changed from fibroblast-like into ellipse after osteoblast-inducing cultivation. After induction for 21 d, alizarin red staining demonstrated the formation of mineralized nods in extracellular matrix. Under the

  6. The effect of Smad6 RNA interference on BMP-2 induced osteogenic differentiation of mesenchymal stem cells%Smad6信号干扰对MSCs骨向分化的影响

    Institute of Scientific and Technical Information of China (English)

    刘猛; 董伟; 冯晓洁; 邓久鹏; 戚孟春; 李金源

    2011-01-01

    Objective To investigate the effect of Smad6 mRNA interference on bone morphogenetic protein 2 (BMP-2) induced osteogenic differentiation of bone marrow mesenchymal stem cells (MSCs). Methods The bone marrow MSCs of mice were cultured and underwent BMP-2 induced osteogenic differentiation. The cells were divided into 3 groups: the cells in group A were transfected with recombinant Smad6 RNA interference vector,which was labeled with green fluorescent protein (GFP) , and the cells in group B were transfected with control vector,and the cells in group C served as controls. The activity of alkaline phosphonate (ALP) and levels of osteocalcin were detected at five days after transfection by ALP staining and radioimmunoassay, respectively. The formation of mineralization nodus was also examined by alizarin red staining. Results The GFP was obviously expressed in MSCs after viral transfection, and viral transfection efficiency reached 98. 5%. As compared with group B,Smad6 RNA interference increased significantly ALP activity and osteocalcin levels in group A ( P < 0.01) .however,both ALP activity and osteocalcin levels in group C were significantly lower than those in the other two groups( P <0.01). The results of alizarin red staining showed that the counts of mineralization nodus in group A were significantly more than those in group B ( P <0.05) ,but no mineralization nodus was found in group C. Conclusion Smad6 mRNA interference. Can promote effectively BMP-2 induced osteogenic differentiation of MSCs,which may be a valuable method for bone regeneration for bone defect in bone tissue engineering.%目的 研究Smad6信号干扰对骨形态发生蛋白2(BMP-2)诱导的骨髓间充质干细胞(MSCs)骨向分化的促进效应.方法 培养小鼠MSCs,用BMP-2诱导骨向分化.细胞分为3组:A组细胞用携带绿色荧光蛋白(GFP)的Smad6重组RNA干扰载体转染;B组细胞用空白载体转染;C组细胞作为对照.结果 病毒转染后GFP在MSCs中有

  7. 不同代次大鼠骨髓间充质干细胞和脂肪间充质干细胞成骨分化比较%Comparison on osteogenic differentiation ability of different passages of the mesenchymal stem cells derived from adipose and bone marrow in rats

    Institute of Scientific and Technical Information of China (English)

    任宇; 马玉珍; 王清莲; 刘东军; 仓明; 温建勋; 靳木子

    2011-01-01

    The aim of study is to investigate the culture characterizations and osteoblast differentiation in vitro of the mesenchymal stem cells derived from adipose and bone marrow. The bone marrow stromal atem ceUs ( BMSCs) and adipose derived stromal cells ( ADSCs) were isolated using density gradient eentrifuge method and cultured in vitro. followed by osteogenic cultivation in DMEM supplemented with dexamethasone, ascorbic acid and β-glycerophosphate. The osteogenic differentiation was assessed with alkaline phosphatase activity (ALP) and alizarin red staining. The results showed that the morphology of BMSCs and ADSCs showed fibroblast-like characterizations. The BMSCs and ADSCs of the 5, 10. 15 and 20 generations showed the positive results of ALP and Alizarin Red. At the same time, with passages raised, AIP activity decreased. Mineralized nodules of osteoblasts in BMSCs of 20 generations were more than those in other passages, but there was no difference in ADSCs. It suggest that the mesenchymal stem cells derived from adipose and bone marrow from rat can differentiate to osteoblasts when cultured in vitro and can be seried as optimal autogenous ceU source for bone tissue engineering.%本研究旨在观察不同代次骨髓间充质干细胞(BMSCs)和脂肪间充质干细胞(ADSCs)体外培养的生长特点和体外诱导成骨能力.通过密度梯度离心和贴壁培养法分离培养大鼠骨髓间充质干细胞和脂肪间充质干细胞,用含地塞米松、抗坏血酸、β-甘油磷酸钠的培养液定向诱导传代细胞向成骨细胞分化,并利用茜素红染色、碱性磷酸酶染色及PCR方法检测成骨细胞.结果表明骨髓及脂肪间充质干细胞呈成纤维细胞样生长,增殖能力强,生长迅速.第5、10、15、20代BMSCs及ADSCs经诱导培养后茜素红染色呈阳性并且出现"矿化"、碱性磷酸酶活性强,随着细胞代次的递增,诱导后细胞碱性磷酸酶活性呈递减趋势;诱导后的两类细胞传代后

  8. 去分化脂肪细胞的生物学特性探究%Biocharacteristics′ study of dedifferentiated fat cells

    Institute of Scientific and Technical Information of China (English)

    程飞飞; 魏翠; 潘文雯; 徐妍

    2012-01-01

    Objective To isolate and culture human dedifferentiated fat (DFAT) cells, and to observe the hioeharaetcristies for the further fundamental researches and clinical applications. Methods We isolated and cx-traeted mature adipoeytes from the fatty portions of antologons liposnction aspirates of human. Mature adipoeytes were cultured and induced to DFAT cells by ceiling adherent culture method. The bioeharaeteristies of DFAT cells were analyzed. The surface antigens of DFAT cells were detected using flow eytomctry, including CD29, CD34, CD7I , and GD90. The adipogenie, chondrogenic and osteogenic ability of DFAT cells were assessed hy oil red 0 staining, alcian blue staining and alizarin bordeaux staining, respectively. Measure the levels of VEGF-A, HGF and TGF-β1 in DFAT cells hy Elisa technique. Results Flow eytomctry assay demonstrated that DFAT cells were positive for the surface markers CD29 and CD90, and negative for CD34 and CD7I. After three weeks of adipogenie differentiation, the lipid droplets could he observed by oil red 0 staining. .After three wrecks of chondrogenic differentiation, matrix of cartilage cells in DFAT cells could he detected hy alcian blue staining.After three weeks of osteogenic differentiation calcium salts mineralization, DFAT cells could he detected hy alizarin bordeaux staining. Elisa assay demonstrated that DFAT cells were positive for VEGF-A ( 765.5 ng/ml), and negative for HGF and TGF-β1. Conclusion Our results demonstrated that DFAT and ASGs have similar characters in morphology, surface marker profiles and differentiation ability, etc. DFAT cells wrere alternative source of ASGs from the adipose tissues and had great potential in tissue engineering.%目的 采用体外分离培养去分化脂肪细胞,探究去分化脂肪细胞的生物学特性.方法 天花板培养法分离培养去分化脂肪细胞,流式细胞术检测细胞表面抗原表型,Elisa法检测细胞分泌的VEGF-A、HGF和TGF-β1量.细胞成脂、成软

  9. 补肾活血方调控 microRNA-210促进 BMSCs 成骨分化的研究%The kidney-tonifying and blood circulation-promoting recipes promote the osteogenic differentiation of bone mesenehymal stem cells by regulating microRNA-210

    Institute of Scientific and Technical Information of China (English)

    张倍源

    2014-01-01

    目的:研究补肾活血方促进骨髓间充质干细胞(Bone Mesenehymal Stem Cells ,BMSCs)成骨分化的分子机制。方法贴壁培养法分离纯化大鼠BMSCs,将培养成功的BMSCs分为空白组、成骨诱导组和中药组,培养7d、14d行矿化结节茜素红染色(Alizarin Red S,ARS),倒置显微镜观察矿化骨结节形成情况,采用real-time PCR检测miR-210的表达变化。结果与空白组比较,成骨诱导组和中药组BMSCs钙化结节数量明显增多,差异有非常显著的统计学意义( P<0.01)。在成骨诱导剂、补肾活血方水提液的作用下,与空白组比较,miR-210表达明显降低,且中药组较成骨诱导组更明显降低miR-210表达。结论补肾活血方通过降低miR-210表达促进BMSCs成骨分化。%Objective To investigate the mechanism of osteogenic differentiation of bone mesenehymal stem cells ( BMSCs) promoted by the kidney-tonifying and blood circulation-promoting recipes.Methods BMSCs of SD rats were isolated with adherent separation method and primarily cultured in vitro.Then BMSCs were divided into blank group, osteo-induced group, and Chinese medicine group.At the 7 th day and the 14 th day, the mineralized nodules were stained using Alizarin red statining ( ARS) . The formation of mineralized nodules was observed using a converted microscope.The expression of miR-210 was detected using real-time PCR.Results Compared with that in blank group, the number of mineralized nodules in osteo-induced group and Chinese medicine group increased significantly ( P <0.01 ) .After the intervention of osteogenic inducer and extraction of the kidney-tonifying and blood circulation-promoting recipes, the expression of miR-210 in both groups decreased significantly compared with that in blank group.And the expression in Chinese medicine group was much lower than that in osteo-induced group. Conclusion The kidney-tonifying and blood circulation-promoting recipes can

  10. Effects of GJIC diminuendo agent 18α-GA on transdifferentiation from adipocytes to osteoblasts in rabbits%G JIC 渐弱剂18α-甘草次酸对兔脂肪细胞向成骨细胞横向分化的影响

    Institute of Scientific and Technical Information of China (English)

    王吉博; 曾旋; 彭浩; 王凤宇; 王兆杰

    2014-01-01

    Objective It is to explore the effects of transdifferentiation from adipocytes to osteoblasts and gap junction in-tercellular communication( GJIC) by gap junction Diminuendo agent 18α-GA.Methods Groin adipose tissue was taken from 3 months old New Zealand White Rabbit and collected mature adipocytes.Made the mature adipocytes dedifferentiate and ob-tained the dedifferentiated adipocytes.The 3rd generation of the dedifferentiated adipocytes were induced and differentiated in-to osteoblasts.There were two experimental groups that the diminuendo group with adding 18α-GA and the control group. The toxic action of 18α-GA in osteoblasts growth and proliferation were detected by MTT assay.Osteoblasts were qualitatively detected by alizarin red staining and collagen typeⅠimmunohistochemistry staining.The expressions of Cx43 were measured by western-blot and sorape-loading and dye transfet ( SLDT) method.Results The results of MTT assay showed that 18α-GA had no significant effect on cell proliferation.The results of Alizarin red staining showed that calcific nodules were dyed red in each group.Immunohistochemistry staining results showed that the expressions of collagen typeⅠwere weakened in the diminuendo group.The results of Western blot showed that the expressions of Cx43 were decreased in the diminuendo group. Conclusion 18α-GA diminuendo the capability of transdifferentiation from adipocytes to osteoblasts with enhancement of GJIC.%目的:探讨缝隙连接通讯渐弱剂18α-甘草次酸( GA)对脂肪细胞向成骨细胞横向分化及细胞间缝隙连接通讯的影响。方法选取3月龄新西兰大白兔腹股沟处脂肪组织,分离提取成熟脂肪细胞,使其去分化得到去分化脂肪细胞,取第三代去分化的脂肪细胞进行成骨诱导,加入GJIC渐弱剂18α-GA设为减弱组,并设立对照组, MTT法观察18α-GA对细胞倍增是否有影响,进行茜素红钙结节染色和I型胶原免疫组化染色

  11. ISOLATION,CULTURE AND IDENTIFICATION OF MESENCHYMAL STEM CELLS FROM THE BONE MARROW OF RABBIT IN VITRO%兔骨髓间充质干细胞的体外分离培养及鉴定

    Institute of Scientific and Technical Information of China (English)

    黄家志; 陈前芬; 肖增明; 李世德

    2011-01-01

    目的:观察兔骨髓间充质干细胞(BMSCs)生长特性及潜在分化潜能,为组织工程中种子细胞选择提供实验基础.方法:应用全骨髓贴壁法分离培养兔BMSCs,相差显微镜下观察其生长特点,应用流式细胞术对第三代细胞进行表面抗原鉴定.经成脂和成骨诱导液体外诱导兔BMSCs向脂肪细胞、成骨细胞分化,对诱导2周后的细胞进行油红O染色、茜素红染色、Vankossa银染染色及碱性磷酸酶染色.结果:经流式细胞术鉴定,全骨髓贴壁法可获得兔BMSCs,第三代兔BMSCs生物特征基本一致并能诱导分化为脂肪细胞及成骨细胞,成脂诱导后油红O染色细胞内出现红色脂滴,成骨诱导后茜素红染色、Vankossa 银染均可观察到矿化结节.碱性磷酸酶活性染色对照组呈弱阳性,诱导组强强阳性.结论:全骨髓贴壁法是分离培养兔BMSCs简便可行的方法.BMSCs来源丰富并有成脂及成骨潜能,是组织工程的优良种子细胞.%Objective:The rabbit bone marrow mesenchymal stem cells(MSCs) were observed the biological characteristics and the differentiation potential, which to provide experimental basis for selection in the seed cells for tissue engineering. Methods:The rabbit bone marrow-derived MSCs were isolated and cultured from rabbit bone marrow by the bone marrow different adherent method. Morphology of MSCs was examined by phase contrast microscopy, surface antigen from the third passage MSCs was detected by flow cytometry. MSCs were treated with adipose inductor and osteogenetic inductor to differentiated into adipocytes and osteoblast in vitro. And the differentiated cells were identified by oil red O staining ,alizarin bordeaux staining, Vankossa silver staining and alkaline phosphatase staining. Results: Through flow cytometry analyzed, the bone marrow-derived MSCs were obtained by the different adherent method. The biological characteristics of 3 passage MSCs were consistent, which were induced

  12. 偏肿革裥菌漆酶基因克隆及不同染料脱色研究%Cloning and different dye decolorization of a laccase gene from white rot fungus Lenzites gibbosa

    Institute of Scientific and Technical Information of China (English)

    郑苗苗; 池玉杰

    2012-01-01

    染料由于具有复杂的化学结构通常难以降解.漆酶粗酶液对终浓度为50 mg/L的不同化学结构的染料的脱色研究,可快速对蒽醌类茜素红进行脱色可达100%;杂环类中性红、偶氮类刚果红和三苯基甲烷类结晶紫的脱色效果低于茜素红,测试的3种染料均可在没有介体存在的条件下被漆酶脱色,显示出偏肿革裥菌(Lenzites gibbosa)漆酶对茜素红染料脱色具有较大的应用潜力,进而对废水处理具有更好的应用前景.利用PCR和RACE技术首次从该菌株中获得编码漆酶基因的cDNA及其基因组全长序列,基因组大小为2 106 bp.通过比较该漆酶基因的cDNA和基因组DNA的全长序列,发现该基因包含11个外显子和10个内含子.cDNA序列的全长为1 982 bp,其中包含一个完整的ORF,长度为1 563 bp,编码520个氨基酸.%Dyes are usually difficult to be decolorized due to their complex chemical structures. Chemical different dyes were used in the dye decolorization test. Laccase crude enzyme solution to a final concentration of 50 mg / L of different chemical structure of the dye decolorization, And fast grace quinones Alizarin Red decolorization of up to 100% ; Heterocyclic neutral red, Azo kind of Congo red and Triphenylmethane crystal violet Decolorizing effect below alizarin red. The result demonstrated the potential application prospects of the laccase from Lenzites gibbosa dye-containing wastewater treatment. Using PCR and RACE technique, We obtain the cDNA and genomic DNA sequence of the laccase gene from Lenzites gibbosa, Genome size for 2 106 bp. By comparing the laccase cDNA gene and genome DNA sequence length, Found that the gene contains 11 exon and 10 introns. cDNA sequence of full length for 1 982 bp, contains a complete ORF, length for 1 563 bp, code 520 amino acids.

  13. Two-layer membranes of calcium phosphate/collagen/PLGA nanofibres: in vitro biomineralisation and osteogenic differentiation of human mesenchymal stem cells

    Science.gov (United States)

    Hild, Nora; Schneider, Oliver D.; Mohn, Dirk; Luechinger, Norman A.; Koehler, Fabian M.; Hofmann, Sandra; Vetsch, Jolanda R.; Thimm, Benjamin W.; Müller, Ralph; Stark, Wendelin J.

    2011-02-01

    The present study evaluates the in vitro biomedical performance of an electrospun, flexible, anisotropic bilayer with one layer containing a collagen to mineral ratio similar to that in bone. The double membrane consists of a poly(lactide-co-glycolide) (PLGA) layer and an amorphous calcium phosphate (a-CaP)/collagen (Col)/PLGA layer. In vitro biomineralisation and a cell culture study with human mesenchymal stem cells (hMSC) were conducted to characterise such membranes for possible application as biomaterials. Nanofibres with different a-CaP/Col/PLGA compositions were synthesised by electrospinning to mimic the actual composition of bone tissue. Immersion in simulated body fluid and in cell culture medium resulted in the deposition of a hydroxyapatite layer. Incubation of hMSC for 4 weeks allowed for assessment of the proliferation and osteogenic differentiation of the cells on both sides of the double membrane. Confocal laser scanning microscopy was used to observe the proper adhesion of the cells. Calcium and collagen content was proven by Alizarin red S and Sirius red assays. Acute cytotoxic effects of the nanoparticles or the chemicals used in the scaffold preparation could be excluded based on viability assays (alamarBlue and alkaline phosphatase activity). The findings suggest possible application of such double membranes is in treatment of bone defects with complex geometries as wound dressing material.The present study evaluates the in vitro biomedical performance of an electrospun, flexible, anisotropic bilayer with one layer containing a collagen to mineral ratio similar to that in bone. The double membrane consists of a poly(lactide-co-glycolide) (PLGA) layer and an amorphous calcium phosphate (a-CaP)/collagen (Col)/PLGA layer. In vitro biomineralisation and a cell culture study with human mesenchymal stem cells (hMSC) were conducted to characterise such membranes for possible application as biomaterials. Nanofibres with different a

  14. Amplification of rabbit adipose-derived stem cells using explants culture method%组织块贴壁法扩增兔脂肪干细胞

    Institute of Scientific and Technical Information of China (English)

    刘琴; 王丽平; 喻晶; 陈芳; 刁波; 张宜

    2014-01-01

    BACKGROUND:The rabbit adipose-derived stem cells are mostly isolated by type I col agenase digestion, but rarely by explants culture method. OBJECTIVE:To isolate rabbit adipose-derived stem cells for adipogenic and osteogenic differentiation. METHODS:The rabbit adipose-derived stem cells were isolated from rabbit adipose by explants culture method, and cultured in vitro fol owed by morphological observation. The grow curve and cellsurface markers CD29, CD44, CD45 of passage 3 cells were analyzed respectively by 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide and flow cytometry;cells from the third passages were induced for adipogenic and osteogenic differentiation by different revulsants, and cells were examined by oil red O staining and alizarin red staining . RESULTS AND CONCLUSION:The rabbit adipose-derived stem cells cultured in vitro exhibited a spindle-shaped appearance and could rapidly expand. Flow cytometry analysis revealed that the third passage of rabbit adipose-derived stem cells was positive for CD29, CD44, but negative for CD45. Rabbit adipose-derived stem cells were positive for oil red O staining at 14 days of adipogenic induction, and positive for alizarin red staining at 14 days of osteogenic induction. In conclusion, we could successful y isolate rabbit adipose-derived stem cells using explants culture method.%背景:研究显示兔脂肪干细胞的体外分离方法大多数为Ⅰ型胶原酶消化法,采用组织块贴壁法扩增兔脂肪干细胞尚不多见。  目的:采用组织块贴壁法从兔脂肪组织中分离培养兔脂肪干细胞,并进行成脂、成骨的诱导分化。  方法:采用组织块贴壁法分离出兔脂肪干细胞,进行体外培养,观察其形态特征。取对数生长期的第3代细胞,用MTT法绘制其生长曲线;流式细胞仪检测其表面抗原CD29、CD44、CD45的表达情况;分别用成脂和成骨诱导培养液诱导其向脂肪细胞和成骨细胞

  15. Morphological observation of nanobacteria in calcified placenta and umbilical cord blood%钙化胎盘及脐带血中纳米细菌的形态学观察

    Institute of Scientific and Technical Information of China (English)

    刘胜男; 张德纯; 张名均; 郭亚楠; 杨晓容; 杨顺杰

    2013-01-01

    目的 研究纳米细菌(nanobacteria,NB)的分布,寻求检测胎盘钙化NB感染的更好组织来源.方法 收集孕妇胎盘钙化组织和脐带血20例,并以20例正常胎盘组织作对照,分离培养NB;用革兰染色和茜素红染色法、扫描电镜和透射电镜进行形态学鉴定.结果 培养沉淀物符合NB的特性.脐带血NB分离率为80%(16/20),钙化胎盘组织为65%(13/20),均明显高于正常胎盘组织的10%(2/20),X2分别为12.07、9.09,P均<0.01;脐带血与钙化胎盘组织比较,差异无统计学意义(x2=1.33,P>0.05).油镜下脐带血和钙化胎盘组织分离的NB为球状或球杆状,形态相似;茜素红染色脐带血中NB较多且聚集成簇;透射电镜和扫描电镜下NB呈球形,大小为80 ~ 500 nm,并可见毛刺结构.结论 胎盘钙化与NB感染有关,脐带血适用于检测钙化胎盘的NB感染.%Objective To study the distribution of nanobacteria (NB) and explore an appropriate tissue as the source for detection of placental calcification infected with nanobacteria. Methods The calcified placental tissues and umbilical cord blood from 20 pregnant women for isolation and culture of NB, and normal placental tissues were used as controls. The morphological identification of NB was performed with Gram and alizarin red staining and scanning electron microscope (SEM) , transmission electron microscope (TEM). Results The precipitates were corresponding to the characteristics of NB. The isolation rate of NB was 65% (13/20) in the specimens of calcified placental tissues and 80% (16/20) in the cord blood, and both of them were higher than that in normal placental tissues (10% ) with a significantly statistical difference ( chi-square values were 12. 07 and 9. 09 respectively, P 0. 05). The shape of NB isolated from calcified placental tissue and umbilical cord blood under oil lens was spherical or rhabdoid with burr edge, and was globoid with size of 80-500 nm and staining of Gram

  16. 人胚胎干细胞生物学特征及成骨诱导分化相关研究%Related research of human embryonic stem cell biology characteristics and ossification differentiation

    Institute of Scientific and Technical Information of China (English)

    胡健鹏

    2013-01-01

    目的:研究体外培养人胚胎干细胞的生物学特征,初步探讨人胚胎干细胞向成骨细胞定向分化的特性,为后续成骨细胞移植治疗研究提供很好的资源。方法:体外传代培养人胚胎干细胞,显微镜观察细胞的贴壁生长情况,细胞染色体检查确认核型,免疫荧光染色检测人胚胎干细胞的异性和细胞周期,进行成骨诱导分化及碱性磷酸酶染色和茜素红染色鉴定。结果:人胚胎干细胞在体外培养体系中增殖迅速,核型检测为46,XX,免疫荧光染色检测细胞表达人胚胎干细胞特异性标志,细胞周期检测结果显示细胞的增殖能力强,大部分细胞处于 G1期。人胚胎干细胞经成骨诱导分化后,碱性磷酸酶染色和茜素红染色阳性。结论:在特殊培养条件下,人胚胎干细胞在体外能维持其多能特性,体外可以诱导分化成为骨细胞,有望促进应用于骨组织工程移植治疗的机制探讨。%Objective:In order to observe the biology characteristics of human embryonic stem cell in vitro culture, investigate the osteoblast directional differentiation characteristics of human embryonic stem cells, to provide perfect resources for the subsequent osteoblast transplant therapy research. Methods:The human embryonic stem cells have been subculture in vitro, microscope observation that the cells attached growth, karyotypes, immunofluorescence staining detection and cell cycle, bone alkaline phosphatase staining and alizarin red dye appraisal. Results: The human embryonic stem cells proliferation rapidly in vitro culture system, keep the normal karyotype of 46, XX. Immunofluorescence staining detection cell express human embryonic stem cells specific mark; cell cycle test results show that cell proliferation ability, most cells in the G1 phase. Human embryonic stem cells have been ossification differentiation, staining of alkaline phosphatase and alizarin red showed positive. Conclusion

  17. The effect of platelet-rich fibrin gel precipitate liquid on mineralization of human dental pulp cells in vitro%富血小板纤维蛋白凝胶析出液对人牙髓细胞体外矿化的影响

    Institute of Scientific and Technical Information of China (English)

    何璇; 韦维; 陈文霞

    2015-01-01

    目的:探索富血小板纤维蛋白(platelet-rich fibrin,PRF)凝胶析出液对人牙髓细胞(human dental pulp cells, hDPCs)体外矿化的影响。方法组织块法培养 hDPCs。采用 Choukroun 一步离心法制备 PRF 凝胶。将新鲜制备的PRF 凝胶浸泡于 DMEM 培养基中,于第7 d 取析出液。用 PRF 凝胶析出液孵育 hDPCs 3 d 后更换矿化诱导液。采用茜素红染色和 RT-PCR 检测人牙髓细胞矿化的潜能。结果矿化诱导21 d 后,茜素红染色观察到实验组有少量钙结节生成,而对照组无钙结节生成;RT-PCR 结果显示,实验组 hDPCs 碱性磷酸酶(ALP)的表达为对照组的1.5倍,差异具有统计学意义(P <0.05)。结论 PRF 凝胶析出液可促进人牙髓细胞矿化。%Objective This study was designed to investigate the effect of platelet-rich fibrin gel (PRF gel)precipitate liquid on the mineralization of human dental pulp cells (hDPCs)in vitro. Methods The hD-PCs were separated and cultured by using tissue block culture method.PRF gel was prepared by Choukroun's protocols.The newly prepared PRF gel was dipped in DMEM culture media,the precipitate liquid of PRF gel was collected on day 7.hDPCs were treated with mineralization induction solution 3 days after being incubated with the precipitate liquid of PRF gel.The capacity of mineralization was measured by using alizarin red stai-ning and RT-PCR. Results Twenty-one days after mineralization induction,a small amount of mineral-ized nodules on alizarin red staining were observed in experimental group while no mineralized nodule was ob-served in control group;RT-PCR revealed that the expression of alkaline phosphatase (ALP)in experimental group was 1.5 times higher than that in control group,comparison yielded statistical difference (P <0.05). Conclusion The precipitate liquid of PRF gel can accelerate the mineralization of hDPCs.

  18. 富血小板纤维蛋白对人牙槽骨成骨细胞增殖和分化的影响%Choukroun's Platelet-Rich Fibrin Promotes Human Alveolar Osteoblasts Differentiation

    Institute of Scientific and Technical Information of China (English)

    张迎娣; 阮征; 张劲娥; 刘天麟; 罗光明; 郭鹏女; 黄远亮; 王磊

    2015-01-01

    目的:研究富血小板纤维蛋白(plate-rich fibrin, PRF)体外对人牙槽骨成骨细胞(human alveolar osteoblasts, HAOB)增殖分化的影响,探讨HAOB与PRF构建临床组织工程骨的可能性。方法:收集临床拔牙过程中的牙槽骨,采用改良酶消化法体外分离培养HAOB,根据成骨细胞形态学特征及成骨特性对所培养出的细胞进行鉴定,后加入志愿者的PRF分组培养;而后在不同的实验时间点进行细胞增殖CCK-8检测、碱性磷酸酶(ALP)定性检测、钙结节茜素红染色以及成骨相关基因RT-PCR检测。结果:HAOB具有典型的成骨细胞的形态;随时间的延长,细胞数目明显增加(P<0.05),实验组(PRF组)细胞数量明显高于对照组。实验组ALP染色较对照组颜色更深,ALP活性相对较高。经茜素红染色,实验组镜下观察形成的钙结节数量较对照组多。在第7和11天发现实验组成骨相关基因的表达量均高于对照组。结论:采用改良酶消化法分离培养的HAOB具有典型的成骨细胞生物学特性,且成分较为单一;PRF体外具有促进HAOB增殖分化的能力。%Objective: The purpose of the study was to evaluate the effects of Choukroun's platelet-rich fibrin (PRF) on proliferation and osteogenic differentiation of human alveolar osteoblasts (HAOB), and to explore the possibility of bone engineering constructs with PRF and HAOB. Methods: Human alveolar bone was collected during clinical wisdom tooth extraction process. Human alveolar osteoblasts were isolated and cultured in vitro by improved enzyme digestion. The cul-tured cells were identified by the morphological characteristics and osteogenic properties of osteoblasts. HAOB was cul-tured with or without PRF obtained from volunteers. Cell proliferation was measured by CCK-8 method. Qualitative detec-tion of alkaline phosphatase (ALP), alizarin red staining of calcium nodules and RT-PCR of osteogenesis related

  19. Retinoic acid induces osteogenic differentiation of periodontal ligament stem cells from miniature swine in vitro%维甲酸诱导小型猪牙周膜干细胞的体外成骨

    Institute of Scientific and Technical Information of China (English)

    张鹏涛; 钟良军; 张远; 张源明; 徐艳

    2011-01-01

    BACKGROUND: Recent studies have found that retinoic acid can induce the osteogenic differentiation of embryonic stem cellsand multiple adult stem cells.OBJECTIVE: To observe the effect of retinoic acid on osteogenic differentiation of porcine periodontal ligament stem cellsCPDLSCs).METHOOS: Porcine PDLCs were harvested by using outgrowth method: PDLSCs were isolated by limited dilution of culture cellsfor single cell clone. Immunofluores cence was used to detect the expression of STRO-1 andimmunocytochemistry to detect theexpression of vimentin and pan-Cytokeratin(PCK) of porcine PDLSCs. Cell counting hits (CCIKB)was applied to evaluated Cellproliferation of PDLSCs. The colony formation of porcine PDLSCs ratio was assayed. Third passage PDLSCs were inducedwithmineralized conditional medium containing retinoic acid, ascorbic acid and β-glycerophBphate.Mineralzednodules werestudiedby Alizarin red S staining. Osteopontin. Osteocalcin. Collagen type I and collagen type III were detected byimmunocytochemistry.RESULTS AND CONCLUSION: Porcine PDLSCs expressed STRO-1 and vimentin. And the result of pan-Cytokeratin wasnegative. Porcine PDLSCs colony formation ratio was 2.8%. PDLSCs induced by retinoic acid showed positive expression ofak aline phosphatase at 14 days and positive expression of Alizarin red S at 21 days. Osteopontin. Osteocalon.colagent)pe Iwere positive but collagen type III was negative at21 days. These finding! Indicate that retinoic acid can be an effective inducerof osteogenic differentiation of porcine PDLSCs.%背景:近期研究发现维甲酸对胚胎干细胞及多种成体干细胞具有成骨方向诱导的作用.目的:观察维甲酸对小型猪牙周膜干细胞体外成骨作用的影响.方法:采用组织块法获得小型猪牙周膜细胞,有限稀释法纯化小型猪牙周膜干细胞,免疫荧光法检测STRO-1、免疫细胞化学法检测波形蛋白、角蛋白鉴定小型猪牙周膜干细胞.CCK8法测定小型猪牙周膜干细

  20. Use of X-ray microprobe to diagnose bone tissue demineralization after caffeine administration Use of X-ray microprobe to diagnose bone tissue demineralization after caffeine administration

    Directory of Open Access Journals (Sweden)

    Marek Tomaszewski

    2012-10-01

    Full Text Available Caffeine is a methylxanthine which permeates the placenta. In studies on animals, it has been
    shown to produce teratogenic and embryotoxic effects in large doses. The objective of this study was to
    assess the influence of caffeine on the development of bone tissue, with particular reference to elemental
    bone composition using an X-ray microprobe. The research was conducted on rats. The fertilized females
    were randomly divided into an experimental and a control group. The experimental group was
    given caffeine orally in 30 mg/day doses from the 8th to the 21st day of pregnancy, while the control group
    was given water. The fetuses were used to assess the growth and mineralization of the skeleton. On the
    basis of double dyeing, a qualitative analysis of the bone morphology and mineralization was conducted.
    For calcium and potassium analysis, an X-ray microprobe was used. In 67 fetuses from the experimental
    group, changes in skeleton staining with the alcian-alizarin method were noticed. The frequency of the
    development of variants in the experimental group was statistically higher. In the experimental group,
    a significant decrease in the calcium level, as well as an increase in the potassium level, was observed.
    The X-ray microprobe’s undoubted advantage is that is offers a quick qualitative and quantitative analysis
    of the elemental composition of the examined samples. Employing this new technique may furnish us
    with new capabilities when investigating the essence of the pathology process.Caffeine is a methylxanthine which permeates the placenta. In studies on animals, it has been
    shown to produce teratogenic and embryotoxic effects in large doses. The objective of this study was to
    assess the influence of caffeine on the development of bone tissue, with particular reference to elemental
    bone composition using an X-ray microprobe. The research was conducted on

  1. PELA microspheres with encapsulated arginine-chitosan/pBMP-2 nanoparticles induce pBMP-2 controlled-release, transfected osteoblastic progenitor cells, and promoted osteogenic differentiation.

    Science.gov (United States)

    Xu, Xiaolong; Qiu, Sujun; Zhang, Yuxian; Yin, Jie; Min, Shaoxiong

    2017-03-01

    Repair of the bone injury remains a challenge in clinical practices. Recent progress in tissue engineering and therapeutic gene delivery systems have led to promising new strategies for successful acceleration of bone repair process. The aim of this study was to create a controlled-release system to slowly release the arginine-chitosan/plasmid DNA nanoparticles encoding BMP-2 gene (Arg-CS/pBMP-2 NPs), efficiently transfect osteoblastic progenitor cells, secrete functional BMP-2 protein, and promote osteogenic differentiation. In this study, chitosan was conjugated with arginine to generate arginine-chitosan polymer (Arg-CS) for gene delivery. Mix the Arg-CS with pBMP-2 to condense pBMP-2 into nano-sized particles. In vitro transfection assays demonstrated that the transfection efficiency of Arg-CS/pBMP-2 nanoparticles and the expression level of BMP-2 was obviously exceed control groups. Further, PELA microspheres as the controlled-release carrier for the nanoparticles were used to encapsulate Arg-CS/pBMP-2 NPs. We demonstrated that the Arg-CS/pBMP-2 NPs could slowly release from the PELA microspheres at least for 42 d. During the co-culture with the PELA microspheres, the content of BMP-2 protein secreted by MC3T3-E1 reached the peak at 7 d. After 21d, the secretion of BMP-2 protein still maintain a higher level. The alkaline phosphatase activity, alizarin red staining, and osteogenesis-related gene expression by real-time quantitative PCR analysis all showed the PELA microspheres entrapping with Arg-CS/pBMP-2 NPs can obviously induce the osteogenic differentiation. The results indicated that the Arg-CS is a suitable gene vector which can promote the gene transfection. And the novel PELA microspheres-nanoparticle controlled-release system has potential clinical application in the future after further research.

  2. In Vitro Osteogenic Potential of Green Fluorescent Protein Labelled Human Embryonic Stem Cell-Derived Osteoprogenitors

    Directory of Open Access Journals (Sweden)

    Intekhab Islam

    2016-01-01

    Full Text Available Cellular therapy using stem cells in bone regeneration has gained increasing interest. Various studies suggest the clinical utility of osteoprogenitors-like mesenchymal stem cells in bone regeneration. However, limited availability of mesenchymal stem cells and conflicting evidence on their therapeutic efficacy limit their clinical application. Human embryonic stem cells (hESCs are potentially an unlimited source of healthy and functional osteoprogenitors (OPs that could be utilized for bone regenerative applications. However, limited ability to track hESC-derived progenies in vivo greatly hinders translational studies. Hence, in this study, we aimed to establish hESC-derived OPs (hESC-OPs expressing green fluorescent protein (GFP and to investigate their osteogenic differentiation potential in vitro. We fluorescently labelled H9-hESCs using a plasmid vector encoding GFP. The GFP-expressing hESCs were differentiated into hESC-OPs. The hESC-OPsGFP+ stably expressed high levels of GFP, CD73, CD90, and CD105. They possessed osteogenic differentiation potential in vitro as demonstrated by increased expression of COL1A1, RUNX2, OSTERIX, and OPG transcripts and mineralized nodules positive for Alizarin Red and immunocytochemical expression of osteocalcin, alkaline phosphatase, and collagen-I. In conclusion, we have demonstrated that fluorescently labelled hESC-OPs can maintain their GFP expression for the long term and their potential for osteogenic differentiation in vitro. In future, these fluorescently labelled hESC-OPs could be used for noninvasive assessment of bone regeneration, safety, and therapeutic efficacy.

  3. KDM6B epigenetically regulates odontogenic differentiation of dental mesenchymal stem cells

    Institute of Scientific and Technical Information of China (English)

    Juan Xu; Bo Yu; Christine Hong; Cun-Yu Wang

    2013-01-01

    Mesenchymal stem cells (MSCs) have been identified and isolated from dental tissues, including stem cells from apical papilla, which demonstrated the ability to differentiate into dentin-forming odontoblasts. The histone demethylase KDM6B (also known as JMJD3) was shown to play a key role in promoting osteogenic commitment by removing epigenetic marks H3K27me3 from the promoters of osteogenic genes. Whether KDM6B is involved in odontogenic differentiation of dental MSCs, however, is not known. Here, we explored the role of KDM6B in dental MSC fate determination into the odontogenic lineage. Using shRNA-expressing lentivirus, we performed KDM6B knockdown in dental MSCs and observed that KDM6B depletion leads to a significant reduction in alkaline phosphate (ALP) activity and in formation of mineralized nodules assessed by Alizarin Red staining. Additionally, mRNA expression of odontogenic marker gene SP7 (osterix, OSX), as well as extracellular matrix genes BGLAP (osteoclacin, OCN) and SPP1 (osteopontin, OPN), was suppressed by KDM6B depletion. When KDM6B was overexpressed in KDM6B-knockdown MSCs, odontogenic differentiation was restored, further confirming the facilitating role of KDM6B in odontogenic commitment. Mechanistically, KDM6B was recruited to bone morphogenic protein 2 (BMP2) promoters and the subsequent removal of silencing H3K27me3 marks led to the activation of this odontogenic master transcription gene. Taken together, our results demonstrated the critical role of a histone demethylase in the epigenetic regulation of odontogenic differentiation of dental MSCs. KDM6B may present as a potential therapeutic target in the regeneration of tooth structures and the repair of craniofacial defects.

  4. Effects of bFGF on Biological Characteristics of Cells Derived from Periodontium%碱性成纤维细胞生长因子对牙周细胞生物学活性的影响

    Institute of Scientific and Technical Information of China (English)

    葛少华; 杨丕山

    2001-01-01

    目的 观察基因重组人碱性成纤维细胞生长因子(rh-bFGF)对人牙龈成纤维细胞(GF)、人牙周韧带成纤维细胞(PDLF)及人牙槽骨细胞(ABC)的增殖、碱性磷酸酶活性、总蛋白含量及对3种细胞矿化结节形成能力的影响。方法 采用细胞培养、MTT比色测定、碱性磷酸酶测定法、考马斯亮蓝法及茜素红染色法。结果 bFGF能促进3种细胞的增殖,但对PDLF和ABC的ALP活性,蛋白含量及矿化结节的形成有抑制作用。结论 bFGF可促进细胞的增殖,抑制细胞的分化成熟,从而促进牙周再生。%Objective To observe effects of basic fibroblast growth factor onthe biological characteristics of cultured human gingival,periodontal ligament fibroblasts and human alveolar bone cells, such as proliferation, alkaline phosphatase activity, protein synthesis and the formation of mineralized nodules. Methods Using cell culture technique, MTT colorimetric assay, ALP activity assay, Commasie brilliant blue staining and Dahl McGec-Russell's alizarin red stain for calcium.Results bFGF enhanced the proliforative responses of the three types of cells. In contrast, bFGF inhabited the induction of alkaline phosphatase activity, protein synthesis and the mineralized nodule formation by PDLF and ABC. Conclusion bFGF can enhance cell proliferation while inhibit cytodifferentiation, thus accelerating periodontal regeneration.

  5. Extractive spectrophotometric determination of some α-adrenergic-antagonists in pure forms and in pharmaceutical formulations

    Directory of Open Access Journals (Sweden)

    El Sheikh Ragaa

    2012-01-01

    Full Text Available A simple, rapid, and extractive spectrophotometric methods was developed for the determination of some postsynaptic α-1 adrenoreceptor antagonist; doxazosin mesylate (DOX, terazosin (TRZ and alfuzosine HCl (ALF in pure forms and pharmaceutical formulations. The developed methods are based on the formation of yellow colored chloroform ion-pair complexes between the basic nitrogen of the drugs and dyes, namely; bromocresol green (BCG, bromothymol blue (BTB, methyl orange (MO and alizarine red S (ARS, in acidic buffer of pH range (3.0-5.0. The formed complexes were extracted with chloroform or dichloromethane and measured at 418, 414, 425 and 426 nm for DOX and at 419, 415, 425 and 428 for TRZ and at 418, 412, 421 and 427 nm for ALF using BCG, BTB, MO and ARS, respectively. The analytical parameters and their effects on the reported systems are investigated. Beer’s law was obeyed in the range 1.0-130 μg mL−1 with correlation coefficient (n = 6 ≥ 0.9991. The molar absorpitivity, Sandell sensitivity, detection and quantification limits were also calculated. The composition of the ion associates was found 1:1 by Job’s method. The proposed methods have been applied successfully for the analysis of the studied drugs in pure forms and in pharmaceutical formulations with percentage recoveries ranges from 99.18-100.61. The results of analysis were validated statistically. The results were in good agreement and compared with those obtained with reported methods.

  6. Short bouts of mechanical loading are as effective as dexamethasone at inducing matrix production by human bone marrow Mesenchymal stem cell

    Directory of Open Access Journals (Sweden)

    A Sittichokechaiwut

    2010-07-01

    Full Text Available Dexamethasone (Dex is used widely to induce differentiation in human mesenchymal stem cells (hMSCs; however, using a pharmaceutical agent to stimulate hMSC differentiation is not the best choice for engineered tissue transplantation due to potential side-effects. The goal of the present study was to investigate the effects of dynamic compressive loading on differentiation and mineralized matrix production of hMSCs in 3D polyurethane scaffolds, using a loading regimen previously shown to stimulate mineralised matrix production of mature bone cells (MLO-A5. hMSCs were seeded in polyurethane scaffolds and cultured in standard culture media with or without Dex. Cell-seeded scaffolds were compressed at 5% global strain for 2 h on day 9 and then every 5 days in a media-filled sterile chamber. Samples were tested for mRNA expression of alkaline phosphatase (ALP, osteopontin (OPN, collagen type 1 (col 1 and runt-related transcription factor-2 (RUNX-212 h after the first loading, cell viability by MTS assay and alkaline phosphatase activity at day 12 of culture and cell viability, collagen content by Sirius red and calcium content by alizarin red at day 24 of culture. Neither Dex nor loading had significant effects on cell viability. Collagen content was significantly higher (p<0.01 in the loaded group compared with the non-loaded group in all conditions. There was no difference in ALP activity or the amount of collagen and calcium produced between the non-loaded group supplemented with Dex and the loaded group without Dex. We conclude that dynamic loading has the ability to stimulate osteogenic differentiation of hMSC in the absence of glucocorticoids.

  7. Collagen osteoid-like model allows kinetic gene expression studies of non-collagenous proteins in relation with mineral development to understand bone biomineralization.

    Science.gov (United States)

    Silvent, Jérémie; Nassif, Nadine; Helary, Christophe; Azaïs, Thierry; Sire, Jean-Yves; Guille, Marie Madeleine Giraud

    2013-01-01

    Among persisting questions on bone calcification, a major one is the link between protein expression and mineral deposition. A cell culture system is here proposed opening new integrative studies on biomineralization, improving our knowledge on the role played by non-collagenous proteins in bone. This experimental in vitro model consisted in human primary osteoblasts cultured for 60 days at the surface of a 3D collagen scaffold mimicking an osteoid matrix. Various techniques were used to analyze the results at the cellular and molecular level (adhesion and viability tests, histology and electron microscopy, RT- and qPCR) and to characterize the mineral phase (histological staining, EDX, ATG, SAED and RMN). On long term cultures human bone cells seeded on the osteoid-like matrix displayed a clear osteoblast phenotype as revealed by the osteoblast-like morphology, expression of specific protein such as alkaline phosphatase and expression of eight genes classically considered as osteoblast markers, including BGLAP, COL1A1, and BMP2. Von Kossa and alizarine red allowed us to identify divalent calcium ions at the surface of the matrix, EDX revealed the correct Ca/P ratio, and SAED showed the apatite crystal diffraction pattern. In addition RMN led to the conclusion that contaminant phases were absent and that the hydration state of the mineral was similar to fresh bone. A temporal correlation was established between quantified gene expression of DMP1 and IBSP, and the presence of hydroxyapatite, confirming the contribution of these proteins to the mineralization process. In parallel a difference was observed in the expression pattern of SPP1 and BGLAP, which questioned their attributed role in the literature. The present model opens new experimental possibilities to study spatio-temporal relations between bone cells, dense collagen scaffolds, NCPs and hydroxyapatite mineral deposition. It also emphasizes the importance of high collagen density environment in bone cell

  8. Uncertainty evaluation for determination of fluorine content by distillation-thorium nitrate volumetric method%蒸馏-硝酸钍容量法测定氟含量不确定度评定

    Institute of Scientific and Technical Information of China (English)

    张小霞

    2013-01-01

    The uncertainty sources in determination of fluorine content of cryolite and aluminium fluoride by distillation-thorium nitrate volumetric method were analyzed.Evaluation for the uncertainty of each component was carried out by establishing the mathematic model,and the expanded uncertainty of fluorine content determination with distillation-thorium nitrate volumetric method was also calculated.Experimental principle:melt the sample with anhydrous sodium carbonate,separate fluoride through sulfuric acid-water vapor distillation,taking sodium alizarin sulfonate and methylene blue as indicators,using thorium nitrate standard solution to titrate fluorine content in the sample.Evaluation results indicated that,the expanded uncertainty of thorium nitrate volumetric method for the determination of fluorine content was 0.42%.%对蒸馏-硝酸钍容量法测定冰晶石及氟化铝中氟含量的不确定度来源进行分析,通过建立数学模型,评定各分量的不确定度,并计算蒸馏-硝酸钍容量法测定氟元素含量的扩展不确定度.实验原理:试样用无水碳酸钠熔融,经硫酸-水蒸气蒸馏分离氟后,以茜素磺酸钠和次甲基蓝作指示剂,用硝酸钍标准溶液滴定测定样品中氟含量.评定结果表明,蒸馏-硝酸钍容量法测定氟含量结果的扩展不确定度为0.42%.

  9. Neuropeptide Y1 Receptor Regulates Glucocorticoid-Induced Inhibition of Osteoblast Differentiation in Murine MC3T3-E1 Cells via ERK Signaling

    Directory of Open Access Journals (Sweden)

    Wei Yu

    2016-12-01

    Full Text Available High dose glucocorticoid (GC administration impairs the viability and function of osteoblasts, thus causing osteoporosis and osteonecrosis. Neuropeptide Y1 receptor (Y1 receptor is expressed in bone tissues and cells, and regulates bone remodeling. However, the role of Y1 receptor in glucocorticoid-induced inhibition of osteoblast differentiation remains unknown. In the present study, osteoblastic cell line MC3T3-E1 cultured in osteogenic differentiation medium was treated with or without of 10−7 M dexamethasone (Dex, Y1 receptor shRNA interference, Y1 receptor agonist [Leu31, Pro34]-NPY, and antagonist BIBP3226. Cell proliferation and apoptosis were assessed by cell counting kit-8 (CCK-8 assay and cleaved caspase expression, respectively. Osteoblast differentiation was evaluated by Alizarin Red S staining and osteogenic marker gene expressions. Protein expression was detected by Western blot analysis. Dex upregulated the expression of Y1 receptor in MC3T3-E1 cells associated with reduced osteogenic gene expressions and mineralization. Blockade of Y1 receptor by shRNA transfection and BIBP3226 significantly attenuated the inhibitory effects of Dex on osteoblastic activity. Y1 receptor signaling modulated the activation of extracellular signal-regulated kinases (ERK as well as the expressions of osteogenic genes. Y1 receptor agonist inhibited ERK phosphorylation and osteoblast differentiation, while Y1 receptor blockade exhibited the opposite effects. Activation of ERK signaling by constitutive active mutant of MEK1 (caMEK abolished Y1 receptor-mediated Dex inhibition of osteoblast differentiation in MC3T3-E1 cells. Taken together, Y1 receptor regulates Dex-induced inhibition of osteoblast differentiation in murine MC3T3-E1 cells via ERK signaling. This study provides a novel role of Y1 receptor in the process of GC-induced suppression in osteoblast survival and differentiation.

  10. Protective effect of quercetin on skeletal and neural tube teratogenicity induced by cyclophosphamide in rat fetuses

    Science.gov (United States)

    Khaksary Mahabady, Mahmood; Gholami, Mohammad Reza; Najafzadeh Varzi, Hossein; Zendedel, Abolfazl; Doostizadeh, Mona

    2016-01-01

    Cyclophosphamide (CP) is a drug commonly used to treat neoplastic disease and some autoimmune diseases. It is also a well-known and well-studied teratogen causing a variety of birth defects in fetuses of pregnant women treated with the drug. There are many reports that show the adverse effects of CP can be decreased by use of antioxidant drugs. It appears that, quercetin has antioxidant effect. The aim of this study was prevention or decrease of teratogenicity of CP in fetuses of rats by quercetin. This study was performed on 35 pregnant rats divided into six groups. Control group was received normal saline (5 mL kg-1, intraperitoneally) and 2-6 groups received a single dose of CP (15 mg kg-1), a single dose of quercetin (75 or 200 mg kg-1), CP plus quercetin (75 or 200 mg kg-1) intraperitoneally at 9th day of gestation, respectively. Fetuses were collected at 20th day of gestation and after determination of weight and crown rump length were stained by alizarin red – alcian blue method and skeletal system were examined by stereomicroscope. The results showed that the cleft palate, exencephaly, spina bifida and omphalocele incidence were 55.56%, 27.77%, 33.34% and 11.11%, in fetuses of rat that received only CP, respectively. However, it decreased to 16.00%, 16.00%, 16.00% and 8.00% by quercetin (75 mg kg-1) and so to 12.90%, 12.90%, 6.45% and 3.28% by quercetin (200 mg kg-1), respectively. On the basis of results, quercetin significantly can decrease teratogenicity induced by CP. PMID:27482358

  11. Wnt3a enhances bone morphogenetic protein 9-induced osteogenic differentiation of C3H10T1/2 cells

    Institute of Scientific and Technical Information of China (English)

    ZHANG Xiao; LIN Liang-bo; XU Dao-jing; CHEN Rong-fu; TAN Ji-xiang; LIANG Xi; HU Ning

    2013-01-01

    Background Bone morphogenetic protein 9 (BMP9) and Wnt/β-catenin signaling pathways are able to induce osteogenic differentiation of mesenchymal stem cells (MSCs),but the role of Wnt/β-catenin signaling pathway in BMP9-induced osteogenic differentiation is not well understood.Thus,our experiment was undertaken to investigate the interaction between BMP9 and Wnt/β-catenin pathway in inducing osteogenic differentiation of MSCs.Methods C3H10T1/2 cells were infected with recombinant adenovirus expressing BMP9,Wnt3a,and BMP9+Wnt3a.ALP,the early osteogenic marker,was detected by quantitative and staining assay.Later osteogenic marker,mineral calcium deposition,was determined by Alizarin Red S staining.The expression of osteopotin (OPN),osteocalcin (OC),and Runx2 was analyzed by Real time PCR and Western blotting.In vivo animal experiment was carried out to further confirm the role of Wnt3a in ectopic bone formation induced by BMP9.Results The results showed that Wnt3a enhanced the ALP activity induced by BMP9 and increased the expressions of OC and OPN,with increase of mineral calcium deposition in vitro and ectopic bone formation in vivo.Furthermore,we also found that Wnt3a increased the level of Runx2,an important nuclear transcription factor of BMP9.Conclusion Canonical Wnt/β-catenin signal pathway may play an important role in BMP9-induced osteogenic differentiation of MSCs,and Runx2 may be a linkage between the two signal pathways.

  12. Enhancement of osteogenic differentiation and proliferation in human mesenchymal stem cells by a modified low intensity ultrasound stimulation under simulated microgravity.

    Directory of Open Access Journals (Sweden)

    Sardar M Z Uddin

    Full Text Available Adult stem cells can differentiate into multiple lineages depending on their exposure to differing biochemical and biomechanical inductive factors. Lack of mechanical signals due to disuse can inhibit osteogenesis and induce adipogenesis of mesenchymal stem cells (MSCs. Long-term bed rest due to both brain/spinal cord injury and space travel can lead to disuse osteoporosis that is in part caused by a reduced number of osteoblasts. Thus, it is essential to provide proper mechanical stimulation for cellular viability and osteogenesis, particularly under disuse conditions. The objective of this study was to examine the effects of low intensity pulsed ultrasound (LIPUS on the osteogenic differentiation of adipose-derived human stem cells (Ad-hMSC in simulated microgravity conditions. Cells were cultured in a 1D clinostat to simulate microgravity (SMG and treated with LIPUS at 30mW/cm(2 for 20 min/day. It was hypothesized that the application of LIPUS to SMG cultures would restore osteogenesis in Ad-hMSCs. The results showed significant increases in ALP, OSX, RANKL, RUNX2, and decreases in OPG in LIPUS treated SMG cultures of Ad-MSC compared to non-treated cultures. LIPUS also restored OSX, RUNX2 and RANKL expression in osteoblast cells. SMG significantly reduced ALP positive cells by 70% (p<0.01 and ALP activity by 22% (p<0.01, while LIPUS treatment restored ALP positive cell number and activity to equivalence with normal gravity controls. Extracellular matrix collagen and mineralization was assessed by Sirius red and Alizarin red staining, respectively. SMG cultures showed little or no collagen or mineralization, but LIPUS treatment restored collagen content to 50% (p<0.001 and mineralization by 45% (p<0.001 in LIPUS treated-SMG cultures relative to SMG-only cultures. The data suggest that LIPUS treatment can restore normal osteogenic differentiation of MSCs from disuse by daily short duration stimulation.

  13. Neuropeptide Y1 Receptor Regulates Glucocorticoid-Induced Inhibition of Osteoblast Differentiation in Murine MC3T3-E1 Cells via ERK Signaling.

    Science.gov (United States)

    Yu, Wei; Zhu, Chao; Xu, Wenning; Jiang, Leisheng; Jiang, Shengdan

    2016-12-21

    High dose glucocorticoid (GC) administration impairs the viability and function of osteoblasts, thus causing osteoporosis and osteonecrosis. Neuropeptide Y1 receptor (Y1 receptor) is expressed in bone tissues and cells, and regulates bone remodeling. However, the role of Y1 receptor in glucocorticoid-induced inhibition of osteoblast differentiation remains unknown. In the present study, osteoblastic cell line MC3T3-E1 cultured in osteogenic differentiation medium was treated with or without of 10(-7) M dexamethasone (Dex), Y1 receptor shRNA interference, Y1 receptor agonist [Leu(31), Pro(34)]-NPY, and antagonist BIBP3226. Cell proliferation and apoptosis were assessed by cell counting kit-8 (CCK-8) assay and cleaved caspase expression, respectively. Osteoblast differentiation was evaluated by Alizarin Red S staining and osteogenic marker gene expressions. Protein expression was detected by Western blot analysis. Dex upregulated the expression of Y1 receptor in MC3T3-E1 cells associated with reduced osteogenic gene expressions and mineralization. Blockade of Y1 receptor by shRNA transfection and BIBP3226 significantly attenuated the inhibitory effects of Dex on osteoblastic activity. Y1 receptor signaling modulated the activation of extracellular signal-regulated kinases (ERK) as well as the expressions of osteogenic genes. Y1 receptor agonist inhibited ERK phosphorylation and osteoblast differentiation, while Y1 receptor blockade exhibited the opposite effects. Activation of ERK signaling by constitutive active mutant of MEK1 (caMEK) abolished Y1 receptor-mediated Dex inhibition of osteoblast differentiation in MC3T3-E1 cells. Taken together, Y1 receptor regulates Dex-induced inhibition of osteoblast differentiation in murine MC3T3-E1 cells via ERK signaling. This study provides a novel role of Y1 receptor in the process of GC-induced suppression in osteoblast survival and differentiation.

  14. Application of chromatography and mass spectrometry to the characterization of cobalt, copper, manganese and molybdenum in Morinda citrifolia.

    Science.gov (United States)

    Rybak, Justyna; Ruzik, Lena

    2013-03-15

    An analytical procedure was proposed to determine the manganese species and to study the fractionation of microelements such as copper, cobalt and molybdenum in Noni juice. Morinda citrifolia is known as a noni fruit, Indian mulberry, nunaakai, dog dumpling, mengkudu, beach mulberry, vomit fruit and cheese fruit. It is a tropical plant with a long tradition of medicinal use in Polynesia and tropical parts of eastern Asia and Australia. This article covers the determination of manganese species in Noni juice and established by fractionation by size exclusion chromatography inductively coupled plasma mass spectrometry (SEC ICP MS) and next characterization of species by electrospray ionization mass spectrometry (ESI MS). Also presented the fractionation analysis of copper, cobalt and molybdenum in Noni juice sample using SEC ICP MS - juice was treated with buffer and enzymatic extraction media and analyzed. For the evaluation of the amounts of the metal fractions distinguished, the ICP MS was used off-line prior to the determination of copper, cobalt, molybdenum and manganese concentrations in the juice. It was established that elements are present in the analyzed samples in different species and their concentration is μg mL(-1) and ng mL(-1) range in fruit. The accuracy of the entire fractionation scheme and sample preparation procedures involved was verified by the performance of the recovery test. For the information about the bioavailability of these elements, in vitro bioavailability investigation was used by SEC ICP MS technique. Two step digestion model simulating gastric (pepsin digestion) and intestinal (pancreatin digestion) juices. In Noni juice, manganese is complexed from flavonoids - rutin, from dye like anthraquinone (alizarin) and glycosides - asperulosidic acid (ESI MS - characterization). The study shows that copper and molybdenum contained in Noni juice are complexed by peptides, and cobalt by organic acids (which are 3.6% of juice). Molybdenum in

  15. Effect of Calcitriol on Differentiation of Periodontal Ligament Stem Cells to Osteoblasts

    Directory of Open Access Journals (Sweden)

    Soheilifar

    2016-02-01

    Full Text Available Background Periodontium may be able to respond to injuries by regeneration via the function of stem cells. Objectives This study sought to assess the differentiation of human periodontal ligament stem cells (PDLSCs into osteoblasts in standard osteogenic medium and in a medium supplemented with 1,25-dihydroxyvitamin D3 (calcitriol. Materials and Methods In this experimental study, PDLSCs were isolated under sterile conditions by scraping the periodontal ligament tissues attached to the middle third of the root surface of extracted teeth, which were obtained from patients who were candidates for orthodontics therapy in the dental faculty at Hamadan University. The collected cells were cultured on four culture plates for 24 hours. Group 1 contained a basic medium (α-MEM, containing 10% fetal bovine serum (FBS, 5 mM β-glycerophosphate, and 50 μg/mL l-ascorbic acid, supplemented with 10 - 8 M dexamethasone. Group 2 contained a basic medium supplemented with vitamin D3. Group 3 contained a basic medium supplemented with vitamin D3 and dexamethasone, Group4 contained negative control cultures. Alizarin red staining (ARS, alkaline phosphatase (ALP activity, and calcium content (CC tests were performed to evaluate osteogenic differentiation of third passage cells in the developing adherent layer. Results Quantitative analysis of ARS demonstrated that mineralized nodule formation was highest in the group supplemented with calcitriol and dexamethasone (P < 0.001. Results of the ALP test on day 28 demonstrated the highest ALP activity in the group supplemented with calcitriol (P < 0.001. The amount of CC was lowest in the control group at all-time points, and was highest in the group supplemented with both calcitriol and dexamethasone on day 28 (P < 0.001. Conclusions The combination of calcitriol with dexamethasone, ascorbic acid, and beta-glycerophosphate (that is, the osteogenic medium may be beneficial for differentiation of PDLSCs into osteoblasts.

  16. The effect of hypoxia on facial shape variation and disease phenotypes in chicken embryos

    Directory of Open Access Journals (Sweden)

    Francis Smith

    2013-07-01

    Craniofacial anomalies can arise from both genetic and environmental factors, including prenatal hypoxia. Recent clinical evidence correlates hypoxia to craniofacial malformations. However, the mechanisms by which hypoxia mediates these defects are not yet understood. We examined the cellular mechanisms underlying malformations induced by hypoxia using a chicken (Gallus gallus embryo model. Eggs were incubated in either hypoxic (7, 9, 11, 13, 15, 17 or 19% O2 or normoxic (21% O2 conditions. Embryos were photographed for morphological analysis at days 3–6. For analysis of skeletal development, 13-day embryos were cleared and stained with alcian blue and alizarin red for cartilage and bone, respectively. Quantitative analysis of facial shape variation was performed on images of embryos via geometric morphometrics. Early-stage embryos (day 2 were analyzed for apoptosis via whole-mount and section TUNEL staining and immunostaining for cleaved caspase-3, whereas later-stage embryos (days 4–6 were sectioned in paraffin for analysis of cell proliferation (BrdU, apoptosis (TUNEL and metabolic stress (phospho-AMPK. Results demonstrate that survival is reduced in a dose-dependent manner. Hypoxic embryos displayed a spectrum of craniofacial anomalies, from mild asymmetry and eye defects to more severe frontonasal and cephalic anomalies. Skull bone development was delayed in hypoxic embryos, with some skeletal defects observed. Morphometric analysis showed facial shape variation relative to centroid size and age in hypoxic groups. Hypoxia disrupted cell proliferation and, in early-stage embryos, caused apoptosis of neural crest progenitor cells. Hypoxic embryos also displayed an increased metabolic stress response. These results indicate that hypoxia during early embryonic craniofacial development might induce cellular oxidative stress, leading to apoptosis of the neural crest progenitor cells that are crucial to normal craniofacial morphogenesis.

  17. Minocycline Loaded Hybrid Composites Nanoparticles for Mesenchymal Stem Cells Differentiation into Osteogenesis

    Directory of Open Access Journals (Sweden)

    Allister Yingwei Tham

    2016-07-01

    Full Text Available Bone transplants are used to treat fractures and increase new tissue development in bone tissue engineering. Grafting of massive implantations showing slow curing rate and results in cell death for poor vascularization. The potentials of biocomposite scaffolds to mimic extracellular matrix (ECM and including new biomaterials could produce a better substitute for new bone tissue formation. A purpose of this study is to analyze polycaprolactone/silk fibroin/hyaluronic acid/minocycline hydrochloride (PCL/SF/HA/MH nanoparticles initiate human mesenchymal stem cells (MSCs proliferation and differentiation into osteogenesis. Electrospraying technique was used to develop PCL, PCL/SF, PCL/SF/HA and PCL/SF/HA/MH hybrid biocomposite nanoparticles and characterization was analyzed by field emission scanning electron microscope (FESEM, contact angle and Fourier transform infrared spectroscopy (FT-IR. The obtained results proved that the particle diameter and water contact angle obtained around 0.54 ± 0.12 to 3.2 ± 0.18 µm and 43.93 ± 10.8° to 133.1 ± 12.4° respectively. The cell proliferation and cell-nanoparticle interactions analyzed using (3-(4,5-dimethyl thiazol-2-yl-5-(3-carboxymethoxyphenyl-2-(4-sulfophenyl-2H-tetrazolium inner salt MTS assay (Promega, Madison, WI, USA, FESEM for cell morphology and 5-Chloromethylfluorescein diacetate (CMFDA dye for imaging live cells. Osteogenic differentiation was proved by expression of osteocalcin, alkaline phosphatase activity (ALP and mineralization was confirmed by using alizarin red (ARS. The quantity of cells was considerably increased in PCL/SF/HA/MH nanoparticles when compare to all other biocomposite nanoparticles and the cell interaction was observed more on PCL/SF/HA/MH nanoparticles. The electrosprayed PCL/SF/HA/MH biocomposite nanoparticle significantly initiated increased cell proliferation, osteogenic differentiation and mineralization, which provide huge potential for bone tissue engineering.

  18. Autophagy protects end plate chondrocytes from intermittent cyclic mechanical tension induced calcification.

    Science.gov (United States)

    Xu, Hong-guang; Yu, Yun-fei; Zheng, Quan; Zhang, Wei; Wang, Chuang-dong; Zhao, Xiao-yn; Tong, Wen-xue; Wang, Hong; Liu, Ping; Zhang, Xiao-ling

    2014-09-01

    Calcification of end plate chondrocytes is a major cause of intervertebral disc (IVD) degeneration. However, the underlying molecular mechanism of end plate chondrocyte calcification is still unclear. The aim of this study was to clarify whether autophagy in end plate chondrocytes could protect the calcification of end plate chondrocytes. Previous studies showed that intermittent cyclic mechanical tension (ICMT) contributes to the calcification of end plate chondrocytes in vitro. While autophagy serves as a cell survival mechanism, the relationship of autophagy and induced end plate chondrocyte calcification by mechanical tension in vitro is unknown. Thus, we investigated autophagy, the expression of the autophagy genes, Beclin-1 and LC3, and rat end plate chondrocyte calcification by ICMT. The viability of end plate chondrocytes was examined using the LIVE/DEAD viability/cytotoxicity kit. The reverse transcription-polymerase chain reaction and western blotting were used to detect the expression of Beclin-1; LC3; type I, II and X collagen; aggrecan; and Sox-9 genes. Immunofluorescent and fluorescent microscopy showed decreased autophagy in the 10- and 20-day groups loaded with ICMT. Additionally, Alizarin red and alkaline phosphatase staining detected the palpable calcification of end plate chondrocytes after ICMT treatment. We found that increased autophagy induced by short-term ICMT treatment was accompanied by an insignificant calcification of end plate chondrocytes. To the contrary, the suppressive autophagy inhibited by long-term ICMT was accompanied by a more significant calcification. The process of calcification induced by ICMT was partially resisted by increased autophagy activity induced by rapamycin, implicating that autophagy may prevent end plate chondrocyte calcification.

  19. Effects of melatonin on the proliferation and differentiation of rat adipose-derived stem cells

    Directory of Open Access Journals (Sweden)

    Zaminy Arash

    2008-01-01

    Full Text Available Background: Osteogenesis driven by adipose-derived stem cells (ADSCs is regulated by physiological and pathological factors. Accumulating evidence from in vitro and in vivo experiments suggests that melatonin may have an influence on bone formation. However, little is known about the effects of melatonin on osteogenesis, which thus remains to be elucidated. This study was performed to determine whether melatonin at physiological concentrations (0.01-10 nM could affect the in vitro proliferation and osteogenic differentiation of rat ADSCs. Materials and Methods: ADSCs were isolated from the fat of adult rats. After cell expansion in culture media and through three passages, osteogenesis was induced in a monolayer culture using osteogenic medium with or without melatonin at physiological concentrations (0.01-10 nM. After four weeks, the cultures were examined for mineralization by Alizarin Red S and von Kossa staining and for alkaline phosphatase (ALP activity using an ALP kit. Cell viability and apoptosis were also assayed by 3-(4, 5-dimethylthiazol-2-yl-5-(3-carboxymethoxyphenyl-2-(4-sulfophenyl-2H-tetrazolium (MTT assay and flow cytometry, respectively. Results: The results indicated that at physiological concentrations, melatonin suppressed proliferation and differentiation of ADSCs. These data indicate that ADSCs exposed to melatonin, had a lower ALP activity in contrast to the cells exposed to osteogenic medium alone. Similarly, mineral deposition (calcium level also decreased in the presence of melatonin. Flow cytometry confirmed that cell growth had decreased and that the numbers of apoptotic cells had increased. Conclusion: These results suggest that the physiological concentration of melatonin has a negative effect on ADSC osteogenesis.

  20. Micro/Nano Multilayered Scaffolds of PLGA and Collagen by Alternately Electrospinning for Bone Tissue Engineering

    Science.gov (United States)

    Kwak, Sanghwa; Haider, Adnan; Gupta, Kailash Chandra; Kim, Sukyoung; Kang, Inn-Kyu

    2016-07-01

    The dual extrusion electrospinning technique was used to fabricate multilayered 3D scaffolds by stacking microfibrous meshes of poly(lactic acid-co-glycolic acid) (PLGA) in alternate fashion to micro/nano mixed fibrous meshes of PLGA and collagen. To fabricate the multilayered scaffold, 35 wt% solution of PLGA in THF-DMF binary solvent (3:1) and 5 wt% solution of collagen in hexafluoroisopropanol (HFIP) with and without hydroxyapatite nanorods (nHA) were used. The dual and individual electrospinning of PLGA and collagen were carried out at flow rates of 1.0 and 0.5 mL/h, respectively, at an applied voltage of 20 kV. The density of collagen fibers in multilayered scaffolds has controlled the adhesion, proliferation, and osteogenic differentiation of MC3T3-E1 cells. The homogeneous dispersion of glutamic acid-modified hydroxyapatite nanorods (nHA-GA) in collagen solution has improved the osteogenic properties of fabricated multilayered scaffolds. The fabricated multilayered scaffolds were characterized using FT-IR, X-ray photoelectron spectroscopy, and transmission electron microscopy (TEM). The scanning electron microscopy (FE-SEM) was used to evaluate the adhesion and spreads of MC3T3-E1 cells on multilayered scaffolds. The activity of MC3T3-E1 cells on the multilayered scaffolds was evaluated by applying MTT, alkaline phosphatase, Alizarin Red, von Kossa, and cytoskeleton F-actin assaying protocols. The micro/nano fibrous PLGA-Col-HA scaffolds were found to be highly bioactive in comparison to pristine microfibrous PLGA and micro/nano mixed fibrous PLGA and Col scaffolds.

  1. The effect of magnetic stimulation on the osteogenic and chondrogenic differentiation of human stem cells derived from the adipose tissue (hASCs)

    Science.gov (United States)

    Lima, João; Gonçalves, Ana I.; Rodrigues, Márcia T.; Reis, Rui L.; Gomes, Manuela E.

    2015-11-01

    The use of magnetic nanoparticles (MNPs) towards the musculoskeletal tissues has been the focus of many studies, regarding MNPs ability to promote and direct cellular stimulation and orient tissue responses. This is thought to be mainly achieved by mechano-responsive pathways, which can induce changes in cell behavior, including the processes of proliferation and differentiation, in response to external mechanical stimuli. Thus, the application of MNP-based strategies in tissue engineering may hold potential to propose novel solutions for cell therapy on bone and cartilage strategies to accomplish tissue regeneration. The present work aims at studying the influence of MNPs on the osteogenic and chondrogenic differentiation of human adipose derived stem cells (hASCs). MNPs were incorporated in hASCs and cultured in medium supplemented for osteogenic and chondrogenic differentiation. Cultures were maintained up to 28 days with/without an external magnetic stimulus provided by a magnetic bioreactor, to determine if the MNPs alone could affect the osteogenic or chondrogenic phenotype of the hASCs. Results indicate that the incorporation of MNPs does not negatively affect the viability nor the proliferation of hASCs. Furthermore, Alizarin Red staining evidences an enhancement in extracellular (ECM) mineralization under the influence of an external magnetic field. Although not as evident as for osteogenic differentiation, Toluidine blue and Safranin-O stainings also suggest the presence of a cartilage-like ECM with glycosaminoglycans and proteoglycans under the magnetic stimulus provided. Thus, MNPs incorporated in hASCs under the influence of an external magnetic field have the potential to induce differentiation towards the osteogenic and chondrogenic lineages.

  2. miRNA-720 controls stem cell phenotype, proliferation and differentiation of human dental pulp cells.

    Directory of Open Access Journals (Sweden)

    Emilio Satoshi Hara

    Full Text Available Dental pulp cells (DPCs are known to be enriched in stem/progenitor cells but not well characterized yet. Small non-coding microRNAs (miRNAs have been identified to control protein translation, mRNA stability and transcription, and have been reported to play important roles in stem cell biology, related to cell reprogramming, maintenance of stemness and regulation of cell differentiation. In order to characterize dental pulp stem/progenitor cells and its mechanism of differentiation, we herein sorted stem-cell-enriched side population (SP cells from human DPCs and periodontal ligament cells (PDLCs, and performed a locked nucleic acid (LNA-based miRNA array. As a result, miR-720 was highly expressed in the differentiated main population (MP cells compared to that in SP cells. In silico analysis and a reporter assay showed that miR-720 targets the stem cell marker NANOG, indicating that miR-720 could promote differentiation of dental pulp stem/progenitor cells by repressing NANOG. Indeed, gain-and loss-of-function analyses showed that miR-720 controls NANOG transcript and protein levels. Moreover, transfection of miR-720 significantly decreased the number of cells positive for the early stem cell marker SSEA-4. Concomitantly, mRNA levels of DNA methyltransferases (DNMTs, which are known to play crucial factors during stem cell differentiation, were also increased by miR-720 through unknown mechanism. Finally, miR-720 decreased DPC proliferation as determined by immunocytochemical analysis against ki-67, and promoted odontogenic differentiation as demonstrated by alizarin red staining, as well as alkaline phosphatase and osteopontin mRNA levels. Our findings identify miR-720 as a novel miRNA regulating the differentiation of DPCs.

  3. Marking of burbot Lota lota (L. juveniles using commercial feed as a vector of fluorochrome application

    Directory of Open Access Journals (Sweden)

    Katarzyna Stańczak

    2015-10-01

    Full Text Available The effectiveness of restocking is an important element of sustainable fishery. In restocking programmes, where larvae and juveniles are used mass marking is carried out with non-toxic dyes such as fluorochromes transferred into the fish body by immersion or with feed. The experiment was carried out to assess the possibility of using commercial feed supplemented Alizarin Red S (ARS as an alternative non-invasive method for the mass marking of burbot fry. Fish of an average body weight of 1.8 (± 0.2 g were reared in separated 10 dm3 tanks and fed for 10 days with extruded feed supplemented ARS in six concentrations, i.e. 0 (control, 20, 40, 60, 80 and 100 g fluorochrome per kg of feed at a daily dose of 5% of biomass. During the next 15 days fish were fed commercial feed without ARS. After 25 days of rearing 30 individuals were sampled from each group, euthanized and total length and wet body weight of each fish were determined. Next, the sampled fish were preserved in 70% ethyl alcohol. Then otoliths were excised from the fish and analysed under a fluorescence microscope to identify fluorescent tags. Marked individuals were identified in each analysed group provided with feed supplemented ARS. The highest rate of marked fish (100% was found in the group that received 40 g ARS per kg of feed, and the lowest (73.3% in the group that received 60 g ARS per kg of feed. Supplementation of feed with ARS had no negative effect on the survival rate and growth parameters (mean body weight and total body length of burbot. The results suggest that ARS applied in commercial feed (40 g/kg for about 7-10 days is an effective method for marking burbot fry and can be recommended in fishery practice.

  4. HPLC-DAD-MS analysis of dyes identified in textiles from Mount Athos.

    Science.gov (United States)

    Mantzouris, Dimitrios; Karapanagiotis, Ioannis; Valianou, Lemonia; Panayiotou, Costas

    2011-03-01

    Organic colorants contained in 30 textiles (16th to early 20th century) from the monastery of Simonos Petra (Mount Athos) have been investigated using high-performance liquid chromatography equipped with diode-array detection and mass spectrometry (HPLC-DAD-MS). The components of natural dyes identified in samples treated by the standard HCl dyestuff extraction method were: alizarin, apigenin, butein, carminic acid, chrysoeriol, dcII, dcIV, dcVII, ellagic acid, emodin, fisetin, flavokermesic acid, fustin, genistein, haematein derivative (Hae'), indigotin, indirubin, isoliquiritigenin, isorhamnetin, kaempferide, kaempferol, kermesic acid, luteolin, naringenin, purpurin, quercetin, rhamnazin, rhamnetin, sulfuretin, and type B and type C compounds (last two are markers for Caesalpinia trees). Early, semi-synthetic dyes, for example indigo carmine, fuchsin components, and rhodamine B were identified in objects dated late 19th to early 20th century. A dyestuff extraction method which involves use of TFA, instead of HCl, was applied to selected historical samples, showing that the mild method enables efficient extraction of weld (Reseda luteola L.) and dyer's broom (Genista tinctoria L.) glycosides. The marker compound (Hae') for logwood (Haematoxylum campechianum L.) identification after treatment with HCl was investigated by liquid chromatography coupled to mass spectrometry (LC-MS) in negative electrospray ionization (LC-MS-ESI(-)) mode. LC-MS in negative atmospheric pressure chemical ionization (LC-MS-APCI(-)) mode was used, probably for the first time, to investigate cochineal (Dactylopius coccus Costa) samples. Positive electrospray ionization (LC-MS-ESI(+)) mode was used for identification of fuchsin components. Detailed HPLC-DAD studies were performed on young fustic (Cotinus coggygria Scop.) and Persian berries (Rhamnus trees).

  5. Neuropeptide Y1 Receptor Regulates Glucocorticoid-Induced Inhibition of Osteoblast Differentiation in Murine MC3T3-E1 Cells via ERK Signaling

    Science.gov (United States)

    Yu, Wei; Zhu, Chao; Xu, Wenning; Jiang, Leisheng; Jiang, Shengdan

    2016-01-01

    High dose glucocorticoid (GC) administration impairs the viability and function of osteoblasts, thus causing osteoporosis and osteonecrosis. Neuropeptide Y1 receptor (Y1 receptor) is expressed in bone tissues and cells, and regulates bone remodeling. However, the role of Y1 receptor in glucocorticoid-induced inhibition of osteoblast differentiation remains unknown. In the present study, osteoblastic cell line MC3T3-E1 cultured in osteogenic differentiation medium was treated with or without of 10−7 M dexamethasone (Dex), Y1 receptor shRNA interference, Y1 receptor agonist [Leu31, Pro34]-NPY, and antagonist BIBP3226. Cell proliferation and apoptosis were assessed by cell counting kit-8 (CCK-8) assay and cleaved caspase expression, respectively. Osteoblast differentiation was evaluated by Alizarin Red S staining and osteogenic marker gene expressions. Protein expression was detected by Western blot analysis. Dex upregulated the expression of Y1 receptor in MC3T3-E1 cells associated with reduced osteogenic gene expressions and mineralization. Blockade of Y1 receptor by shRNA transfection and BIBP3226 significantly attenuated the inhibitory effects of Dex on osteoblastic activity. Y1 receptor signaling modulated the activation of extracellular signal-regulated kinases (ERK) as well as the expressions of osteogenic genes. Y1 receptor agonist inhibited ERK phosphorylation and osteoblast differentiation, while Y1 receptor blockade exhibited the opposite effects. Activation of ERK signaling by constitutive active mutant of MEK1 (caMEK) abolished Y1 receptor-mediated Dex inhibition of osteoblast differentiation in MC3T3-E1 cells. Taken together, Y1 receptor regulates Dex-induced inhibition of osteoblast differentiation in murine MC3T3-E1 cells via ERK signaling. This study provides a novel role of Y1 receptor in the process of GC-induced suppression in osteoblast survival and differentiation. PMID:28009825

  6. Encapsulation of bone morphogenic protein-2 with Cbfa1-overexpressing osteogenic cells derived from human embryonic stem cells in hydrogel accelerates bone tissue regeneration.

    Science.gov (United States)

    Kim, Min Jung; Park, Ji Sun; Kim, Sinae; Moon, Sung-Hwan; Yang, Han Na; Park, Keun-Hong; Chung, Hyung-Min

    2011-08-01

    Bone tissue defects caused by trauma and disease are significant problems in orthopedic surgery. Human embryonic stem cells (hESCs) hold great promise for the treatment of bone tissue disease in regenerative medicine. In this study, we have established an effective method for the differentiation of osteogenic cells derived from hESCs using a lentiviral vector containing the transcription factor Cbfa1. Differentiation was initiated in embryoid body formation of Cbfa1-expressing hESCs, resulting in a highly purified population of osteogenic cells based on flow cytometric analysis. These cells also showed characteristics of osteogenic cells in vitro, as determined by reverse-transcription (RT)-polymerase chain reaction and immunocytochemistry using osteoblast-specific markers. We also evaluated the regenerative potential of Cbfa1-expressing cells derived from hESCs (hESC-CECs) compared with hESCs and the osteogenic effects of bone morphogenic protein-2 (BMP2) encapsulated in thermoreversible hydrogel in vivo. hESC-CECs were embedded in hydrogel constructs enriched with BMP2 to promote bone regeneration. We observed prominent mineralization and the formation of nodule-like structures using von Kossa and alizarin red S staining. In addition, the expression patterns of osteoblast-specific genes were verified by RT-polymerase chain reaction, and immunohistochemical analysis revealed that collagen type 1 and Cbfa1 were highly expressed in hESC-CECs compared with other cell types. Taken together, our results suggest that encapsulation of hESC-CECs with BMP2 in hydrogel constructs appears to be a promising method to enhance the in vitro osteoblastic differentiation and in vivo osteogenic activity of hESC-CECs.

  7. Tissue level material composition and mechanical properties in Brtl/+ mouse model of Osteogenesis Imperfecta after sclerostin antibody treatment

    Science.gov (United States)

    Lloyd, William R.; Sinder, Benjamin P.; Salemi, Joseph; Ominsky, Michael S.; Marini, Joan C.; Caird, Michelle S.; Morris, Michael D.; Kozloff, Kenneth M.

    2015-02-01

    Osteogenesis imperfecta (OI) is a genetic disorder resulting in defective collagen or collagen-associated proteins and fragile, brittle bones. To date, therapies to improve OI bone mass, such as bisphosphonates, have increased bone mass in the axial skeleton of OI patients, but have shown limited effects at reducing long bone fragility. Sclerostin antibody (Scl- Ab), currently in clinical trials for osteoporosis, stimulates bone formation and may have the potential to reduce long bone fracture rates in OI patients. Scl-Ab has been investigated as an anabolic therapy for OI in the Brtl/+ mouse model of moderately severe Type IV OI. While Scl-Ab increases long bone mass in the Brtl/+ mouse, it is not known whether material properties and composition changes also occur. Here, we report on the effects of Scl-Ab on wild type and Brtl/+ young (3 week) and adult (6 month) male mice. Scl-Ab was administered over 5 weeks (25mg/kg, 2x/week). Raman microspectroscopy and nanoindentation are used for bone composition and biomechanical bone property measurements in excised bone. Fluorescent labels (calcein and alizarin) at 4 time points over the entire treatment period are used to enable measurements at specific tissue age. Differences between wild type and Brtl/+ groups included variations in the mineral and matrix lattices, particularly the phosphate v1, carbonate v1, and the v(CC) proline and hydroxyproline stretch vibrations. Results of Raman spectroscopy corresponded to nanoindentation findings which indicated that old bone (near midcortex) is stiffer (higher elastic modulus) than new bone. We compare and contrast mineral to matrix and carbonate to phosphate ratios in young and adult mice with and without treatment.

  8. Analysis of the Fgfr2C342Y mouse model shows condensation defects due to misregulation of Sox9 expression in prechondrocytic mesenchyme

    Science.gov (United States)

    Peskett, Emma; Kumar, Samin; Baird, William; Jaiswal, Janhvi; Li, Ming; Patel, Priyanca; Britto, Jonathan A.

    2017-01-01

    ABSTRACT Syndromic craniosynostosis caused by mutations in FGFR2 is characterised by developmental pathology in both endochondral and membranous skeletogenesis. Detailed phenotypic characterisation of features in the membranous calvarium, the endochondral cranial base and other structures in the axial and appendicular skeleton has not been performed at embryonic stages. We investigated bone development in the Crouzon mouse model (Fgfr2C342Y) at pre- and post-ossification stages to improve understanding of the underlying pathogenesis. Phenotypic analysis was performed by whole-mount skeletal staining (Alcian Blue/Alizarin Red) and histological staining of sections of CD1 wild-type (WT), Fgfr2C342Y/+ heterozygous (HET) and Fgfr2C342Y/C342Y homozygous (HOM) mouse embryos from embryonic day (E)12.5-E17.5 stages. Gene expression (Sox9, Shh, Fgf10 and Runx2) was studied by in situ hybridisation and protein expression (COL2A1) by immunohistochemistry. Our analysis has identified severely decreased osteogenesis in parts of the craniofacial skeleton together with increased chondrogenesis in parts of the endochondral and cartilaginous skeleton in HOM embryos. The Sox9 expression domain in tracheal and basi-cranial chondrocytic precursors at E13.5 in HOM embryos is increased and expanded, correlating with the phenotypic observations which suggest FGFR2 signalling regulates Sox9 expression. Combined with abnormal staining of type II collagen in pre-chondrocytic mesenchyme, this is indicative of a mesenchymal condensation defect. An expanded spectrum of phenotypic features observed in the Fgfr2C342Y/C342Y mouse embryo paves the way towards better understanding the clinical attributes of human Crouzon–Pfeiffer syndrome. FGFR2 mutation results in impaired skeletogenesis; however, our findings suggest that many phenotypic aberrations stem from a primary failure of pre-chondrogenic/osteogenic mesenchymal condensation and link FGFR2 to SOX9, a principal regulator of skeletogenesis

  9. Boron Induces Early Matrix Mineralization via Calcium Deposition and Elevation of Alkaline Phosphatase Activity in Differentiated Rat Bone Marrow Mesenchymal Stem Cells

    Directory of Open Access Journals (Sweden)

    Bent-al-hoda Movahedi Najafabadi

    2016-04-01

    Full Text Available Objective: Boron (B is essential for plant development and might be an essential micronutrient for animals and humans. This study was conducted to characterize the impact of boric acid (BA on the cellular and molecular nature of differentiated rat bone marrow mesenchymal stem cells (BMSCs. Materials and Methods: In this experimental study, BMSCs were extracted and expanded to the 3rd passage, then cultured in Dulbecco’s Modified Eagle’s Medium (DMEM complemented with osteogenic media as well as 6 ng/ml and 6 μg/ml of BA. After 5, 10, 15 and 21 days the viability and the level of mineralization was determined using MTT assay and alizarin red respectively. In addition, the morphology, nuclear diameter and cytoplasmic area of the cells were studied with the help of fluorescent dye. The concentration of calcium, activity of alanine transaminase (ALT, aspartate transaminase (AST, lactate dehydrogenase (LDH and alkaline phosphatase (ALP as well as sodium and potassium levels were also evaluated using commercial kits and a flame photometer respectively. Results: Although 6 μg/ml of BA was found to be toxic, a concentration of 6 ng/ml increased the osteogenic ability of the cell significantly throughout the treatment. In addition it was observed that B treatment caused the early induction of matrix mineralization compared to controls. Conclusion: Although more investigation is required, we suggest the prescription of a very low concentration of B in the form of BA or foods containing BA, in groups at high risk of osteoporosis or in the case of bone fracture.

  10. Confocal laser scanning microscopy in study of bone calcification

    Energy Technology Data Exchange (ETDEWEB)

    Nishikawa, Tetsunari, E-mail: tetsu-n@cc.osaka-dent.ac.jp [Department of Oral Pathology, Osaka Dental University, Osaka (Japan); Kokubu, Mayu; Kato, Hirohito [Department of Oral Pathology, Osaka Dental University, Osaka (Japan); Imai, Koichi [Department of Biomaterials, Osaka Dental University, Osaka (Japan); Tanaka, Akio [Department of Oral Pathology, Osaka Dental University, Osaka (Japan)

    2012-12-01

    Highlights: Black-Right-Pointing-Pointer High-magnification images with depth selection, and thin sections were observed using CLSM. Black-Right-Pointing-Pointer The direction and velocity of calcification of the bone was observed by administration of 2 fluorescent dyes. Black-Right-Pointing-Pointer In dog femora grafted with coral blocks, newly-formed bone was observed in the coral block space with a rough surface. Black-Right-Pointing-Pointer Twelve weeks after dental implant was grafted in dog femora, the space between screws was filled with newly-formed bones. - Abstract: Bone regeneration in mandible and maxillae after extraction of teeth or tumor resection and the use of rough surface implants in bone induction must be investigated to elucidate the mechanism of calcification. The calcified tissues are subjected to chemical decalcification or physical grinding to observe their microscopic features with light microscopy and transmission electron microscopy where the microscopic tissue morphology is significantly altered. We investigated the usefulness of confocal laser scanning microscopy (CLSM) for this purpose. After staggering the time of administration of calcein and alizarin red to experimental rats and dogs, rat alveolar bone and dog femur grafted with coral as scaffold or dental implants were observed with CLSM. In rat alveolar bone, the calcification of newly-formed bone and net-like canaliculi was observed at the mesial bone from the roots progressed at the rate of 15 {mu}m/day. In dog femur grafted with coral, newly-formed bones along the space of coral were observed in an orderly manner. In dog femur with dental implants, after 8 weeks, newly-formed bone proceeded along the rough surface of the implants. CLSM produced high-magnification images of newly-formed bone and thin sections were not needed.

  11. In vitro and in vivo biocompatibility and osteogenesis of graphene-reinforced nanohydroxyapatite polyamide66 ternary biocomposite as orthopedic implant material

    Science.gov (United States)

    Zhang, Shiyang; Yang, Qiming; Zhao, Weikang; Qiao, Bo; Cui, Hongwang; Fan, Jianjun; Li, Hong; Tu, Xiaolin; Jiang, Dianming

    2016-01-01

    Graphene and its derivatives have been receiving increasing attention regarding their application in bone tissue engineering because of their excellent characteristics, such as a vast specific surface area and excellent mechanical properties. In this study, graphene-reinforced nanohydroxyapatite/polyamide66 (nHA/PA66) bone screws were prepared. The results of scanning electron microscopy observation and X-ray diffraction data showed that both graphene and nHA had good dispersion in the PA66 matrix. In addition, the tensile strength and elastic modulus of the composites were significantly improved by 49.14% and 21.2%, respectively. The murine bone marrow mesenchymal stem cell line C3H10T1/2 exhibited better adhesion and proliferation in graphene reinforced nHA/PA66 composite material compared to the nHA/PA66 composites. The cells developed more pseudopods, with greater cell density and a more distinguishable cytoskeletal structure. These results were confirmed by fluorescent staining and cell viability assays. After C3H10T1/2 cells were cultured in osteogenic differentiation medium for 7 and 14 days, the bone differentiation-related gene expression, alkaline phosphatase, and osteocalcin were significantly increased in the cells cocultured with graphene reinforced nHA/PA66. This result demonstrated the bone-inducing characteristics of this composite material, a finding that was further supported by alizarin red staining results. In addition, graphene reinforced nHA/PA66 bone screws were implanted in canine femoral condyles, and postoperative histology revealed no obvious damage to the liver, spleen, kidneys, brain, or other major organs. The bone tissue around the implant grew well and was directly connected to the implant. The soft tissues showed no obvious inflammatory reaction, which demonstrated the good biocompatibility of the screws. These observations indicate that graphene-reinforced nHA/PA66 composites have great potential for application in bone tissue

  12. BMP7 gene transfer via gold nanoparticles into stroma inhibits corneal fibrosis in vivo.

    Directory of Open Access Journals (Sweden)

    Ashish Tandon

    Full Text Available This study examined the effects of BMP7 gene transfer on corneal wound healing and fibrosis inhibition in vivo using a rabbit model. Corneal haze in rabbits was produced with the excimer laser performing -9 diopters photorefractive keratectomy. BMP7 gene was introduced into rabbit keratocytes by polyethylimine-conjugated gold nanoparticles (PEI2-GNPs transfection solution single 5-minute topical application on the eye. Corneal haze and ocular health in live animals was gauged with stereo- and slit-lamp biomicroscopy. The levels of fibrosis [α-smooth muscle actin (αSMA, F-actin and fibronectin], immune reaction (CD11b and F4/80, keratocyte apoptosis (TUNEL, calcification (alizarin red, vonKossa and osteocalcin, and delivered-BMP7 gene expression in corneal tissues were quantified with immunofluorescence, western blotting and/or real-time PCR. Human corneal fibroblasts (HCF and in vitro experiments were used to characterize the molecular mechanism mediating BMP7's anti-fibrosis effects. PEI2-GNPs showed substantial BMP7 gene delivery into rabbit keratocytes in vivo (2×10(4 gene copies/ug DNA. Localized BMP7 gene therapy showed a significant corneal haze decrease (1.68±0.31 compared to 3.2±0.43 in control corneas; p88%; p<0.0001, and immunoblotting of BMP7-transefected HCFs grown in the presence of TGFβ demonstrated significantly enhanced pSmad-1/5/8 (95%; p<0.001 and Smad6 (53%, p<0.001, and decreased αSMA (78%; p<0.001 protein levels. These results suggest that localized BMP7 gene delivery in rabbit cornea modulates wound healing and inhibits fibrosis in vivo by counter balancing TGFβ1-mediated profibrotic Smad signaling.

  13. Relationships between fetal body weight of Wistar rats at term and the extent of skeletal ossification

    Directory of Open Access Journals (Sweden)

    I. Chahoud

    2005-04-01

    Full Text Available We investigated the relationship between fetal body weight at term (pregnancy day 21 and the extent of ossification of sternum, metacarpus, metatarsus, phalanges (proximal, medial and distal of fore- and hindlimbs and cervical and coccygeal vertebrae in Wistar rats. The relationships between fetal body weight and sex, intrauterine position, uterine horn, horn size, and litter size were determined using historical control data (7594 fetuses; 769 litters of untreated rats. Relationships between body weight and degree of ossification were examined in a subset of 1484 historical control fetuses (154 litters which were subsequently cleared and stained with alizarin red S. Fetal weight was independent of horn size, uterine horn side (left or right or intrauterine position. Males were heavier than females and fetal weight decreased with increasing litter size. Evaluation of the skeleton showed that ossification of sternum, metacarpus and metatarsus was extensively complete and independent of fetal weight on pregnancy day 21. In contrast, the extent of ossification of fore- and hindlimb phalanges and of cervical and sacrococcygeal vertebrae was dependent on fetal body weight. The strongest correlation between body weight and degree of ossification was found for hindlimb, medial and proximal phalanges. Our data therefore suggest that, in full-term rat fetuses (day 21, reduced ossification of sternum, metacarpus and metatarsus results from a localized impairment of bone calcification (i.e., a malformation or variation rather than from general growth retardation and that ossification of hindlimb (medial and proximal phalanges is a good indicator of treatment-induced fetal growth retardation.

  14. Determination of Dye Concentration Using a Synergistic TiO2-Ce(SO4) 2 Photocatalytic Oxidation System%TiO2-Ce(SO4)2光催化氧化体系荧光光度法测定染料浓度

    Institute of Scientific and Technical Information of China (English)

    张一平

    2014-01-01

    As Ce( SO4 ) 2 can obtain the photo⁃induced electrons and thereby enhance the photocatalytic oxidation ability of semiconductor, we established a simple method of dye concentration determination by using a nano TiO2⁃Ce( SO4 ) 2 synergistic photocatalytic oxidation system and fluorescence spectrophotometric method.As alizarin red was used as simulated dyeing wastewater,the optimum reaction conditions for dye degradation were studied.The experimental results show that the optimal digestion conditions were as follows:pH=0.4,TiO2 dosage = 1.0 g/L,the initial concentration of Ce( SO4 ) 2= 6.0 mmol/L,solution temperature= 80 °C,and UV irradiation time = 12 min.%基于Ce( SO4)2可以捕获光生电子从而提高半导体光催化氧化能力的原理,建立了一种用纳米TiO2-Ce( SO4)2协同体系光催化氧化,然后采用荧光光度法测定染料浓度的简便方法。以茜素红为模拟染料废水,对染料消解的最佳反应条件进行了考察。实验结果表明,pH值0�4,TiO2用量1�0g/L,Ce( SO4)2初始浓度6�0mmol/L,溶液温度80℃,紫外光照时间12min为最佳的消解条件。

  15. Procaine Inhibits Osteo/Odontogenesis through Wnt/β-Catenin Inactivation

    Science.gov (United States)

    Herencia, Carmen; Diaz-Tocados, Juan Miguel; Jurado, Lidia; Montes de Oca, Addy; Rodríguez-Ortiz, Maria Encarnación; Martín-Alonso, Carmen; Martínez-Moreno, Julio M.; Vergara, Noemi; Rodríguez, Mariano; Almadén, Yolanda; Muñoz-Castañeda, Juan R.

    2016-01-01

    Introduction Periodontitis is a complex pathology characterized by the loss of alveolar bone. The causes and the mechanisms that promote this bone resorption still remain unknown. The knowledge of the critical regulators involved in the alteration of alveolar bone homeostasis is of great importance for developing molecular therapies. Procaine is an anesthetic drug with demethylant properties, mainly used by dentists in oral surgeries. The inhibitor role of Wnt signaling of procaine was described in vitro in colon cancer cells. Methods In this work we evaluated the role of procaine (1 uM) in osteo/odontogenesis of rat bone marrow mesenchymal stem cells. Similarly, the mechanisms whereby procaine achieves these effects were also studied. Results Procaine administration led to a drastic decrease of calcium content, alkaline phosphatase activity, alizarin red staining and an increase in the expression of Matrix Gla Protein. With respect to osteo/odontogenic markers, procaine decreased early and mature osteo/odontogenic markers. In parallel, procaine inhibited canonical Wnt/β-catenin pathway, observing a loss of nuclear β-catenin, a decrease in Lrp5 and Frizzled 3, a significant increase of sclerostin and Gsk3β and an increase of phosphorylated β-catenin. The combination of osteo/odontogenic stimuli and Lithium Chloride decreased mRNA expression of Gsk3β, recovered by Procaine. Furthermore it was proved that Procaine alone dose dependently increases the expression of Gsk3β and β-catenin phosphorylation. These effects of procaine were also observed on mature osteoblast. Interestingly, at this concentration of procaine no demethylant effects were observed. Conclusions Our results demonstrated that procaine administration drastically reduced the mineralization and osteo/odontogenesis of bone marrow mesenchymal stem cells inhibiting Wnt/β-catenin pathway through the increase of Gsk3β expression and β-catenin phosphorylation. PMID:27257912

  16. In vitro differentiation profile of osteoblasts derived from patients with Saethre-Chotzen syndrome.

    Science.gov (United States)

    Ratisoontorn, Chootima; Seto, Marianne L; Broughton, Kristen M; Cunningham, Michael L

    2005-04-01

    Seathre-Chotzen syndrome (SCS) is an autosomal dominant craniosynostosis syndrome, associated with loss-of-function mutations in the basic helix-loop-helix transcription factor, TWIST1. The biologic activity of TWIST1 has been implicated in the inhibition of differentiation of multiple cell lineages. Therefore, premature fusion of cranial sutures (craniosynostosis) in SCS may be mediated by altered differentiation of calvarial osteoblasts. In this study, we evaluated osteoblasts derived from calvarial bone of three patients with SCS and three unaffected individuals as controls to investigate the principle stages of osteoblast differentiation: (1) proliferation, (2) matrix maturation, and (3) mineralization. Using a BrdU-Hoechst flow cytometry assay, we found that the percent of proliferating cells was significantly reduced in cells derived from patients with SCS compared with those derived from controls (P < or = 0.05). In the matrix maturation stage, alkaline phosphatase (ALP) enzyme activity and the expression of extracellular matrix genes, collagen I alpha 2 (COL1A2), osteopontin (OPN), osteocalcin (OC), and the runt-related transcription factor RUNX2 were examined by enzymatic assay and real-time quantitative RT-PCR, respectively. We identified no significant differences in the expression of matrix related transcripts. However, we found significant reductions in ALP activity on days 3 and 7 and in RUNX2 expression on days 14 and 21 (P < or = 0.05). Quantitative alizarin red S mineralization assays showed a trend toward increased mineralization in osteoblasts derived from patients with SCS at days 21 and 28, although not statistically significant. Our results demonstrated that loss-of-function mutations of TWIST1 led to reduced proliferation regardless of the functional domain affected. We did not find any conclusive differences in matrix maturation or mineralization in these primary osteoblasts. It is plausible that mutations in different functional domains of

  17. Hypoxia Promotes Osteogenesis of Human Placental-Derived Mesenchymal Stem Cells.

    Science.gov (United States)

    Gu, Qiaoli; Gu, Yanzheng; Shi, Qin; Yang, Huilin

    2016-01-01

    Placental-derived mesenchymal stem cells (pMSCs) are promising candidates for regenerative medicine because they possess high proliferative capacity and multi-differentiation potential. Human pMSCs are residing in an environment with low oxygen tension in the body. Heme oxygenase-1 (HO-1) is known to participate in the regulation of MSC differentiation. The present study aimed to investigate the impact of hypoxia on the osteogenic differentiation of human pMSCs, and to elucidate the role of HO-1 in the osteogenic differentiation of hypoxic pMSCs. Human pMSCs were cultured under normoxia (21% O2) or hypoxia (5% O2) for 3 days. We found that hypoxia maintained the morphology and immunophenotype of human pMSCs. The expression of stemness markers Oct4, Nanog, and Sox2 was increased under hypoxia. After a 5-day hypoxic culture, the proliferation ability of pMSCs was increased, which might be correlated with the increased expression of stem cell factor. During osteogenic induction, hypoxia increased the expression of osteogenic genes including osteopontin, osteocalcin, and alkaline phosphatase (ALP). Moreover, hypoxia increased the mineralization and ALP levels of human pMSCs as evidenced by Alizarin Red staining and ALP staining. Upregulation of HO-1 by cobalt-protoporphyrin treatment increased the osteogenic differentiation of pMSCs under hypoxia, while inhibition of HO-1 by Zn-protoporphyrin reduced the osteogenic differentiation of hypoxic pMSCs. Taken together, our data suggest that hypoxia can promote the osteogenic differentiation of human pMSCs. Upregulation of HO-1 can further increase the osteogenesis of human pMSCs under hypoxia. Our findings will highlight the therapeutic potential of MSCs in the tissue engineering of bones.

  18. Cellular and Matrix Response of the Mandibular Condylar Cartilage to Botulinum Toxin

    Science.gov (United States)

    Dutra, Eliane H.; O’ Brien, Mara H.; Lima, Alexandro; Kalajzic, Zana; Tadinada, Aditya; Nanda, Ravindra; Yadav, Sumit

    2016-01-01

    Objectives To evaluate the cellular and matrix effects of botulinum toxin type A (Botox) on mandibular condylar cartilage (MCC) and subchondral bone. Materials and Methods Botox (0.3 unit) was injected into the right masseter of 5-week-old transgenic mice (Col10a1-RFPcherry) at day 1. Left side masseter was used as intra-animal control. The following bone labels were intraperitoneally injected: calcein at day 7, alizarin red at day 14 and calcein at day 21. In addition, EdU was injected 48 and 24 hours before sacrifice. Mice were sacrificed 30 days after Botox injection. Experimental and control side mandibles were dissected and examined by x-ray imaging and micro-CT. Subsequently, MCC along with the subchondral bone was sectioned and stained with tartrate resistant acid phosphatase (TRAP), EdU, TUNEL, alkaline phosphatase, toluidine blue and safranin O. In addition, we performed immunohistochemistry for pSMAD and VEGF. Results Bone volume fraction, tissue density and trabecular thickness were significantly decreased on the right side of the subchondral bone and mineralized cartilage (Botox was injected) when compared to the left side. There was no significant difference in the mandibular length and condylar head length; however, the condylar width was significantly decreased after Botox injection. Our histology showed decreased numbers of Col10a1 expressing cells, decreased cell proliferation and increased cell apoptosis in the subchondral bone and mandibular condylar cartilage, decreased TRAP activity and mineralization of Botox injected side cartilage and subchondral bone. Furthermore, we observed reduced proteoglycan and glycosaminoglycan distribution and decreased expression of pSMAD 1/5/8 and VEGF in the MCC of the Botox injected side in comparison to control side. Conclusion Injection of Botox in masseter muscle leads to decreased mineralization and matrix deposition, reduced chondrocyte proliferation and differentiation and increased cell apoptosis in the

  19. Translationally controlled tumor protein supplemented chitosan modified glass ionomer cement promotes osteoblast proliferation and function

    Energy Technology Data Exchange (ETDEWEB)

    Sangsuwan, Jiraporn [Department of Molecular Biology and Bioinformatics, Center for Genomics and Bioinformatics Research, Faculty of Science, Prince of Songkla University, Hat Yai, Songkhla 90112 (Thailand); Department of Oral Biology and Occlusion, Faculty of Dentistry, Prince of Songkla University, Hat Yai, Songkhla 90112 (Thailand); Wanichpakorn, Supreya; Kedjarune-Leggat, Ureporn [Department of Oral Biology and Occlusion, Faculty of Dentistry, Prince of Songkla University, Hat Yai, Songkhla 90112 (Thailand)

    2015-09-01

    The objective of this study was to evaluate the effect of translationally controlled tumor protein (TCTP) supplemented in a novel glass ionomer cement (BIO-GIC) on normal human osteoblasts (NHost cells). BIO-GIC was a glass ionomer cement (GIC) modified by adding chitosan and albumin to promote the release of TCTP. NHost cells were seeded on specimens of GIC, GIC + TCTP, BIO-GIC and BIO-GIC + TCTP. Cell proliferation was determined by BrdU assay. It was found that BIO-GIC + TCTP had significantly higher proliferation of cells than other specimens. Bone morphogenetic protein-2 (BMP-2) and osteopontin (OPN) gene expressions assessed by quantitative real time PCR and alkaline phosphatase (ALP) activity were used to determine cell differentiation. Bone cell function was investigated by calcium deposition using alizarin assay. Both BMP-2 and OPN gene expressions of cells cultured on specimens with added TCTP increased gradually up-regulation after day 1 and reached the highest on day 3 then down-regulation on day 7. The ALP activity of cells cultured on BIO-GIC + TCTP for 7 days and calcium content after 14 days were significantly higher than other groups. BIO-GIC + TCTP can promote osteoblast cells proliferation, differentiation and function. - Highlights: • Developed a new GIC by supplementing TCTP in BIO-GIC (GIC with chitosan and albumin) • BIO-GIC + TCTP released a higher amount of TCTP than GIC + TCTP. • BIO-GIC + TCTP promoted cell proliferation higher than other specimens and control. • BIO-GIC + TCTP promoted osteoblasts differentiation and function.

  20. Silorane resin supports proliferation, differentiation, and mineralization of MLO-A5 bone cells in vitro and bone formation in vivo.

    Science.gov (United States)

    Eick, J David; Barragan-Adjemian, Cielo; Rosser, Jennifer; Melander, Jennifer R; Dusevich, Vladimir; Weiler, Rachel A; Miller, Bradley D; Kilway, Kathleen V; Dallas, Mark R; Bi, Lianxing; Nalvarte, Elisabet L; Bonewald, Lynda F

    2012-04-01

    Methyl methacrylate used in bone cements has drawbacks of toxicity, high exotherm, and considerable shrinkage. A new resin, based on silorane/oxirane chemistry, has been shown to have little toxicity, low exotherm, and low shrinkage. We hypothesized that silorane-based resins may also be useful as components of bone cements as well as other bone applications and began testing on bone cell function in vitro and in vivo. MLO-A5, late osteoblast cells, were exposed to polymerized silorane (SilMix) resin (and a standard polymerized bisGMA/TEGDMA methacrylate (BT) resin and compared to culture wells without resins as control. A significant cytotoxic effect was observed with the BT resin resulting in no cell growth, whereas in contrast, SilMix resin had no toxic effects on MLO-A5 cell proliferation, differentiation, nor mineralization. The cells cultured with SilMix produced increasing amounts of alkaline phosphatase (1.8-fold) compared to control cultures. Compared to control cultures, an actual enhancement of mineralization was observed in the silorane resin-containing cultures at days 10 and 11 as determined by von Kossa (1.8-2.0 fold increase) and Alizarin red staining (1.8-fold increase). A normal bone calcium/phosphate atomic ratio was observed by elemental analysis along with normal collagen formation. When used in vivo to stabilize osteotomies, no inflammatory response was observed, and the bone continued to heal. In conclusion, the silorane resin, SilMix, was shown to not only be non cytototoxic, but actually supported bone cell function. Therefore, this resin has significant potential for the development of a nontoxic bone cement or bone stabilizer.

  1. Bone morphogenetic protein 2 promotes transforming growth factor β3-induced chondrogenesis of human osteoarthritic synovium-derived stem cells

    Institute of Scientific and Technical Information of China (English)

    RUI Yun-feng; DU Lin; WANG You; WANG Yang; LUI Pauline po-yee; TANG Ting-ting; CHAN Kai-ming; DAI Ke-rong

    2010-01-01

    Background Synovium-derived stem cells (SDSCs) with higher chondrogenic potential are attracting considerable attention as a cell source for cartilage regeneration. We investigated the effect of bone morphogenetic protein 2 (BMP-2) on transforming growth factor beta3 (TGF-β3)-induced chondrogenesis of SDSCs isolated from human osteoarthritic synovium in a pellet culture system. Methods The clonogenicity, stem cell marker expression and multi-differentiation potential of isolated SDSCs were determined by colony forming unit assay, flow cytometry and specific staining including alizarin red S, Oil red O and alcian blue staining, respectively. SDSCs pellet was cultured in chondrogenic medium with or without TGF-β3 or/and BMP-2. At day 21, the diameter and the weight of the pellets were measured. Chondrogenic differentiation of SDSCs was evaluated by Safranin O staining, immunohistochemical staining of collagen type Ⅱ, sulfated glycosaminoglycan (sGAG) synthesis and mRNA expression of collagen type Ⅱ, aggrecan, SOX9, link-protein, collagen type X and BMP receptor Ⅱ. Results Cells isolated under the optimized culturing density (104/60 cm2) showed clonogenicity and multi-differentiation potential. These cells were positive (>99%) for CD44, CD90, CD105 and negative (<10%) for CD34 and CD71. SDSCs differentiated to a chondrocytic phenotype in chondrogenic medium containing TGF-β3 with or without BMP-2. Safranin O staining of the extracellular matrix was positive and the expression of collagen type Ⅱ was detected. Cell pellets treated with TGF-β3 and BMP-2 were larger in diameter and weight, produced more sGAGs, and expressed higher levels of collagen type Ⅱ and other chondrogenic markers, except COL10A1, than medium with TGF-β3 alone. Conclusions SDSCs could be isolated from human osteoarthritic synovium. Supplementation with BMP-2 significantly promoted the in vitro TGF-β3-induced chondrogenic differentiation of SDSCs.

  2. Hyperbaric oxygen promotes osteogenic differentiation of bone marrow stromal cells by regulating Wnt3a/β-catenin signaling—An in vitro and in vivo study

    Directory of Open Access Journals (Sweden)

    Song-Shu Lin

    2014-01-01

    Full Text Available We hypothesized that the effect of hyperbaric oxygen (HBO on bone formation is increased via osteogenic differentiation of bone marrow stromal cells (BMSCs, which is regulated by Wnt3a/β-catenin signaling. Our in vitro data showed that HBO increased cell proliferation, Wnt3a production, LRP6 phosphorylation, and cyclin D1 expression in osteogenically differentiated BMSCs. The mRNA and protein levels of Wnt3a, β-catenin, and Runx2 were upregulated while those of GSK-3β were downregulated after HBO treatment. The relative density ratio (phospho-protein/protein of Akt and GSK-3β was both up-regulated while that of β-catenin was down-regulated after HBO treatment. We next investigated whether HBO affects the accumulation of β-catenin. Our Western blot analysis showed increased levels of translocated β-catenin that stimulated the expression of target genes after HBO treatment. HBO increased TCF-dependent transcription, Runx2 promoter/Luc gene activity, and the expression of osteogenic markers of BMSCs, such as alkaline phosphatase activity, type I collagen, osteocalcin, calcium, and the intensity of Alizarin Red staining. HBO dose dependently increased the bone morphogenetic protein (BMP2 and osterix production. We further demonstrated that HBO increased the expression of vacuolar-ATPases, which stimulated Wnt3a secretion from BMSCs. Finally, we showed that the beneficial effects of HBO on bone formation were related to Wnt3a/β-catenin signaling in a rabbit model by histology, mechanical testing, and immunohistochemical assays. Accordingly, we concluded that HBO increased the osteogenic differentiation of BMSCs by regulating Wnt3a secretion and signaling.

  3. Osteogenic cell cultures cannot utilize exogenous sources of synthetic polyphosphate for mineralization.

    Science.gov (United States)

    Ariganello, Marianne B; Omelon, Sidney; Variola, Fabio; Wazen, Rima M; Moffatt, Pierre; Nanci, Antonio

    2014-12-01

    Phosphate is critical for mineralization and deficiencies in the regulation of free phosphate lead to disease. Inorganic polyphosphates (polyPs) may represent a physiological source of phosphate because they can be hydrolyzed by biological phosphatases. To investigate whether exogenous polyP could be utilized for mineral formation, mineralization was evaluated in two osteogenic cell lines, Saos-2 and MC3T3, expressing different levels of tissue non-specific alkaline phosphatase (tnALP). The role of tnALP was further explored by lentiviral-mediated overexpression in MC3T3 cells. When cells were cultured in the presence of three different phosphate sources, there was a strong mineralization response with β-glycerophosphate (βGP) and orthophosphate (Pi) but none of the cultures sustained mineralization in the presence of polyP (neither chain length 17-Pi nor 42-Pi). Even in the presence of mineralizing levels of phosphate, low concentrations of polyP (50 μM) were sufficient to inhibit mineral formation. Energy-dispersive X-ray spectroscopy confirmed the presence of apatite-like mineral deposits in MC3T3 cultures supplemented with βGP, but not in those with polyP. While von Kossa staining was consistent with the presence or absence of mineral, an unusual Alizarin staining was obtained in polyP-treated MC3T3 cultures. This staining pattern combined with low Ca:P ratios suggests the persistence of Ca-polyP complexes, even with high residual ALP activity. In conclusion, under standard culture conditions, exogenous polyP does not promote mineral deposition. This is not due to a lack of active ALP, and unless conditions that favor significant processing of polyP are achieved, its mineral inhibitory capacity predominates.

  4. Cytotoxicity and Osteogenic Potential of Silicate Calcium Cements as Potential Protective Materials for Pulpal Revascularization

    Science.gov (United States)

    Bortoluzzi, Eduardo A.; Niu, Li-na; Palani, Chithra D.; El-Awady, Ahmed R.; Hammond, Barry D.; Pei, Dan-dan; Tian, Fu-cong; Cutler, Christopher W.; Pashley, David H.; Tay, Franklin R.

    2016-01-01

    Objectives In pulpal revascularization, a protective material is placed coronal to the blood clot to prevent recontamination and to facilitate osteogenic differentiation of mesenchynal stem cells to produce new dental tissues. Although mineral trioxide aggregate (MTA) has been the material of choice for clot protection, it is easily displaced into the clot during condensation. The present study evaluated the effects of recently-introduced calcium silicate cements (Biodentine and TheraCal LC) on the viability and osteogenic differentiation of human dental pulp stem cells (hDPSCs) by comparing with MTA Angelus. Methods Cell viability was assessed using XTT assay and flow cytometry. The osteogenic potential of hDPSCs exposed to calcium silicate cements was examined using qRT-PCR for osteogeic gene expressions, alkaline phosphatase enzyme activity, Alizarin red S staining and transmission electron microscopy of extracellular calcium deposits. Parametric statistical methods were employed for analyses of significant difference among groups, with α=0.05. Results The cytotoxic effects of Biodentine and TheraCal LC on hDPSCs were time- and concentration-dependent. Osteogenic differentiation of hDPSCs was enhanced after exposure to Biodentine that was depleted of its cytotoxic components. This effect was less readily observed in hDPSCs exposed to TheraCal LC, although both cements supported extracelluar mineralization better than the positive control (zinc oxide-eugenol–based cement). Significance A favorable tissue response is anticipated to occur with the use of Biodentine as a blood clot-protecting material for pulpal revascularizaiton. Further investigations with the use of in vivo animal models are required to validate the potential adverse biological effects of TheraCal LC on hDPSCs. PMID:26494267

  5. Activation of the FGF signaling pathway and subsequent induction of mesenchymal stem cell differentiation by inorganic polyphosphate

    Directory of Open Access Journals (Sweden)

    Yumi Kawazoe, Shinichi Katoh, Yuichiro Onodera, Takao Kohgo, Masanobu Shindoh, Toshikazu Shiba

    2008-01-01

    Full Text Available Inorganic polyphosphate [poly(P] is a biopolymer existing in almost all cells and tissues, although its biological functions in higher eukaryotes have not been completely elucidated. We previously demonstrated that poly(P enhances the function of fibroblast growth factors (FGFs by stabilizing them and strengthening the affinity between FGFs and their cell surface receptors. Since FGFs play crucial roles in bone regeneration, we further investigated the effect of poly(P on the cell differentiation of human stem cells via FGF signaling systems. Human dental pulp cells (HDPCs isolated from human dental pulp show the characteristics of multipotent mesenchymal stem cells (MSCs. HDPCs secreted FGFs and the proliferation of HDPCs was shown to be enhanced by treatment with poly(P. Cell surface receptor-bound FGF-2 was stably maintained for more than 40 hours in the presence of poly(P. The phosphorylation of ERK1/2 was also enhanced by poly(P. The effect of poly(P on the osteogenic differentiation of HDPCs and human MSCs (hMSCs were also investigated. After 5 days of treatment with poly(P, type-I collagen expression of both cell types was enhanced. The C-terminal peptide of type-I collagen was also released at higher levels in poly(P-treated HDPCs. Microarray analysis showed that expression of matrix metalloproteinase-1 (MMP1, osteopontin (OPN, osteocalcin (OC and osteoprotegerin was induced in both cell types by poly(P. Furthermore, induced expression of MMP1, OPN and OC genes in both cells was confirmed by real-time PCR. Calcification of both cell types was clearly observed by alizarin red staining following treatment with poly(P. The results suggest that the activation of the FGF signaling pathway by poly(P induces both proliferation and mineralization of stem cells.

  6. Matrix stiffness regulation of integrin-mediated mechanotransduction during osteogenic differentiation of human mesenchymal stem cells.

    Science.gov (United States)

    Shih, Yu-Ru V; Tseng, Kuo-Fung; Lai, Hsiu-Yu; Lin, Chi-Hung; Lee, Oscar K

    2011-04-01

    Mesenchymal stem cells (MSCs) cultured on extracellular matrices with different stiffness have been shown to possess diverse lineage commitment owing to the extracellular mechanical stimuli sensed by the cells. The aim of this study was to further delineate how matrix stiffness affects intracellular signaling through the mechanotransducers Rho kinase (ROCK) and focal adhesion kinase (FAK) and subsequently regulates the osteogenic phenotype of MSCs. MSCs were cultured in osteogenic medium on tunable polyacrylamide hydrogels coated with type I collagen with elasticities corresponding to Young's modulus of 7.0 ± 1.2 and 42.1 ± 3.2 kPa. Osteogenic differentiation was increased on stiffer matrices, as evident by type I collagen, osteocalcin, and Runx2 gene expressions and alizarin red S staining for mineralization. Western blot analysis demonstrated an increase in kinase activities of ROCK, FAK, and ERK1/2 on stiffer matrices. Inhibition of FAK, an important mediator of osteogenic differentiation, and inhibition of ROCK, a known mechanotransducer of matrix stiffness during osteogenesis, resulted in decreased expression of osteogenic markers during osteogenic induction. In addition, FAK affects osteogenic differentiation through ERK1/2, whereas ROCK regulates both FAK and ERK1/2. Furthermore, α(2)-integrin was upregulated on stiffer matrices during osteogenic induction, and its knockdown by siRNA downregulated the osteogenic phenotype through ROCK, FAK, and ERK1/2. Taken together, our results provide evidence that the matrix rigidity affects the osteogenic outcome of MSCs through mechanotransduction events that are mediated by α(2)-integrin.

  7. Modulation of the Differentiation of Dental Pulp Stem Cells by Different Concentrations of β-Glycerophosphate

    Directory of Open Access Journals (Sweden)

    Weiping Hu

    2012-01-01

    Full Text Available Dentinogenesis is a necessary prerequisite for dental tissue engineering. One of the steps for dentinogenesis is to obtain large quantities of highly purified odontoblasts. Therefore, we have undertaken an experiment applying different concentrations of β-glycerophosphate (β-GP to induce the differentiation of dental pulp stem cells (DPSCs in a long-term 28-day culture. In the meanwhile, we have studied the time- and maturation-dependent expression of matrix extracellular phosphoglycoprotein (MEPE and that of the odontoblast-like marker-dentin sialoprotein (DSP, in order to investigate an optimized mineralized condition. Western blot results revealed that the expression of DSP became lower when accompanied by the increase of the β-GP concentration, and there was also an influence on MEPE expression when different concentrations of β-GP were applied. Meanwhile, the mineralized groups had an inhibitory function on the expression of MEPE as compared with the control group. Above all, all experimental groups successfully generated mineralized nodules by Alizarin Red S and the 5 mM β-GP group formed more mineralized nodules quantitated using the CPC extraction method. In conclusion, there is a significant modulation of the β-GP during the differentiation of the DPSCs. The degree of odontoblast differentiation is β-glycerophosphate concentration dependent. A low concentration of β-GP (5 mM has been shown to be the optimal concentration for stimulating the maturation of the DPSCs. Moreover, MEPE accompanied with DSP clearly demonstrates the degree of the differentiation.

  8. Housekeeping gene stability influences the quantification of osteogenic markers during stem cell differentiation to the osteogenic lineage.

    Science.gov (United States)

    Quiroz, Felipe Garcia; Posada, Olga M; Gallego-Perez, Daniel; Higuita-Castro, Natalia; Sarassa, Carlos; Hansford, Derek J; Agudelo-Florez, Piedad; López, Luis E

    2010-04-01

    Real-time reverse transcription PCR (RT-qPCR) relies on a housekeeping or normalizer gene whose expression remains constant throughout the experiment. RT-qPCR is commonly used for characterization of human bone marrow mesenchymal stem cells (hBMSCs). However, to the best of our knowledge, there are no studies validating the expression stability of the genes used as normalizers during hBMSCs differentiation. This work aimed to study the stability of the housekeeping genes beta-actin, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and ribosomal protein L13A (RPL13A) during the osteogenic differentiation of hBMSCs. Their stability was evaluated via RT-qPCR in 14 and 20 day differentiation assays to the osteogenic lineage. Different normalization strategies were evaluated to quantify the osteogenic markers collagen type I, bone sialoprotein and osteonectin. Cell differentiation was confirmed via alizarin red staining. The results demonstrated up-regulation of beta-actin with maximum fold changes (MFC) of 4.38. GAPDH and RPL13A were not regulated by osteogenic media after 14 days and presented average fold changes lower than 2 in 20 day cultures. RPL13A (MFC < 2) had a greater stability when normalizing as a function of culture time compared with GAPDH (MFC

  9. Growth Kinetics, Characterization, and Plasticity of Human Menstrual Blood Stem Cells

    Directory of Open Access Journals (Sweden)

    Davood Mehrabani

    2016-03-01

    Full Text Available One of the readily available sources of mesenchymal stem cells (MSCs is menstrual blood-derived stem cells (Men-SCs, which exhibit characteristics similar to other types of MSCs. This study was performed to determine the growth kinetics, plasticity, and characterization of Men-SCs in women. During spring 2014 in the southern Iranian city of Shiraz, menstrual blood (5 mL was obtained from 10 women on their third day of menstruation in 2 age groups of 30 to 40 and 40 to 50 years old. Ficoll was used to separate the mononuclear cell fraction. After the Men-SCs were cultured, they were subcultured up to passage 4. Growth behavior and population doubling time were evaluated by seeding 5×104 cells into 12- and 24-well culture plates, and the colonies were enumerated. The expression of CD44, CD90, and CD34 was evaluated. The osteogenic potential was assessed by alizarin red staining. The Men-SCs were shown to be plastic adherent and spindle-shaped. Regarding the growth curves in the 12- and 24-well culture plates, it was demonstrated that in the women aged between 30 and 40 years, population doubling time was 55.5 and 62 hours, respectively, while these values in the women aged between 40 and 50 years were 70.4 and 72.4 hours, correspondingly. Positive expression of CD44 and CD90 and negative expression of CD34 were noted. In the osteogenic differentiation medium, the cells differentiated toward osteoblasts. As human Men-SCs are easily collectable without any invasive procedure and are a safe and rapid source of MSCs, they can be a good candidate for stem cell banking and cell transplantation in women.

  10. Effect of Hypergravity on Carbonanhydrase Reactivity in inner Ear Ioncytes of developing Cichlid Fish

    Science.gov (United States)

    Beier, M.; Anken, R.; Rahmann, H.

    It has been shown earlier that hypergravity slows down inner ear otolith growth in developing fish. Otolith growth in terms of mineralisation mainly depends on the enzyme carboanhydrase (CAH), which is responsible for the provision of the pH- value necessary for calcium carbonate deposition and thus also is presumed to play a prominent role in Ménière's disease (a sensory - motor disorder inducing vertigo and kinetosis). Larval siblings of cichlid fish (Oreochromis mossambicus) were subjected to hypergravity (3g; 6 hours) during development and separated into normally and kinetotically swimming individuals following the transfer to 1g (i.e., stopping the centrifuge; kinetotically behaving fish performed spinning movements). Subsequently, CAH was histochemically demonstrated in inner ear ionocytes (cells involved in the endolymphatic ion exchange) and enzyme reactivity was determined densitometrically. The results showed that CAH-reactivity was significantly increased in normally behaving hyper-g specimens as compared to controls kept at 1g, whereas no difference in enzyme reactivity was evident between the controls and kinetotically behaving fish. On the background of earlier studies, according to which (1) hypergravity induces a decrease of otolith growth and (2) the otolithic calcium incorporation (visualized using the calcium -tracer alizarin complexone) of kinetotically swimming hyper - g fish was lower as compared to normally behaving hyper - g animals, the present study strongly supports the concept that an increase in CAH-reactivity may result in a decrease of otolithic calcium deposition. The mechanism regulating CAH-activity hitherto remains to be determined. Acknowledgement: This work was financially supported by the German Aerospace Center (DLR) (FKZ: 50 WB 9997).

  11. Romanian Words of Arabic Origin: Scientific and Technical Vocabulary

    Directory of Open Access Journals (Sweden)

    Georgeta Rata

    2016-10-01

    Full Text Available There are 141 Romanian words of Arabic origin acquired either directly from Arabic or else indirectly by passing from Arabic into other languages and then into Romanian. Most entered one or more of the Romance languages before entering Romanian. To qualify for this list, a word must be reported in etymology dictionaries as having descended from Arabic. Words associated with the Islamic religion are omitted. Archaic and rare words are also omitted. Given the nature of the journal in which the paper is to be published, the author selected for analysis only about 126 terms belonging to the scientific and technical vocabulary: Adobe, alambic, albatros, alcalin, alchimie, alcool, alfalfa, algebră, algoritm, alidadă, alizarină, amalgam, ambră, anil, antimoniu, azimuth, azur, benjoin, bezoar, bor, cafea, calibre, camfor, carat, carciofoi, caric, cârmâz, carob, chimie, cifru, coton, curcuma, cuşcuş, erg, falafel, fanfară, felucă, fenec, gazelă, gerbil, girafă, halva, hamada, humus, iasomie, jar, julep, kaliu, lac, lămâie, lazurit, liliac, lime, marcasit, masicot, mizenă, muson, nadir, natriu, papagal, rachetă, realgar, sabkha, safari, şah, sandarac, şaorma, şerbet, sirop, sodium, şofran, sorbet, spanac, sumac, tabac, tahân, taifun, talc, tamarin(d, tangerină, tar, tară, tarhon, tarif, tasă, ţechin, ton, varan, zahăr, zenith, zero, zircon, etc. Some of them are obsolescent, but a large number are in everyday use and have been so well assimilated into Romanian that they have produced other words through derivation and composition, or they have acquired new meanings.

  12. The effect of magnetic stimulation on the osteogenic and chondrogenic differentiation of human stem cells derived from the adipose tissue (hASCs)

    Energy Technology Data Exchange (ETDEWEB)

    Lima, João; Gonçalves, Ana I.; Rodrigues, Márcia T.; Reis, Rui L. [3Bs Research Group–Biomaterials, Biodegradables and Biomimetics, University of Minho, Guimarães (Portugal); ICVS/3Bs–PT Government Associate Laboratory, Braga/Guimarães (Portugal); Gomes, Manuela E., E-mail: megomes@dep.uminho.pt [3Bs Research Group–Biomaterials, Biodegradables and Biomimetics, University of Minho, Guimarães (Portugal); ICVS/3Bs–PT Government Associate Laboratory, Braga/Guimarães (Portugal)

    2015-11-01

    The use of magnetic nanoparticles (MNPs) towards the musculoskeletal tissues has been the focus of many studies, regarding MNPs ability to promote and direct cellular stimulation and orient tissue responses. This is thought to be mainly achieved by mechano-responsive pathways, which can induce changes in cell behavior, including the processes of proliferation and differentiation, in response to external mechanical stimuli. Thus, the application of MNP-based strategies in tissue engineering may hold potential to propose novel solutions for cell therapy on bone and cartilage strategies to accomplish tissue regeneration. The present work aims at studying the influence of MNPs on the osteogenic and chondrogenic differentiation of human adipose derived stem cells (hASCs). MNPs were incorporated in hASCs and cultured in medium supplemented for osteogenic and chondrogenic differentiation. Cultures were maintained up to 28 days with/without an external magnetic stimulus provided by a magnetic bioreactor, to determine if the MNPs alone could affect the osteogenic or chondrogenic phenotype of the hASCs. Results indicate that the incorporation of MNPs does not negatively affect the viability nor the proliferation of hASCs. Furthermore, Alizarin Red staining evidences an enhancement in extracellular (ECM) mineralization under the influence of an external magnetic field. Although not as evident as for osteogenic differentiation, Toluidine blue and Safranin-O stainings also suggest the presence of a cartilage-like ECM with glycosaminoglycans and proteoglycans under the magnetic stimulus provided. Thus, MNPs incorporated in hASCs under the influence of an external magnetic field have the potential to induce differentiation towards the osteogenic and chondrogenic lineages. - Highlights: • Cellular viability was not negatively influenced by the nanoparticles. • Chondrogenic medium influences more the synthesis of cartilage-like ECM than MNPs. • Synergetic effect among

  13. Preparation and characterization of polylactide/poly(ε-caprolactone-poly(ethylene glycol-poly(ε-caprolactone hybrid fibers for potential application in bone tissue engineering

    Directory of Open Access Journals (Sweden)

    Wang YL

    2014-04-01

    Full Text Available YueLong Wang,1,2,* Gang Guo,1,* HaiFeng Chen,2 Xiang Gao,1 RangRang Fan,1 DongMei Zhang,1 LiangXue Zhou2 1State Key Laboratory of Biotherapy and Cancer Center, 2Department of Neurosurgery, West China Hospital, West China Medical School, Sichuan University, Chengdu, People's Republic of China *These authors contributed equally to this paper Abstract: The aim of this study was to develop a kind of osteogenic biodegradable composite graft consisting of human placenta-derived mesenchymal stem cell (hPMSC material for site-specific repair of bone defects and attenuation of clinical symptoms. The novel nano- to micro-structured biodegradable hybrid fibers were prepared by electrospinning. The characteristics of the hybrid membranes were investigated by a range of methods, including Fourier transform infrared spectroscopy, X-ray diffraction, and differential scanning calorimetry. Morphological study with scanning electron microscopy showed that the average fiber diameter and the number of nanoscale pores on each individual fiber surface decreased with increasing concentration of poly(ε-caprolactone-poly(ethylene glycol-poly(ε-caprolactone (PCEC. The prepared polylactide (PLA/PCEC fibrous membranes favored hPMSC attachment and proliferation by providing an interconnected, porous, three-dimensional mimicked extracellular environment. What is more, hPMSCs cultured on the electrospun hybrid PLA/PCEC fibrous scaffolds could be effectively differentiated into bone-associated cells by positive alizarin red staining. Given the good cellular response and excellent osteogenic potential in vitro, the electrospun PLA/PCEC fibrous scaffolds could be one of the most promising candidates for bone tissue engineering. Keywords: electrospinning, PLA, PCEC, hPMSCs, bone tissue engineering

  14. Antibacterial properties of essential oils and methanol extracts of sweet basil Ocimum basilicum occurring in Bangladesh.

    Science.gov (United States)

    Hossain, M Amzad; Kabir, M J; Salehuddin, S M; Rahman, S M Mizanur; Das, A K; Singha, Sandip Kumar; Alam, Md Khorshed; Rahman, Atiqur

    2010-05-01

    The antibacterial potential of essential oils and methanol extracts of sweet basil Ocimum basilicum L. (Lamiaceae) was evaluated for controlling the growth range of food-borne pathogenic bacteria. Essential oils extracted by hydrodistillation from the leaves and stems were analyzed by GC-MS. Fifty-seven compounds representing 94.9 and 96.1% of the total leaf and stem oils, respectively, were identified, of which methyl chavicol (36.7 and 29.9%), gitoxigenin (9.3 and 10.2%), trimethoquinol (10.3 and 8.4%), beta-guaiene (3.7 and 4.1%), aciphyllene (3.4 and 3.0%), alizarin (3.2 and 4.4%), naphthaline (2.2 and 3.8%), (-)-caryophyllene (2.0 and 1.9%), and mequinol (1.6 and 1.8%) were the major compounds. The essential oils (10 microL/disc of 1:5, v/v dilution with methanol) and methanol extracts (300 microg/disc) of O. basilicum displayed a great potential of antibacterial activity against Bacillius cereus, B. subtilis, B. megaterium, Staphylococcus aureus, Listeria monocytogenes, Escherichia coli, Shigella boydii, S. dysenteriae, Vibrio parahaemolyticus, V. mimicus, and Salmonella typhi with their respective zones of inhibition of 11.2-21.1 mm and MIC values of 62.5-500 microg/mL. The results of this study suggest that the natural products derived from O. basilicum may have potential use in the food and/or pharmaceutical industries as antimicrobial agents.

  15. Surgical anatomy of the innervation of pylorus in human and Suncus murinus, in relation to surgical technique for pylorus- preserving pancreaticoduodenectomy

    Institute of Scientific and Technical Information of China (English)

    Shuang-Qin Yi; Shigenori Tanaka; Masahiro Itoh; Fei Ru; Tetsuo Ohta; Hayato Terayama; Munekazu Naito; Shogo Hayashi; Sichen Buhe; Nozomi Yi; Takayoshi Miyaki

    2006-01-01

    AIM: To clarify the innervation of the antro-pyloric region in humans from a dinico-anatomical perspective.METHODS: The stomach, duodenum and surrounding structures were dissected in 10 cadavers, and immersed in a 10mg/L solution of alizarin red S in ethanol to stain the peripheral nerves. The distribution details were studied to confirm innervations in the above areas using a binocular microscope. Similarly, innervations in 10Suncus murinus were examined using the method of whole-mount im munohistochemistry.RESULTS: The innervation of the pyloric region in humans involved three routes: One arose from the anterior hepatic plexus via the route of the suprapyloric/supraduodenal branch of the right gastric artery; the second arose from the anterior and posterior gastric divisions, and the third originated from the posteriorlower region of the pyloric region, which passed via the infrapyloric artery or retroduodenal branches and was related to the gastroduodenal artery and right gastroepiploic artery. For Suncus murinus, results similar to those in humans were observed.CONCLUSION: There are three routes of innervation of the pyloric region in humans, wherein the route of the right gastric artery is most important for preserving pyloric region innervation. Function will be preserved by more than 80% by preserving the artery in pyloruspreserving pancreaticoduodenectomy (PPPD). However,the route of the infrapyloric artery should not be disregarded. This route is related to several arteries (the right gastroepiploic and gastroduodenal arteries),and the preserving of these arteries is advantageous for preserving pyloric innervation in PPPD. Concurrently,the nerves of Latarjet also play an important role in maintaining innervation of the antro-pyloric region in PPPD. This is why pyloric function is not damaged in some patients when the right gastric artery is dissected or damaged in PPPD.

  16. Modulation of Isoflavones on Bone-nodule Formation in Rat Calvaria Osteoblasts in vitro

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    Objective To observe the effects of two main isoflavones, daidzein and genistein on the bone-nodule formation in rat calvaria osteoblasts in vitro. Methods Osteoblasts obtained from newborn Sprague-dawley rat calvarias were cultured for several generations. The second generation cells were cultured in Minimum Essential Medium supplemented with ascorbic acid and Na-beta-glycerophosphate for several days, in the presence of daidzein and genistein, with or without the estrogen receptor antagonist ICI 182780. Number of nodules was counted at the end of the incubation period (day 20) by staining with Alizarin Red S calcium stain. The release of osteocalcin, as a marker of osteoblast activity, was also determined on day 7 and day 12 during the incubation period. Results Compared with the control, the numbers of nodules were both increased by incubation with daidzein and genistein. 17a-estradiol was used as a positive control and proved to be a more effective inducer of the increase in bone-nodules formation than daidzein and genistein. The release of osteocalcin into culture media was also increased in the presence of daidzein and genistein, as well as 17a-estradiol on day 7 and day 12 (day 12 were higher). The estrogen receptor antagonist ICI 182780 completely blocked the genistein- and 17a-estradiol-induced increase of nodule numbers and osteocalcin release in osteoblasts. However, the effects induced by daidzein could not be inhibited by ICI 182780. Conclusion These findings suggest that geinistein can stimulate bone-nodule formation and increase the release of osteocalcin in rat osteoblasts. The effects, like those induced by 17a-estradiol, are mediated by the estrogen receptor dependent pathway. Daidzein also can stimulate bone-nodule formation and increase the release of osteocalcin in rat osteoblasts, but it is not, at least not merely, mediated by the estrogen receptor dependent pathway.

  17. Osteogenic-related gene expression profiles of human dental follicle cells induced by dexamethasone

    Institute of Scientific and Technical Information of China (English)

    Zuo-lin JIN; Yong-kuan ZHANG; Hai-yan SUN; Zhu LIN; Ying-chun BI; Yin-zhong DUAN; Yin DING

    2008-01-01

    Aim:Human dental follicle cells (hDFC) have the ability to differentiate into mineralized tissue-forming cells during root and periodontal development or os-teogenic induction in vitro. The present study aimed to validate the osteogenic induction of hDFC by dexamethasone (DEX) and to explore the changes of related genes responsible for the osteogenic differentiation process. Methods: Passage-cultured hDFC were induced by DEX and analyzed for mineralization activity by morphological observation, alkaline phosphatase (ALP) activity, and alizarin red S staining. GEArray Q series human osteogenesis gene array was used to describe large-scale gene expression in treated hDFC compared to the control group. Quantitative real-time RT-PCR was performed to confirm the microarray data by analyzing the expression of 7 critical transcripts. Results: Osteogenic differentiation of hDFC was confirmed by morphological change, elevated ALP activity and calcified nodules. In 96 genes investigated through the microarray analysis, 20 genes were upregulated and 8 genes were downregn-lated more than 2-fold. The results of the real-time RT-PCR correlated with the microarray analysis. The expression of the transforming growth factor-β superfamily showed varying degrees of increase, and fibroblast growth factors exhibited a differential changing trend of expression. The expression of most types of collagen genes representative of extracellular matrixes increased under DEX treatment while small mothers against decapentaplegic 6 and 7 expressions significantly decreased. Conclusion: Our results demonstrated that hDFC dis-played osteoblastic features in both phenotypic and genotypic traits induced by DEX in vitro.

  18. miRNA expression profile during fluid shear stress-induced osteogenic differentiation in MC3T3-E1 cells

    Institute of Scientific and Technical Information of China (English)

    MAI Zhi-hui; PENG Zhu-li; ZHANG Jing-lan; CHEN Lin; LIANG Huan-you; CAI Bin; AI Hong

    2013-01-01

    Background Mechanical stress plays an important role in the maintenance of bone homeostasis.Current hypotheses suggest that interstitial fluid flow is an important component of the system by which tissue level strains are amplified in bone.This study aimed to test the hypothesis that the short-term and appropriate fluid shear stress (FSS) is expected to promote the terminal differentiation of pre-osteoblasts and detect the expression profile of microRNAs in the FSS-induced osteogenic differentiation in MC3T3-E1 cells.Methods MC3T3-E1 cells were subjected to 1 hour of FSS at 12 dyn/cm2 using a parallel plate flow system.After FSS treatment,cytoskeleton immunohistochemical staining and microRNAs (miRNAs) were detected immediately.Osteogenic gene expression and immunohistochemical staining for collagen type Ⅰ were tested at the 24th hour after treatment,alkaline phosphatase (ALP) activity assay was performed at 24th,48th,and 72th hours after FSS treatment,and Alizarin Red Staining was checked at day 12.Results One hour of FSS at 12 dyn/cm2 induced actin stress fiber formation and rearrangement,up-regulated osteogenic gene expression,increased ALP activity,promoted synthesis and secretion of type Ⅰ collagen,enhanced nodule formation,and promoted terminal differentiation in MC3T3-E1 cells.During osteogenic differentiation,expression levels of miR-20a,-21,-19b,-34a,-34c,-140,and-200b in FSS-induced cells were significantly down-regulated.Conclusion The short-term and appropriate FSS is sufficient to promote terminal differentiation of pre-osteoblasts and a group of miRNAs may be invovled in FSS-induced pre-osteoblast differentiation.

  19. Polyetheretherketone Hybrid Composites with Bioactive Nanohydroxyapatite and Multiwalled Carbon Nanotube Fillers

    Directory of Open Access Journals (Sweden)

    Chen Liu

    2016-12-01

    Full Text Available Polyetheretherketone (PEEK hybrid composites reinforced with inorganic nanohydroxyapatite (nHA and multiwalled carbon nanotube (MWNT were prepared by melt-compounding and injection molding processes. The additions of nHA and MWNT to PEEK were aimed to increase its elastic modulus, tensile strength, and biocompatibility, rendering the hybrids suitable for load-bearing implant applications. The structural behavior, mechanical property, wettability, osteoblastic cell adhesion, proliferation, differentiation, and mineralization of the PEEK/nHA-MWNT hybrids were studied. X-ray diffraction and SEM observation showed that both nHA and MWNT fillers are incorporated into the polymer matrix of PEEK-based hybrids. Tensile tests indicated that the elastic modulus of PEEK can be increased from 3.87 to 7.13 GPa by adding 15 vol % nHA and 1.88 vol % MWNT fillers. The tensile strength and elongation at break of the PEEK/(15% nHA-(1.88% MWNT hybrid were 64.48 MPa and 1.74%, respectively. Thus the tensile properties of this hybrid were superior to those of human cortical bones. Water contact angle measurements revealed that the PEEK/(15% nHA-(1.88% MWNT hybrid is hydrophilic due to the presence of nHA. Accordingly, hydrophilic PEEK/(15% nHA-(1.88% MWNT hybrid promoted the adhesion, proliferation, differentiation, and mineralization of murine MC3T3-E1 osteoblasts on its surface effectively on the basis of cell culture, fluorescence microscopy, MTT assay, WST-1 assay, alkaline phosphatase activity, and Alizarin red staining tests. Thus the PEEK/(15% nHA-(1.88% MWNT hybrid has the potential to be used for fabricating load-bearing bone implants.

  20. Isolation of adipose and bone marrow mesenchymal stem cells using CD29 and CD90 modifies their capacity for osteogenic and adipogenic differentiation

    Directory of Open Access Journals (Sweden)

    Owen G Davies

    2015-06-01

    Full Text Available Mesenchymal stem cells isolated from rats are frequently used for tissue engineering research. However, considerable differences have been identified between rat mesenchymal stem cells and those derived from humans, and no defined panel of markers currently exists for the isolation of these cells. The aim of this study was to examine the effects of cell sorting for CD29+/CD90+ cells from rat adipose and bone marrow tissues on their differentiation and expression of stem cell–associated genes. Flow cytometry showed 66% and 78% CD29+/CD90+ positivity within passage 1 of adipose and bone marrow cultures, respectively. CD29+/CD90+ cells showed a reduction in both osteogenic and adipogenic differentiation when compared with unsorted cells, as determined by alizarin red and Oil Red-O staining, respectively. These findings could not entirely be explained by fluorescence-activated cell sorting–induced cell injury as sort recovery was only modestly affected in adipose-derived cells. Maintaining cells in fluorescence-activated cell sorting buffer did not affect adipose-derived cell viability, but a significant (p < 0.05 reduction was found in bone marrow–derived cell viability. Additionally, CD29+/CD90+ selection was associated with a significant decrease in the expression of Lin28, Sox2, Nanog and CD73 in adipose-derived cell cultures, whereas differences in stem cell–associated gene expression were not observed in sorted bone marrow–derived cell cultures. In summary, this study demonstrated that fluorescence-activated cell sorting had differential effects on adipose-derived cells and bone marrow–derived cells, and both CD29+/CD90+ cells displayed a significantly reduced capacity for osteogenic/adipogenic differentiation. In conclusion, we identify that maintaining heterogeneity within the mesenchymal stem cell population may be important for optimal differentiation.

  1. Anti-cytomegalovirus activity of the anthraquinone atanyl blue PRL.

    Science.gov (United States)

    Alam, Zohaib; Al-Mahdi, Zainab; Zhu, Yali; McKee, Zachary; Parris, Deborah S; Parikh, Hardik I; Kellogg, Glen E; Kuchta, Alison; McVoy, Michael A

    2015-02-01

    Human cytomegalovirus (CMV) causes significant disease in immunocompromised patients and serious birth defects if acquired in utero. Available CMV antivirals target the viral DNA polymerase, have significant toxicities, and suffer from resistance. New drugs targeting different pathways would be beneficial. The anthraquinone emodin is proposed to inhibit herpes simplex virus by blocking the viral nuclease. Emodin and related anthraquinones are also reported to inhibit CMV. In the present study, emodin reduced CMV infectious yield with an EC50 of 4.9μM but was cytotoxic at concentrations only twofold higher. Related anthraquinones acid blue 40 and alizarin violet R inhibited CMV at only high concentrations (238-265μM) that were also cytotoxic. However, atanyl blue PRL inhibited infectious yield of CMV with an EC50 of 6.3μM, significantly below its 50% cytotoxic concentration of 216μM. Atanyl blue PRL reduced CMV infectivity and inhibited spread. When added up to 1h after infection, it dramatically reduced CMV immediate early protein expression and blocked viral DNA synthesis. However, it had no antiviral activity when added 24h after infection. Interestingly, atanyl blue PRL inhibited nuclease activities of purified CMV UL98 protein with IC50 of 4.5 and 9.3μM. These results indicate that atanyl blue PRL targets very early post-entry events in CMV replication and suggest it may act through inhibition of UL98, making it a novel CMV inhibitor. This compound may provide valuable insights into molecular events that occur at the earliest times post-infection and serve as a lead structure for antiviral development.

  2. Alpha-adrenergic blocker mediated osteoblastic stem cell differentiation

    Energy Technology Data Exchange (ETDEWEB)

    Choi, Yoon Jung [Craniomaxillofacial Reconstructive Sciences Major, College of Dentistry, Seoul National University, Seoul 110-749 (Korea, Republic of); Lee, Jue Yeon [Craniomaxillofacial Reconstructive Sciences Major, College of Dentistry, Seoul National University, Seoul 110-749 (Korea, Republic of); Research Center, Nano Intelligent Biomedical Engineering Corporation (NIBEC), Seoul (Korea, Republic of); Lee, Seung Jin [Department of Industrial Pharmacy, College of Pharmacy, Ewha Womans University, Seoul (Korea, Republic of); Research Center, Nano Intelligent Biomedical Engineering Corporation (NIBEC), Seoul (Korea, Republic of); Chung, Chong-Pyoung [Department of Periodontology, School of Dentistry, Seoul National University, Seoul (Korea, Republic of); Research Center, Nano Intelligent Biomedical Engineering Corporation (NIBEC), Seoul (Korea, Republic of); Park, Yoon Jeong, E-mail: parkyj@snu.ac.kr [Craniomaxillofacial Reconstructive Sciences Major, College of Dentistry, Seoul National University, Seoul 110-749 (Korea, Republic of); Research Center, Nano Intelligent Biomedical Engineering Corporation (NIBEC), Seoul (Korea, Republic of)

    2011-12-16

    Highlights: Black-Right-Pointing-Pointer Doxazocin directly up-regulated bone metabolism at a low dose. Black-Right-Pointing-Pointer Doxazocin induced osteoblastic stem cell differentiation without affecting cell proliferation. Black-Right-Pointing-Pointer This osteogenic stem cell differentiation is mediated by ERK-signal dependent pathway. -- Abstract: Recent researches have indicated a role for antihypertensive drugs including alpha- or beta-blockers in the prevention of bone loss. Some epidemiological studies reported the protective effects of those agents on fracture risk. However, there is limited information on the association with those agents especially at the mechanism of action. In the present study, we investigated the effects of doxazosin, an alpha-blocker that is clinically used for the treatment of benign prostatic hyperplasia (BPH) along with antihypertensive medication, on the osteogenic stem cell differentiation. We found that doxazosin increased osteogenic differentiation of human mesenchymal stem cells, detected by Alizarin red S staining and calcein. Doxazosin not only induced expression of alkaline phosphatase, type I collagen, osteopontin, and osteocalcin, it also resulted in increased phosphorylation of extracellular signal-regulated kinase (ERK1/2), a MAP kinase involved in osteoblastic differentiation. Treatment with U0126, a MAP kinase inhibitor, significantly blocked doxazosin-induced osteoblastic differentiation. Unrelated to activation of osteogenic differentiation by doxazosin, we found that there were no significant changes in adipogenic differentiation or in the expression of adipose-specific genes, including peroxisome proliferator-activated receptor {gamma}, aP2, or LPL. In this report, we suggest that doxazosin has the ability to increase osteogenic cell differentiation via ERK1/2 activation in osteogenic differentiation of adult stem cells, which supports the protective effects of antihypertensive drug on fracture risk and

  3. Prophylactic Effects of Melatonin on Sodium Valproate-Induced Neural Tube Defects and Skeletal Malformations in Rat Embryos

    Directory of Open Access Journals (Sweden)

    Omolbanin Rahgazar

    2011-01-01

    Full Text Available Problem statement: Some reports showed the teratogenic effects of sodium valproate can be prevented by application of antioxidant drugs and stimulation of the maternal immune system. Therefore, in this study, the prophylactic effect of melatonin on teratogenic effects of sodium valproate was compared. Approach: This study was performed on 31 pregnant rats that were divided into five groups. Control group received normal saline and test groups received sodium valproate (300 mg kg−1, sodium valproate (300 mg kg−1 plus melatonin (5 mg kg−1 and sodium valproate (300 mg kg−1 plus melatonin (10 mg kg−1 and melatonin (10 mg kg−1, intraperitonealy at 8-9th days of gestation, respectively. Fetuses were collected at 20th day of gestation and after determination of weight and length; they were stained by Alizarin red-Alcian blue method. Results: Cleft palate, spina bifida and exencephaly incidence were 17.70, 20 and 20% in fetuses of rats that received only sodium valproate. Cleft palate, spina bifida and exencephaly incidence were 4.16, 8.33 and 8.33% range in group which received sodium valproate plus melatonin (5 mg kg−1, respectively. However, Cleft palate, spina bifida and excencaphaly incidence were 4/76, 0 and 0% in group which received sodium valproate plus melatonin (10 mg kg−1, respectively. The mean of weight and length of animals fetuses that received melatonin were significantly greater than those received only sodium valproate. Conclusion: It is concluded that melatonin with dose of 10 mg kg−1 had significantly more prophylactic effect than melatonin with dose of 5 mg kg−1 on incidence of sodium valproate-induced skeletal malformations.

  4. The susceptive alendronate-treatment timing and dosage for osteogenesis enhancement in human bone marrow-derived stem cells.

    Directory of Open Access Journals (Sweden)

    Chih-Hsiang Chang

    Full Text Available Recent studies indicated that alendronate enhanced osteogenesis in osteoblasts and human bone marrow-derived stem cells. However, the time- and dose-dependent effects of Aln on osteogenic differentiation and cytotoxicity of hBMSCs remain undefined. In present study, we investigated the effective dose range and timing of hBMSCs. hBMSCs were treated with various Aln doses (1, 5 and 10 µM according to the following groups: group A was treated with Aln during the first five days of bone medium, groups B, C and D were treated during the first, second, and final five days of osteo-induction medium and group E was treated throughout the entire experiment. The mineralization level and cytotoxicity were measured by quantified Alizarin Red S staining and MTT assay. In addition, the reversal effects of farnesyl pyrophosphate and geranylgeranyl pyrophosphate replenishment in group B were also investigated. The results showed that Aln treatment in groups A, B and E enhanced hBMSC mineralization in a dose-dependent manner, and the most pronounced effects were observed in groups B and E. The higher dose of Aln simultaneously enhanced mineralization and caused cytotoxicity in groups B, C and E. Replenishment of FPP or GGPP resulted in partial or complete reverse of the Aln-induced mineralization respectively. Furthermore, the addition of FPP or GGPP also eliminated the Aln-induced cytotoxicity. We demonstrated that hBMSCs are susceptible to 5 µM Aln during the initiation stage of osteogenic differentiation and that a 10 µM dose is cytotoxic.

  5. Cryopreservation of Rat Bone Marrow Derived Mesenchymal Stem Cells by Two Conventional and Open-pulled Straw Vitrification Methods

    Directory of Open Access Journals (Sweden)

    Mohammad Hadi Bahadori

    2009-01-01

    Full Text Available Objective: Mesenchymal stem cells (MSCs are obtained from a variety of sources, mainlythe bone marrow. These cells have a great potential for clinical research, however they cannotstay alive for long periods in culture. The aim of this study is to determine whether vitrificationcan be a useful freezing method for the storage of MSCs.Materials and Methods: Mesenchymal stem cells were isolated from rat bone marrow basedon their capacity to adhere to plastic culture surfaces. MSCs were cryopreserved using boththe vitrification method and open-pulled straw (OPS vitrification and stored in liquid nitrogenwith ethylene glycol ficoll (EFS as a cryoprotectant for two months. The morphology andviability of thawed MSCs were evaluated by trypan blue staining. Furthermore, pre and postcryopreserved MSCs were induced to osteocyte and adipocyte with corresponding osteogenicand adipogenic medium.Results: After thawing, the viability rates were 81.33% ± 6.83 for the vitrification method and80.83% ± 6.4 for OPS vitrification, while the values in the pre-vitrification control group were88.16% ± 6.3 (Mean ± SD, n = 6. Post-cryopreserved cells from both the vitrification methodand OPS vitrification also had a similar cellular morphology and colony-formation that wasindistinguishable from non-vitrified fresh MSCs. In addition, the resuscitated cells cultured ininduction medium showed osteogenesis. Mineral production and deposition was detectableby alizarine red S staining. Moreover, by applying an adipogenic differentiation condition,both pre and post cryopreserved cells differentiated into adipocyte and lipid vacuole accumulationthat was stained by oil red O.Conclusion: Vitrification is a reliable and effective method for the cryopreservation of MSCs.

  6. BONES, TEETH AND GENES: A Genomic Homage to Harry Sicher’s “Axial Movement of Teeth”

    Science.gov (United States)

    Holliday, Sean; Schneider, Bernard; Galang, Maria Therese S.; Fukui, Tadayoshi; Yamane, Akira; Luan, Xianghong; Diekwisch, Thomas G.H.

    2009-01-01

    Aims We have used the model of the un-opposed rodent molar to study morphologic and genetic mechanisms of tooth eruption. Methods Left maxillary molar teeth of 12-day old Swiss-Webster mice were extracted under anesthesia and mandibular molars were allowed to super-erupt. To trace areas of tissue remodeling and to determine areas of new tissue formation, mice were injected with fluorescent dyes, tetracycline, alizarin red, and calcein blue. Subsequent to sacrifice, mandible tissue blocks were prepared for ultrathin ground sections, fluorescent microscopy, and von Kossa’s mineral detection procedure. A second set of specimen was prepared for RNA extraction and microarray analysis. Results Our data established significant eruption of first and second mandibular mouse molars following complete extraction of antagonists by 0.13 mm after 12 days. Labeled tissue sections revealed significant amounts of new bone and cementum apposition on the un-opposed side compared to the control side as revealed by fluorescent markers and ultrathin ground sections. Microarray transcript level comparisons between the experimental and the control group demonstrated significant (more than 2-fold) increase in gene expression of elastin and tenascin C extracellular matrix proteins; brevican, lumican, and biglycan proteoglycans; as well as fibroblast growth factor 9. Conclusion In this study we have established the un-opposed mouse molar as a model to study tissue dynamics during the axial movement of teeth. Our data indicated significant new formation of bone and cementum in tandem with increased expression of extracellular matrix-related genes. PMID:15794043

  7. Preparation of laponite bioceramics for potential bone tissue engineering applications.

    Directory of Open Access Journals (Sweden)

    Chuanshun Wang

    Full Text Available We report a facile approach to preparing laponite (LAP bioceramics via sintering LAP powder compacts for bone tissue engineering applications. The sintering behavior and mechanical properties of LAP compacts under different temperatures, heating rates, and soaking times were investigated. We show that LAP bioceramic with a smooth and porous surface can be formed at 800°C with a heating rate of 5°C/h for 6 h under air. The formed LAP bioceramic was systematically characterized via different methods. Our results reveal that the LAP bioceramic possesses an excellent surface hydrophilicity and serum absorption capacity, and good cytocompatibility and hemocompatibility as demonstrated by resazurin reduction assay of rat mesenchymal stem cells (rMSCs and hemolytic assay of pig red blood cells, respectively. The potential bone tissue engineering applicability of LAP bioceramic was explored by studying the surface mineralization behavior via soaking in simulated body fluid (SBF, as well as the surface cellular response of rMSCs. Our results suggest that LAP bioceramic is able to induce hydroxyapatite deposition on its surface when soaked in SBF and rMSCs can proliferate well on the LAP bioceramic surface. Most strikingly, alkaline phosphatase activity together with alizarin red staining results reveal that the produced LAP bioceramic is able to induce osteoblast differentiation of rMSCs in growth medium without any inducing factors. Finally, in vivo animal implantation, acute systemic toxicity test and hematoxylin and eosin (H&E-staining data demonstrate that the prepared LAP bioceramic displays an excellent biosafety and is able to heal the bone defect. Findings from this study suggest that the developed LAP bioceramic holds a great promise for treating bone defects in bone tissue engineering.

  8. Preparation of laponite bioceramics for potential bone tissue engineering applications.

    Science.gov (United States)

    Wang, Chuanshun; Wang, Shige; Li, Kai; Ju, Yaping; Li, Jipeng; Zhang, Yongxing; Li, Jinhua; Liu, Xuanyong; Shi, Xiangyang; Zhao, Qinghua

    2014-01-01

    We report a facile approach to preparing laponite (LAP) bioceramics via sintering LAP powder compacts for bone tissue engineering applications. The sintering behavior and mechanical properties of LAP compacts under different temperatures, heating rates, and soaking times were investigated. We show that LAP bioceramic with a smooth and porous surface can be formed at 800°C with a heating rate of 5°C/h for 6 h under air. The formed LAP bioceramic was systematically characterized via different methods. Our results reveal that the LAP bioceramic possesses an excellent surface hydrophilicity and serum absorption capacity, and good cytocompatibility and hemocompatibility as demonstrated by resazurin reduction assay of rat mesenchymal stem cells (rMSCs) and hemolytic assay of pig red blood cells, respectively. The potential bone tissue engineering applicability of LAP bioceramic was explored by studying the surface mineralization behavior via soaking in simulated body fluid (SBF), as well as the surface cellular response of rMSCs. Our results suggest that LAP bioceramic is able to induce hydroxyapatite deposition on its surface when soaked in SBF and rMSCs can proliferate well on the LAP bioceramic surface. Most strikingly, alkaline phosphatase activity together with alizarin red staining results reveal that the produced LAP bioceramic is able to induce osteoblast differentiation of rMSCs in growth medium without any inducing factors. Finally, in vivo animal implantation, acute systemic toxicity test and hematoxylin and eosin (H&E)-staining data demonstrate that the prepared LAP bioceramic displays an excellent biosafety and is able to heal the bone defect. Findings from this study suggest that the developed LAP bioceramic holds a great promise for treating bone defects in bone tissue engineering.

  9. Prophylactic Effects of Levamisole and Vitamin E on Phenobarbital-induced Cleft Palate and Spina Bifida in Rat Embryos

    Science.gov (United States)

    Khaksary Mahabady, Mahmood; Najafzadeh Varzi, Hossein

    2011-01-01

    There are many reports that show the teratogenic effects of phenobarbital can be decreased by stimulation of maternal immune system. Therefore, in this study, the prophylactic effects of levamisole and vitamin E on teratogenic effects of phenobarbital were compared. This study was performed on 20 pregnant rats that were divided into four groups. Control group received normal saline and test groups received phenobarbital (120 mg/kg), phenobarbital (120 mg/kg) plus levamisole (10 mg/kg) and phenobarbital (120 mg/kg) plus vitamin E (100 mg/kg) intraperitoneally at 9-11th days of gestation, respectively. Fetuses were collected at 20th day of gestation and after determination of weight and length; they were stained by Alizarin red - Alcian blue method. Cleft palate and spina bifida incidence were 66.66% and 69.44% in fetuses of rats that had received only phenobarbital. Cleft palate and spina bifida incidence were 65.45% and 38.18% had in the group which had received phenobarbital plus levamisole. However, Cleft palate and spina bifida incidence were 54.54% and 27.27% in the group which had received phenobarbital plus vitamin E. The arithmetic means of the weight and length of fetuses the rats that had received levamisole and vitamin E were significantly greater than that of those that had received only phenobarbital. Vitamin E had a greater prophylactic effect than levamisole on the incidence of phenobarbital-induced cleft palate and spina bifida. However, the difference was not significant. PMID:24363692

  10. Oxidized alginate hydrogels for bone morphogenetic protein-2 delivery in long bone defects.

    Science.gov (United States)

    Priddy, Lauren B; Chaudhuri, Ovijit; Stevens, Hazel Y; Krishnan, Laxminarayanan; Uhrig, Brent A; Willett, Nick J; Guldberg, Robert E

    2014-10-01

    Autograft treatment of large bone defects and fracture non-unions is complicated by limited tissue availability and donor site morbidity. Polymeric biomaterials such as alginate hydrogels provide an attractive tissue engineering alternative due to their biocompatibility, injectability, and tunable degradation rates. Irradiated RGD-alginate hydrogels have been used to deliver proteins such as bone morphogenetic protein-2 (BMP-2), to promote bone regeneration and restoration of function in a critically sized rat femoral defect model. However, slow degradation of irradiated alginate hydrogels may impede integration and remodeling of the regenerated bone to its native architecture. Oxidation of alginate has been used to promote degradation of alginate matrices. The objective of this study was to evaluate the effects of alginate oxidation on BMP-2 release and bone regeneration. We hypothesized that oxidized-irradiated alginate hydrogels would elicit an accelerated release of BMP-2, but degrade faster in vivo, facilitating the formation of higher quality, more mature bone compared to irradiated alginate. Indeed, oxidation of irradiated alginate did accelerate in vitro BMP-2 release. Notably, the BMP-2 retained within both constructs was bioactive at 26days, as observed by induction of alkaline phosphatase activity and positive Alizarin Red S staining of MC3T3-E1 cells. From the in vivo study, robust bone regeneration was observed in both groups through 12weeks by radiography, micro-computed tomography analyses, and biomechanical testing. Bone mineral density was significantly greater for the oxidized-irradiated alginate group at 8weeks. Histological analyses of bone defects revealed enhanced degradation of oxidized-irradiated alginate and suggested the presence of more mature bone after 12weeks of healing.

  11. Effects of BMP2 and VEGF165 on the osteogenic differentiation of rat bone marrow-derived mesenchymal stem cells.

    Science.gov (United States)

    Lin, Zhaowei; Wang, Jiang-Sheng; Lin, Lijun; Zhang, Jingwen; Liu, Yunlong; Shuai, Ming; Li, Qi

    2014-03-01

    Bone marrow-derived mesenchymal stem cells (MSCs) are dominant seed cell sources for bone regeneration. Bone morphogenetic proteins (BMPs) initiate cartilage and bone formation in a sequential cascade. Vascular endothelial growth factor (VEGF) is an essential coordinator of extracellular matrix remodeling, angiogenesis and bone formation. In the present study, the effects of the vascular endothelial growth factor 165 (VEGF165) and bone morphogenetic protein 2 (BMP2) genes on bone regeneration were investigated by the lentivirus-mediated cotransfection of the two genes into rat bone marrow-derived MSCs. The successful co-expression of the two genes in the MSCs was confirmed using quantitative polymerase chain reaction (qPCR) and western blot analysis. The results of alizarin red and alkaline phosphatase (ALP) staining at 14 days subsequent to transfection showed that the area of staining in cells transfected with BMP2 alone was higher than that in cells transfected with BMP2 and VEGF165 or untransfected control cells, while the BMP2 + VEGF165 group showed significantly more staining than the untransfected control. This indicated that BMP2 alone exhibited a stronger effect in bone regeneration than BMP2 in combination with VEGF165. Similarly, in inducing culture medium, the ALP activity of the BMP2 + VEGF165 group was notably suppressed compared with that of the BMP2 group. The overexpression of VEGF165 inhibited BMP2-induced MSC differentiation and osteogenesis in vitro. Whether or not local VEGF gene therapy is likely to affect bone regeneration in vivo requires further investigation.

  12. Proliferation and odontogenic differentiation of BMP2 gene‑transfected stem cells from human tooth apical papilla: an in vitro study.

    Science.gov (United States)

    Zhang, Wen; Zhang, Xiaolei; Ling, Junqi; Liu, Wei; Zhang, Xinchun; Ma, Jinglei; Zheng, Jianmao

    2014-10-01

    Stem cells from the apical papilla (SCAP) have odontogenic potential, which plays a pivotal role in the root dentin development of permanent teeth. Human bone morphogenetic protein 2 (BMP2) is a well-known gene that participates in regulating the odontogenic differentiation of dental tissue‑derived stem cells. However, little is known regarding the effects of the BMP2 gene on the proliferation and odontogenic differentiation of SCAP. This study aimed to evaluate the odontogenic differentiation potential of lentiviral‑mediated BMP2 gene‑transfected human SCAP (SCAP/BMP2) in vitro. SCAP were isolated by enzymatic dissociation of human teeth apical papillae. The multipotential of SCAP was verified by their osteogenic and adipogenic differentiation characteristics. The phenotype of SCAP was evaluated by flow cytometry (FCM). The proliferation status of the blank vector‑transfected SCAP (SCAP/Vector) and SCAP/BMP2 was analyzed by a cell counting kit-8 (CCK‑8). Odontogenic genes, including alkaline phosphatase (ALP), osteocalcin (OCN), dentin sialophosphoprotein (DSPP) and dentin matrix protein 1 (DMP1) of the two groups of cells were evaluated by quantitative polymerase chain reaction (qPCR). ALP staining and alizarin red (AR) staining of the cells was performed on the 16th day after transfection. In vitro results of CCK-8, qPCR, ALP and AR staining demonstrated that: ⅰ) SCAP/BMP2 had a comparable proliferation rate to SCAP/Vector; ⅱ) SCAP/BMP2 presented significantly better potential to differentiate into odontoblasts compared to SCAP/Vector by upregulating ALP, OCN, DSPP and DMP1 genes; ⅲ) more ALP granules and mineralized deposits were formed by SCAP/BMP2 as compared to SCAP/Vector. The results suggested that lentiviral-mediated BMP2 gene transfection enhances the odontogenic differentiation capacity of human SCAP in vitro.

  13. Estrogen withdrawal from osteoblasts and osteocytes causes increased mineralization and apoptosis.

    Science.gov (United States)

    Brennan, M Á; Haugh, M G; O'Brien, F J; McNamara, L M

    2014-07-01

    Recent studies have demonstrated increased bone mineral heterogeneity following estrogen withdrawal in vivo. Such changes likely contribute to fracture risk during post-menopausal osteoporosis since tissue mineralization is correlated with bone strength and stiffness. However, the cellular mechanisms responsible for increased mineral variability have not yet been distinguished. The objective of this study is to elucidate how alterations in mineral distribution are initiated during estrogen depletion. Specifically, we tested two separate hypotheses; (1) estrogen deficiency directly alters osteoblast mineralization and (2) estrogen deficiency increases bone cell apoptosis. Osteoblast-like cells (MC3T3-E1) and osteocyte-like cells (MLO-Y4) were pretreated with or without estrogen (17β-estradiol) for 14 days. Estrogen deficiency was subsequently induced by either withdrawing estrogen from cells or blocking estrogen receptors using an estrogen antagonist, fulvestrant (ICI 182,780). Cell number (Hoechst DNA), alkaline phosphatase activity (p-NPP), mineralization (alizarin red) and apoptosis (Caspase 3/7) were evaluated. Whether estrogen withdrawal altered apoptosis rates in the presence of an apoptosis promoting agent (etoposide) was also determined. Interestingly, estrogen withdrawal from cells accustomed to estrogen exposure caused significantly increased osteoblast mineralization and osteocyte apoptosis compared with continued estrogen treatment. In contrast, blocking estrogen receptors with fulvestrant abrogated the mineralization induced by estrogen treatment. When apoptosis was induced using etoposide, cells undergoing estrogen withdrawal increased apoptosis compared to cells with continued estrogen treatment. Recognizing the underlying mechanisms regulating bone cell mineralization and apoptosis during estrogen deficiency and their consequences is necessary to further our knowledge of osteoporosis.

  14. The influence of MicroRNA-150 in Osteoblast Matrix Mineralization.

    Science.gov (United States)

    Dong, Chun-Ling; Liu, Hao-Zhi; Zhang, Zhen-Chun; Zhao, Huan-Li; Zhao, Hui; Huang, Yan; Yao, Jian-Hua; Sun, Tian-Sheng

    2015-12-01

    This study investigated the influence of miR-150 expression on osteoblast matrix mineralization and its mechanisms. The mouse osteoblast cell line MC3T3-E1 was used as an in vitro model of bone formation. On the fifth day of mineralization, transfection experiments using agomiR-150, agomiR-NC, antagomiR-150 antagomiR-NC, and mock groups were set up to test the effects of miR-150 in MC3T3-E1 model. The mRNA and protein levels of OC, ALP, type I collagen, and OPN were measured by qRT-PCR and ELISA. Matrix mineralization was detected by alizarin red S (ARS) staining and flow cytometry was employed to quantify apoptosis in each group. RT-PCR and Western blot were applied to detect the expression of target gene MMP14. Our results demonstrated that the endogenous expression levels of miR-150, OC, ALP, type I collagen, and OPN in MC3T3-E1 cells increased steadily. Exogenous expressions of agomiR-150 and antagomiR-150 can significantly up-/down-regulate, respectively, the expression level of miR-150 in MC3T3-E1 cells. Compared with the mock group, higher expression levels of OC, ALP, type I collagen, and OPN mRNA were observed in the agomiR-150 group, while lower mRNA expression levels of OC, ALP, type I collagen, and OPN were found in the antagomiR-150 group. Based on these results, potential miR-150 targeted genes are discussed. Our results showed that miR-150 supports the osteoblastic phenotype related to osteoblast function and bone mineralization. Thus, miR-150 may have potential therapeutic applications in promoting bone formation in certain disease settings, such as in osteoporosis and in elderly patients.

  15. Nacre extract restores the mineralization capacity of subchondral osteoarthritis osteoblasts.

    Science.gov (United States)

    Brion, A; Zhang, G; Dossot, M; Moby, V; Dumas, D; Hupont, S; Piet, M H; Bianchi, A; Mainard, D; Galois, L; Gillet, P; Rousseau, M

    2015-12-01

    Osteoarthritis (OA) is the most common cause of joint chronic pain and involves the entire joints. Subchondral osteoarthritic osteoblasts present a mineralization defect and, to date, only a few molecules (Vitamin D3 and Bone Morphogenetic Protein2) could improve the mineralization potential of this cell type. In this context, we have tested for the first time the effect of nacre extract on the mineralization capacity of osteoblasts from OA patients. Nacre extract is known to contain osteogenic molecules which have demonstrated their activities notably on the MC3T3 pre-osteoblastic cell line. For this goal, molecules were extracted from nacre (ESM, Ethanol Soluble Matrix) and tested on osteoblasts of the subchondral bone from OA patients undergoing total knee replacement and on MC3T3 cells for comparison. We chose to investigate the mineralization with Alizarin Red staining and with the study of extracellular matrix (ECM) structure and composition. In a complementary way the structure of the ECM secreted during the mineralization phase was investigated using second harmonic generation (SHG). Nacre extract was able to induce the early presence (after 7 days) of precipitated calcium in cells. Raman spectroscopy and electron microscopy showed the presence of nanograins of an early crystalline form of calcium phosphate in OA osteoblasts ECM and hydroxyapatite in MC3T3 ECM. SHG collagen fibers signal was present in both cell types but lower for OA osteoblasts. In conclusion, nacre extract was able to rapidly restore the mineralization capacity of osteoarthritis osteoblasts, therefore confirming the potential of nacre as a source of osteogenic compounds.

  16. Evaluation of 3D printed PCL/PLGA/β-TCP versus collagen membranes for guided bone regeneration in a beagle implant model.

    Science.gov (United States)

    Won, J-Y; Park, C-Y; Bae, J-H; Ahn, G; Kim, C; Lim, D-H; Cho, D-W; Yun, W-S; Shim, J-H; Huh, J-B

    2016-10-07

    Here, we compared 3D-printed polycaprolactone/poly(lactic-co-glycolic acid)/β-tricalcium phosphate (PCL/PLGA/β-TCP) membranes with the widely used collagen membranes for guided bone regeneration (GBR) in beagle implant models. For mechanical property comparison in dry and wet conditions and cytocompatibility determination, we analyzed the rate and pattern of cell proliferation of seeded fibroblasts and preosteoblasts using the cell counting kit-8 assay and scanning electron microscopy. Osteogenic differentiation was verified using alizarin red S staining. At 8 weeks following implantation in vivo using beagle dogs, computed tomography and histological analyses were performed after sacrifice. Cell proliferation rates in vitro indicated that early cell attachment was higher in collagen than in PCL/PLGA/β-TCP membranes; however, the difference subsided by day 7. Similar outcomes were found for osteogenic differentiation, with approximately 2.5 times greater staining in collagen than PCL/PLGA/β-TCP, but without significant difference by day 14. In vivo, bone regeneration in the defect area, represented by new bone formation and bone-to-implant contact, paralleled those associated with collagen membranes. However, tensile testing revealed that whereas the PCL/PLGA/β-TCP membrane mechanical properties were conserved in both wet and dry states, the tensile property of collagen was reduced by 99% under wet conditions. Our results demonstrate in vitro and in vivo that PCL/PLGA/β-TCP membranes have similar levels of biocompatibility and bone regeneration as collagen membranes. In particular, considering that GBR is always applied to a wet environment (e.g. blood, saliva), we demonstrated that PCL/PLGA/β-TCP membranes maintained their form more reliably than collagen membranes in a wet setting, confirming their appropriateness as a GBR membrane.

  17. Effect of lamin A/C knockdown on osteoblast differentiation and function.

    Science.gov (United States)

    Akter, Rahima; Rivas, Daniel; Geneau, Graziello; Drissi, Hicham; Duque, Gustavo

    2009-02-01

    Recent studies have associated mutations in lamin A/C, a component of the nuclear lamina, with premature aging and severe bone loss. In this study, we hypothesized that reduced expression of lamin A/C has a negative impact on osteoblastogenesis and bone formation in vitro. We inhibited lamin A/C using increasing doses of lamin A/C siRNA in normal human osteoblasts and differentiating mesenchymal stem cells (MSCs). Untreated cells and cells treated with vehicle but without the siRNA-oligo were used as control. The level of effectiveness of siRNA was determined by RT-PCR, Western blot, and immunofluorescence. Nuclear blebbing, a typical finding of lamin A/C inhibition, was quantified using propidium iodine staining, and its effect on cell survival was determined using MTS-formazan. Furthermore, alizarin red and alkaline phosphatase staining were correlated with osteocalcin secretion and levels of expression of osteocalcin, osterix, bone sialoprotein, and Runx2. Finally, the nuclear binding activity of Runx2, an essential transcription factor for osteoblast differentiation, was assessed using ELISA and EMSA. A successful inhibitory effect on the lamin A/C gene at doses of 400-800 nM oligo was obtained without affecting cell survival. Whereas osteoblast function was significantly affected by lamin A/C inhibition, siRNA-treated MSC showed a higher incidence of nuclear changes, lower osteoblast differentiation, and enhanced adipocyte differentiation. Finally, lamin A/C knockdown reduced Runx2 nuclear binding activity without affecting Runx2 expression. In summary, our results indicate that lamin A/C is a new factor needed for osteoblast differentiation that plays an important role in the cellular mechanisms of age-related bone loss.

  18. The role of muscle loading on bone (Remodeling at the developing enthesis.

    Directory of Open Access Journals (Sweden)

    Alexander M Tatara

    Full Text Available Muscle forces are necessary for the development and maintenance of a mineralized skeleton. Removal of loads leads to malformed bones and impaired musculoskeletal function due to changes in bone (remodeling. In the current study, the development of a mineralized junction at the interface between muscle and bone was examined under normal and impaired loading conditions. Unilateral mouse rotator cuff muscles were paralyzed using botulinum toxin A at birth. Control groups consisted of contralateral shoulders injected with saline and a separate group of normal mice. It was hypothesized that muscle unloading would suppress bone formation and enhance bone resorption at the enthesis, and that the unloading-induced bony defects could be rescued by suppressing osteoclast activity. In order to modulate osteoclast activity, mice were injected with the bisphosphonate alendronate. Bone formation was measured at the tendon enthesis using alizarin and calcein fluorescent labeling of bone surfaces followed by quantitative histomorphometry of histologic sections. Bone volume and architecture was measured using micro computed tomography. Osteoclast surface was determined via quantitative histomorphometry of tartrate resistant acid phosphatase stained histologic sections. Muscle unloading resulted in delayed initiation of endochondral ossification at the enthesis, but did not impair bone formation rate. Unloading led to severe defects in bone volume and trabecular bone architecture. These defects were partially rescued by suppression of osteoclast activity through alendronate treatment, and the effect of alendronate was dose dependent. Similarly, bone formation rate was increased with increasing alendronate dose across loading groups. The bony defects caused by unloading were therefore likely due to maintained high osteoclast activity, which normally decreases from neonatal through mature timepoints. These results have important implications for the treatment of

  19. Adiponectin and AMP kinase activator stimulate proliferation, differentiation, and mineralization of osteoblastic MC3T3-E1 cells

    Directory of Open Access Journals (Sweden)

    Yamauchi Mika

    2007-11-01

    Full Text Available Abstract Background Adiponectin is a key mediator of the metabolic syndrome that is caused by visceral fat accumulation. Adiponectin and its receptors are known to be expressed in osteoblasts, but their actions with regard to bone metabolism are still unclear. In this study, we investigated the effects of adiponectin on the proliferation, differentiation, and mineralization of osteoblastic MC3T3-E1 cells. Results Adiponectin receptor type 1 (AdipoR1 mRNA was detected in the cells by RT-PCR. The adenosine monophosphate-activated protein kinase (AMP kinase was phosphorylated by both adiponectin and a pharmacological AMP kinase activator, 5-amino-imidazole-4-carboxamide-riboside (AICAR, in the cells. AdipoR1 small interfering RNA (siRNA transfection potently knocked down the receptor mRNA, and the effect of this knockdown persisted for as long as 10 days after the transfection. The transfected cells showed decreased expressions of type I collagen and osteocalcin mRNA, as determined by real-time PCR, and reduced ALP activity and mineralization, as determined by von Kossa and Alizarin red stainings. In contrast, AMP kinase activation by AICAR (0.01–0.5 mM in wild-type MC3T3-E1 cells augmented their proliferation, differentiation, and mineralization. BrdU assay showed that the addition of adiponectin (0.01–1.0 μg/ml also promoted their proliferation. Osterix, but not Runx-2, appeared to be involved in these processes because AdipoR1 siRNA transfection and AICAR treatments suppressed and enhanced osterix mRNA expression, respectively. Conclusion Taken together, this study suggests that adiponectin stimulates the proliferation, differentiation, and mineralization of osteoblasts via the AdipoR1 and AMP kinase signaling pathways in autocrine and/or paracrine fashions.

  20. Micro/Nano Multilayered Scaffolds of PLGA and Collagen by Alternately Electrospinning for Bone Tissue Engineering.

    Science.gov (United States)

    Kwak, Sanghwa; Haider, Adnan; Gupta, Kailash Chandra; Kim, Sukyoung; Kang, Inn-Kyu

    2016-12-01

    The dual extrusion electrospinning technique was used to fabricate multilayered 3D scaffolds by stacking microfibrous meshes of poly(lactic acid-co-glycolic acid) (PLGA) in alternate fashion to micro/nano mixed fibrous meshes of PLGA and collagen. To fabricate the multilayered scaffold, 35 wt% solution of PLGA in THF-DMF binary solvent (3:1) and 5 wt% solution of collagen in hexafluoroisopropanol (HFIP) with and without hydroxyapatite nanorods (nHA) were used. The dual and individual electrospinning of PLGA and collagen were carried out at flow rates of 1.0 and 0.5 mL/h, respectively, at an applied voltage of 20 kV. The density of collagen fibers in multilayered scaffolds has controlled the adhesion, proliferation, and osteogenic differentiation of MC3T3-E1 cells. The homogeneous dispersion of glutamic acid-modified hydroxyapatite nanorods (nHA-GA) in collagen solution has improved the osteogenic properties of fabricated multilayered scaffolds. The fabricated multilayered scaffolds were characterized using FT-IR, X-ray photoelectron spectroscopy, and transmission electron microscopy (TEM). The scanning electron microscopy (FE-SEM) was used to evaluate the adhesion and spreads of MC3T3-E1 cells on multilayered scaffolds. The activity of MC3T3-E1 cells on the multilayered scaffolds was evaluated by applying MTT, alkaline phosphatase, Alizarin Red, von Kossa, and cytoskeleton F-actin assaying protocols. The micro/nano fibrous PLGA-Col-HA scaffolds were found to be highly bioactive in comparison to pristine microfibrous PLGA and micro/nano mixed fibrous PLGA and Col scaffolds.

  1. Human amnion mesenchymal stem cells promote proliferation and osteogenic differentiation in human bone marrow mesenchymal stem cells.

    Science.gov (United States)

    Wang, Yuli; Yin, Ying; Jiang, Fei; Chen, Ning

    2015-02-01

    Human amnion mesenchymal stem cells (HAMSCs) can be obtained from human amniotic membrane, a highly abundant and readily available tissue. HAMSC sources present fewer ethical issues, have low immunogenicity, anti-inflammatory properties, considerable advantageous characteristics, and are considered an attractive potential treatment material in the field of regenerative medicine. We used a co-culture system to determine whether HAMSCs could promote osteogenesis in human bone marrow mesenchymal stem cells (HBMSCs). We isolated HAMSCs from discarded amnion samples and collected them using pancreatin/collagenase digestion. We cultured HAMSCs and HBMSCSs in basal medium. Activity of alkaline phosphatase (ALP), an early osteogenesis marker, was increased in the co-culture system compared to the control single cultures, which we also confirmed by ALP staining. We used immunofluorescence testing to investigate the effects of co-culturing with HAMSCs on HBMSC proliferation, which revealed that the co-culturing enhanced EdU expression in HBMSCs. Western blotting and quantitative real-time PCR indicated that co-culturing promoted osteogenesis in HBMSCs. Furthermore, Alizarin red S staining revealed that extracellular matrix calcium levels in mineralized nodule formation produced by the co-cultures were higher than that in the controls. Using the same co-culture system, we further observed the effects of HAMSCs on osteogenic differentiation in primary osteoblasts by Western blotting, which better addressed the mechanism for HAMSCs in bone regeneration. The results showed HAMSCs are osteogenic and not only play a role in promoting HBMSC proliferation and osteogenic differentiation but also in osteoblasts, laying the foundation for new regenerative medicine methods.

  2. Transport of ARS-labeled hydroxyapatite nanoparticles in saturated granular media is influenced by surface charge variability even in the presence of humic acid

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Dengjun [Key Laboratory of Soil Environment and Pollution Remediation, Institute of Soil Science, Chinese Academy of Sciences, No. 71 East Beijing Road, Nanjing 210008 (China); Graduate School of the Chinese Academy of Sciences, Beijing 100049 (China); Bradford, Scott A. [U.S. Salinity Laboratory, Agricultural Research Service, United States Department of Agriculture, 450 W. Big Springs Road, Riverside, CA 92507 (United States); Harvey, Ronald W. [U.S. Geological Survey, 3215 Marine Street, Boulder, CO 80303 (United States); Hao, Xiuzhen [Key Laboratory of Soil Environment and Pollution Remediation, Institute of Soil Science, Chinese Academy of Sciences, No. 71 East Beijing Road, Nanjing 210008 (China); Zhou, Dongmei, E-mail: dmzhou@issas.ac.cn [Key Laboratory of Soil Environment and Pollution Remediation, Institute of Soil Science, Chinese Academy of Sciences, No. 71 East Beijing Road, Nanjing 210008 (China)

    2012-08-30

    Highlights: Black-Right-Pointing-Pointer The transport and retention kinetics of ARS-labeled hydroxyapatite nanoparticles (ARS-nHAP) were investigated over a range of ionic strengths in the presence of humic acid. Black-Right-Pointing-Pointer A two-site kinetic attachment model predicted both the breakthrough curves and retention profiles of ARS-nHAP quite well. Black-Right-Pointing-Pointer The retention profiles of ARS-nHAP exhibited hyperexponential shapes for all the test conditions. Black-Right-Pointing-Pointer Surface charge heterogeneities on the collector surfaces and especially within the ARS-nHAP population contributed to hyperexponential retention profiles. - Abstract: Hydroxyapatite nanoparticle (nHAP) is increasingly being used to remediate soils and water polluted by metals and radionuclides. The transport and retention of Alizarin red S (ARS)-labeled nHAP were investigated in water-saturated granular media. Experiments were carried out over a range of ionic strength (I{sub c}, 0-50 mM NaCl) conditions in the presence of 10 mg L{sup -1} humic acid. The transport of ARS-nHAP was found to decrease with increasing suspension I{sub c} in part, because of enhanced aggregation and chemical heterogeneity. The retention profiles (RPs) of ARS-nHAP exhibited hyperexponential shapes (a decreasing rate of retention with increasing transport distance) for all test conditions, suggesting that some of the attachment was occurring under unfavorable conditions. Surface charge heterogeneities on the collector surfaces and especially within the ARS-nHAP population were contributing causes for the hyperexponential RPs. Consideration of the effect(s) of I{sub c} in the presence of HA is needed to improve the efficacy of nHAP for scavenging metals and actinides in real soils and groundwater environments.

  3. Fucoidan promotes osteoblast differentiation via JNK- and ERK-dependent BMP2-Smad 1/5/8 signaling in human mesenchymal stem cells.

    Science.gov (United States)

    Kim, Beom Su; Kang, Hyo-Jin; Park, Ji-Yun; Lee, Jun

    2015-01-09

    Fucoidan has attracted attention as a potential drug because of its biological activities, which include osteogenesis. However, the molecular mechanisms involved in the osteogenic activity of fucoidan in human alveolar bone marrow-derived mesenchymal stem cells (hABM-MSCs) remain largely unknown. We investigated the action of fucoidan on osteoblast differentiation in hABM-MSCs and its impact on signaling pathways. Its effect on proliferation was determined using the crystal violet staining assay. Osteoblast differentiation was evaluated based on alkaline phosphatase (ALP) activity and the mRNA expression of multiple osteoblast markers. Calcium accumulation was determined by Alizarin red S staining. We found that fucoidan induced hABM-MSC proliferation. It also significantly increased ALP activity, calcium accumulation and the expression of osteoblast-specific genes, such as ALP, runt-related transcription factor 2, type I collagen-α 1 and osteocalcin. Moreover, fucoidan induced the expression of bone morphogenetic protein 2 (BMP2) and stimulated the activation of extracellular signal-related kinase (ERK), c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase by increasing phosphorylation. However, the effect of fucoidan on osteogenic differentiation was inhibited by specific inhibitors of ERK (PD98059) and JNK (SP600125) but not p38 (SB203580). Fucoidan enhanced BMP2 expression and Smad 1/5/8, ERK and JNK phosphorylation. Moreover, the effect of fucoidan on osteoblast differentiation was diminished by BMP2 knockdown. These results indicate that fucoidan induces osteoblast differentiation through BMP2-Smad 1/5/8 signaling by activating ERK and JNK, elucidating the molecular basis of the osteogenic effects of fucoidan in hABM-MSCs.

  4. Pericyst may be a new pharmacological and therapeutic target for hydatid disease

    Institute of Scientific and Technical Information of China (English)

    WU Xiang-wei; CHEN Xue-ling; ZHANG Shi-jie; ZHANG Xi; SUN Hong; PENG Xin-yu

    2011-01-01

    Background Most hydatid cysts with calcified walls are biologically and clinically silent and inactive. Transforming growth factor-beta 1 (TGF-β1) plays a critical role in the calcification process of cells. The aim of this study was to assess the effect of modulating TGF-β1 signaling on the calcification of hydatid cysts.Methods Pericyst cells isolated from hepatic hydatid cysts were cultured with osteogenic media. These cells were assessed for alkaline phosphatase activity and mineralization capacity using Alizarin Red staining. Cells were also treated with recombinant human TGF-β1 and TGF-β inhibitor, and the expression profiles of osteoblast markers (RUNX2,osterix, and osteocalcin) were analyzed using Western blotting. The effects of inhibiting TGF-β1 signaling on calcification of pericyst walls were assessed using different doses of TGF-β inhibitor for 7 weeks in a preclinical disease model of liver cystic echinococcosis.Results Cells within the pericyst displayed high levels of alkaline phosphatase activity and mineralized nodule formation, as induced by osteogenic media. These activities, as well as expression profiles of osteoblast markers (RUNX2, osterix, and osteocalcin) could be inhibited by addition of recombinant human TGF-β1 (rhTGF-β1) and enhanced by TGF-β inhibitor. In the animal model of cystic echinococcosis, inhibition of TGF-β1 signaling increased calcification of the pericyst wall, which was associated with decreased cyst load index and lower viability of protoscoleces.Conclusions Cells within the pericysts adopt an osteoblast-like phenotype and have osteogenic potential. Inhibition of TGF-β1 signaling increases hydatid cyst calcification. Pharmacological modulation of calcification in pericysts may be a new therapeutic target in the treatment of hydatid disease.

  5. Isolation of mesenchymal stem cells from equine umbilical cord blood

    Directory of Open Access Journals (Sweden)

    Thomsen Preben D

    2007-05-01

    Full Text Available Abstract Background There are no published studies on stem cells from equine cord blood although commercial storage of equine cord blood for future autologous stem cell transplantations is available. Mesenchymal stem cells (MSC have been isolated from fresh umbilical cord blood of humans collected non-invasively at the time of birth and from sheep cord blood collected invasively by a surgical intrauterine approach. Mesenchymal stem cells isolation percentage from frozen-thawed human cord blood is low and the future isolation percentage of MSCs from cryopreserved equine cord blood is therefore expectedly low. The hypothesis of this study was that equine MSCs could be isolated from fresh whole equine cord blood. Results Cord blood was collected from 7 foals immediately after foaling. The mononuclear cell fraction was isolated by Ficoll density centrifugation and cultured in a DMEM low glucose based media at 38.5°C in humidified atmosphere containing 5% CO2. In 4 out of 7 samples colonies with MSC morphology were observed. Cellular morphology varied between monolayers of elongated spindle-shaped cells to layered cell clusters of cuboidal cells with shorter cytoplasmic extensions. Positive Alizarin Red and von Kossa staining as well as significant calcium deposition and alkaline phosphatase activity confirmed osteogenesis. Histology and positive Safranin O staining of matrix glycosaminoglycans illustrated chondrogenesis. Oil Red O staining of lipid droplets confirmed adipogenesis. Conclusion We here report, for the first time, the isolation of mesenchymal-like stem cells from fresh equine cord blood and their differentiation into osteocytes, chondrocytes and adipocytes. This novel isolation of equine cord blood MSCs and their preliminary in vitro differentiation positions the horse as the ideal pre-clinical animal model for proof-of-principle studies of cord blood derived MSCs.

  6. Investigation of the optimal timing for chondrogenic priming of MSCs to enhance osteogenic differentiation in vitro as a bone tissue engineering strategy.

    Science.gov (United States)

    Freeman, F E; Haugh, M G; McNamara, L M

    2016-04-01

    Recent in vitro tissue engineering approaches have shown that chondrogenic priming of human bone marrow mesenchymal stem cells (MSCs) can have a positive effect on osteogenesis in vivo. However, whether chondrogenic priming is an effective in vitro bone regeneration strategy is not yet known. In particular, the appropriate timing for chondrogenic priming in vitro is unknown albeit that in vivo cartilage formation persists for a specific period before bone formation. The objective of this study is to determine the optimum time for chondrogenic priming of MSCs to enhance osteogenic differentiation by MSCs in vitro. Pellets derived from murine and human MSCs were cultured in six different media groups: two control groups (chondrogenic and osteogenic) and four chondrogenic priming groups (10, 14, 21 and 28 days priming). Biochemical analyses (Hoechst, sulfate glycosaminoglycan (sGAG), Alkaline Phosphate (ALP), calcium), histology (Alcian Blue, Alizarin Red) and immunohistochemistry (collagen types I, II and X) were performed on the samples at specific times. Our results show that after 49 days the highest amount of sGAG production occurred in MSCs chondrogenically primed for 21 days and 28 days. Moreover we found that chondrogenic priming of MSCs in vitro for specific amounts of time (14 days, 21 days) can have optimum influence on their mineralization capacity and can produce a construct that is mineralized throughout the core. Determining the optimum time for chondrogenic priming to enhance osteogenic differentiation in vitro provides information that might lead to a novel regenerative treatment for large bone defects, as well as addressing the major limitation of core degradation and construct failure.

  7. Acetylcholinesterase Regulates Skeletal In Ovo Development of Chicken Limbs by ACh-Dependent and -Independent Mechanisms

    Science.gov (United States)

    Spieker, Janine; Ackermann, Anica; Salfelder, Anika; Vogel-Höpker, Astrid; Layer, Paul G.

    2016-01-01

    Formation of the vertebrate limb presents an excellent model to analyze a non-neuronal cholinergic system (NNCS). Here, we first analyzed the expression of acetylcholinesterase (AChE) by IHC and of choline acetyltransferase (ChAT) by ISH in developing embryonic chicken limbs (stages HH17-37). AChE outlined formation of bones, being strongest at their distal tips, and later also marked areas of cell death. At onset, AChE and ChAT were elevated in two organizing centers of the limb anlage, the apical ectodermal ridge (AER) and zone of polarizing activity (ZPA), respectively. Thereby ChAT was expressed shortly after AChE, thus strongly supporting a leading role of AChE in limb formation. Then, we conducted loss-of-function studies via unilateral implantation of beads into chicken limb anlagen, which were soaked in cholinergic components. After varying periods, the formation of cartilage matrix and of mineralizing bones was followed by Alcian blue (AB) and Alizarin red (AR) stainings, respectively. Both acetylcholine (ACh)- and ChAT-soaked beads accelerated bone formation in ovo. Notably, inhibition of AChE by BW284c51, or by the monoclonal antibody MAB304 delayed cartilage formation. Since bead inhibition of BChE was mostly ineffective, an ACh-independent action during BW284c51 and MAB304 inhibition was indicated, which possibly could be due to an enzymatic side activity of AChE. In conclusion, skeletogenesis in chick is regulated by an ACh-dependent cholinergic system, but to some extent also by an ACh-independent aspect of the AChE protein. PMID:27574787

  8. Osteoadherin accumulates in the predentin towards the mineralization front in the developing tooth.

    Directory of Open Access Journals (Sweden)

    Hero Nikdin

    Full Text Available BACKGROUND: Proteoglycans (PG are known to be involved in the organization and assembly of the extracellular matrix (ECM prior to mineral deposition. Osteoadherin (OSAD, a keratan sulphate PG is a member of the small leucine-rich (SLRP family of PGs and unlike other SLRPs, OSAD expression is restricted to mineralized tissues. It is proposed to have a high affinity for hydroxyapatite and has been shown to be expressed by mature osteoblasts but its exact role remains to be elucidated. METHODOLOGY/PRINCIPAL FINDINGS: We investigated the protein distribution of OSAD in the developing mouse tooth using immunohistochemistry and compared its expression with other SLRPs, biglycan (BGN, decorin (DCN and fibromodulin (FMD. OSAD was found to be specifically localized in the predentin layer of the tooth and focused at the mineralization front. These studies were confirmed at the ultrastructural level using electron microscopy (iEM, where the distribution of immunogold labeled OSAD particles were quantified and significant amounts were found in the predentin, forming a gradient towards the mineralization front. In addition, iEM results revealed OSAD to lie in close association with collagen fibers, further suggesting an important role for OSAD in the organization of the ECM. The expression profile of mineralization-related SLRP genes by rat dental pulp cells exposed to mineralization inducing factors, showed an increase in all SLRP genes. Indeed, OSAD expression was significantly increased during the mineralization process, specifically following, matrix maturation, and finally mineral deposition. Alizarin Red S staining for calcium deposition showed clear bone-like nodules, which support matrix maturation and mineralization. CONCLUSIONS: These studies provide new evidence for the role of OSAD in the mineralization process and its specific localization in the predentin layer accumulating at the mineralization front highlighting its role in tooth development.

  9. Silorane resin supports proliferation, differentiation, and mineralization of MLO-A5 bone cells in vitro and bone formation in vivo

    Science.gov (United States)

    Eick, J. David; Barragan-Adjemian, Cielo; Rosser, Jennifer; Melander, Jennifer R.; Dusevich, Vladimir; Weiler, Rachel A.; Miller, Bradley D.; Kilway, Kathleen V.; Dallas, Mark R.; Bi, Lianxing; Nalvarte, Elisabet L.; Bonewald, Lynda F.

    2015-01-01

    Methyl methacrylate used in bone cements has drawbacks of toxicity, high exotherm, and considerable shrinkage. A new resin, based on silorane/oxirane chemistry, has been shown to have little toxicity, low exotherm, and low shrinkage. We hypothesized that silorane-based resins may also be useful as components of bone cements as well as other bone applications and began testing on bone cell function in vitro and in vivo. MLO-A5, late osteoblast cells, were exposed to polymerized silorane (SilMix) resin (and a standard polymerized bisGMA/TEGDMA methacrylate (BT) resin and compared to culture wells without resins as control. A significant cytotoxic effect was observed with the BT resin resulting in no cell growth, whereas in contrast, SilMix resin had no toxic effects on MLO-A5 cell proliferation, differentiation, nor mineralization. The cells cultured with SilMix produced increasing amounts of alkaline phosphatase (1.8-fold) compared to control cultures. Compared to control cultures, an actual enhancement of mineralization was observed in the silorane resin-containing cultures at days 10 and 11 as determined by von Kossa (1.8–2.0 fold increase) and Alizarin red staining (1.8-fold increase). A normal bone calcium/phosphate atomic ratio was observed by elemental analysis along with normal collagen formation. When used in vivo to stabilize osteotomies, no inflammatory response was observed, and the bone continued to heal. In conclusion, the silorane resin, SilMix, was shown to not only be non cytototoxic, but actually supported bone cell function. Therefore, this resin has significant potential for the development of a nontoxic bone cement or bone stabilizer. PMID:22278990

  10. L-carnitine significantly decreased aging of rat adipose tissue-derived mesenchymal stem cells.

    Science.gov (United States)

    Mobarak, Halimeh; Fathi, Ezzatollah; Farahzadi, Raheleh; Zarghami, Nosratollah; Javanmardi, Sara

    2017-03-01

    Mesenchymal stem cells are undifferentiated cells that have the ability to divide continuously and tissue regeneration potential during the transplantation. Aging and loss of cell survival, is one of the main problems in cell therapy. Since the production of free radicals in the aging process is effective, the use of antioxidant compounds can help in scavenging free radicals and prevent the aging of cells. The aim of this study is evaluate the effects of L-carnitine (LC) on proliferation and aging of rat adipose tissue-derived mesenchymal stem cells (rADSC). rADSCs were isolated from inguinal region of 5 male Rattus rats. Oil red-O, alizarin red-S and toluidine blue staining were performed to evaluate the adipogenic, osteogenic and chondrogenic differentiation of rADSCs, respectively. Flow cytometric analysis was done for investigating the cell surface markers. The methyl thiazol tetrazolium (MTT) method was used to determine the cell proliferation of rADSCs following exposure to different concentrations of LC. rADSCs aging was evaluated by beta-galactosidase staining. The results showed significant proliferation of rADSCs 48 h after treatment with concentrations of 0.2 mM LC. In addition, in the presence of 0.2 mM LC, rADSCs appeared to be growing faster than control group and 0.2 mM LC supplementation could significantly decrease the population doubling time and aging of rADSCs. It seems that LC would be a good antioxidant to improve lifespan of rADSCs due to the decrease in aging.

  11. Endosulfan Impacts on the Developing Chick Embryos: Morphological, Morphometric and Skeletal Changes

    Directory of Open Access Journals (Sweden)

    Y.M. Mobarak

    2011-01-01

    Full Text Available This study aims to explore the effects of the organochlorine pesticide Endosulfan (35% EC on the developing chick embryos. After 24 h of eggs incubation, a single dose of 7 or 14 or 21 mg Endosulfan/egg was administered through the egg air space at once. The eggs were opened on embryonic days 6 and 12 and the embryos were evaluated for viability, wet body weights and various morphological, morphometric and skeletal changes. Skeletons of 12-day-old embryos were stained by alizarin red S and Alcian blue using a whole mount double cartilage and bone staining technique. Comparing the three doses with control and with each others, the high dose treatment resulted in statistically significant more embryonic deaths, while the mid-dose caused statistically more malformed embryos. On both embryonic days, the treated embryos exhibited dose-related growth retardation, as reflected by significant reductions of embryonic wet body weight, anterior-posterior head and crown-rump lengths as well as generalized edema and hematomas formations. Also, on embryonic day 12 significant reductions of beak length, eye diameters and measurements of wing and hind-limb parts were recorded. Abnormal survivors showed high percentages of limb deformities (as limb paralysis, clinodactyly, flexion and shortness of limbs or digits, microphthalmia, microtia and omphalocele. The skeleton of treated embryos showed anomalies and incomplete chondrification and/or ossification of some skull parts (interorbital septum, frontals, parietals, palatines and external auditory apertures, cervicals, scapulae, ribs, sacrals and caudals. These findings suggest that Endosulfan exhibits embryotoxic and teratogenic effects on the developing chick embryos in terms of growth retardation, external and skeletal malformations.

  12. The ossification of the pelvic girdle and leg skeleton of the quail (Coturnix coturnix japonica).

    Science.gov (United States)

    Pourlis, A F; Antonopoulos, J

    2014-08-01

    The onset of ossification centres of the pelvic girdle and leg skeleton of the quail in embryos and juvenile birds were studied. Specimens, which were cleared and were stained with Alcian Blue and Alizarin Red S, were examined at the stereomicroscope. The ilium and the pubis began to ossify at the 8th day (E8), whereas the ischium at E9. Perichondral ossification was observed at E6 in the femur, tibia and fibula. A secondary ossification centre was detected in the proximal epiphysis of the tibiotarsus at the 15th post-hatching day (P15). The patella began to ossify at P30. Regarding the tarsal bones tibiale, pre-tibiale and fibulare, ossification was observed at the E15, E12 and E16, respectively. The metatarsals II, III, IV ossified at E7, whereas the metatarsal I at E11. The centres of ossification of the 1st phalanges of all digits were observed at E9. At the same day, the ossification centres of the 2nd phalanx of digits II and III as well as the 3rd phalanx of digit III appeared. At E10, ossification was observed in the 2nd phalanx of digit I, in the 3rd phalanx of digit II and in the 2nd and 3rd phalanx of digit IV. In the 4th phalanx of digit III and in the terminal phalanges of digit IV, ossification was observed at E11. The data presented here provide useful baseline information on the normal sequential pattern of ossification in the pelvic girdle and leg skeleton in this species.

  13. mTORC1 signaling promotes osteoblast differentiation of preosteoblast MC3 T3-E1%mTORC1信号促进前成骨细胞MC3 T3-E1的分化成熟

    Institute of Scientific and Technical Information of China (English)

    张婧婧; 张楠; 吴伟; 王鹏; 杨景瑞; 段巨洪; 于海川

    2016-01-01

    目的:研究mTORC1信号对前成骨细胞MC3T3-E1向成骨细胞分化成熟的调控作用。方法:通过向MC3T3-E1转染pcDNA3.1-Raptor,对mTORC1信号相关蛋白Raptor进行过表达。向MC3T3-E1转染Raptor siRNA,对mTORC1信号蛋白Raptor进行基因沉默。通过Real-time PCR方法测定Raptor的基因表达,通过蛋白免疫印迹法测定Raptor蛋白水平,并通过茜素红染色检测成骨矿化情况,以测定成骨分化程度。通过Real-time PCR检测成骨分化指标的基因表达。结果:与对照组相比,Raptor过表达组的Raptor mRNA和蛋白水平明显增加;茜素红染色结果显示Raptor过表达组染色更深,说明成骨矿化程度更高;荧光定量PCR结果显示,Raptor过表达组的成骨分化标记基因以及成骨转录因子的表达量均高于对照组。与对照组相比,Raptor siRNA组的Raptor mRNA和蛋白水平明显降低;茜素红染色结果显示Raptor siRNA组染色更浅,说明成骨矿化程度更低;荧光定量PCR结果显示,Raptor siRNA组的成骨分化标记基因以及成骨转录因子的表达量均低于对照组。结论:mTORC1信号促进前成骨细胞MC3T3-E1向成骨细胞分化成熟。%Objective:To investigate the regulatory role of mTORC 1 signaling on osteoblast differentiation from preosteoblast cell MC3T3-E1.Methods:To overexpress the gene Raptor ,which was a critical component of mTORC1 signaling,pcDNA3.1-Raptor was transfected to preosteoblast cell MC 3T3-E1.The Raptor overexpression was confirmed by Real-time PCR and Western blot.For the gene silence of Raptor ,Raptor siRNA was transfected to preosteoblast cell MC 3T3-E1.Alizarin red staining was used to detect the min-eralization of MC3T3-E1.Real-time PCR was used to detect the gene expression of osteoblast specific marker genes.Western blot was used to detect the protein level of Raptor.Results: Compared to the control group ,the levels of Raptor mRNA and proteins increased

  14. 比较3种形态学方法观察成骨细胞矿化结节的应用价值%Comparison of three morphology methods for observing mineralization nodules of osteoblasts

    Institute of Scientific and Technical Information of China (English)

    廖乃顺; 李钻芳; 林如辉; 陈文列; 黄云梅; 黄美雅

    2014-01-01

    背景:矿化结节是成骨细胞分化成熟的标志,以往观察方法多采用茜素红等特殊染色法。  目的:比较茜素红染色-光镜、四环素荧光标记-激光扫描共聚焦显微镜、扫描电镜3种形态学观察成骨细胞矿化结节的方法,分析各自的特点及在骨病研究中的应用价值。  方法:将大鼠成骨细胞株UMR-106正常培养、每天换液,连续培养14 d,分别采用茜素红染色-光镜、四环素荧光标记-激光扫描共聚焦显微镜、扫描电镜观察矿化结节的形态结构,并利用扫描电镜结合能谱仪原位定量分析钙元素;此外,在培养中加入可抑制成骨细胞增殖、分化的肿瘤坏死因子α作为对照实验。  结果与结论:3种观察方法均可观察到正常成骨细胞矿化结节的形态结构。对于肿瘤坏死因子α抑制成骨细胞产生矿化结节的情况,经茜素红染色-光镜观察法,未见明显的矿化结节;而四环素荧光染色-激光扫描共聚焦显微镜观察法,可见清晰、稀少的矿化结节;扫描电镜观察法明显可见较少而细小的矿化结节,提示后二种方法灵敏度较高。此外,扫描电镜可将观察成骨细胞分泌钙质、形成矿化结节的亚细胞结构,与能谱元素分析结合,可实现矿化结节的定位与定量,在骨病研究中值得推广应用。%BACKGROUND:Mineralized nodules are the mature marker of osteoblast differentiation, and the observation methods mainly use alizarin red staining. OBJECTIVE:To compare the observation results of mineralized nodules by three methods, and to explore their characteristics and advantages, as wel as further application in the research of bone disease. METHODS:The rat osteoblast-like cellline UMR-106 were cultured in the fresh medium that was changed every day, for 14 days. Alizarin red staining-light microscope, tetracycline fluorescence labeling-laser confocal scanning

  15. Chondrogenic potential of subpopulations of cells expressing mesenchymal stem cell markers derived from human synovial membranes.

    Science.gov (United States)

    Arufe, M C; De la Fuente, A; Fuentes, I; de Toro, F J; Blanco, F J

    2010-11-01

    In this study we analyzed the chondrogenic potential of subpopulations of mesenchymal stem cells (MSCs) derived from human synovial membranes enriched for CD73, CD106, and CD271 markers. Subpopulations of human synovial membrane MSCs enriched for CD73, CD106, and CD271 markers were isolated using a cytometry sorter and characterized by flow cytometry for MSC markers. The expression of Sox9, Nanog, and Runx2 genes by these cells was measured by reverse transcriptase-polymerase chain reaction. The chondrogenesis of each subpopulation was assessed by culturing the cells in a defined medium to produce spontaneous spheroid formation and differentiation towards chondrocyte-like cells. The examination of the spheroids by histological and immunohistochemical analyses for collagen type II (COL2), aggrecan, collagen type I (COL1), metalloprotease 13 (MMP13), and collagen type X (COLX) levels were performed to assess their chondrogenesis capacity. The adipogenesis and osteogenesis potential of each subpopulation was determined using commercial media; the resulting cells were stained with oil red O or red alizarin to test the degree of differentiation. The subpopulations had different profiles of cells positive for the MSC markers CD44, CD69, CD73, CD90, and CD105 and showed different expression levels of the genes Sox9, Nanog, and Runx2 involved in chondrogenesis, undifferentiation, and osteoblastogenesis, respectively. Immunohistochemical analysis demonstrated that COL1, COL2, COLX, MMP13, and aggrecan were expressed in the spheroids as soon as 14 days of culture. The CD271(+) subpopulation expressed the highest levels of COL2 staining compared to the other subpopulations. CD105 and Runx2 were shown by immunohistochemistry and genetic analysis to have significantly higher expression CD271(+) subpopulation than the other subpopulations. Spheroids formed from CD271-enriched and CD73-enriched MSCs from normal human synovial membranes mimic the native cartilage extracellular

  16. Teratogenic effects of Origanum Vulgare extract in mice fetals

    Directory of Open Access Journals (Sweden)

    Iraj Ragerdi Kashani

    2013-11-01

    Full Text Available Background: A number of studies on reproduction have mentioned Origanum Vulgare extract’s ability to reduce mortality rates and improve fertility rates. However, other studies have suggested that it is possible to use Origanum Vulgare extract to induce abortion. The aim of this study was to investigate the effect of different doses of Origanum Vulgare on embryo survival and macroscopic abnormalities in mice.Methods: In this study, 24 mice Balb/c female weighting approximately 25-30 g were divided into 4 groups. Origanum Vulgare extract was prepared; different concentrations (2.5, 12.5, and 25 mg in 0.25 ml distilled water were administered, by oral gavage, to three experimental groups of mice between day 6 (starting gastrulation until day 15 of pregnancy (end of organogenesis. The control group consisted of six mice that received 0.25 ml of distilled water daily. On day 16 of study, pregnant mice were anesthetized by chloroform and fetuses were removed and stained with Alcian Blue, Alizarin Red s and microwave irradiation. Morphological and skeletal abnormalities were investigated by light and stereomicroscopes.Results: The results of this study showed that high doses of the Origanum Vulgare extract significantly decreased the mean number of embryos (100.5, P>0.05, mean number of live embryos (70.5, P>0.05 in each mouse and resulted in significant reduction in mean weight(11848 mg, P>0.05 and crown-rump length(11.90.23 mm, P>0.05 and the overall size of fetuses compared to control group, whereas there was no significant difference between the groups receiving low dose of Origanum Vulgare extract with control group. In addition, under the effect of the Origanum Vulgare extract the subcutaneous bleeding seemed (20.1, P>0.05 significantly more frequent compared to the control group. Conclusion: Origanum Vulgare extract did not have any positive effect on fetal development; and high dosages led to an increased incidence rate of

  17. Effects of Mechanical Stimulation on Proliferation and Differentiation in MG-63 Osteoblast-like Cells%力学刺激对MG-63成骨样细胞增殖分化的影响

    Institute of Scientific and Technical Information of China (English)

    杨敏; 黄凌云; 肖立伟; 廖二元

    2012-01-01

    This paper is aimed to explore the effects oi mechanical stimulation on proliferation and differentiation of osteoblasts. Cultured MG-63 osteoblast-like cells were strained by the four-point bending cell mechanics loader. In the study, we observed the effects of different magnitudes and duration of mechanical strain on the markers of proliferation and differentiation in osteoblasts. The protein levels and Alkaline phosphatase (ALP) activity were determined by western blot and a-nitrophenyl phosphate assay respectively. The mineralization nodules were stained using Alizarin Red-S method. We found: (1) the expression of proliferating cell nuclear antigen (PCNA), ALP activity in strained group were significantly increased compared to those in the control group, but the role did not increase with the increase of the magnitude of the stimulation; and (2) under appropriate stimulation (2000μstrain), the expression of PCNA, COL I protein and ALP activity increased gradually with the increase of loading time, and appropriate stimulation promoted the formation of mineralization nodules. It indicated that appropriate mechanical stimulation could promote proliferation and differentiation of osteoblasts.%为探讨力学刺激对成骨细胞增殖分化的影响,应用根据四点弯曲原理设计的加力装置对MG-63成骨样细胞进行力学刺激,观察不同大小和作用时间的机械应变对成骨细胞增殖分化的影响.本研究中蛋白表达、碱性磷酸酶(ALP)活性测定分别采用Western blot、α-磷酸奈酚法,采用茜素红染色观察矿化结节的形成.结果发现:(1)与对照组相比,加力组增殖细胞核抗原(PCNA)蛋白表达、ALP活性均明显增加,但这种作用并不随着力刺激大小的增加而增加.(2)在适宜的力刺激(2000 μstrain)下,随着加力时间的增加,PCNA、Ⅰ型胶原(COLⅠ)蛋白表达及ALP活性逐渐增加,并且适宜的力刺激也促进了矿化结节的形成.该结果提示适宜的力学

  18. 二甲双胍减轻糖基化终末产物对成骨细胞功能损害的观察%Metformin ameliorating the deleterious effects of advanced glycation end products on osteoblastic cells

    Institute of Scientific and Technical Information of China (English)

    甄东户; 汤旭磊; 成建国; 韩婕; 刘丽娟; 傅松波

    2012-01-01

    Objective To explore the effect of metformin on proliferation, differentiation and mineralization in the advanced glycation end products(AGEs)-induced rat cranium osteoblasts. Methods Rat cranium osteoblasts were isolated and cultured. The proliferation of osteoblasts was assayed by MTT method, the activity of alkaline phosphatase(ALP) was measured by biochemical method, the number of mineralized nodules was assessed by Alizarin red S staining, and the calcium deposition in the mineralized nodules was detected by hydrochloric acid decalcification method. Results With 500 fig/ml AGEs, the cellular proliferation, ALP activity, number of mineralized nodules, and calcium deposition were reduced significantly. Metformine (100-500 μmol/L) increased the cellular proliferation and ALP activity, and promoted the formation of mineralized nodules and calcium deposition in both control and AGEs groups. Conclusion AGEs inhibit the proliferation, differentiation and mineralization of the primary osteoblasts. Metformine strengthens the osteogenesis of osteoblasts and relieves the deleterious effect of AGEs on osteoblasts.%目的 观察二甲双胍对晚期糖基化终末产物(AGEs)诱导的大鼠颅骨成骨细胞增殖、分化、矿化的影响.方法 分离培养大鼠颅骨成骨细胞,四氮唑蓝(MTT)比色分析法测定细胞增殖,生化法测定碱性磷酸酶(ALP)活性、茜素红S钙染法检测矿化结节形成,盐酸脱钙法检测矿化结节中钙含量.结果 500μg/ml AGEs抑制成骨细胞增殖、ALP活性、钙化结节形成、钙沉积;给予二甲双胍(100~500 μmol/L)可不同程度上提高成骨细胞数量和ALP活性,促进矿化结节形成及钙沉积,减轻AGEs对成骨细胞增殖、ALP活性、钙化结节形成及钙沉积的抑制.结论 AGEs对原代成骨细胞增殖、分化与矿化产生抑制作用,二甲双胍提高成骨细胞的成骨能力,减轻AGEs对成骨细胞功能的损害.

  19. Preparation of nano b-tricalcuim phosphate, gelatin and velvet antler polypeptide composites and its effect on the proliferation, differentiation and mineralization in hFOB1.19 cells%纳米磷酸三钙/明胶/鹿茸多肽复合材料的制备及对人成骨细胞增殖、分化及矿化的影响

    Institute of Scientific and Technical Information of China (English)

    刘晓峰; 张郑瑶; 田秀丽; 张瑾; 史祺云; 胡进平; 郭燕川; 邓旭明; 周秋丽

    2013-01-01

    探讨反相微乳液法制备的β-纳米磷酸三钙/明胶/鹿茸多肽复合材料(简称纳米复合材料)对人成骨细胞功能的影响.利用反相微乳液法制备纳米复合材料,扫描电镜观察其形态,EDX与SELDI-TOF-MS分析对复合材料进行表征.采用MTT比色法、茜素红染色法检测纳米复合材料对体外培养的人成骨细胞hFOB1.19的增殖、矿化结节形成的影响;采用全自动生化分析仪检测hFOB细胞碱性磷酸酶活性(ALP).结果表明该纳米复合材料可促进人成骨细胞增殖,提高ALP活性并增加矿化结节形成的数量,可促进人成骨细胞的增殖、分化成熟及矿化.%Abstract:The nano tricalcium phosphate/ gelatin/velvet antler polypeptide (VAP)composite materials (referred as nanocomposites) were prepared by water-in-oil (W/O) emulsion process.The effect of nanocomposites on human osteoblasts was studied in hFOB1.19 cell lines.Nanocomposites were prepared through emulsification-coacervation method in water-in-oil emulsions.The morphology was observed by scanning electron microscope.Characterization of composite spheres was examined by EDX and SELDI-TOF-MS.MTT assay and alizarin red staining were employed to detect the effects of nanocomposites on the proliferation and formation of mineralization nodules formation in hFOB cells,respectively.The alkaline phosphatase(ALP) activity was determined by automatic clinical chemistry analyzer.The results showed that nanocomposites could stimulate human osteoblasts cells proliferation increase ALP activity,and promote the formation of mineralization nodules.

  20. Potential of l-thyroxine to differentiate osteoblast-like cells via Angiopoietin1.

    Science.gov (United States)

    Park, See-Hyoung; Lee, Jongsung; Kang, Mi-Ae; Moon, Young Jae; Wang, Sung Il; Kim, Kyoung Min; Park, Byung-Hyun; Jang, Kyu Yun; Kim, Jung Ryul

    2016-09-23

    Angiogenesis is closely associated with osteoblast differentiation. Previously, we demonstrated that bone formation can be accelerated by treatment with COMP-Angiopoietin1, a known angiogenic factor. Angiopoietin1 (Ang1) is a specific growth factor that generates stable and mature vasculature through the Tie2 receptor. In this study, we aimed to identify a novel drug that can activate endogenous Ang1 expression as a pharmacological treatment for bone formation. Therefore, Ang1 expression was examined in U2OS osteoblast-like cells treated with 770 drugs from a library of Food and Drug Administration (FDA)-approved drugs by using ELISA for Ang1. l-thyroxine was selected as a novel drug candidate. l-Thyroxine is a synthetic form of the hormone thyroxine, which is used to treat patients with hypothyroidism. Enzyme-linked immunosorbent assays (ELISAs) were performed to test whether Ang1 is induced in a dose-dependent manner in human osteoblast-like cell lines, U2OS and MG63. The effects of l-thyroxine on osteoblast differentiation and mineralization were evaluated by alkaline phosphatase (ALP) activity and Alizarin red s staining. To determine the molecular mechanism, the expression of proteins related to bone formation and differentiation, such as type I collagen (COL1A1), osteocalcin (OC), bone sialoprotein (BSP), distal-less homeobox 5 (Dlx5), Runt-related transcription factor 2 (Runx2), osterix (OSX), and ALP, was tested by Western blotting analysis. Consequently, l-thyroxine induced Ang1 expression in a dose-dependent manner in both U2OS and M63 cells, which was confirmed by ELISA and Western blotting. Also, l-thyroxine activated ALP activity in U2OS and MG63 cells as well as ALP expression. Furthermore, l-thyroxine enhanced the expression of COL1A1, Runx2, OC, BSP, Dlx5, and OSX mRNA and proteins. Taken together, we demonstrated that l-thyroxine increased Ang1 expression and induces bone formation, differentiation, and mineralization in U2OS and MG63 cell lines

  1. Preparation, preservation, and morphological evaluation of the donor graft for descemet membrane endothelial keratoplasty: an experimental study

    Institute of Scientific and Technical Information of China (English)

    Sun Yiqian; Peng Rongmei; Hong Jing

    2014-01-01

    Background Though there have been various methods for harvesting and preserving descemet membrane (DM) and intact endothelium,there is no literature about the morphological evaluation of endothelium after graft preparation for descemet membrane endothelial keratoplasty (DMEK).The aim of this study was to establish and improve a simple method for preparing,preserving,and morphologically evaluating the donor graft for DMEK.Methods To obtain a donor graft,an air bubble was formed by injecting a 29 G needle with 1 ml sterile air into a small edge created outside the Schwalbe line.Another needle was inserted into the bubble through the stroma to aspirate the air or replace half the air with organ culture medium.Trypan blue was used to mark the location for small incision to improve the success rate.Frozen sections were stained with hematoxylin and eosin (HE).Based on the air bubble,DM grafts were divided into four groups:group A (normal control),graft without any operative technique; group B,graft with zero-pressure air bubble; group C,graft with full-pressure air bubble; group D,graft with half-pressure air bubble.The four groups of grafts were preserved for 24 hours to observe the effect of bubbles on cells.The gross and ultrastructure morphologies were evaluated using alizarin red and scanning electron microscopy (SEM),respectively.Results Donor grafts were harvested via the air bubble technique,facilitated by prior trypan blue staining.HE-stained sections revealed a pure graft without stroma.There were no significant changes under light microscope.In group A,SEM revealed a confluent layer of polygonal endothelium with distributed microvilli exhibiting characteristics of interdigitating junctions.In group B,intercellular borders became thinner.In group C,interdigitations were almost flat and microvilli were observed less frequently.In group D,other than less microvilli,there were minimal changes.Conclusions The donor graft preparation method appears to be effective

  2. Effects of demineralized dentin matrix on the odontogenic differentiation potential of bone marrow mesenchymal stem cells in vitro%脱矿牙本质基质诱导骨髓间充质干细胞向成牙本质样细胞分化的体外实验研究

    Institute of Scientific and Technical Information of China (English)

    张红梅; 张晓梅; 闫志伟; 王江; 荀文兴; 李蓉; 庞楠; 杨海珍; 金岩

    2011-01-01

    AIM; To observe the adhesion and growth of rat bone marrow mesenehymal stem ceils (rBMSCs)seeded on demineralized dentin matrix (DDM) in vitro and the odontogenic induction of DDM. METHODS; BMSCs were successfully isolated and cultured. The compatibility of the biomaterial was evaluated by MTT assay. The odontogenic differentiation of BMSCs induced by DDM was measured using Alizarin red staining, the alkaline phosphatase (ALP) activity and reverse transcription-polymerase chain reaction (RT-PCR) assay. RESULTS; The proliferation of BMSCs in DDM groups was obviously higher than that in the control groups. The higher levels of ALP activity and larger mineralizing nodes were also detected in the experiment groups. RT-PCR demonstrated mRNA expression of dentin sialophospho-protein ( DSPP ) and dentin matrix proteinl ( DMP-1) in BMSCs induced by DDM. CONCLUSION; DDM possesses a good cellular compatibility with BMSCs. It can induce the odontogenic differentiation of BMSCs.%目的:观察脱矿牙本质基质(demineralized dentin matrix,DDM)与大鼠骨髓间充质干细胞(rat bone marrow mesenehymal stem ceils,rBMSCs)的生物相容性及其作为支架材料对BMSCs的牙向诱导作用.方法:分离培养BMSCs,分别用四唑盐比色法(MTS)、茜素红染色法、碱性磷酸酶(alkaline phosphatase,ALP)和逆转录聚合酶链式反应(reverse transcription-polymerase chain reaction,RT-PCR)方法观察DDM生物材料对BMSCs增殖活性和牙向分化能力的影响.结果:DDM能显著促进体外培养的BMSCs的增殖、诱导细胞的矿化和提高细胞ALP活性;同时还能诱导BMSCs在mRNA水平表达牙本质涎磷蛋白(dentin sialophosphoprotein,DSPP)和牙本质基质蛋白(dentin matrix proteinl,DMP-1).结论:体外培养条件下,DDM与BMSCs有良好的生物相容性,能够诱导BMSCs牙向分化.

  3. 小肠黏膜下层对牙周膜干细胞体外增殖、骨向分化的影响%Effects of small intestinal submucosa on proliferation and osteogenic differentiation of human periodontal ligament stem cells in vitro

    Institute of Scientific and Technical Information of China (English)

    张红梅; 荀文兴; 王江; 张晓梅; 李蓉

    2014-01-01

    目的:观察小肠黏膜下层(SIS)与人牙周膜干细胞(PDLSCs)的生物相容性及其作为支架材料对 PDLSCs 的骨向分化诱导作用。方法体外分离培养 PDLSCs,分别用四唑盐比色法(MTS)、茜素红染色法、碱性磷酸酶(ALP)和逆转录聚合酶链式反应(RT -PCR)方法观察 SIS 生物材料对 PDLSCs 增殖活性和骨向分化能力的影响。结果SIS 显著地刺激体外培养的PDLSCs 增殖,诱导了细胞的矿化和提高细胞 ALP 活性。RT -PCR 结果显示 SIS 诱导后细胞 mRNA 水平表达骨涎蛋白(BSP)和骨钙素(OCN)。结论体外培养条件下,SIS 与 PDLSCs 有良好的生物相容性,能够诱导 PDLSCs 骨向分化。%Objective To observe the adhesion and growth of human periodontal ligament stem cells (PDLSCs)seeded on small in-testinal submucosa (SIS)in vitro and the osteogenic induction of SIS.Methods PDLSCs were successfully isolated and cultured. The compatibility of the biomaterial was evaluated by MTS assay.The osteogenic differentiation of PDLSCs induced by SIS was meas-ured by using Alizarin red staining,the alkaline phosphatase (ALP)activity and reverse transcription-polymerase chain reaction (RT-PCR)assay.Results The proliferation of PDLSCs in SIS groups was obviously higher than that in the control groups.The higher lev-els of ALP activity,the larger mineralizing nods were also detected in the experiment groups.RT-PCR demonstrated mRNA expression of bone sialoprotein (BSP)and osteocalcin (OCN)in PDLSCs induced by SIS.Conclusions SIS possesses a good cellular compati-bility with PDLSCs.It can induce the osteogenic differentiation of PDLSCs.

  4. Effects of demineralized dentin matrix on proliferation and odontogenic differentiation of human dental pulp stem cells in vitro%脱矿牙本质基质对人牙髓干细胞体外增殖、牙向分化能力的影响

    Institute of Scientific and Technical Information of China (English)

    张红梅; 金岩; 王永刚; 张晓梅; 谭家莉; 荀文兴; 杨海珍; 李蓉; 庞楠; 曾光

    2012-01-01

    目的 观察脱矿牙本质基质(demineralized dentin matrix,DDM)与人牙髓干细胞(human dental pulp stem cells,hDP-SCs)的生物相容性及其作为支架材料的牙向诱导作用.方法 体外分离培养DPSCs,分别用四唑盐比色法(MTS)、茜素红染色法、碱性磷酸酶(alkaline phosphatase,ALP)和逆转录聚合酶链式反应(reverse transcription- polymerase chain reaction,RTPCR)方法观察DDM生物材料对DPSCs增殖活性和牙向分化能力的影响.结果 DDM显著地刺激体外培养的DPSCs增殖,诱导了细胞的矿化和提高细胞ALP活性.RT- PCR结果显示DDM诱导后细胞mRNA水平表达牙本质涎磷蛋白(dentin sialophosphoprotein,DSPP)和牙本质基质蛋白(dentin matrix protein1,DMP -1).结论 体外培养条件下,DDM与DPSCs有良好的生物相容性,作为支架材料对于DPSCs有较好牙向诱导作用.%Objective To observe the adhesion and growth of human dental pulp stem cells seeded on demineralized dentin matrix ( DDM) in vitro and the adontogenic induction of DOM. Methods OPSCs were successfully isolated and cultured. The compatibility of the biomaterial was evaluated by MTS assay. The odontogenic differentiation of DPSCs induced by DDM was measured using Alizarin red staining, the alkaline phosphatase (ALP) activity and reverse transcription-polymerase chain reaction (RT-PCR) assay. Results The proliferation of DPSCs in DDM groups was obviously higher than that in the control groups. The higher levels of ALP activity and larger mineralizing nods were also detected in the experimental groups. RT-PCR demonstrated mRNA expression of dentin sialophosphoprotein ( DSPP ) and dentin matrix protein 1 (DMP-1) in DPSCs induced by DDM. Conclusions DDM possesses a good cellular compatibility with DPSCs, As an engineering scaffold.it can induce the odontogenic differentiation of DPSCs.

  5. 氯化锂对人颌骨来源的骨髓间充质干细胞增殖及骨向分化能力的影响%Effects of lithium chloride on the proliferation and osteogenic differentiation of human orofacial bone marrow mesenchymal stem cells

    Institute of Scientific and Technical Information of China (English)

    高丽娜; 安莹; 杨昊; 陈发明; 金岩

    2013-01-01

    Objective: To investigate the effects of lithium chloride ( LiCl) on the proliferation and osteogenic differentiation of human orofacial-derived bone marrow mesenchymal cells ( OF-BMMSCs) in vitro. Methods: Human OF-BMMSCs were isolated and cultured by limited dilution method. The cloned OF-BMMSCs were stimulated with LiCl at different concentrations(0, 5, 10, 20, 40 mmol/L) respectively. After 3 days stimulation, proliferation of OF-BMMSCs was examined by MTT assay. The osteogenic differentiation of OF-BMMSCs was evaluated by Alizarin red staining, ALP staining and real-time quantitative PCR for the mRNA expression of ALP, Runx-2 and OCN after treatment by LiCl and followed by osteogenic induction. Results: LiCl at 5 mmol/L promoted the proliferation of OF-BMMSCs(P 0.05). However, LiCl at 20 and 40 mmol/L inhibited the cells proliferation (P >0.05) , while, promoted the mRNA expression of the genes(P<0.05). Conclusion: LiCl may promote the osteogenic differentiation of OF-BMMSCs.%目的:探讨氯化锂(LiCl)对颌骨来源的骨髓间充质干细胞(OF-BMMSCs)增殖及骨性分化的影响.方法:采用有限稀释法克隆化培养人颌骨来源的骨髓间充质干细胞;正常培养基中加入不同浓度LiCl(5、10、20、40 mmol/L),同时设立对照组(0 mmol/L);运用MTT法分析不同浓度LiC1对人OF-BMMSCs增殖活性的影响;采用组织化学染色法、RT-PCR法测定不同浓度LiC1处理人OF-BMMSCs后的ALP活性及相关骨向分化基因(ALP,Runx-2,OCN)的表达.结果:5 mmol/L LiCl促进OF-BMMSCs增殖,对ALP、Runx-2、OCN表达的影响作用不明显(P>0.05),20和40 mmol/L LiC1抑制细胞增殖(P<0.05),促进ALP、Runx-2、OCN基因表达(P<0.05).结论:LiCl能够促进人OF-BMMSCs的骨向分化.

  6. 模拟微重力对人牙髓干细胞-PLGA复合物矿化的影响%Mineralization of Human Dental Pulp Stem Cells on PLGA Scaffolds in Simulated Microgravity Environments

    Institute of Scientific and Technical Information of China (English)

    费晓磊; 张巍巍; 李艳萍; 潘爽; 牛玉梅

    2013-01-01

    目的:研究模拟微重力(SMG)对人牙髓干细胞(hDPSCs)在聚乳酸-羟基乙酸(PLGA)支架上矿化的影响.方法:采用酶消化法分离、培养人牙髓干细胞,并进行鉴定.hDPSCs常规接种于PLGA支架,24 h后,模拟微重力环境和普通环境下分别矿化诱导.矿化诱导第3、5、7、10、14天时进行碱性磷酸酶活性检测,第21天时进行茜素红染色以观察人牙髓干细胞在PLGA支架上矿化结节形成情况.结果:模拟微重力环境下的碱性磷酸酶活性明显低于普通环境(P<0.01),茜素红染色显示模拟微重力环境下形成的矿化结节较普通环境下的小且数量少.结论:模拟微重力环境抑制hDPSCs在PLGA支架上的矿化.%Objective: To investigate the effect of Simulated Microgravity on mineralization of human dental pulp stem cells (hDPSCs) on PLGA (Poly lactic acid glycolic acid) scaffolds.Methods: The hDPSCs were inoculated on PLGA scaffolds for 24h after routine isolation, culture and identification, then cultured in simulated microgravity environment or conventional mineralization-induced environment, respectively.Alkaline phosphatase testing by 3, 5, 7, 10, 14 days and Alizarin red staining by 21 days were detected.Results: ALPase testing demonstrated lower ALPase activity in simulated microgravity environment at various time points compared with the control group.A-lizarin red staining showed positive granules were hardly detected in simulated microgravity group, however, noted in the control group.Conclusion: Together, these results indicate that simulated microgravity inhibits mineralization of hDPSCs on PLGA scaffolds.

  7. Biological Behavior of Human Periodontal Ligament Stem Cells in Simulated Microgravity Environment MA%模拟微重力培养环境下牙周膜干细胞生长状态的研究

    Institute of Scientific and Technical Information of China (English)

    马兆峰; 李石; 牛忠英

    2011-01-01

    Objective To investigate the growth status of human periodontal ligament stem cells (hPDLSCs) in simulated microgravity in vitro. Methods HPDLSCs were isolated and cultivated, then characterized by immunohistochemis-try of stromal cell antigen-1 ( STRO-1). After 21 days of induction, the results were evaluated by Alizarin red staining and oil' 0' staining. HPDLSCs were co-incubated with microcarrier beads of Cytodex-3, and were placed in rotary cell culture system. Cells morphology and proliferation potential were examined. Results HPDLSCs were cultivated, and growth characteristics and multipotent differentiation were assessed. The results showed that hPDLSCs can be cultured in simulated microgravity environment. On the 1st day (t =5. 590, P =0. 005), the 3rd day ( t = 12. 238, P =0.000) , the 5th day (t = 19.124, P = 0.000), the 7th day (t=35. 103, P =0.000), simulated microgravity statistically promoted the proliferation potential compared with cells in normal gravity environment. Conclusion The simulated microgravity culture system has the potential to be used for the bioengineering reconstruction of the periodontal tissues.%目的 探讨模拟微重力培养体系下人牙周膜干细胞(human periodontal ligament stem cells,HPDLSCs)的生长特点.方法 在体外用有限稀释法克隆化生长获得HPDLSCs,接种于葡聚糖微载体,观察在旋转微重力细胞培养环境下与普通重力环境下细胞生长状态的差异.结果 利用克隆生长法成功获取具备多向分化潜能的HPDLSCs,在微重力环境下的微载体表面细胞多呈半球形,少数铺展为不规则扁平形或长梭形,与普通重力环境相比,细胞生长速度明显加快.结论 三维微重力培养环境可以迅速获得大量的HPDLSCs,为构建工程化牙周组织奠定了实验基础.

  8. Critical evaluation of branch polarity and apical dominance as dictators of colony astogeny in a branching coral.

    Directory of Open Access Journals (Sweden)

    Lee Shaish

    Full Text Available The high morphological resemblance between branching corals and trees, can lead to comparative studies on pattern formation traits, best exemplified in plants and in some cnidarians. Here, 81 branches of similar size of the hermatypic coral Stylophora pistillata were lopped of three different genets, their skeletons marked with alizarin red-S, and divided haphazardly into three morphometric treatment groups: (I upright position; (II horizontal position, intact tip; and (III horizontal position, cut tip. After 1 y of in-situ growth, the 45 surviving ramets were brought to the laboratory, their tissues removed and their architectures analyzed by 22 morphological parameters (MPs. We found that within 1 y, isolated branches developed into small coral colonies by growing new branches from all branch termini, in all directions. No architectural dissimilarity was assigned among the three studied genets of treatment I colonies. However, a major architectural disparity between treatment I colonies and colonies of treatments II and III was documented as the development of mirror structures from both sides of treatments II and III settings as compared to tip-borne architectures in treatment I colonies. We did not observe apical dominance since fragments grew equally from all branch sides without documented dominant polarity along branch axis. In treatment II colonies, no MP for new branches originating either from tips or from branch bases differed significantly. In treatment III colonies, growth from the cut tip areas was significantly lower compared to the base, again, suggesting lack of apical dominance in this species. Changes in branch polarity revealed genet associated plasticity, which in one of the studied genets, led to enhanced growth. Different genets exhibited canalization flexibility of growth patterns towards either lateral growth, or branch axis extension (skeletal weight and not porosity was measured. This study revealed that colony

  9. Advanced glycation end products promote human aortic smooth muscle cell calcification in vitro via activating NF-κB and down-regulating IGF1R expression

    Institute of Scientific and Technical Information of China (English)

    Yi WANG; Zhen-yu ZHANG; Xiao-qing CHEN; Xiang WANG; Heng CAO; Shao-wen LIU

    2013-01-01

    Aim:To investigate the effects of advanced glycation end products (AGEs) on calcification in human aortic smooth muscle cells (HASMCs) in vitro and the underlying mechanisms.Methods:AGEs were artificially prepared.Calcification of HASMCs was induced by adding inorganic phosphate (Pi,2 mmol/L) in the media,and observed with Alizarin red staining.The calcium content in the supernatant was measured using QuantiChrome Calcium Assay Kit.Expression of the related mRNAs and proteins was analyzed using real-time PCR and Western blot,respectively.Chromatin immunoprecipitation (ChIP) assay was used to detect the binding of NF-κB to the putative IGF1R promoter.Results:AGEs (100 μg/mL) significantly enhanced Pi-induced calcification and the levels of osteocalcin and Cbfα1 in HASMCs.Furthermore,the treatment decreased the expression of insulin-like growth factor 1 receptor (IGF1R).Over-expression of IGF1R in HASMCs suppressed the AGEs-induced increase in calcium deposition.When IGF1R expression was knocked down in HASMCs,AGEs did not enhance the calcium deposition.Meanwhile,AGEs time-dependently decreased the amounts of IκBα and Flag-tagged p65 in the cytoplasmic extracts,and increased the amount of nuclear p65 in HASMCs.In the presence of NF-κB inhibitor PDTC (50 μmol/L),the AGEs-induced increase in calcium deposition was blocked.Over-expression of p65 significantly enhanced Pi-induced mineralization,but suppressed IGF1R mRNA level.Knockdown of p65 suppressed the AGEs-induced increase in calcium deposition,and rescued the IGF1R expression.The ChIP analysis revealed that NF-κB bound the putative IGF1R promoter at position-230 to-219 bp.The inhibition of IGF1R by NF-κB was abolished when IGF1R reporter plasmid contained mutated binding sequence for NF-κB or an NF-κB reporter vector.Conclusion:The results demonstrate that AGEs promote calcification of human aortic smooth muscle cells in vitro via activation of NF-κB and down-regulation of IGF1R expression.

  10. 鱿鱼皮胶原蛋白水解肽对镉抑制MC3T3-E1增殖、分化及钙化的影响%Effect of Collagen Peptide Extracted from Dosidicus gigas Skin on Proliferation, Differentiation and Calcification of MC3T3-E1 Cell Induced by Cd

    Institute of Scientific and Technical Information of China (English)

    蔡江佳; 李晔; 全晶晶; 蔺佳良; 张云云; 王峰; 苏秀榕

    2015-01-01

    目的:探究秘鲁鱿鱼皮胶原蛋白水解肽增强MC3T3-E1细胞抗骨质疏松的作用.方法:将培养的MC3T3-E1细胞分为正常对照组、氯化镉损伤组和胶原蛋白水解肽干预组;利用MTT法确定氯化镉的半数抑制浓度,建立细胞损伤模型;根据各组细胞增殖率差异,确定最佳胶原蛋白水解肽的添加量;通过细胞周期、凋亡,碱性磷酸酶活性的测定及Alizarin red染色法分析胶原蛋白水解肽在细胞增殖、分化、钙化阶段拮抗氯化镉的抑制作用.结果:与氯化镉损伤组相比,胶原蛋白水解肽干预组细胞活性升高(P<0.01);处于G1期的细胞数量减小(P<0.05),S期细胞数量增大(P<0.05);凋亡率降低(P<0.05);9,12,15 d时AKP活性升高(P<0.05,P<0.01,P<0.01).结论:摄入一定剂量的氯化镉会抑制MC3T3-E1成骨细胞增殖、分化和钙化.胶原蛋白水解肽在一定程度上可改善此类损伤对MC3T3-E1细胞的影响.

  11. Effect of the HDAC inhibitor vorinostat on the osteogenic differentiation of mesenchymal stem cells in vitro and bone formation in vivo

    Institute of Scientific and Technical Information of China (English)

    Song XU; Kim DE VEIRMAN; Holly EVANS; Gaia Cecilia SANTINI; Isabelle VANDE BROEK; Xavier LELEU; Ann DE BECKER

    2013-01-01

    Vorinostat,a histone deacetylase (HDAC) inhibitor currently in a clinical phase III trial for multiple myeloma (MM) patients,has been reported to cause bone loss.The purpose of this study was to test whether,and to what extent,vorinostat influences the osteogenic differentiation of mesenchymal stem cells (MSCs) in vitro and bone formation in vivo.Methods:Bone marrow-derived MSCs were prepared from both normal donors and MM patients.The MSCs were cultured in an osteogenic differentiation induction medium to induce osteogenic differentiation,which was evaluated by alkaline phosphatase (ALP) staining,Alizarin Red S staining and the mRNA expression of osteogenic markers.Naive mice were administered vorinostat (100 mg/kg,ip) every other day for 3 weeks.After the mice were sacrificed,bone formation was assessed based on serum osteocalcin level and histomorphometric analysis.Results:Vorinostat inhibited the viability of hMSCs in a concentration-dependent manner (the IC50 value was 15.57 μmol/L).The low concentration of vorinostat (1 μmol/L) did not significantly increase apoptosis in hMSCs,whereas pronounced apoptosis was observed following exposure to higher concentrations of vorinostat (10 and 50 μmol/L).In bone marrow-derived hMSCs from both normal donors and MM patients,vorinostat (1 μmol/L) significantly increased ALP activity,mRNA expression of osteogenic markers,and matrix mineralization.These effects were associated with upregulation of the bone-specifying transcription factor Runx2 and with the epigenetic alterations during normal hMSCs osteogenic differentiation.Importantly,the mice treated with vorinostat did not show any bone loss in response to the optimized treatment regimen.Conclusion:Vorinostat,known as a potent anti-myeloma drug,stimulates MSC osteogenesis in vitro.With the optimized treatment regimen,any decrease in bone formation was not observed in vivo.

  12. 偏最小二乘法-分光光度法同时检测6种合成色素的研究%A Study on Simultaneous Determination of Six Synthetic Colorants by Partial Least Squares-Spectrophotometric Method

    Institute of Scientific and Technical Information of China (English)

    赵丽

    2015-01-01

    By means of multivariate calibration curve,a simultaneous determination of six synthetic colorants including pon⁃ceau 2R,sunset yellow,quinoline yellow,basic orange,alizarin green and diamine green B has been achieved by using the partial least squares-spectrophotometric method. Effects of primary parameters such as pH values ,additivity of the ab⁃sorbance of different components,number of principal components in the mathematic model and the wavelength range for spectrophotometric measurement have also been investigated. Under the optimal conditions,the simultaneous determina⁃tion of six synthetic colorants mentioned above was obtained and the detection limits of 0.2,0.3,0.07,0.2,2.6,2.1μg/mL were gained respectively. Applied to simulation samples,the recoveries were found in the range from 98.2%to 104.4%. The method in this work is simple and might provide a new approach to multi-component statistic analysis of these six colo⁃rants without any preliminary chemical separation.%基于多元校正曲线分辨技术,建立了立春红2R、日落黄、喹啉黄、碱性橙、酸性绿和直接绿B等6种合成色素的偏最小二乘法-分光光度法分析方法。研究了溶液pH值、吸光度加和性、计量模型主成分数及计量波段等主要因素的影响,实现了立春红2R等6种合成色素的偏最小二乘法-分光光度法同时分析,检测限分别为0.2、0.3、0.07、0.2、2.6、2.1μg/mL,测定相对标准偏差RSD小于3%;应用于样品分析,立春红2R等合成色素的加标回收率在98.2%~104.4%范围内。所建立的方法简单、快速,为分光光度法不经分离而直接测定立春红2R等合成色素提供一种新的途径。

  13. Compound soft regenerated skull material for repairing dog skull defects using bone morphogenetic protein as an inductor and nanohydroxyapatite as a scaffold

    Institute of Scientific and Technical Information of China (English)

    Zhidong Shi; Mingwang Liu; Zhongzong Qin; Qinmei Wang; Ying Guo; Haiyong He; Zhonghe Yu

    2008-01-01

    BACKGROUND: In previous studies of skull defects and regeneration, bone morphogenetic protein as an inductor and nanohydroxyapatite as a scaffold have been cocultured with osteoblasts.OBJECTIVE: To verify the characteristics of the new skull regenerated material after compound soft regenerated skull material implantatiom.DESIGN, TIME AND SETTING: The self-control and inter-group control animal experiment was perfurmed at the Sun Yat-sen University, China from February to July 2007.MATERIALS: Twenty-tour healthy adult dogs of both genders weighing 15-20 kg were used in this study. Nanohydroxyapatite as a scaffold was cocultured with osteoblasts. Using demineralized canine bone matrix as a carrier, recombinant human bone morphogenetic protein-2 was employed to prepare compound soft regenerated skull material. Self-designed compound soft regenerated skull material was implanted in models of skull defects.METHODS: Animals were randomly assigned into two groups, Group A (n = 16) and Group B (n = 8).Bilateral 2.5-cm-diameter full-thickness parietal skull defects were made in all animals. In Group A, the right side was reconstructed with calcium alginate gel, osteoblasts, and nanomcter bone meal composite;the left side was reconstructed with calcium alginate gel, osteoblasts, nanometer bone meal and recombinant human bone morphogenetic protein-2 composite. In Group B, the right side was kept as a simple skull detect, and the left side was reconstructed with calcium alginate gel, osteoblasts, nanometer bone meal and recombinant human bone morphogenetic protein-2 composite.MAIN OUTCOME MEASURES: Bone regeneration and histopathological changes at the site of the skull defect were observed with an optical microscope and a scanning electron microscope after surgery.The ability to form bone was measured by alizarin red S staining. In vitro cultured osteoblasts were observed for morphology.RESULTS: One month following surgery, newly formed bone trabeculae mostly covered the

  14. Study of the embryofeto-toxicity of Crown-of-Thorns (Euphorbia milii latex, a natural molluscicide

    Directory of Open Access Journals (Sweden)

    Souza C.A.M.

    1997-01-01

    Full Text Available The crude latex of Crown-of-Thorns (Euphorbia milii var. hislopii is a potent plant molluscicide and a promising alternative to the synthetic molluscicides used in schistosomiasis control. The present study was undertaken to investigate the embryofeto-toxic potential of E. milii latex. The study is part of a comprehensive safety evaluation of this plant molluscicide. Lyophilized latex (0, 125, 250 and 500 mg/kg body weight in corn oil was given by gavage to Wistar rats (N = 100 from days 6 to 15 of pregnancy and cesarean sections were performed on day 21 of pregnancy. The numbers of implantation sites, living and dead fetuses, resorptions and corpora lutea were recorded. Fetuses were weighed, examined for external malformations, and fixed for visceral examination, or cleared and stained with Alizarin red S for skeleton evaluation. A reduction of body weight minus uterine weight at term indicated that E. milii latex was maternally toxic over the dose range tested. No latex-induced embryolethality was noted at the lowest dose (125 mg/kg but the resorption rate was markedly increased at 250 mg/kg (62.5% and 500 mg/kg (93.4%. A higher frequency of fetuses showing signs of delayed ossification (control: 17.4%; 125 mg/kg: 27.4% and 250 mg/kg: 62.8%; P<0.05 vs control indicated that fetal growth was retarded at doses ³ 125 mg latex/kg body weight. No increase in the proportion of fetuses with skeletal anomalies was observed at the lowest dose but the incidence of minor skeletal malformations was higher at 250 mg/kg body weight (control: 13.7%; 125 mg/kg: 14.8%; 250 mg/kg: 45.7%; P<0.05 vs control. Since a higher frequency of minor malformations was noted only at very high doses of latex which are embryolethal and maternally toxic, it is reasonable to conclude that this plant molluscicide poses no teratogenic hazard or, at least, that this possibility is of a considerably low order of magnitude

  15. Ketoconazole- and fluconazole-induced embryotoxicity and skeletal anomalies in wistar rats: a comparative study

    Directory of Open Access Journals (Sweden)

    Vanessa Cristiane de Santana Amaral

    2008-12-01

    Full Text Available Ketoconazole and fluconazole are two broad-spectrum azole antifungals used for the treatment of superficial and systemic mycoses. Embryotoxicity and teratogenicity have been reported in some studies when those drugs are administered at high doses to pregnant rats. The aim of this study was to present a comparative study of embryotoxic effects as well as the skeletal anomalies in fetuses of Wistar rats which received ketoconazole and fluconazole at teratogenic doses on gestational days (GD 6 through 15 (organogenesis period. On gestational day (GD 21, the dams were euthanized and examined for standard parameters of reproductive outcome. Fetuses were stained with alizarin red and the bones of the head, trunk, forelimb and hindlimb were examined for detection of skeletal anomalies. The frequency of skeletal anomalies in the ketoconazole-treated group was significant when compared to the fluconazole and the control group.O cetoconazol e o fluconazol são dois antifúngicos azólicos, de amplo espectro, utilizados no tratamento de micoses superficiais e sistêmicas. Alguns estudos relatam a embriotoxicidade e teratogenicidade induzidas por estes fármacos quando os mesmos são administrados em altas doses a ratas prenhes. O objetivo deste trabalho foi apresentar um estudo comparativo dos efeitos embriotóxicos e das anomalias esqueléticas em fetos de ratas Wistar que receberam cetoconazol e fluconazol em doses teratogênicas do 6º ao 15º dia gestacional (GD (período da organogênese. No 21º GD as ratas foram eutanaziadas e examinadas quanto aos parâmetros padrões de performance reprodutiva. Os fetos foram corados com vermelho de alizarina e os ossos da cabeça, do tronco e dos membros anteriores e posteriores foram examinados para a verificação de anomalias esqueléticas. A freqüência de anomalias esqueléticas no grupo tratado com cetoconazol foi significante quando comparada à dos grupos fluconazol e controle.

  16. [Osteological development of the vertebral column and caudal complex of Lujanus guttatus (Perciformes: Lutjanidae) larvae under rearing conditions].

    Science.gov (United States)

    Rodríguez-Ibarra, Luz Estela; Abdo-de la Parra, María Isabel; Aguilar-Zárate, Gabriela; Valasco-Blanco, Gabriela; Ibarra-Castro, Leonardo

    2015-03-01

    The spotted rose snapper (Lutjanus guttatus) is an important commercial species in Mexico with good culture potential. The osteological study at early stages in this species is an important tool to confirm normal bone structure and for the detection of malformations that may occur during early development. This study was carried out in order to evaluate and describe the normal osteological development of the vertebral column and caudal complex of this species grown under controlled conditions. For this, a total of 540 larvae of L. guttatus, between 2.1 and 17.5 mm of total length (TL), were cultured during 36 days; culture conditions were 28 degrees C, 5.74 mg/L oxygen and 32.2 ups salinity with standard feeding rates. To detect growth changes, a sample of 15 organisms was daily taken from day one until day 36 of post-hatch (DPH). Samples were processed following standard techniques of clearing, and cartilage (alcian blue) and bone staining (alizarin red). Results showed that the vertebral column is composed of ten vertebrae in the abdominal region, and 14 vertebrae including the urostyle in the caudal region. The development of the axial skeleton starts with the neural arches and haemal arches at 3.8 mm TL. Caudal elements such as the hypurals and parahypural began to develop at 4.1 mm TL. Pre-flexion and flexion of the notochord and the formation of all hypurals were observed between 5.3 and 5.8 mm TL. Ossification of the vertebrae in the abdominal region and in some neural arches initiated at 9.5mm TL. In the caudal region, all the neural and haemal arches ossified at 10.2 mm TL. All the abdominal vertebrae and their respective neural arches and parapophyses ossified at 11.2 mm TL, while the elements of the caudal complex that ossified were the hypurals, parahypurals and modified haemal spines. All caudal fm rays, 12 neural spines and 3 haemal arches were ossified by 15.5 mm. The complete ossification process of this specie under laboratory culture conditions

  17. Preparation of poly(ethylene glycol/polylactide hybrid fibrous scaffolds for bone tissue engineering

    Directory of Open Access Journals (Sweden)

    Ni P

    2011-11-01

    Full Text Available PeiYan Ni, ShaoZhi Fu, Min Fan, Gang Guo, Shuai Shi, JinRong Peng, Feng Luo, ZhiYong QianState Key Laboratory of Biotherapy and Cancer Center, West China Hospital, West China Medical School, Sichuan University, Chengdu, Sichuan, People's Republic of ChinaAbstract: Polylactide (PLA electrospun fibers have been reported as a scaffold for bone tissue engineering application, however, the great hydrophobicity limits its broad application. In this study, the hybrid amphiphilic poly(ethylene glycol (PEG/hydrophobic PLA fibrous scaffolds exhibited improved morphology with regular and continuous fibers compared to corresponding blank PLA fiber mats. The prepared PEG/PLA fibrous scaffolds favored mesenchymal stem cell (MSC attachment and proliferation by providing an interconnected porous extracellular environment. Meanwhile, MSCs can penetrate into the fibrous scaffold through the interstitial pores and integrate well with the surrounding fibers, which is very important for favorable application in tissue engineering. More importantly, the electrospun hybrid PEG/PLA fibrous scaffolds can enhance MSCs to differentiate into bone-associated cells by comprehensively evaluating the representative markers of the osteogenic procedure with messenger ribonucleic acid quantitation and protein analysis. MSCs on the PEG/PLA fibrous scaffolds presented better differentiation potential with higher messenger ribonucleic acid expression of the earliest osteogenic marker Cbfa-1 and mid-stage osteogenic marker Col I. The significantly higher alkaline phosphatase activity of the PEG/PLA fibrous scaffolds indicated that these can enhance the differentiation of MSCs into osteoblast-like cells. Furthermore, the higher messenger ribonucleic acid level of the late osteogenic differentiation markers OCN (osteocalcin and OPN (osteopontin, accompanied by the positive Alizarin red S staining, showed better maturation of osteogenic induction on the PEG/PLA fibrous scaffolds at the

  18. Ca-ALC络合物和鱼精子DNA反应的电化学与光谱研究%Studies of Ca-ALC interacting with herring sperm DNA by electrochemical,fluorimetric and UV-spectrophotometric method

    Institute of Scientific and Technical Information of China (English)

    刘炜; 杨丽珠; 蒋丽萍; 李新民; 陆光汉

    2005-01-01

    In the buffer solution of (CH2)6N4(pH=6.0), Ca-ALC(alizarin complexone) complex can react with herring sperm DNA(hsDNA) to form an electrochemically non-active supermolecular complex DNA-Ca-ALC within 2 min at room temperature, which results in the maximum decrease of the peak current of Ca-ALC and a negative shift of the peak potential. The Ip is proportional to hsDNA concentration from 0.1 to 3 μg·mL-1. So this method can be used for the determination of DNA. Compared with some other methods, the procedure is simple, rapid, selective and practical. In the presence of DNA, the electrochemical parameters(the electron transfer coefficient α, the standard rate constant Ks) show significant change. In addition, by the use of fluorimetric and UV-spectrophotometric methods, it is assumed that the binding mode is intercalation.%在pH为6.0的六次甲基四胺缓冲溶液中,Ca-ALC(茜素)络合物与鱼精子DNA在室温下2 min内结合生成了非电活性的超分子化合物,导致络合物的峰电流降低和峰电位的负移,峰电流值在一定范围内与DNA的浓度成线性关系,线性范围为0.1 ~ 3 μg/mL,方法可以用于DNA的测定.和其它方法相比,该方法简便,快速,有较强的选择性和实用性.在加入DNA的条件下,电化学参数(电子转移系数α,标准速率常数Ks)显示了较大的改变.同时,结合荧光法和UV光谱法,推断反应的结合方式可能为嵌入结合.

  19. Microsphere-Based Hierarchically Juxtapositioned Biphasic Scaffolds Prepared from Poly(Lactic-co-Glycolic Acid and Nanohydroxyapatite for Osteochondral Tissue Engineering

    Directory of Open Access Journals (Sweden)

    K. T. Shalumon

    2016-12-01

    Full Text Available This study aims to prepare biphasic osteochondral scaffolds based on seamless joining of sintered polymer and polymer/ceramic microspheres for co-culture of chondrocytes and bone marrow stem cells (BMSCs. Poly(lactide-co-glycolide (PLGA microspheres and 10% nanohydroxyapatite (nHAP-incorporated PLGA (PGA/nHAP microspheres were prepared through the oil-in-water precipitation method. Virgin (V and composite (C scaffolds were prepared from 250–500 µm PLGA and PLGA/nHAP microspheres, respectively, while osteochondral (OC scaffolds were fabricated through the combination of V and C scaffolds. Physico-chemical properties of scaffolds were characterized through microscopic-spectroscopic evaluations. The effect of nHAP in scaffolds was investigated through thermogravimetric analysis and mechanical testing, while surface hydrophobicity was tested through contact angle measurements. Rabbit chondrocytes and BMSCs were used for cell culture, and cell morphology and proliferation were determined from SEM and DNA assays. Alizarin red and Alcian blue stains were used to identify the in vitro bone and cartilage tissue-specific regeneration, while cetylpyridinium chloride was used to quantitatively estimate calcium in mineralized bone. For co-culture in OC scaffolds, BMSCs were first seeded in the bone part of the scaffold and cultured in osteogenic medium, followed by seeding chondrocytes in the cartilage part, and cultured in chondrocyte medium. High cell viability was confirmed from the Live/Dead assays. Actin cytoskeleton organization obtained by DAPI-phalloidin staining revealed proper organization of chondrocytes and BMSCs in OC scaffolds. Immunofluorescent staining of bone (type I collagen and osteocalcin (OCN and cartilage marker proteins (type II collagen (COL II confirmed cellular behavior of osteoblasts and chondrocytes in vitro. Using an ectopic osteochondral defect model by subcutaneous implantation of co-cultured OC scaffolds in nude mice

  20. Fabrication and evaluation of osteoblastic differentiation of human mesenchymal stem cells on novel CaO-SiO2-P2O5-B2O3 glass-ceramics.

    Science.gov (United States)

    Lee, Jae Hyup; Seo, Jun-Hyuk; Lee, Kyung Mee; Ryu, Hyun-Seung; Baek, Hae-Ri

    2013-07-01

    Apatite-wollastonite glass-ceramics have high mechanical strength, and CaO-SiO2 -B2 O3 glass-ceramics showed excellent bioactivity and high biodegradability. A new type of CaO-SiO2 -P2 O5 -B2 O3 system of bioactive glass-ceramics (BGS-7) was fabricated, and the effect and usefulness was evaluated via bioactivity using simulated body fluid and human mesenchymal stem cells (hMSCs). The purpose of this study was to compare BGS-7 and hydroxyapatite (HA) using hMSCs in order to evaluate the bioactivity of BGS-7 and its possibility as a bone graft extender. Alkaline phosphatase (ALP) staining, ALP activity, cell proliferation 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt (MTS) assay, Alizarin Red-S (AR-S) staining, calcium levels, the mRNA expression of ALP, osteocalcin, osteopontin, and runt-related transcription factor 2 (runx-2) using reverse-transcription polymerase chain reaction (RT-PCR) and the protein expression of osteocalcin and runx-2 using Western blot were measured by transplanting hMSC onto a tissue culture plate, HA, and BGS-7. The ALP staining and AR-S staining of BGS-7 was greater than that of HA and control. The ALP value of BGS-7 was significantly higher than that of HA and control. The MTS results showed that BGS-7 had a higher value than the groups transplanted onto HA and control on day 15. The calcium level was higher than the control in both HA and BGS-7, and was especially high in BGS-7. There were more mineral products on BGS-7 than on the HA when analyzed by scanning electron microscopy. The mRNA expression of ALP, osteopontin, osteocalcin, and runx-2 were higher on BGS-7 than on HA and the control when analyzed by RT-PCR. The relative gene expression of osteopontin and runx-2 were found to be higher on BGS-7 than on HA and the control by Western blot. Accordingly, it is predicted that BGS-7 would have high biocompatibility and good osteoconductivity, and presents a possibility as a new

  1. Pathogenic role of basic calcium phosphate crystals in destructive arthropathies.

    Directory of Open Access Journals (Sweden)

    Hang-Korng Ea

    Full Text Available basic calcium phosphate (BCP crystals are commonly found in osteoarthritis (OA and are associated with cartilage destruction. BCP crystals induce in vitro catabolic responses with the production of metalloproteases and inflammatory cytokines such as interleukin-1 (IL-1. In vivo, IL-1 production induced by BCP crystals is both dependant and independent of NLRP3 inflammasome. We aimed to clarify 1/ the role of BCP crystals in cartilage destruction and 2/ the role of IL-1 and NLRP3 inflammasome in cartilage degradation related to BCP crystals.synovial membranes isolated from OA knees were analysed by alizarin Red and FTIR. Pyrogen free BCP crystals were injected into right knees of WT, NLRP3 -/-, ASC -/-, IL-1α -/- and IL-1β-/- mice and PBS was injected into left knees. To assess the role of IL-1, WT mice were treated by intra-peritoneal injections of anakinra, the IL-1Ra recombinant protein, or PBS. Articular destruction was studied at d4, d17 and d30 assessing synovial inflammation, proteoglycan loss and chondrocyte apoptosis. BCP crystals were frequently found in OA synovial membranes including low grade OA. BCP crystals injected into murine knee joints provoked synovial inflammation characterized by synovial macrophage infiltration that persisted at day 30, cartilage degradation as evidenced by loss of proteoglycan staining by Safranin-O and concomitant expression of VDIPEN epitopes, and increased chondrocyte apoptosis. BCP crystal-induced synovitis was totally independent of IL-1α and IL-1β signalling and no alterations of inflammation were observed in mice deficient for components of the NLRP3-inflammasome, IL-1α or IL-1β. Similarly, treatment with anakinra did not prevent BCP crystal effects. In vitro, BCP crystals elicited enhanced transcription of matrix degrading and pro-inflammatory genes in macrophages.intra-articular BCP crystals can elicit synovial inflammation and cartilage degradation suggesting that BCP crystals have a direct

  2. Sea-surface temperature reconstruction from trace elements variations of tropical coralline red algae

    Science.gov (United States)

    Darrenougue, Nicolas; De Deckker, Patrick; Eggins, Stephen; Payri, Claude

    2014-06-01

    We used laser ablation inductively coupled plasma mass spectrometry (LA-ICPMS) to obtain high-resolution variations of the Mg/Ca, Sr/Ca and Li/Ca composition of free-living forms (i.e. rhodoliths) of the coralline red algal species Sporolithon durum in order to test their potential to archive seawater temperature information. A monitoring experiment was conducted based on alizarin red S (ARS) staining of rhodoliths specimens collected in various locations across a ˜1 km2 rhodolith bed in the vicinity of Nouméa, New Caledonia, where in situ temperature (IST) variations were recorded for 22 months between November 2009 and August 2011. A >45-year comparison of Mg and trace elements with sea-surface temperature (SST) was established from the analysis of 5 different branches belonging to three of the largest (7.4-8.5 cm in diameter) rhodolith specimens observed at the site. Consistent mean Mg/Ca, Sr/Ca and Li/Ca concentrations and seasonal patterns are found for the rhodoliths' last living years (2009-2011) across 43 branches and for the full 1963-2008 period across the 5 branches. Average elemental concentrations (Mg/Ca: 0.31 ± 0.04 mol/mol; Sr/Ca: 3.5 ± 0.4 mmol/mol and Li/Ca: 0.08 ± 0.02 mmol/mol) fall within range of those found in the literature. Individual element variations show good reproducibility between records and Mg/Ca, Sr/Ca and Li/Ca co-vary systematically. Combined records of Mg/Ca, Sr/Ca and Li/Ca are highly correlated with the IST monthly pattern for the 2009-2011 period (0.82 < r < 0.91; p < 0.001) and with local variations of monthly SST for the 1963-2008 period (0.65 < r < 0.85; p < 0.001), with Mg/Ca systematically being the best fit to monthly seawater temperature variations. Inter-annual Mg/Ca anomalies show significant correlation with the Oceanic Nino Index (ONI), indicating that S. durum rhodoliths also have the capacity to record the regional climate pattern in the tropical Pacific. Finally, consistent variations between the combined Mg

  3. Biphasic influence of dexamethasone exposure on embryonic vertebrate skeleton development

    Energy Technology Data Exchange (ETDEWEB)

    Cheng, Xin; Chen, Jian-long; Ma, Zheng-lai; Zhang, Zhao-long; Lv, Shun; Mai, Dong-mei; Liu, Jia-jia [Department of Histology and Embryology, Key Laboratory for Regenerative Medicine of the Ministry of Education, School of Medicine, Jinan University, Guangzhou 510632 (China); Chuai, Manli [Division of Cell and Developmental Biology, University of Dundee, Dundee DD1 5EH (United Kingdom); Lee, Kenneth Ka Ho; Wan, Chao [Stem Cell and Regeneration Thematic Research Programme, School of Biomedical Sciences, Chinese University of Hong Kong, Shatin (Hong Kong); Yang, Xuesong, E-mail: yang_xuesong@126.com [Department of Histology and Embryology, Key Laboratory for Regenerative Medicine of the Ministry of Education, School of Medicine, Jinan University, Guangzhou 510632 (China); Institute of Fetal-Preterm Labor Medicine, Jinan University, Guangzhou 510632 (China)

    2014-11-15

    Dexamethasone (Dex) has anti-inflammatory and immunomodulatory properties against many conditions. There is a potential teratogenic risk, however, for pregnant women receiving Dex treatment. It has been claimed that Dex exposure during pregnancy could affect osteogenesis in the developing embryo, which still remains highly controversial. In this study, we employed chick embryos to investigate the effects of Dex exposure on skeletal development using combined in vivo and in vitro approach. First, we demonstrated that Dex (10{sup −8}–10{sup −6} μmol/egg) exposure resulted in a shortening of the developing long bones of chick embryos, and it accelerated the deposition of calcium salts. Secondly, histological analysis of chick embryo phalanxes exhibited Dex exposure inhibited the proliferation of chondrocytes, increased apoptosis of chondrocytes and osteocytes, and led to atypical arranged hypertrophic chondrocytes. The expression of genes related to skeletogenesis was also analyzed by semi-quantitative RT-PCR. The expression of ALP, Col1a2 and Col2a1 was decreased in the Dex treated phalanxes. A detectable increase was observed in Runx-2 and Mmp-13 expression. We next examined how Dex affected the different stages of skeletogenesis in vitro. Utilizing limb bud mesenchyme micromass cultures, we determined that Dex exposure exerted no effect on apoptosis but impaired chondrogenic cell proliferation. Interestingly, low dose of Dex moderately prompted nodule formation as revealed by alcian blue staining, but higher doses of Dex significantly inhibited similar chondrogenic differentiation. Dex exposure did not induce apoptosis when the chondrogenic precursors were still at the mesenchymal stage, however, cell viability was suppressed when the mesenchyme differentiated into chondrocytes. Alizarin red staining revealed that the capacity to form mineralized bone nodules was correspondingly enhanced as Dex concentrations increased. The mRNA level of Sox-9 was slightly

  4. Selective laser sintering fabrication of nano-hydroxyapatite/poly-ε-caprolactone scaffolds for bone tissue engineering applications

    Directory of Open Access Journals (Sweden)

    Xia Y

    2013-11-01

    Full Text Available Yan Xia,1,* Panyu Zhou,1,* Xiaosong Cheng,1,* Yang Xie,1,* Chong Liang,2 Chao Li,1 Shuogui Xu1,2 1Department of Orthopedics, Changhai Hospital, Second Military Medical University, Shanghai, People's Republic of China; 2Department of Neurosurgery, The 81 Hospital of People's Liberation Army of China, Nanjing, People's Republic of China *These authors contributed equally to this work Abstract: The regeneration of functional tissue in osseous defects is a formidable challenge in orthopedic surgery. In the present study, a novel biomimetic composite scaffold, here called nano-hydroxyapatite (HA/poly-ε-caprolactone (PCL was fabricated using a selective laser sintering technique. The macrostructure, morphology, and mechanical strength of the scaffolds were characterized. Scanning electronic microscopy (SEM showed that the nano-HA/PCL scaffolds exhibited predesigned, well-ordered macropores and interconnected micropores. The scaffolds have a range of porosity from 78.54% to 70.31%, and a corresponding compressive strength of 1.38 MPa to 3.17 MPa. Human bone marrow stromal cells were seeded onto the nano-HA/PCL or PCL scaffolds and cultured for 28 days in vitro. As indicated by the level of cell attachment and proliferation, the nano-HA/PCL showed excellent biocompatibility, comparable to that of PCL scaffolds. The hydrophilicity, mineralization, alkaline phosphatase activity, and Alizarin Red S staining indicated that the nano-HA/PCL scaffolds are more bioactive than the PCL scaffolds in vitro. Measurements of recombinant human bone morphogenetic protein-2 (rhBMP-2 release kinetics showed that after nano-HA was added, the material increased the rate of rhBMP-2 release. To investigate the in vivo biocompatibility and osteogenesis of the composite scaffolds, both nano-HA/PCL scaffolds and PCL scaffolds were implanted in rabbit femur defects for 3, 6, and 9 weeks. The wounds were studied radiographically and histologically. The in vivo results showed

  5. Maxillary sinus floor elevation using a tissue-engineered bone with calcium-magnesium phosphate cement and bone marrow stromal cells in rabbits.

    Science.gov (United States)

    Zeng, Deliang; Xia, Lunguo; Zhang, Wenjie; Huang, Hui; Wei, Bin; Huang, Qingfeng; Wei, Jie; Liu, Changsheng; Jiang, Xinquan

    2012-04-01

    The objective of this study was to assess the effects of maxillary sinus floor elevation with a tissue-engineered bone constructed with bone marrow stromal cells (bMSCs) and calcium-magnesium phosphate cement (CMPC) material. The calcium (Ca), magnesium (Mg), and phosphorus (P) ions released from calcium phosphate cement (CPC), magnesium phosphate cement (MPC), and CMPC were detected by inductively coupled plasma atomic emission spectroscopy (ICP-AES), and the proliferation and osteogenic differentiation of bMSCs seeded on CPC, MPC, and CMPC or cultured in CPC, MPC, and CMPC extracts were measured by MTT analysis, alkaline phosphatase (ALP) activity assay, alizarin red mineralization assay, and real-time PCR analysis of the osteogenic genes ALP and osteocalcin (OCN). Finally, bMSCs were combined with CPC, MPC, and CMPC and used for maxillary sinus floor elevation in rabbits, while CPC, MPC, or CMPC without cells served as control groups. The new bone formation in each group was detected by histological finding and fluorochrome labeling at weeks 2 and 8 after surgical operation. It was observed that the Ca ion concentrations of the CMPC and CPC scaffolds was significantly higher than that of the MPC scaffold, while the Mg ions concentration of CMPC and MPC was significantly higher than that of CPC. The bMSCs seeded on CMPC and MPC or cultured in their extracts proliferated more quickly than the cells seeded on CPC or cultured in its extract, respectively. The osteogenic differentiation of bMSCs seeded on CMPC and CPC or cultured in the corresponding extracts was significantly enhanced compared to that of bMSCs seeded on MPC or cultured in its extract; however, there was no significant difference between CMPC and CPC. As for maxillary sinus floor elevation in vivo, CMPC could promote more new bone formation and mineralization compared to CPC and MPC, while the addition of bMSCs could further enhance its new bone formation ability significantly. Our data suggest that

  6. Effect of Aminated Mesoporous Bioactive Glass Nanoparticles on the Differentiation of Dental Pulp Stem Cells.

    Directory of Open Access Journals (Sweden)

    Jung-Hwan Lee

    Full Text Available Mesoporous bioactive nanoparticles (MBNs have been developed as promising additives to various types of bone or dentin regenerative material. However, biofunctionality of MBNs as dentin regenerative additive to dental materials have rarely been studied. We investigated the uptake efficiency of MBNs-NH2 with their endocytosis pathway and the role of MBNs-NH2 in odontogenic differentiation to clarify inherent biofunctionality. MBNs were fabricated by sol-gel synthesis, and 3% APTES was used to aminate these nanoparticles (MBNs-NH2 to reverse their charge from negative to positive. To characterize the MBNs-NH2, TEM, XRD, FTIR, zeta(ξ-potential measurements, and Brunauer-Emmett-Teller analysis were performed. After primary cultured rat dental pulp stem cells (rDPSCs were incubated with various concentrations of MBNs-NH2, stem cell viability (24 hours with or without differentiated media, internalization of MBNs-NH2 in rDPSCs (~4 hours via specific endocytosis pathway, intra or extracellular ion concentration and odontoblastic differentiation (~28 days were investigated. Incubation with up to 50 μg/mL of MBNs-NH2 had no effect on rDPSCs viability with differentiated media (p>0.05. The internalization of MBNs-NH2 in rDPSCs was determined about 92% after 4 hours of incubation. Uptake was significantly decreased with ATP depletion and after 1 hour of pre-treatment with the inhibitor of macropinocytosis (p<0.05. There was significant increase of intracellular Ca and Si ion concentration in MBNs-NH2 treated cells compared to no-treated counterpart (p<0.05. The expression of odontogenic-related genes (BSP, COL1A, DMP-1, DSPP, and OCN and the capacity for biomineralization (based on alkaline phosphatase activity and alizarin red staining were significantly upregulated with MBNs-NH2. These results indicate that MBNs-NH2 induce odontogenic differentiation of rDPSCs and may serve as a potential dentin regenerative additive to dental material for promoting

  7. Induced differentiation of C2C12 to osteoblast via adenovirus-mediated Cbfa1 in vitro%体外诱导C2C12细胞向成骨细胞的分化

    Institute of Scientific and Technical Information of China (English)

    张勇; 杨彤涛; 胡运生; 廖博; 文艳华; 范清宇

    2013-01-01

    目的 成骨细胞特异性转录因子a1(core binding factor a1,Cbfa1)通过调节生长因子和骨特异性细胞外基质蛋白的基因表达而参与成骨细胞的分化和骨发育过程.文中构建成Cbfa1,以腺病毒载体转染成肌细胞C2C12,为种子细胞构建组织工程化骨.方法 体外培养小鼠成肌细胞C2C12,用重组腺病毒质粒pAd-IL-31介导Cbfa1/Osf2基因瞬时转染小鼠成肌C2C12细胞,Western blot检测Cbfa1蛋白表达.结果 Cbfa1蛋白表达、碱性磷酸酶(alkaline phosphatase,ALP)活性测定、骨钙素(osteocalcin,OCN)分泌量以及茜素红染色感染组明显高于对照组.结论 成肌细胞C2C12可以作为种子细胞构建组织工程化骨.%Objective Osteoblast core binding factor a 1 ( Cbfal) plays a role in osteoblast differentiation and development by regulating the gene of growth factor and extracellular matrix proteins . Recombinant adenovirus vector mediated Cbfa 1 was transferred to myoblast C2C12 to construct the tissue-engineered bone. Methods The myoblast C2C12 was cultured in vitro, and then transiently transfected with recombinant adenovirus vector pAd -IL-31 mediated-Cbfal/Osf2. Western blot was used to detect the expression of Cbfal. Results Compared with the control group , the expression of Cbfal, activity of alkaline phosphtase (ALP) , secretory volume of osteocalcin (OCN) and staining via alizarin bordeaux were higher in the transfection group . Conclusion Myoblast C2C12 acts as a seed cell for constructing tissue -engineered bone.

  8. Exploring the critical dependence of adsorption of various dyes on the degradation rate using Ln{sup 3+}-TiO{sub 2} surface under UV/solar light

    Energy Technology Data Exchange (ETDEWEB)

    Devi, L. Gomathi, E-mail: gomatidevi_naik@yahoo.co.in [Department of Post Graduate Studies in Chemistry, Central College City Campus, Dr. Ambedkar Street, Bangalore University, Bangalore 560001 (India); Kumar, S. Girish [Department of Post Graduate Studies in Chemistry, Central College City Campus, Dr. Ambedkar Street, Bangalore University, Bangalore 560001 (India)

    2012-11-15

    Graphical abstract: The surface reactive acidic sites enhances on doping with rare earth ions which facilitates efficient adsorption of the dye molecules on the catalyst surface. In addition, the nature of the dopant, its concentration and electronic configuration additionally contributes to the overall efficiency. Highlights: Black-Right-Pointing-Pointer The degradation of structurally different anionic dyes under different pH conditions is reported. Black-Right-Pointing-Pointer Pre adsorption of pollutant on catalyst surface is vital for efficient photocatalysis. Black-Right-Pointing-Pointer Adsorption of dye on the catalyst surface depends on the substituent's attached to it. Black-Right-Pointing-Pointer The dopant with half filled electronic configuration served as shallow traps for charge carriers. - Abstract: The degradation of structurally different anionic dyes like Alizarin Red S (ARS) Amaranth (AR), Brilliant Yellow (BY), Congo Red (CR), Fast Red (FR), Methyl Orange (MO), and Methyl Red (MR) were carried out using Ln{sup 3+} (Ln{sup 3+} = La{sup 3+}, Ce{sup 3+} and Gd{sup 3+}) doped TiO{sub 2} at different pH conditions under UV/solar light. All the anionic dyes underwent rapid degradation at acidic pH, while resisted at alkaline conditions due to the adsorptive tendency of these dyes on the catalyst surface at different pH conditions. Gd{sup 3+} (0.15 mol%)-TiO{sub 2} exhibited better activity compared to other photocatalyst ascribed to half filled electronic configuration of Gd{sup 3+} ions. It is proposed that Ln{sup 3+} serves only as charge carrier traps under UV light, while it also act as visible light sensitizers under solar light. Irrespective of the catalyst and excitation source, the dye degradation followed the order: AR > FR > MO > MR > ARS > BY > CR. The results suggest that pre-adsorption of the pollutant is vital for efficient photocatalysis which is dependent on the nature of the substituent's group attached to the dye molecule.

  9. Petrology of upper silurian reservoir rocks from the Kudirka Atoll, Lithuania. On the spatiotemporal relations between different diagenetic events that have affected the reservoir quality of the rocks

    Energy Technology Data Exchange (ETDEWEB)

    Stentoft, N.; Lapinskas, P.; Musteikis, P.; Kristensen, L.; Hoelstad, T.

    1998-08-01

    The purpose of the study was to provide an understanding of the diagnetic development of the different reservoir rocks of the Kudirka Atoll, and to make out to what degree the present reservoir qualities (permeability and porosity) of the reef rocks are dependent on the preceding post depositional diagenetic events. Beside that the scope was to get a better understanding of the architecture of the reef complex by combining macroscopic core descriptions with log interpretations of selected wells. The spatial distribution of both porosity, water saturation, lithology and texture is included. The project encompasses investigation of core and plug material from following wells of the Kudirka Atoll field: Vilkaviskis-126, -131, -132, -135, -136, -137 and -139. From 50 small core pieces (cf. Appendix A) a total of 100 one inch plug samples were drilled (that is two plus from each core piece) and cleaned by use of soxhlet extractor. Porosity, air permeability and grain density were then measured on each plug (cf. Appendix E). Wireline logs from the seven wells, plus from Vilkaviskis-133 and Kudirka-146, were digitized and used in the log interpretation. To ensure that characteristics essential to both the log interpretation and the diagenetic history were called attention to, the macroscopic core descriptions were carried out in accordance to the procedure described in the `introductory remarks` of Appendix A. The microscopic investigations, that form the basis of the description of the diagenetic history, are based on 50 thin-section (30 {mu}m thick) prepared from one inch plugs (cf Appenix B). To see whether the carbonate crystals, that were assumed to be dolomite from their reaction with Alizarin Red S, are in fact pure, or close to pure CaMg(CO-3){sub 2}, two thin sections were polished and examined by SEM-ED (cf. Appendix F). A total of 15 plugs were picked out for capillary pressure measurements (the porous plate method using 8 pressure steps) including

  10. Petrology of upper silurian reservoir rocks from the Kudirka Atoll, Lithuania. On the spatiotemporal relations between different diagenetic events that have affected the reservoir quality of the rocks

    Energy Technology Data Exchange (ETDEWEB)

    Stentoft, N.; Lapinskas, P.; Musteikis, P.; Kristensen, L.; Hoelstad, T.

    1998-08-01

    The purpose of the study was to provide an understanding of the diagenetic development of the different reservoir rocks of the Kudirka Atoll, and to make out to what degree the present reservoir qualities (permeability and porosity) of the reef rocks are dependent on the preceding post depositional diagenetic events. Beside that the scope was to get a better understanding of the architecture of the reef complex by combining macroscopic core descriptions with log interpretations of selected wells. The spatial distribution of both porosity, water saturation, lithology and texture is included. The project encompasses investigation of core and plug material from following wells of the Kudirka Atoll field: Vilkaviskis-126, -131, -132, -135, -136, -137 and -139. From 50 small core pieces (cf. Appendix A) a total of 100 one inch plug sampels were drilled (that is two plus from each core piece) and cleaned by use of soxhlet extractor. Porosity, air permeability and grain density were then measured on each plug (cf. Appendix E). Wireline logs from the seven wells, plus from Vilkaviskis-133 and Kudirka-146, were digitized and used in the log interpretation. To ensure that characteristics essential to both the log interpretation and the diagnetic history were called attention to, the macroscopic core descriptions were carried out in accordance to the procedure described in the `introductory remarks` of Appendix A. The microscopic investigations, that form the basis of the description of the diagenetic history, are based on 50 thin-section (30 {mu}m thick) prepared from one inch plugs (cf Appenix B). To see whether the carbonate crystals, that were assumed to be dolomite from their reaction with Alizarin Red S, are in fact pure, or close to pure CaMg(CO-3){sub 2}, two thin sections were polished and examined by SEM-ED (cf. Appencix F). A total of 15 plugs were picked out for capillary pressure measurements (the porous plate method using 8 pressure steps) including

  11. The experimental investigation of glioma-trophic capacity of human umbilical cord-derived mesenchymal stem cells after intraventricular administration

    Directory of Open Access Journals (Sweden)

    FAN Cun-gang

    2013-07-01

    Full Text Available Objective To explore the glioma-trophic migration capacity of human umbilical cord-derived mesenchymal stem cells (hUC-MSCs by intraventricular administration. Methods The umbilical cord tissue were obtained during full-term pregnancy cesarean section under sterile conditions. This study was approved by Ethics Committee and got the informed consent of patient. The hUC-MSCs were isolated by trypsin and collagenase digestion, followed by adherent culture methods. The characteristics of isolated hUC-MSCs were demonstrated by cell morphylogy, phenotype analysis and multi-differentiation potentials into adipocytes, osteoblasts and neural cells. Then the hUC-MSCs were labeled with CM-DiI and injected into contralateral ventricle of glioma of the C6 glioma-bearing Sprague-Dawley (SD rats. Two weeks later, the rats were sacrificed and the brains were taken out to examine the migration and distribution of hUC-MSCs in the tumor bed, at the interface of tumor and cerebral parenchyma as well as the tumor satelites infiltrating into the normal brain. Results The hUC-MSCs demonstrated plastic-adherent characterization and homogeneous fibroblastic-like morphylogy in culture, expression of specific surface phenotypes of MSCs (CD13, CD29, CD44, CD90 but not endothelial or hematopoietic markers (CD14, CD31, CD34, CD38, CD45, CD133, and muti-differentiatiation potentials into Oil red O stained adipocytes, Alizarin red S stained osteoblasts, neuron-specific enolase (NSE-positive neurons and glial fibrillary acidic protein (GFAP-positive astrocytes in permissive inducive conditions. Importantly, after labeled hUC-MSCs injection into contralateral ventricle of glioma, the hUC-MSCs migrated from initial injection site to the glioma mass and along the interface of tumor and brain, and some of them "chasing" the glioma satellites infiltrated into the normal parenchyma. Conclusion The hUC-MSCs possess prominent tumor-specific targeting capacity and extensive intratumoral

  12. Trehalose maintains bioactivity and promotes sustained release of BMP-2 from lyophilized CDHA scaffolds for enhanced osteogenesis in vitro and in vivo.

    Directory of Open Access Journals (Sweden)

    Jun Zhao

    Full Text Available Calcium phosphate (Ca-P scaffolds have been widely employed as a supportive matrix and delivery system for bone tissue engineering. Previous studies using osteoinductive growth factors loaded Ca-P scaffolds via passive adsorption often experience issues associated with easy inactivation and uncontrolled release. In present study, a new delivery system was fabricated using bone morphogenetic protein-2 (BMP-2 loaded calcium-deficient hydroxyapatite (CDHA scaffold by lyophilization with addition of trehalose. The in vitro osteogenesis effects of this formulation were compared with lyophilized BMP-2/CDHA construct without trehalose and absorbed BMP-2/CDHA constructs with or without trehalose. The release characteristics and alkaline phosphatase (ALP activity analyses showed that addition of trehalose could sufficiently protect BMP-2 bioactivity during lyophilization and achieve sustained BMP-2 release from lyophilized CDHA construct in vitro and in vivo. However, absorbed BMP-2/CDHA constructs with or without trehalose showed similar BMP-2 bioactivity and presented a burst release. Quantitative real-time PCR (RT-qPCR and enzyme-linked immunosorbent assay (ELISA demonstrated that lyophilized BMP-2/CDHA construct with trehalose (lyo-tre-BMP-2 promoted osteogenic differentiation of bone marrow stromal cells (bMSCs significantly and this formulation could preserve over 70% protein bioactivity after 5 weeks storage at 25°C. Micro-computed tomography, histological and fluorescent labeling analyses further demonstrated that lyo-tre-BMP-2 formulation combined with bMSCs led to the most percentage of new bone volume (38.79% ± 5.32% and area (40.71% ± 7.14% as well as the most percentage of fluorochrome stained bone area (alizarin red S: 2.64% ± 0.44%, calcein: 6.08% ± 1.37% and mineral apposition rate (4.13 ± 0.62 µm/day in critical-sized rat cranial defects healing. Biomechanical tests also indicated the maximum stiffness (118.17 ± 15.02 Mpa and

  13. Short-term effects of calcium ions on the apoptosis and onset of mineralization of human dental pulp cells in vitro and in vivo.

    Science.gov (United States)

    An, Shaofeng; Gao, Yan; Huang, Yihua; Jiang, Xiaoqiong; Ma, Ke; Ling, Junqi

    2015-07-01

    Calcium ions (Ca2+) are a major constituent of most pulp-capping materials and have an important role in the mineralization of human dental pulp cells (hDPCs). A previous study by our group has shown that increased levels of Ca2+ can promote hDPC-mediated mineralization in long-term cultures (21 days). However, the initiation of mineralization occurs in the early stage of osteogenic inductive culture, and the effects of Ca2+ on the mineralization of hDPCs in short-term cultures (five days) have not been studied in detail. Furthermore, the underlying mechanism by which Ca2+ stimulates the mineralization of hDPCs has remained controversial. A strong correlation between mineralization and cell apoptosis and/or death has been identified. Thus, the present study hypothesized that Ca2+ may promote the onset of hDPC-mediated mineralization through inducing their apoptosis and/or death. To verify this hypothesis, Ca2+ was added to the growth culture medium and osteogenic culture medium at various concentrations. Alizarin Red S staining and reverse transcription-polymerase chain reaction analysis were used to evaluate the onset of mineralization. Furthermore, the cell counting kit-8 and fluorescein isothiocyanate-Annexin V/propidium iodide double-staining method were adopted to detect the proliferation and apoptosis of hDPCs in the growth culture medium. An animal experiment and scanning electron microscopic observation of ceramic graft implants were applied to measure the mineralization in vivo. The results showed that 5.4 and 9.0 mM Ca2+ accelerated the onset of mineralized matrix nodule formation, promoted osteopontin mRNA expression and induced marked cell apoptosis and necrosis, but had no obvious effect on cell proliferation. These findings indicated a positive association between cell apoptosis and/or death and the timing of formation as well as the quantity of extracellular mineralization induced by Ca2+ in short-term cultured hDPCs.

  14. In situ osteoblast mineralization mediates post-injection mechanical properties of osteoconductive material.

    Science.gov (United States)

    Bialorucki, Callan; Subramanian, Gayathri; Elsaadany, Mostafa; Yildirim-Ayan, Eda

    2014-10-01

    The objective of this study was to understand the temporal relationship between in situ generated calcium content (mineralization) and the mechanical properties of an injectable orthobiologic bone-filler material. Murine derived osteoblast progenitor cells were differentiated using osteogenic factors and encapsulated within an injectable polycaprolactone nanofiber-collagen composite scaffold (PN-COL +osteo) to evaluate the effect of mineralization on the mechanical properties of the PN-COL scaffold. A comprehensive study was conducted using both an experimental and a predictive analytical mechanical analysis for mechanical property assessment as well as an extensive in vitro biological analysis for in situ mineralization. Cell proliferation was evaluated using a PicoGreen dsDNA quantification assay and in situ mineralization was analyzed using both an alkaline phosphatase (ALP) assay and an Alizarin Red stain-based assay. Mineralized matrix formation was further evaluated using energy dispersive x-ray spectroscopy (EDS) and visualized using SEM and histological analyses. Compressive mechanical properties of the PN-COL scaffolds were determined using a confined compression stress-relaxation protocol and the obtained data was fit to the standard linear solid viscoelastic material mathematical model to demonstrate a relationship between increased in situ mineralization and the mechanical properties of the PN-COL scaffold. Cell proliferation was constant over the 21 day period. ALP activity and calcium concentration significantly increased at day 14 and 21 as compared to PN-COL -osteo with undifferentiated osteoblast progenitor cells. Furthermore, at day 21 EDS, SEM and von Kossa histological staining confirmed mineralized matrix formation within the PN-COL scaffolds. After 21 days, compressive modulus, peak stress, and equilibrium stress demonstrate significant increases of 3.4-fold, 3.3-fold, and 4.0-fold respectively due to in situ mineralization. Viscoelastic

  15. The effect of low static magnetic field on osteogenic and adipogenic differentiation potential of human adipose stromal/stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Marędziak, Monika, E-mail: monika.maredziak@gmail.com [Faculty of Veterinary Medicine, University of Environmental and Life Sciences, Wrocław (Poland); Wroclaw Research Centre EIT+, Wrocław (Poland); Śmieszek, Agnieszka, E-mail: smieszek.agnieszka@gmail.com [Wroclaw Research Centre EIT+, Wrocław (Poland); Faculty of Biology, University of Environmental and Life Sciences, Wrocław (Poland); Tomaszewski, Krzysztof A., E-mail: krtomaszewski@gmail.com [Department of Anatomy, Jagiellonian University Medical College, Krakow (Poland); Lewandowski, Daniel, E-mail: daniel.lewandowski@pwr.wroc.pl [Institute of Materials Science and Applied Mechanics, Wroclaw University of Technology, Wroclaw (Poland); Marycz, Krzysztof, E-mail: krzysztofmarycz@interia.pl [Wroclaw Research Centre EIT+, Wrocław (Poland); Faculty of Biology, University of Environmental and Life Sciences, Wrocław (Poland)

    2016-01-15

    The aim of this work was to investigate the effects of static magnetic field (SMF) on the osteogenic properties of human adipose derived mesenchymal stem cells (hASCs). In this study in seven days viability assay we examined the impact of SMF on cells proliferation rate, population doubling time, and ability to form single-cell derived colonies. We have also examined cells' morphology, ultrastructure and osteogenic properties on the protein as well as mRNA level. We established a complex approach, which enabled us to obtain information about SMF and hASCs potential in the context of differentiation into osteogenic and adipogenic lineages. We demonstrated that SMF enhances both viability and osteogenic properties of hASCs through higher proliferation factor and shorter population doubling time. We have also observed asymmetrically positioned nuclei and organelles after SMF exposition. With regards to osteogenic properties we observed increased levels of osteogenic markers i.e. osteopontin, osteocalcin and increased ability to form osteonodules with positive reaction to Alizarin Red dye. We have also shown that SMF besides enhancing osteogenic properties of hASCs, simultaneously decreases their ability to differentiate into adipogenic lineage. Our results clearly show a direct influence of SMF on the osteogenic potential of hASCs. These results provide key insights into the role of SMF on their cellular fate and properties. - Graphical abstract: Influence of static magnetic field on viability and differentiation properties of human adipose derived mesenchymal stem cells. Abbreviations: SMF – static magnetic field; hASCs – human adipose derived mesenchymal stem cells; PF – proliferation factor; PDT – population doubling time; CFU-E –> colony forming unit efficiency; OPN – osteopontin; OCL – osteocalcin; Col – collagen type I; BMP-2 – bone morphogenetic protein 2; Ca – calcium; P – phosphorus. - Highlights: • Effects of static

  16. L-type calcium channels play a crucial role in the proliferation and osteogenic differentiation of bone marrow mesenchymal stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Wen, Li [Department of Orthodontics, School of Stomatology, Fourth Military Medical University, Xi' an 710032 (China); Wang, Yu [Department of Oncology, Xijing Hospital, Fourth Military Medical University, Xi' an 710032 (China); Wang, Huan [Department of Orthodontics, School of Stomatology, Fourth Military Medical University, Xi' an 710032 (China); Kong, Lingmin [Department of Fundamental Medicine, Cell Engineering Research Centre, Fourth Military Medical University, Xi' an 710032 (China); Zhang, Liang [Department of Orthodontics, School of Stomatology, Fourth Military Medical University, Xi' an 710032 (China); Chen, Xin [Department of General Dentistry, The 174th Hospital of Chinese PLA, Xiamen 361003 (China); Ding, Yin, E-mail: dingyin@fmmu.edu.cn [Department of Orthodontics, School of Stomatology, Fourth Military Medical University, Xi' an 710032 (China)

    2012-08-03

    Highlights: Black-Right-Pointing-Pointer We detect the functional Ca{sup 2+} currents and mRNA expression of VDCC{sub L} in rMSCs. Black-Right-Pointing-Pointer Blockage of VDCC{sub L} exert antiproliferative and apoptosis-inducing effects on rMSCs. Black-Right-Pointing-Pointer Inhibiting VDCC{sub L} can suppress the ability of rMSCs to differentiate into osteoblasts. Black-Right-Pointing-Pointer {alpha}1C of VDCC{sub L} may be a primary functional subunit in VDCC{sub L}-regulating rMSCs. -- Abstract: L-type voltage-dependent Ca{sup 2+} channels (VDCC{sub L}) play an important role in the maintenance of intracellular calcium homeostasis, and influence multiple cellular processes. They have been confirmed to contribute to the functional activities of osteoblasts. Recently, VDCC{sub L} expression was reported in mesenchymal stem cells (MSCs), but the role of VDCC{sub L} in MSCs is still undetermined. The aim of this study was to determine whether VDCC{sub L} may be regarded as a new regulator in the proliferation and osteogenic differentiation of rat MSC (rMSCs). In this study, we examined functional Ca{sup 2+} currents (I{sub Ca}) and mRNA expression of VDCC{sub L} in rMSCs, and then suppressed VDCC{sub L} using nifedipine (Nif), a VDCC{sub L} blocker, to investigate its role in rMSCs. The proliferation and osteogenic differentiation of MSCs were analyzed by MTT, flow cytometry, alkaline phosphatase (ALP), Alizarin Red S staining, RT-PCR, and real-time PCR assays. We found that Nif exerts antiproliferative and apoptosis-inducing effects on rMSCs. ALP activity and mineralized nodules were significantly decreased after Nif treatment. Moreover, the mRNA levels of the osteogenic markers, osteocalcin (OCN), bone sialoprotein (BSP), and runt-related transcription factor 2 (Runx2), were also down-regulated. In addition, we transfected {alpha}1C-siRNA into the cells to further confirm the role of VDCC{sub L} in rMSCs, and a similar effect on osteogenesis was found. These

  17. Study of the effects of ß-myrcene on rat fertility and general reproductive performance

    Directory of Open Access Journals (Sweden)

    F.J.R. Paumgartten

    1998-07-01

    Full Text Available ß-Myrcene (MYR is a monoterpene found in the oils of a variety of aromatic plants including lemongrass, verbena, hop, bay, and others. MYR and essential oils containing this terpenoid compound are used in cosmetics, household products, and as flavoring food additives. This study was undertaken to investigate the effects of MYR on fertility and general reproductive performance in the rat. MYR (0, 100, 300 and 500 mg/kg in peanut oil was given by gavage to male Wistar rats (15 per dose group for 91 days prior to mating and during the mating period, as well as to females (45 per dose group continuously for 21 days before mating, during mating and pregnancy, and throughout the period of lactation up to postnatal day 21. On day 21 of pregnancy one-third of the females of each group were submitted to cesarean section. Resorption, implantation, as well as dead and live fetuses were counted. All fetuses were examined for external malformations, weighed, and cleared and stained with Alizarin Red S for skeleton evaluation. The remaining dams were allowed to give birth to their offspring. The progeny was examined at birth and subsequently up to postnatal day 21. Mortality, weight gain and physical signs of postnatal development were evaluated. Except for an increase in liver and kidney weights, no other sign of toxicity was noted in male and female rats exposed to MYR. MYR did not affect the mating index (proportion of females impregnated by males or the pregnancy index (ratio of pregnant to sperm-positive females. No sign of maternal toxicity and no increase in externally visible malformations were observed at any dose level. Only at the highest dose tested (500 mg/kg did MYR induce an increase in the resorption rate and a higher frequency of fetal skeleton anomalies. No adverse effect of MYR on postnatal weight gain was noted but days of appearance of primary coat, incisor eruption and eye opening were slightly delayed in the exposed offspring. On the

  18. High glucose mediates endothelial-to-chondrocyte transition in human aortic endothelial cells

    Directory of Open Access Journals (Sweden)

    Tang Rining

    2012-09-01

    Full Text Available Abstract Background Vascular calcification is one of the common complications in diabetes mellitus. Many studies have shown that high glucose (HG caused cardiovascular calcification, but its underlying mechanism is not fully understood. Recently, medial calcification has been most commonly described in the vessels of patients with diabetes. Chondrocytes were involved in the medial calcification. Recent studies have shown that the conversion into mesenchymal stem cells (MSCs via the endothelial-to-mesenchymal transition (EndMT could be triggered in chondrocytes. Our previous research has indicated that HG induced EndMT in human aortic endothelial cells (HAECs. Therefore, we addressed the question of whether HG-induced EndMT could be transitioned into MSCs and differentiated into chondrocytes. Methods HAECs were divided into three groups: a normal glucose (NG group, HG group (30 mmol/L, and mannitol (5.5 mmol/L NG + 24.5 mmol/L group. Pathological changes were investigated using fluorescence microscopy and electron microscopy. Immunofluorescence staining was performed to detect the co-expression of endothelial markers, such as CD31, and fibroblast markers, such as fibroblast-specific protein 1 (FSP-1. The expression of FSP-1 was detected by real time-PCR and western blots. Endothelial-derived MSCs were grown in MSC medium for one week. The expression of the MSCs markers STRO-1, CD44, CD10 and the chondrocyte marker SOX9 was detected by immunofluorescence staining and western blots. Chondrocyte expression was detected by alcian blue staining. Calcium deposits were analyzed by alizarin red staining. Results The incubation of HAECs exposed to HG resulted in a fibroblast-like phenotype. Double staining of the HAECs indicated a co-localization of CD31 and FSP-1. The expression of FSP-1 was significantly increased in the HG group, and the cells undergoing EndMT also expressed STRO-1, CD44 and SOX9 compared with the controls (P  Conclusions Our

  19. Treatment of pesticide rinsate towards reuse by photosensitized Fenton-like process.

    Science.gov (United States)

    Kuo, W S; Ho, Y Y

    2010-01-01

    A Fenton-like process with combination of dye has been used to enhance the treatment of carbofuran (2,3-dihydro-2,2-dimethylbenzofuran -7-yl methylcarbamate) pesticide rinsate. Results showed that as compared to Fenton-like process, this photosensitization Fenton-like process improved the degradation efficiency of carbofuran rinsate significantly. Among the conditions studied, the optimum dosage for the complete destruction of carbofuran molecular structure was found under a [H2O2]0/[Fe3+]0 ratio of 30-35 and a [Dye]0/[Fe3+]0 ratio of 2%, respectively, after an irradiance of 500 W/m2 for 20 min. As a result, the COD degradation efficiency of rinsate could be promoted from 37.1 to 61.2% and 66.0% by an addition of methylene blue (MB) and alizarin red S (ARS), respectively. Nevertheless, ARS showed a much more effective acceleration effect on the mineralization and microtoxicity reduction of carbofuran than MB. A mineralization efficiency of 57.2% and a microtoxicity reduction of 90% could be achieved with the addition of ARS. Because of its quinone structure unit, the dye ARS could play a role like hydroquinone to recycle Fe2+ from Fe3+, resulting in one more catalytic effect on the reduction of Fe3+ and thus the mineralization and microtoxicity reduction of carbofuran was greatly promoted in the presence of ARS. In addition, it was found that carbofuran molecules could be decomposed quickly to lower-molecular-weight intermediates and even mineralized by attacking of hydroxyl radicals. Carbofuran was found to be decomposed to carbofuran phenol, 3-oxo carbofuran phenol, and 3-hydroxyl carbofuran phenol initially, and then further be degraded to smaller molecules, such as NO3-, CH3COOH, (COOH)2 and CO2. Accordingly, it was believed that the Fenton-like process along with the aid of a photosensitizer, such as ARS, under an appropriate ratio could be a feasible and potential technology for the treatment of pesticide rinsate.

  20. Multiple Directional Differentiation Difference of Neonatal Rat Fibroblasts from Six Organs

    Directory of Open Access Journals (Sweden)

    Yuqiao Chang

    2016-06-01

    Full Text Available Background/Aims: Fibroblasts are abundantly distributed throughout connective tissues in the body and are very important in maintaining the structural and functional integrity. Recent reports have proved that fibroblasts and mesenchymal stem cells share much more in common than previously recognized. The aim of this study was to investigate comparative studies in fibroblasts on the differences in the expression of molecular markers and differentiation capacity from different organs. Methods: Combined trypsin/collagenase enzymes digestion method was used to isolate and culture the fibroblasts derived from heart, liver, spleen, lung, kidney and skin. Cell activity was determined by methyl thiazolyl tetrazolium (MTT assay. Common molecular markers for fibroblasts such as vimentin, DDR2 and FSP1, stem cell markers nanog, c-kit and sca-1 were detected by RT-PCR, immunofluorescence and western blotting. The osteogenic, adipogenic and cardiogenic differentiations of fibroblasts were performed by inductive culture in special mediums, and analyzed by Alizarin red, Oil red O and immunofluorescence staining of cTnT respectively. Results: The proliferation rate of fibroblasts in lung was faster than in other five organs. Common molecular markers for fibroblasts were expressed differently in different organs. DDR2 was strongly expressed in fibroblasts in the heart, partly expressed in the heart, skin, liver and spleen. Interestingly, no expression of DDR2 was detected in liver and kidney. However, vimentin and FSP1 were consistently expressed in fibroblasts from skin, liver, kidney, spleen and lung. nanog expression in fibroblasts from lung was less than that from heart, skin, liver and spleen (P . c-kit expression in fibroblasts from heart, skin and kidney was higher than that from spleen (P , while the c-kit positive fibroblasts from liver was obviously higher than that from spleen (P . But sca-1 expression in fibroblasts from lung was the lowest among six

  1. Histological and histochemical analysis of the gastrointestinal tract of the common pipistrelle bat (Pipistrellus pipistrellus).

    Science.gov (United States)

    Strobel, S; Encarnação, J A; Becker, N I; Trenczek, T E

    2015-01-01

    Bats have a very high mass-specific energy demand due to small size and active flight. European bat species are mostly insectivorous and the morphology of the gastrointestinal tract should be adapted accordingly. This study investigated the general anatomy by histology and the function by analysing carbohydrate distribution in particular of the mucus of the GI tract of the insectivorous bat Pipistrellus pipistrellus. The GI tracts of three individuals were dissected, fixed in formaldehyde, and embedded in paraffin wax. The tissues and cells of the GI tract of P. pipistrellus were analysed by classical (Acid Alizarin Blue, Haematoxylin-Eosin, and Masson Goldner Trichrome), histochemical (periodic acid-Schiff, Alcian blue at pH 2.5) and lectin histochemical (lectins WGA and HPA) staining procedures. The GI tract of P. pipistrellus was organised into the typical mammalian layers. The short, narrow, and thin-walled esophagus was simple with a folded stratified squamous epithelium without glands but mucous surface cells secreting neutral mucus. The stomach was globular shaped without specialisation. Mucous surface cells produced neutral mucus whereas neck and parietal cells secreted a mixture of neutral and acid mucus. Chief cell surface was positive for N-acetylglucosamine and the cytoplasm for N-acetylgalactosamine residues. The intestine lacked a caecum and appendix. The small intestine was divided into duodenum, jejunum‑ileum and ileum‑colon. The epithelium consisted of columnar enterocytes and goblet cells. The large intestine was short, only represented by the descending colon-rectum. It lacked villi and the mucosa had only crypts of Lieberkühn. Towards the colon-rectum, goblet cells produced mucus with N-acetylglucosamine residues increasing in acidity except in colon-rectum where acidity was highest in the base of crypts. Along the tube the surface of enterocytes was positive for N-acetylglucosamine and N-acetylgalactosamine. All over the mucus filling

  2. Histological and histochemical analysis of the gastrointestinal tract of the common pipistrelle bat (Pipistrellus pipistrellus

    Directory of Open Access Journals (Sweden)

    S. Strobel

    2015-04-01

    Full Text Available Bats have a very high mass-specific energy demand due to small size and active flight. European bat species are mostly insectivorous and the morphology of the gastrointestinal tract should be adapted accordingly. This study investigated the general anatomy by histology and the function by analysing carbohydrate distribution in particular of the mucus of the GI tract of the insectivorous bat Pipistrellus pipistrellus. The GI tracts of three individuals were dissected, fixed in formaldehyde, and embedded in paraffin wax. The tissues and cells of the GI tract of P. pipistrellus were analysed by classical (Acid Alizarin Blue, Haematoxylin-Eosin, and Masson Goldner Trichrome, histochemical (periodic acid-Schiff, Alcian blue at pH 2.5 and lectin histochemical (lectins WGA and HPA staining procedures. The GI tract of P. pipistrellus was organised into the typical mammalian layers. The short, narrow, and thin-walled esophagus was simple with a folded stratified squamous epithelium without glands but mucous surface cells secreting neutral mucus. The stomach was globular shaped without specialisation. Mucous surface cells produced neutral mucus whereas neck and parietal cells secreted a mixture of neutral and acid mucus. Chief cell surface was positive for N-acetylglucosamine and the cytoplasm for N-acetylgalactosamine residues. The intestine lacked a caecum and appendix. The small intestine was divided into duodenum, jejunum‑ileum and ileum‑colon. The epithelium consisted of columnar enterocytes and goblet cells. The large intestine was short, only represented by the descending colon-rectum. It lacked villi and the mucosa had only crypts of Lieberkühn. Towards the colon-rectum, goblet cells produced mucus with N-acetylglucosamine residues increasing in acidity except in colon-rectum where acidity was highest in the base of crypts. Along the tube the surface of enterocytes was positive for N-acetylglucosamine and N-acetylgalactosamine. All over the

  3. WHI-131 Promotes Osteoblast Differentiation and Prevents Osteoclast Formation and Resorption in Mice.

    Science.gov (United States)

    Cheon, Yoon-Hee; Kim, Ju-Young; Baek, Jong Min; Ahn, Sung-Jun; Jun, Hong Young; Erkhembaatar, Munkhsoyol; Kim, Min Seuk; Lee, Myeung Su; Oh, Jaemin

    2016-02-01

    The small molecule WHI-131 is a potent therapeutic agent with anti-inflammatory, antiallergic, and antileukemic potential. However, the regulatory effects of WHI-131 on osteoblast and osteoclast activity are unclear. We examined the effects of WHI-131 on osteoblast and osteoclast differentiation with respect to bone remodeling. The production of receptor activator of nuclear factor kappa-B ligand (RANKL) by osteoblasts in response to interleukin (IL)-1 or IL-6 stimulation decreased by 56.8% or 50.58%, respectively, in the presence of WHI-131. WHI-131 also abrogated the formation of mature osteoclasts induced by IL-1 or IL-6 stimulation. Moreover, WHI-131 treatment decreased RANKL-induced osteoclast differentiation of bone marrow-derived macrophages, and reduced the resorbing activity of mature osteoclasts. WHI-131 further decreased the mRNA and protein expression levels of c-Fos and nuclear factor of activated T cells, cytoplasmic 1 (NFATc1) by almost twofold, and significantly downregulated the mRNA expression of the following genes: tartrate-resistant acid phosphatase (TRAP), osteoclast-associated receptor (OSCAR), DC-STAMP, OC-STAMP, ATP6v0d2, and cathepsin K (CtsK) compared with the control group. WHI-131 further suppressed the phosphorylation of protein kinase B (Akt) and degradation of inhibitor of kappa B (IκB); Ca(2+) oscillation was also affected, and phosphorylation of the C-terminal Src kinase (c-Src)-Bruton agammaglobulinemia tyrosine kinase (Btk)-phospholipase C gamma 2 (PLCγ2) (c-Src-Btk-PLCg2 calcium signaling pathway) was inhibited following WHI-131 treatment. The Janus kinase-signal transducer and activator of transcription (JAK-STAT) signaling pathway was activated by WHI-131, accompanied by phosphorylation of STAT3 Ser727 and dephosphorylation of STAT6. In osteoblasts, WHI-131 caused an approximately fourfold increase in alkaline phosphatase activity and Alizarin Red staining intensity. Treatment with WHI-131 increased the mRNA expression

  4. The investigation of human periodontal ligament cell-sheet in vitro%人牙周韧带细胞膜片的体外实验研究

    Institute of Scientific and Technical Information of China (English)

    张剑英; 付云; 俞少杰

    2013-01-01

    Objective To investigate human periodontal ligament cells' capacities of osteogenesis and adipogenesis,and to identify the expression of type-Ⅰ collagen,laminin and fibronectin which are the main components of extracellular matrix in human periodontal ligament cell sheet through establish cell sheet model in vitro experiment.Methods Human periondontal ligament cells were purified and cultured by using enzymes digesting/tissue culture method.Immunocytochemistry was applied to identify the source and putative stem-cell marker STRO-1 of hPDLCs.The second to fourth passages of hPDLCs were cultured in the osteogenic medium and adipogenic medium for indicated days.Alizarin red and Oil red O staining method were applied to identify capabilities of osteogenesis and adipogenesis of hPDLCs.Haematoxylin-eosin staining and immunohistochemistry were applied to identify expression of type-Ⅰ collagen,laminin and fibronectin in hPDLCs cell sheet.Results The cultured hPDLCs demonstrated extensive proliferation and could be expanded in vitro.Immunostaining demonstrated that the cells expressed intermediate filament protein vimentin and the mesenchymal stem-cell marker STRO-1.hPDLCs in the osteogenic medium formed alizarin red-positive nodules.When hPDLCs were incubated in adipogenic medium,the cells developed into oil red O-positive lipid clusters and a significant high expression of type-Ⅰ collagen,laminin and fibronectin in human periodontalligament cell sheet.Conclusions Our findings confirm that hPDLCs contained stem cells population from culture,source identification,expression of the mesenchymai stem cell marker STRO-1,multilineage differentiation potent under defined culture conditions.Our study indicates that the hPDLCs turn out to be an efficient source and cell sheet to be an excellent bio-material that can be used for periodontal tissue regeneration.%目的 观察体外培养人牙周韧带细胞(hPDLCs)的成骨、成脂能力;构建hPDLCs膜片并鉴定膜片

  5. 成年比格犬骨髓间充质干细胞体外定向成骨诱导实验研究%An in vitro experimental study on the induction of bone marrow mesenchymal stem cells of adult beagle dogs into osteoblasts

    Institute of Scientific and Technical Information of China (English)

    许蕾; 李家锋; 唐巍; 韩建国; 孙晋虎

    2015-01-01

    Objective To cultivate the bone marrow mesenchmyal stem cells ( BMSCs) of adult beagle dogs in vitro and observe their morphology.These cells were then induced into osteoblasts, facilitating the development of bone tissue engineering in the future.Methods The BMSCs were first collected from adult beagle dogs and then cultivated in vitro using the semi-marrow adherence method.The second generation of BMSCs were divided into the following two groups:an experimental group with addition of fetal bovine serum ( FBS) as well as equivalent amounts of Inducer 1 ( dexametha-sone,β-glycerol phosphate disodium salt pentahydrate, and L-ascorbic acid) and Inducer 2 ( BMP-2) , and a con-trol group cultured in media containing FBS alone.Then, on Days 3, 7, 14, and 21 of induction, both groups were de-termined for alkaline phosphatase ( ALP) activity, followed by alizarin red staining and ALP staining three weeks later. The differentiation of induced osteoblasts was identified by Von-Kossa staining.Results On Days 3, 7, 14, and 21 of induction, the activity of ALP in the experimental group became weakened, despite after initial enhancement.In con-trast, no significant changes were found in the control group.After three weeks of induction, positive results were shown for alizarin red staining, ALP staining and Von-Kossa staining in the experimental group, in comparison with negative staining in the control group.These findings were consistent with the characteristics of osteoblasts, indicating the forma-tion of osteoblasts in the experimental group.Conclusion The BMSCs of adult beagle dogs can directionally differentiate into osteoblasts in the presence of an inducing agent.%目的:通过将成年比格犬骨髓间充质干细胞(BMSCs)建立体外培养体系,观察及掌握细胞生长形态,运用诱导剂体外定向诱导分化为成骨细胞,为后期建立骨组织工程提供种子细胞。方法提取成年比格犬的BMSCs,运用半骨髓直接贴壁法进

  6. Preliminary study of influence of bone tissue from osteonecrosis of femoral head on the proliferation and differentiation of canine bone marrow mesenchymal stem cells%股骨头坏死骨组织对骨髓间充质干细胞增殖分化的影响

    Institute of Scientific and Technical Information of China (English)

    王萌; 廖琦; 周斌; 仇志强; 程立明

    2013-01-01

    Objective To examine the effects of bone tissue from osteonecrosis of femoral head on the proliferation and differentiation of canine bone marrow mesenchymal stem cells in vitro culture.Methods A canine model of femoral head osteonecrosis was induced by liquid nitrogen freezing.BMSC were isolated from dog ilium bone marrow by a combination of gradient centrifugation and adherent wall culture.Different bone tissues and BMSC were cultivated indirectly in vitro by co-cultured in Transwell plate.According to the culture media,3 groups were established:blank group (10% FBS/DMEM),control group (10% FBS/DMEM + bone tissue from natural femoral head) and experimental group(10% FBS/DMEM + bone tissue from osteonecrosis of femoral head).Cell proliferation was measured by methylthiazol tetrazolium (MTT)method.Cell differentiation was examined by alkaline phosphatase (ALP) staining and its concentration examined.Alizarin red staining method was used to study the calcification effects and Oil red O staining method was used to detect if there was fat emergence.Results As compared with the blank group,the proliferation in the control and experiment groups were significantly promoted after culturing for Days 1,3 and 5 (P < 0.05).The proliferation of the experiment group was higher than the control group at Day 5 and 7 day (P < 0.05).After a 7-day co-culturing,ALP staining was positive in the control and experiment groups.At Day 7 and 9,the ALP activity in culture fluid was in this order:control group > experiment group >blank group(P <0.05).Alizarin red staining show control group had the most calcium nodules(12.17 ±2.48,P < 0.05) and the number of calcium nodules in the experiment group was more than the blank group (P <0.05).Oil red O staining show there was no fat emergence after 21 days in every group.Conclusion Both natural and osteonecrotic bone tissue of femoral head could promoted the proliferation of canine BMSC and induces them osteogenic

  7. Repair of calvarial defects with human umbilical cord blood derived mesenchymal stem cells and demineralized bone matrix in athymic rats%人脐血间充质干细胞修复颅骨缺损的实验研究

    Institute of Scientific and Technical Information of China (English)

    刘广鹏; 李宇琳; 孙剑; 崔磊; 张文杰; 曹谊林

    2010-01-01

    Objective To investigate the feasibility of using human umbilical cord blood derived mesenchymal stem cells (UCB-MSCs) and demineralized bone matrix (DBM) scaffolds to repair critical-sized calvarial defects in athymic rats. Methods Human UCB-MSCs were isolated, expanded and osteogenically induced in vitro. Osteogenic differentiation of UCB-MSCs was evaluated by Alizarin Red staining and measurement of calcium content respectively, and then the cells were seeded onto DBM scaffolds. Bilateral full-thickness defects (5 mm in diameter) of parietal bone were created in an athymic rat model. The defects were either repaired with UCB-MSC/DBM constructs (experimental group) or with DBM scaffolds alone (control group). Animals were harvested at 6 and 12 weeks post-implantation respectively, and defect repair was evaluated with gross observation, micro-CT measurement and histological analysis. Results Micro-CT showed that new bone was formed in the experimental group at 6 weeks post-implantation, while no sign of new bone formation was observed in the control group. At 12 weeks post-transplantation, scaffolds had been degraded almost completely in both sides. It was shown that an average of (78.19±6.45)% of each defect volume had been repaired in experimental side; while in the control side, only limited bone formed at the periphery of the defect. Histological examination revealed that the defect was repaired by trabecular bone tissue in experimental side at 12 weeks, while only fibrous connection was observed in the control group. Conclusions Tissue-engineered bone composed of osteogenically-induced human UCB-MSCs on DBM scaffolds could successfully repair the critical-sized calvarial defects in athymic rat models.%目的 应用人脐血间充质干细胞(umbilical cord blood derived mesenchymal stem cells,UCB-MSCs)复合脱钙骨材料构建组织工程化骨,修复裸大鼠颅骨标准缺损.方法 体外扩增培养、成骨诱导人UCB-MSCs,采用Alizarin Red染色

  8. 模拟微重力影响人牙髓干细胞的矿化能力与RhoA-Rho激酶信号通路相关性研究%Effects of simulated microgravity on mineralization of hDPSCs via RhoA-Rho signaling pathway

    Institute of Scientific and Technical Information of China (English)

    牛玉梅; 张巍巍; 曹涛; 李艳萍; 刘会梅; 贾丛辉

    2016-01-01

    sialoprotetin) and DSPP (dentin sialophosphoprotein) were detected by Western Blot for 10d, and mineralized nodules were detected by Alizarin red staining for 21d. Results The expression level of RhoA was down-regulated under microgravity. Treated with Y-27632 in normal gravity after 3, 5, 7 and 10 days, the AKPase activity was significantly lower than control. After 21 days’ cul-ture, the Alizarin red staining showed that the positive granules were hardly detected. The expression levels of DSP, DSPP and DMP-1 were lower than those in the control group after 10 days of culture. Conclusion Simulated microgravity inhibited the expression of RhoA. Inactivation of Rho kinase by addition of inhibitor Y-27632 caused lower mineralization ability of hDPSCs. It is possible that RhoA-Rho kinase signaling pathway is involved in the effect of simulated microgravity on mineralization process of hDPSCs.

  9. Effect of microgravity on bone mesenchymal stem cells differentiation to osteogenesis%微重力对骨髓间充质干细胞向成骨细胞分化影响的研究

    Institute of Scientific and Technical Information of China (English)

    李煜; 孙明林; 张新昌; 韩标; 赵滨; 李瑞欣; 李昊; 张西正

    2015-01-01

    Objective To study the effect of microgravity on bone mesenchymal stem cells (BMSCs) differentiating to osteoblast (OB).Methods BMSCs were harvested from femur of mouse and then subcultured in vitro to the 4th generation.The cells were then divided into four groups:(1) normal gravity group(NG group),normal gravity without osteogenic induction.(2) microgravity group (MG group),microgravity without osteogenic induction.(3) normal gravity induction group (NG+Ⅰ group),normal gravity with osteogenic induction.(4) microgravity induction group (MG+Ⅰ group),microgravity with osteogenic induction,and all cells were cultured for 10 d.Cell proliferation was tested by thiazolyl blue tetrazolium bromide (MTT) assay,alizarin red staining was used to determine extracellular matrix mineralization,and alkaline phosphatase activity (ALP) was measured to calculate its expression in the extracellular matrix.Real-time quantitative PCR (qPCR) and Western Blot were used to evaluate the expression of osteoblast-related gene and protein.Results The numbers of BMSCs in microgravity condition were more than that in normatgravity condition after 4 days’ culture (P<0.05).Alizarin red staining results showed that after 10 days’ induction,lots of calcium nodules were found in NG+Ⅰ group,while barely nodules could be found in microgravity group at the same time.Some nodules also could be found in NG group and MG+Ⅰ group.The qPCR demonstrated that microgravity condition inhibits the expression of osteogenic gene (COL Ⅰ,ALP,RUNX-2).The relative values of the three genes mentioned above in NG group and MG group were 2.41 ±0.30 vs 0.56±0.19,2.15±0.12 vs 0.16±0.15,0.98±0.12 vs 0.16±0.05 (P<0.05).The relative values of the three genes in MG+Ⅰ group were also less than that in NG+Ⅰ group:1.79±0.10 vs 0.59±0.04,1.57±0.19 vs 0.57±0.08,1.30±0.14 vs 0.74±0.05 (P<0.05).The Western Blot results were nearly consistence with qPCR,and the osteogenesis-related protein was

  10. Embryonic skeleton development and neonatal learning and memory ability of rats anesthetized with pentobarbital sodium: Differences of administration occasion and time

    Institute of Scientific and Technical Information of China (English)

    Changling Peng; Yuhua Zhu; Ankang Hu; Xiaorong Zhu

    2006-01-01

    BACKGROUND : Generally speaking, anesthesia is often used in gravid body and it has been already proved that many kind of medicine can result in malformation.OBJECTIVE: To explore embryonic skeleton development and neonatal learning and memory of rats anesthetized with pentobarbital sodium in gravid rats.DESIGN: A randomized control trial.SETTING: Laboratory Animal Center of Xuzhou Medical College.MATERIALS: A total of 80 adult female SD rats, of clean grade and weighing 220-240 g, were selected in this study. The main reagents were detailed as follows: pentobarbital sodium (Shanghai Xingzhi Chemical Plant,batch number: 921019); MG-2 maze test apparatus (Zhangjiagang Biomedical Instrument Factory);somatotype microscope (Beijing Taike Instrument Co., Ltd.).METHODS: ① A total of 160 SD rats of half males and females were selected in this study. All rats were copulated. The day that the plug was checked out in the vagina next day was looked as the first day of pregnancy. Gravid rats were divided randomly into four groups, including early anesthesia group, second anesthesia group, late anesthesia group and control group with 20 in each group. Rats in the early anesthesia group were injected with 25 mg/kg soluble pentobarbitone on the 7th day of pregnancy for once; rats in the second anesthesia group were anesthetized with 25 mg/kg soluble pentobarbitone on the 7th and the 14th days of pregnancy for once; rats in the late anesthesia group were anesthetized with 25 mg/kg soluble pentobarbitone on the 14th day of pregnancy for once; rats in the control group did not treat with anything. The time of anesthetizing was controlled in 3 to 4 hours and ether was absorbed while the time was not enough. ②Half of each group was sacrificed on day 20th of pregnancy and the fetus was taken out to be stained with alizarin red S. After stained, the fetal skeleton was examined. The learning and memorizing of one-month rats that were given birth by the rest gravid rats were tested

  11. Osteogenic and adipogenic differentiation of rabbit adipose-derived mesenchymal stem cells in vitro%体外培养兔脂肪源性间充质干细胞的成骨成脂分化

    Institute of Scientific and Technical Information of China (English)

    李受珉; 吴子征; 王泽; 李智; 张键

    2014-01-01

    wel as to investigate the osteogenic and adipogenic potentials of adipose-derived mesenchymal stem cells in vitro. METHODS:Primary adipose-derived mesenchymal stem cells were isolated from the subcutaneous adipose tissue of posterior cervical region from New Zealand white rabbits and digested by 0.1%col agenase I. The cells were passaged and amplified by the trypsin digestion. The passage 4 adipose-derived mesenchymal stem cells were induced to differentiate after exposure to adipogenic or osteogenic medium. The oil red O staining, alkaline phosphatase and alizarin red staining were used to detect the results. The cellviability was detected by the cellcounting kit 8 method to drawn the growth curve. cellsurface markers were examined using flow cytometry. RESULTS AND CONCLUSION:The adipose-derived mesenchymal stem cells isolated from the subcutaneous adipose tissue of rabbits exhibited a fusiform adherent growth in a vortex pattern, and had a strong capability of proliferation that could be passaged stably to the 10th generation. Flow cytometry results showed that the cells highly expressed CD29, CD90, CD44, but lowly expressed CD45 and CD34. After adipogenic induction, the adipose-derived mesenchymal stem cells were positive for oil red O staining;after osteogenic induction, the cells were both positive for alkaline phosphatase and alizarin red staining. These findings suggest that the adipose-derived mesenchymal stem cells were successful y isolated and cultured from the subcutaneous adipose tissue of rabbits, and these cells are pluripotent with the potential to differentiate into adipocytes and osteoblasts, which are expected to be ideal seed cells for bone tissue engineering.

  12. 骨关节炎患者膝关节滑膜组织来源间充质干细胞的增殖与分化%Proliferation and differentiation of mesenchymal stem cells from the synovial tissue in patients with osteoarthritis

    Institute of Scientific and Technical Information of China (English)

    陈路; 夏先学; 蒋科

    2015-01-01

    BACKGROUND:With respect to mesenchymal stem cels from other sources, synovial mesenchymal stem cels are rich in source, and moreover, the synovial tissue can regenerate quickly after partial hepatectomy and lead to fewer complications, in recent year, which have become a hot spot in stem cel research. OBJECTIVE:To observe the proliferation and directional differentiation of synovial mesenchymal stem cels from osteoarthritis patients. METHODS:Synovial mesenchymal stem cels were isolated and cultured. MTT assay was used to detect cel proliferation ability. Alkaline phosphatase activity was detected quantitatively at 7 days of osteogenic induction, and osteogensis-related gene expression was measured at 7, 14, 21 days of osteogenic induction. Alizarin red staining was performed at 21 days of induction. RESULTS AND CONCLUSION:(1) Passage 3 synovial mesenchymal stem cels proliferated faster, which were in latent period at 1, 2 days after inoculation, in logarithmic growth phase at 3-6 days, and then entered into the plateau phase at 7 days. (2) The activity of alkaline phosphatase was significantly higher in the induction group than the control group at 3, 7, 10 days after osteogenic induction (P < 0.05). The cels were positive for alizarin red staining at 21 days of osteogenic induction, and there were calcium deposits and calcium nodules in the extracelular matrix. (3) Bone-binding protein and Runx2 were visible at 7 days of osteogenic induction, and reached the peak at 21 days. These findings indicate that synovial mesenchymal stem cels from patients with advanced osteoarthritis have strong proliferation ability, which can differentiate into osteoblasts under in vitro induction.%背景:相对于其他来源的间充质干细胞,滑膜间充质干细胞来源丰富,并且滑膜组织可以在部分切除后迅速再生,并发症少,已逐渐成为近年来干细胞研究的热点。目的:观察骨关节炎患者膝关节滑膜组织来源间充质干细

  13. 眼内麻醉剂对兔角膜内皮细胞影响的实验研究%Effect of intracameral anesthetics on rabbit cornea endothelium

    Institute of Scientific and Technical Information of China (English)

    马立威; 许军; 许明林; 张劲松

    2005-01-01

    目的:研究不同眼内麻醉剂对离体兔角膜内皮细胞的作用.方法:将实验用兔角膜取下后,分别用10g/L利多卡因、1g/L塞罗卡因及BSS处理.通过台盼蓝-茜素红联合染色观察兔角膜内皮细胞的形态学改变,并用计算机自动图像分析系统对兔角膜内皮细胞的损伤情况进行定量分析;对处理后的兔角膜内皮细胞进行扫描电镜观察,了解其超微形态学改变.结果:①各实验组中,10g/L利多卡因作用后兔角膜内皮细胞的损伤率分别为(0.91±0.12)%,(1.23±0.27)%,(2.42±0.31)%,(3.61±0.14)%;10g/L塞罗卡因作用后的兔角膜内皮细胞的损伤率分别为(0.68±0.16)%,(0.89±0.17)%,(1.84±0.34)%,(2.58±0.34)%.二者差异均具有显著性.(P<0.05,P<0.01).②眼内麻醉剂的作用时间与兔角膜内皮细胞的损伤率呈正相关,相关系数分别为0.974、0.976.③眼内麻醉剂作用后,10g/L利多卡因造成的兔角膜内皮细胞的损伤情况较10g/L塞罗卡因更为严重.结论:10g/L利多卡因及10g/L塞罗卡因都会对兔角膜内皮细胞造成损伤.比较而言,不含防腐剂的塞罗卡因更安全.%· AIM: To investigate the effects of different intraocular anesthetics on rabbit cornea endothelial cells in vitro.· METHODS: The excised rabbit corneae were treated with 10g/L lidocaine or 10g/L xylocaine for different durations. The morphological change was studied under a light microscope using dual staining of Typan-blue and Alizarine-red, and under a scanning electric microscope as well. The cornea endothelial cell (CEC) lost rate was calculated by computerized image analysis system.(Metamorph Image System V 4.6, Olympus Japan).· RESULTS: ①The CEC lost rate in 10g/L lidocaine groups for different durations was(0.91±0.12)%, (1.23±0.27)%, (2.42±0.31)% and (3.61±0.14)% respectively;while in 10g/L xylocaine groups was (0.68±0.16)%,(0.89±0.17)%, (1.84±0.34)% and (2.58±0.34)%, respectively. The difference was significant (P

  14. 小鼠脂肪来源干细胞向成骨及软骨细胞的诱导分化★%Adipose-derived stem cells differentiate into osteoblasts and chondrocytes

    Institute of Scientific and Technical Information of China (English)

    刘晓潭; 徐海斌; 路坦

    2013-01-01

    BACKGROUND: Adipose-derived stem cel s can be separated and obtained from fat tissue. Fat tissue distributes in the whole body, and can be easily obtained in large quantities and has less damage to the donor site when drawing. OBJECTIVE: To identify the methods of in vitro isolating and culturing of mice adipose-derived stem cel s, to induce the adipose-derived stem cel s to differentate into chondrocytes and osteoblasts and to investigate the feasibility of being seed cel s in tissue engineering. METHODS: The adipose-derived stem cel s were isolated from the epididymal fat tissue of Kunming mice. Primary adipose-derived stem cel s were obtained and purified by col agenase Ⅰ digestion and differential adherence method. The adipose-derived stem cel s were induced with osteogenic induction medium, and then gomori alkaline phosphatase staining and alizarin red calcium nodules staining were performed to detect the differentiation of adipose-derived stem cel s; the adipose-derived stem cel s were induced with cartilage induction medium, and the toluidine blue staining, safranin-O staining and type Ⅱ col agen immunohistochemistry testing were performed to detect the differentiation of adipose-derived stem cel s. RESULTS AND CONCLUSION: The adipose-derived stem cel s were spindle-shaped and in adherent growth. After primary cultured for 7-9 days, the cel s could reach 90% confluence. After passaged to the third generation, the cel morphology was in consistency, and the growth curve of the passaged adipose-derived stem cel s presented “S” shape. The expressions of CD29 and CD44 antigens were positive detected with cel -specific antigen test, but the expressions of CD34 and CD45 were negative. After osteoblast-inducing culture, the differentiation of adipose-derived stem cel s towards osteoblasts was verified positively by alkaline phosphatase staining and alizarin red staining. After chondrocyte-inducing culture, the differentiation of adipose-derived stem cel s

  15. Comparison of the effects of human mesenchymal stem cells cultured by autologous serum into osteoblasts by extracorporeal shock waves versus dexamethasone%冲击波与地塞米松对自体血清培养的骨髓间充质干细胞成骨分化的比较研究

    Institute of Scientific and Technical Information of China (English)

    安佰京; 邢更彦

    2011-01-01

    ) and 10% FBS (the control group), respectively.The cell morphology, the cell adhesion rate for 24 - hour, the cell doubling time were observed by inverted microscope.The cellular surface antigen and the generation cycle were detected by flow cytometry and immunocytochemical staining,the hMSCs cultured by AS medium were induced to osteeblasts by method of ESWT ( extracorporeal shock waves therapy) and the traditional method of dexamethasone ( 10 nM), ascorbic acid (50 uM) and sodium glycerophosphate ( 10 mM), respectively.The ALP staining and Alizarin Red staining were used to detect the activity of ALP and the the expression differences of oteogenic genes ( including ALP, OP, OC, ON, Col Ⅰ) were analyzed by RT-PCR after inducion.[Results]The two groups of hMSCs had no significant differences in cell morphology, cell adhesion rate, cell cycle and immunocytochemical staining during cell proliferation in AS or FBS medium.But the cell doubling time and cell proliferation rate of the hMSCs in AS medium was better than that of the hMSCs in FBS medium.The results of ALP quantitative, ALP staining, alizarin red staining and RT -PCR of oteogenic genes showed that the ESWT method was superior to the traditional method.[Conclusion]The ESWT is a better method to promote the osteoblastic differentiation of hMSCs than the traditional dexamethasone method.

  16. Osteogenic differentiation of nucleus puplousus cells co-cultured with autologous periosteal cells%骨膜细胞与髓核细胞共培养向成骨方向的分化

    Institute of Scientific and Technical Information of China (English)

    杨宇明; 袁峰; 陆海涛; 张峻伟; 盛晓磊; 李智多

    2015-01-01

    BACKGROUND:Periosteal cel s have been used in bone repair, but whether nucleus puplousus cel s co-cultured with autologous periosteal cel s can differentiate into osteoblasts in spinal fusion is rarely reported. OBJECTIVE:To isolate nucleus puplousus cel s and periosteal cel s so as to observe the osteogenic ability of nucleus puplousus cel s co-cultured with periosteal cel s or not. METHODS:Type II col agenase digestion method was used to isolate and purify nucleus pulposus cel s, which were confirmed by toluidine blue and immunohistochemical staining. Periosteal cel s were isolated histological y and cultured in complete medium, and cel surface antigens CD90, CD105 were identified by immunofluorescence staining. According to the experimental needs, the cel s were assigned into two groups. Nucleus pulposus cel s and periosteal cel s were co-cultured by osteogenic induction medium in the experimental group. Nucleus pulposus cel s in the control group were cultured alone in osteogenic induction medium. Cel morphology was observed by inverted microscopy, and cel proliferation was detected by cel counting kit-8. The osteogenic differentiation indexes of cel s in each group were measured using alkaline phosphatase staining, alizarin red staining, and type I col agen immunohistoehemical staining. The expression of osteopontin was tested by western blot assay. RESULTS AND CONCLUSION:CD105 and CD90 expressions of the periosteal cel s were positive. Nucleus puplousus cel s were positive for toluidine blue and col agen type II immunohistochemical staining. The proliferative ability of nucleus puplousus cel s was significantly higher in the experimental group than the control group at days 1, 3, 5, 7, 9. After 2 weeks of induction, the cel s were positive for alkaline phosphatase staining, alizarin red staining, and type I col agen immunohistoehemical staining, but the experimental group showed higher positive expressions than the control group (P  目的:分离培

  17. 兔骨膜细胞分离培养的方法改进%A modified method forin vitroisolation and cultivation of periosteal cells in rabbits

    Institute of Scientific and Technical Information of China (English)

    张峻玮; 陆海涛; 袁峰; 杨宇明

    2016-01-01

    BACKGROUND:Periosteum is considered as a source of seed cels for cel therapydue toits biological features. OBJECTIVE:To seek the optimal way to isolate and culture rabbit periosteal cels and identify their biological features. METHODS:Rabbit periosteum on facies medialis tibiae was taken out under aseptic conditions. Periosteal cels isolated through the digestion of type II colagenase with the explants culture method were cultured in DMEM/F12 complete medium. Cel ultrastructure was observedunderan inverted microscope. Periosteal cel proliferation was determined bycel counting kit-8assay. Cel surface antigensCD90 and CD105 were determined using flow cytometry. Osteogenic andlipogenic induction mediums were applied to induce periosteal cels to differentiate into osteocytes and adipocytes, respectively. After 2 weeks of induction, cels were harvestedfor alizarin red staining and oil red O staining to assay the calciumnodules and lipid droplet. RESULTS AND CONCLUSION:The digestion of type II colagenase with the explants culture method shortened the period of primary cels culture and enhanced the survival rate, which causedhigher purity and stronger reproductive activity of harvested periosteal cels. Primary cultured periosteal cels grew in form of spindle spiral or paralel. Alizarin red andOil red O staining verified the multi-directional differentiation potentiality of periosteal cels. These findings suggest that the periosteal cels with high purity,strong reproductive activity,andmulti-directional differentiation potentialitycanbe harvested in short time using digestion of type II colagenase with the explants culture method.%背景:近年来,骨膜作为细胞治疗的种子细胞来源,因其自身的优越性而备受关注。目的:探讨兔骨膜细胞分离培养的最佳方法及其生物学特性。方法:无菌条件下取出兔胫骨内侧面骨膜,以Ⅱ型胶原酶消化结合组织块贴壁法分离培养兔骨膜细胞,置于DMEM/F12完全

  18. Dermis-derived cell subpopulation is used to repair mouse calvarial defects%真皮来源细胞亚群修复小鼠颅骨缺损

    Institute of Scientific and Technical Information of China (English)

    王庭亮; 何金光; 张阳; 李丹; 董佳生; 祝联

    2015-01-01

    BACKGROUND:In consideration of skin as the largest organ al over the body and its abundant vessels and vessel plexuses, there would be sufficient adult stem cels for tissue engineering. OBJECTIVE:To investigate the osteogenic potential of dermis-derived bone morphogenetic protein receptor subtype IB (BMPR-IB) positive cels. METHODS:In current study, histochemical analysis was adopted to study the localization and expression of BMPR-IB+ cels in skin. Fresh skin samples were digested into single cel suspension. Then, the surface marker BMPR-IB was used to isolate cel subpopulation by magnetic activated cel sorting from freshly prepared single cel suspension. After that, the osteogenic potential in vitro andin vivo was tested. Alkaline phosphatase staining and alizarin red staining were performed after osteogenic inductionin vitro. The BMPR-IB+ cels were seeded onto coral scaffolds, and the scaffolds were used to repair critical-sized calvarial defects of mice. Histochemical analysis was performed at 6 weeks postoperatively and micro-CT analysis was carried out at 24 weeks postoperatively to evaluate the ability of bone repairment. RESULTS AND CONCLUSION:We localized BMPR-IB cels in situ by immunohistochemistry that turned out to be expressed in the reticular layer of dermis and by single cels. Cel subpopulation which expressed BMPR-IB could be sorted by magnetic activated cel sorting. Alkaline phosphatase staining was obviously positive and lots of calcium modules were confirmed by alizarin red staining after osteogenic induction, indicating that BMPR-IB+ cels had the osteogenic potentialin vitro. Histochemical analysis demonstrated that plenty of new bone formation was found in BMPR-IB+ cels group after 6 weeks in vivo. Micro-CT analysis revealed that BMPR-IB+ cels-coral scaffold complex could repair calvarial defects successfuly after 24 weeksin vivo. These results indicated that dermis-derived BMPR-IB+ cels possessed adequate osteogenic potential. Moreover, they

  19. Time Effect of Sinusoidal Electromagnetic Field on Enhancing the Maturation and Mineralization of Osteoblasts in Vitro%正弦交变电磁场促进体外培养成骨细胞成熟矿化的时间效应

    Institute of Scientific and Technical Information of China (English)

    周建; 葛宝丰; 陈克明; 李志锋; 程国政; 王嘉琪; 明磊国; 韦哲

    2011-01-01

    The present research was to investigate the time effect of sinusoidal electromagnetic fields(SEMFs)at different exposure time on the proliferation and differentiation of osteoblasts (OB) in vitro. The newborn rat calvarial OB were isolated by enzyme digestion and divided randomly into 7 groups after one passage. The exposure times of the SEMFs were 0. 5h, 1. Oh, 1. 5h, 2. Oh, 2. 5h and 3. Oh, respectively, and the frequency was 50Hz. The cells were exposed in the SEMFs of 1.8mT. Those without SEMFs exposure were used as the control group. They were observed under the contrast phase microscope each day. After 48h, cell proliferation was assayed by MTT method. The alkaline phosphatase (Alkaline Phosphatase, ALP) activities were measured after the exposure of SEMFs for 3d, 6d, 9d and 12d, respectively. The calcified nodules were stained by Alizarin Bordeaux after lOd. The cells exposed in the SEMFs were arranged in Spiral appearance after 8d. The SEMFs exposure time at 2. Oh, 2. 5h and 3. Oh significantly inhibited cell proliferation (P<0. 01) and 0. 5h, 1. 0h,l. 5h groups more significantly than control groups (P<0. 05). When the 3d, 6dand 12 d the ALP activities of the 0. 5h, 1. Oh, 1. 5h and 2. Oh, times group were significantly higher than those in the control group (P<0. 05), and after 9d the 1. 0h,l. 5h and 2. Oh activity of ALP higher significantly than control and other groups(P<0. 01). Other groups had no effect on the ALP activity. Alizarin Bordeaux staining result showed the amounts of calcified nodules 1. 0h,l. 5h and 2. Oh higher than control groups. The SEMFs at 50Hz, 1. 8mTdifferent time exposure groups inhibits the proliferation of OB, but they enhances the maturation and mineralization of the OB and SEMFs at 1. 8mT of the 1. 5h has the strongest activity.%研究正弦交变电磁场(SEMFs)促进体外培养成骨细胞(OB)成熟矿化的时间效应.OB培养后随机分成7组,其中6组用50 Hz、1.8 mT SEMFs处理,处理时间为每天0.5、1.0、1

  20. Study of proliferation,osteogenesis and senescence of bone marrow stromal cells from a cleidocranial dysplasia patient%颅骨锁骨发育不全患者骨髓基质细胞的增殖、成骨和衰老

    Institute of Scientific and Technical Information of China (English)

    张娟; 戈杰; 李光南; 周培培; 江宏兵

    2015-01-01

    目的:骨髓基质细胞(bone marrow stromal cells,BMSCs)在调节颅骨锁骨发育不全(cleidocranial dysplasia,CCD)患者骨结构中发挥关键作用,本研究通过与正常 BMSCs 比较分析,探讨 CCD 患者 BMSCs 的体外增殖、成骨分化、干性及衰老特征。方法:分离培养 CCD 患者及正常同龄人 BMSCs;甲基噻唑基四唑(MTT)法及流式细胞周期分析其增殖能力;成骨诱导后采用Western blot 及茜素红染色分析其成骨能力;检测多潜能转录因子及克隆形成,分析其干性能力;检测衰老调控关键基因 p16、p21表达及通过β-gal 衰老染色分析其衰老特征。结果:与正常 BMSCs 相比,BMSCs-CCD 增殖活性低,且细胞周期中处于 S期、G2的细胞比例较低;两组细胞成骨诱导1、3、7 d 后,BMSCs-CCD 组中 Runt 相关转录因子2(Runt-related transcription factor 2,Runx2)、成骨细胞特异性转录因子 Osterix、骨桥蛋白(Osteopontin,Opn)表达水平较正常组低;成骨诱导14 d 后,BMSCs-CCD组形成的钙化结节较正常组少;BMSCs-CCD 组中多潜能转录因子 Oct4、Nanog、Sox2表达及克隆形成率均较正常组低;相反, BMSCs-CCD 组中 p16、p21等衰老标记分子表达及衰老细胞阳性染色比例均较正常组高。结论:同正常 BMSCs 比较,CCD 患者 BMSCs 的增殖能力、成骨能力、干性强度均较差,且更易衰老。这些特征可能是 CCD 患者易发骨质疏松及骨折的生物学机制之一。%Objective:To study the in vitro biologic characteristics of bone marrow stromal cells with cleidocranial dysplasia (BM-SCs-CCD),including osteogenesis,proliferation ability,stemness and senescence.Methods:MTT and cell cycle detection for prolifera-tion ability,Western blot and alizarin red staining for osteogenesis ability,Western blot and clony-formation for stemness ability,Western blot and senescence staining for senescence characteristics

  1. Extraction and identification of human adipose-derived stem cells%人脂肪干细胞的提取和鉴定

    Institute of Scientific and Technical Information of China (English)

    吴尉; 梁芳; 宋小琴; 胡平安; 刘敏

    2015-01-01

    BACKGROUND:Adipose-derived stem cel s are totipotent stem cel s in the adipose tissue, and have the function of self-renewal and multi-directional differentiation. Human adipose-derived stem cel s are ideal seed cel s with stable genetic milieu and few rejections. OBJECTIVE:To extract human adipose-derived stem cel s from human omental adipose tissue and to identify the cel s by adipogenic and osteogenic induction. METHODS:Omental adipose tissues were col ected from surgical patients to isolate and culture adipose-derived stem cel s using type I col agenase digestion, filtration and centrifugation. Cel growth was observed and proliferative curve of human adipose-derived stem cel s were drawn by cel counting method to calculate the doubling time at logarithmic growth phase. After adipogenic and osteogenic induction, induced cel s were identified using oil red O and alizarin red staining, respectively. RESULTS AND CONCLUSION:Human adipose-derived stem cel s were successful y isolated from the omentum tissues of surgical patients. Adherent cel s were fusiform-shaped and like fibroblasts. The growth curve of passage 3 cel s was in S shape, and the doubling time was 45.90 hours. After adipogenic and osteogenic induction for 2 and 3 hours, respectively, oil red O staining showed unequal-sized orange fat droplets, and alizarin red staining showed typical calcified nodules that were in orange. These findings indicate that adipose-derived stem cel s have the adipogenic and osteogenic capacity.%背景:脂肪干细胞是存在于脂肪中的全能干细胞,具备自我更新能力与多向分化潜能,遗传背景相当稳定,体内植入后免疫排斥少,是一种比较理想的种子细胞。目的:提取人大网膜脂肪干细胞,并进行成脂和成骨分化能力鉴定。  方法:收集手术患者大网膜的脂肪组织,经Ⅰ型胶原酶消化、过滤、离心后进行原代培养,观察细胞生长状态;用细胞计

  2. 绿色荧光蛋白基因转染示踪比格犬体内骨髓间充质干细胞分化%Green fluorescent protein genes transdution and tracer the differentiation of bone mesenchymal stem cells in beagles

    Institute of Scientific and Technical Information of China (English)

    张海峰; 杜子婧; 赵丹阳; 韩修国; 韩冬

    2016-01-01

    Objective To observe differentiation and transformation of bone mesenchymal stem cells (BMSCs)labeled with green fluorescent protein (GFP)technology in bone tissue engineering.Methods Adenoviral delivery of GFP genes (Ad-GFP)transfected to canine BMSCs.The morphology of BMSCs,alkaline phosphatase (ALP)and alizarin red staining were observed with the help of invert light microscope,and the expression of GFP was scrutinized by using fluorescence microscope.Once succeeding,the centrifuged BMSCs were collected and mixed with beta-tricalcium phosphate (β-TCP),then they were transplanted under tibial periosteum.The beagles using the above way to construct tissue engineering bone were marked group A,which only withβ-TCP transplated into tibial periosteum were marked group B, and which withβ-TCP subcutaneously placed were marked group C.In the postoperative 2 weeks,histology and immunohistochemistry results were studied and the expression of GFP was observed by laser scanning confocal microscope.Results After induced differentiation in vitro, observing BMSCs found that ALP staining and alizarin red mineralization nodules staining were positive.Histology observation revealed that compared with group B,abundant bone formed and neovascularized significantly in group A,and bone formation did not exist in group C. Immunohistochemistry staining showed that type Ⅰ collagen and platelet endothelial cell adhesion molecule-1 (CD31)were positive in group A, type Ⅰ collagen and CD31 were positive partly in group B rather than group C.The expression of typeⅠcollagen and CD31 were analyzed in group A and group B by the way of semi-quantitative immunohistochemical staining.The value of type Ⅰ collagen and CD31 had significant differences between group A and group B.The expression of GFP was observed by laser scanning confocal microscope in group A not in group B nor group C.Conclusion BMSCs is the important factor to accelerate the generation of tissue engineering bone instead

  3. Isolation of human adipose-derived stem cells and the identification of biological characteristics%人脂肪源性干细胞的分离及生物学性状的鉴定

    Institute of Scientific and Technical Information of China (English)

    王洁晴; 柏树令; 侯伟健; 佟浩; 田晓红; 徐赫

    2011-01-01

    Objective To establish a method to isolate and culture adipose-derived stem cells (ASCs) from the human liposuction aspirates, and conduct observations of the cell morphology、 growth kinetics、 surface markers and differentiating capacity. Method Adipose tissues were obtained from 4 healthy adult women who were experienced abdominal liposuction. ASCs, from liposuction aspirates, were isolated by enzymatic digestion, and were cultured to passage 20, the morphology of the cultured cells was observed. The cell viability was evaluated with MTT, and compared among passage 3,9, 15 and 20. Cell growth curve was generated. The cell cycle and the surface marker profiles were detected by flow cytometry. Adipogenic differentiation and osteogenic differentiation of ASCs was assessed by oil red O and Alizarin Red staining respectively. Results The ASCs present a vortex pattern growth with a fibroblast-like appearance,and as shown by MTT, proliferation activity was strong when they subcultured to passage 15, then gradually slowed down,significantly reduced when they passed to passage 20. Statistical analysis showed that passage 20 and passage 3,9,15 were significantly different (P < 0.05 ). ASCs also showed characteristics of stem cell cycle. The positive expression of mesenchymal stem cell markers CD90, CD44 and negative expression of hematopoietic stem cell marker CD34, the blood cell marker CD45 ,the endothelial cell marker CD31 were observed in ASCs by flow cytometry. In addition, the expression of CD49d was low and of CD106 was negative. Oil red O staining of ASCs after adipogenic induction demonstrated numerous intracellular lipid droplets. Calcium nodules could seen after osteogenic induction and Alizarin red staining was positive. Conclusion ASCs can be isolated from human liposuction aspirates and expressing cell surface markers of stem cells with strong proliferative ability. ASCs also can be induced to differentiate into adipose tissue and osseous tissue under

  4. Study of cis- and trans-uranium elements by paper chromatography and electrophoresis; Contribution a l'etude des elements cis- et trans-uraniens par chromatographie sur papier et electrophorese

    Energy Technology Data Exchange (ETDEWEB)

    Clanet, F. [Commissariat a l' Energie Atomique, 92 - Fontenay-aux-Roses (France). Centre d' Etudes Nucleaires

    1968-01-01

    In this work, the field of application of paper chromatography and electrophoresis in inorganic chemistry has been extended to elements 90 to 96 in hydrochloric and nitric acid solution. Results obtained concern the following points: 1) - Characterization of the valency states of Np and of Pu using coloured reactions on chromatograms and electrophoregrams. The valency IV is characterized by alizarin, arsenazo-I and thorin-I, whilst diphenylcarbazide is used for the hexavalent state. 2) - Paper chromatography: by using as eluent, mixtures of equal parts of aqueous HCl and HNO{sub 3} solutions and of alcohols (methanol, ethanol and n-butanol), the R{sub f} values of elements 90 to 96 have been determined. It has been possible to deduce certain conclusions concerning the complexing of these elements by Cl{sup -} and NO{sub 3}{sup -} ions. 3) - We have developed an electrophoretic technique on cellulose acetate membranes in order to separate the charged species formed by the elements 90 to 96 in HCl and HNO{sub 3} solutions from 1 to 12 M. Mobility curves have been obtained. It appears from our results that the tendency for the elements considered to form anionic complexes follows the order of the ionic potentials when the valency state is four; this order is reversed for the valency three. The ions Cl{sup -} have a smaller tendency to form complexes than the NO{sub 3}{sup -} ions with respe