Multi-scale cell/surface interaction on modified titanium aluminum vanadium surfaces

Science.gov (United States)

Chen, Jianbo

This dissertation presents a series of experimental studies of the effects of multi-scale cell/surface interactions on modified Ti-6Al-4V surfaces. These include laser-grooved surfaces; porous structures and RGD-coated laser-grooved surfaces. A nano-second DPSS UV lasers with a Gaussian pulse energy profile was used to introduce the desired micro-groove geometries onto Ti-6Al-4V surfaces. This was done without inducing micro-cracks or significant changes in surface chemistry within the heat affected zones. The desired 8-12 mum groove depths and widths were achieved by the control of pulse frequency, scan speed, and the lens focal length that controls spot size. The interactions between human osteosarcoma (HOS) cells and laser-grooved Ti-6Al-4V surfaces were investigated after 48 hours of cell culture. The cell behavior, including cell spreading, alignment and adhesion, was elucidated using scanning electronic microscopy (SEM), immuno-fluorescence staining and enzymatic detachment. Contact guidance was shown to increase as grooved spacing decreased. For the range of micro-groove geometries studied, micro-grooves with groove spacings of 20 mum provided the best combination of cell orientation and adhesion. Short-term adhesion experiments (15 mins to 1 day) also revealed that there is a positive correlation between cell orientation and cell adhesion. Contact guidance on the micro-grooved surfaces is shown to be enhanced by nano- and micro-scale asperities that provide sites for the attachment of lamellopodia during cell locomotion and spreading. Contact guidance is also promoted by the geometrical confinement provided by laser grooves. An experimental study of initial cell spreading and ingrowth into Ti-6Al-4V porous structures was also carried out on porous structures with different pore sizes and geometries. A combination of SEM, the tetrazolium salt (MTT) colorimetric assay and enzymatic detachment were used to study cell spreading and adhesion. The extent of cell

Surface modified superparamagnetic nanoparticles: Interaction with fibroblasts in primary cell culture

Energy Technology Data Exchange (ETDEWEB)

Chapa Gonzalez, Christian; Roacho Pérez, Jorge A.; Martínez Pérez, Carlos A.; Olivas Armendáriz, Imelda [Instituto de Ingeniería y Tecnología, Universidad Autónoma de Ciudad Juárez, Ave. Del Charro #610 norte, Col. Partido Romero, C.P. 32320 Cd. Juárez, Chihuahua, México (Mexico); Jimenez Vega, Florinda [Instituto de Ciencias Biomédicas, Universidad Autónoma de Ciudad Juárez, Anillo envolvente del PRONAF y Estocolmo, C.P. 32320 Cd. Juárez, Chihuahua, México (Mexico); Castrejon Parga, Karen Y. [Instituto de Ingeniería y Tecnología, Universidad Autónoma de Ciudad Juárez, Ave. Del Charro #610 norte, Col. Partido Romero, C.P. 32320 Cd. Juárez, Chihuahua, México (Mexico); Garcia Casillas, Perla E., E-mail: pegarcia@uacj.mx [Instituto de Ingeniería y Tecnología, Universidad Autónoma de Ciudad Juárez, Ave. Del Charro #610 norte, Col. Partido Romero, C.P. 32320 Cd. Juárez, Chihuahua, México (Mexico)

2014-12-05

Highlights: • An inorganic layer before an organic material shell onto MNPs improves cell viability. • The coating type and the concentration of nanoparticles directly affect cell viability. • Modified magnetite nanoparticles with organic and inorganic materials was developed. - Abstract: The development of a variety of medical applications such as drug delivery, cell labeling, and medical imaging have been possible owing to the unique features exhibited by magnetic nanoparticles. Nanoparticle–cell interaction is related to the surface aspects of nanoparticle, which may be described based on their chemistry or inorganic/organic characteristics. The coating on particle surface reduces the inter-particle interactions and provides properties such as biocompatibility. Among the coating materials used for nanoparticles employed in biomedical applications, oleic acid is one of the most utilized due to its biocompatibility. However, a major drawback with this naturally occurring fatty acid is that it is easily oxidized by cells and this reduces their performance in biomedical applications. In order to avoid the direct contact of the cell with the magnetite particle, coating with an inorganic material prior to the oleic acid shell would be effective. This would retard the magnetite dissociation thereby improve the cell viability. Here we report our investigation on the effect of surface modified magnetite nanoparticles (MNPs) on the cell viability using primary cultures incubated with those particles. We prepared magnetite nanoparticles by chemical co-precipitation method; nanoparticle surface was first modified by silanol condensation followed by chemisorption of oleic acid. All nanostructures have a particle size less than 100 nm, depending on the material coating and superparamagnetic behavior. The saturated magnetizations (M{sub s}) of the magnetite samples coated with oleic acid (MAO; 49.15 emu/g) and double shell silica-oleic acid (MSAO; 46.16 emu/g) are

Bio-active molecules modified surfaces enhanced mesenchymal stem cell adhesion and proliferation

International Nuclear Information System (INIS)

Mobasseri, Rezvan; Tian, Lingling; Soleimani, Masoud; Ramakrishna, Seeram; Naderi-Manesh, Hossein

2017-01-01

Surface modification of the substrate as a component of in vitro cell culture and tissue engineering, using bio-active molecules including extracellular matrix (ECM) proteins or peptides derived ECM proteins can modulate the surface properties and thereby induce the desired signaling pathways in cells. The aim of this study was to evaluate the behavior of human bone marrow mesenchymal stem cells (hBM-MSCs) on glass substrates modified with fibronectin (Fn), collagen (Coll), RGD peptides (RGD) and designed peptide (R-pept) as bio-active molecules. The glass coverslips were coated with fibronectin, collagen, RGD peptide and R-peptide. Bone marrow mesenchymal stem cells were cultured on different substrates and the adhesion behavior in early incubation times was investigated using scanning electron microscopy (SEM) and confocal microscopy. The MTT assay was performed to evaluate the effect of different bio-active molecules on MSCs proliferation rate during 24 and 72 h. Formation of filopodia and focal adhesion (FA) complexes, two steps of cell adhesion process, were observed in MSCs cultured on bio-active molecules modified coverslips, specifically in Fn coated and R-pept coated groups. SEM image showed well adhesion pattern for MSCs cultured on Fn and R-pept after 2 h incubation, while the shape of cells cultured on Coll and RGD substrates indicated that they might experience stress condition in early hours of culture. Investigation of adhesion behavior, as well as proliferation pattern, suggests R-peptide as a promising bio-active molecule to be used for surface modification of substrate in supporting and inducing cell adhesion and proliferation. - Highlights: • Bioactive molecules modified surface is a strategy to design biomimicry scaffold. • Bi-functional Tat-derived peptide (R-pept) enhanced MSCs adhesion and proliferation. • R-pept showed similar influences to fibronectin on FA formation and attachment.

Preparation of immobilized glucose oxidase wafer enzyme on calcium-bentonite modified by surfactant

Science.gov (United States)

Widi, R. K.; Trisulo, D. C.; Budhyantoro, A.; Chrisnasari, R.

2017-07-01

Wafer glucose oxidase (GOx) enzymes was produced by addition of PAH (Poly-Allyamine Hydrochloride) polymer into immobilized GOx enzyme on modified-Tetramethylammonium Hydroxide (TMAH) 5%-calsium-bentonite. The use of surfactant molecul (TMAH) is to modify the surface properties and pore size distribution of the Ca-bentonite. These properties are very important to ensure GOx molecules can be bound on the Ca-bentonit surface to be immobilized. The addition of the polymer (PAH) is expected to lead the substrates to be adsorbed onto the enzyme. In this study, wafer enzymes were made in various concentration ratio (Ca-bentonite : PAH) which are 1:0, 1:1, 1:2 and 1:3. The effect of PAH (Poly-Allyamine Hydrochloride) polymer added with various ratios of concentrations can be shown from the capacitance value on LCR meter and enzyme activity using DNS method. The addition of the polymer (PAH) showed effect on the activity of GOx, it can be shown from the decreasing of capacitance value by increasing of PAH concentration.

Engineering the cell surface display of cohesins for assembly of cellulosome-inspired enzyme complexes on Lactococcus lactis

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Wieczorek Andrew S

2010-09-01

Full Text Available Abstract Background The assembly and spatial organization of enzymes in naturally occurring multi-protein complexes is of paramount importance for the efficient degradation of complex polymers and biosynthesis of valuable products. The degradation of cellulose into fermentable sugars by Clostridium thermocellum is achieved by means of a multi-protein "cellulosome" complex. Assembled via dockerin-cohesin interactions, the cellulosome is associated with the cell surface during cellulose hydrolysis, forming ternary cellulose-enzyme-microbe complexes for enhanced activity and synergy. The assembly of recombinant cell surface displayed cellulosome-inspired complexes in surrogate microbes is highly desirable. The model organism Lactococcus lactis is of particular interest as it has been metabolically engineered to produce a variety of commodity chemicals including lactic acid and bioactive compounds, and can efficiently secrete an array of recombinant proteins and enzymes of varying sizes. Results Fragments of the scaffoldin protein CipA were functionally displayed on the cell surface of Lactococcus lactis. Scaffolds were engineered to contain a single cohesin module, two cohesin modules, one cohesin and a cellulose-binding module, or only a cellulose-binding module. Cell toxicity from over-expression of the proteins was circumvented by use of the nisA inducible promoter, and incorporation of the C-terminal anchor motif of the streptococcal M6 protein resulted in the successful surface-display of the scaffolds. The facilitated detection of successfully secreted scaffolds was achieved by fusion with the export-specific reporter staphylococcal nuclease (NucA. Scaffolds retained their ability to associate in vivo with an engineered hybrid reporter enzyme, E. coli β-glucuronidase fused to the type 1 dockerin motif of the cellulosomal enzyme CelS. Surface-anchored complexes exhibited dual enzyme activities (nuclease and β-glucuronidase, and were

Recent Insights into Cell Surface Heparan Sulphate Proteoglycans and Cancer

DEFF Research Database (Denmark)

Couchman, John R; Multhaupt, Hinke; Sanderson, Ralph D

2016-01-01

behaviour. Here, we review some recent advances, emphasising that many tumour-related functions of proteoglycans are revealed only after their modification in processes subsequent to synthesis and export to the cell surface. These include enzymes that modify heparan sulphate structure, recycling of whole...

Growth and Functionality of Cells Cultured on Conducting and Semi-Conducting Surfaces Modified with Self-Assembled Monolayers (SAMs

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Rajendra K. Aithal

2016-02-01

Full Text Available Bioengineering of dermal and epidermal cells on surface modified substrates is an active area of research. The cytotoxicity, maintenance of cell phenotype and long-term functionality of human dermal fibroblast (HDF cells on conducting indium tin oxide (ITO and semi-conducting, silicon (Si and gallium arsenide (GaAs, surfaces modified with self-assembled monolayers (SAMs containing amino (–NH2 and methyl (–CH3 end groups have been investigated. Contact angle measurements and infrared spectroscopic studies show that the monolayers are conformal and preserve their functional end groups. Morphological analyses indicate that HDFs grow well on all substrates except GaAs, exhibiting their normal spindle-shaped morphology and exhibit no visible signs of stress or cytoplasmic vacuolation. Cell viability analyses indicate little cell death after one week in culture on all substrates except GaAs, where cells died within 6 h. Cells on all surfaces proliferate except on GaAs and GaAs-ODT. Cell growth is observed to be greater on SAM modified ITO and Si-substrates. Preservation of cellular phenotype assessed through type I collagen immunostaining and positive staining of HDF cells were observed on all modified surfaces except that on GaAs. These results suggest that conducting and semi-conducting SAM-modified surfaces support HDF growth and functionality and represent a promising area of bioengineering research.

Infrared Analysis Of Enzymes Adsorbed Onto Model Surfaces

Science.gov (United States)

Story, Gloria M.; Rauch, Deborah S.; Brode, Philip F.; Marcott, Curtis A.

1989-12-01

The adsorption of the enzymes, subtilisin BPN' and lysozyme, onto model surfaces was examined using attenuated total reflectance (ATR) infrared (IR) spectroscopy. Using a cylindrical internal reflection (CIRcle) cell with a Germanium (Ge) internal reflection element (IRE), model hydrophilic surfaces were made by plasma cleaning the IRE and model hydrophobic surfaces were made by precoating the IRE with a thin film of polystyrene. Gas chromatography (GC)-IR data collection software was used to monitor adsorption kinetics during the first five minutes after injection of the enzyme into the CIRcle cell. It was found that for both lysozyme and BPN', most of the enzyme that was going to adsorb onto the model surface did so within ten seconds after injection. Nearly an order-of-magnitude more BPN' adsorbed on the hydrophobic Ge surface than the hydrophilic one, while lysozyme adsorbed somewhat more strongly to the hydrophilic Ge surface. Overnight, the lysozyme layer continued to increase in thickness, while BPN' maintained its initial coverage. The appearance of carboxylate bands in some of the adsorbed BPN' spectra suggests the occurrence of peptide bond hydrolysis. A Au/Pd coating on the CIRcle cell o-rings had a significant effect on the adsorption of BPN'. (This coating was applied in an attempt to eliminate interfering Teflon absorption bands.) An apparent electrochemical reaction occurred, involving BPN', Ge, Au/Pd, and the salt solution used to stabilize BPN'. The result of this reaction was enhanced adsorption of the enzyme around the coated o-rings, etching of the Ge IRE at the o-ring site, and some autolysis of the enzyme. No such reaction was observed with lysozyme.

Optimization of Phospholipase A1 Immobilization on Plasma Surface Modified Chitosan Nanofibrous Mat

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Zahra Beig Mohammadi

2016-01-01

Full Text Available Phospholipase A1 is known as an effective catalyst for hydrolysis of various phospholipids in enzymatic vegetable oil degumming. Immobilization is one of the most efficient strategies to improve its activity, recovery and functional properties. In this study, chitosan-co-polyethylene oxide (90:10 nanofibrous mat was successfully fabricated and modified with atmospheric plasma at different times (2, 6 and 10 min to interact with enzyme molecules. Scanning electron microscopy images revealed that the membranes retained uniform nanofibrous and open porous structures before and after the treatment. PLA1 was successfully immobilized onto the membrane surfaces via covalent bonds with the functional groups of chitosan nanofibrous mat. Response surface methodology was used to optimize the immobilization conditions for reaching the maximum immobilization efficiency. Enzyme concentration, pH, and immobilization time were found to be significant key factors. Under optimum conditions (5.03 h, pH 5.63, and enzyme dosage 654.36 UI, the atmospheric plasma surface modified chitosan nanofibers reached the highest immobilization efficiency (78.50%. Fourier transform infrared spectroscopy of the control and plasma surface-modified chitosan nanofibers revealed the functional groups of nanofibers and their reaction with the enzyme. The results indicated that surface modification by atmospheric plasma induced an increase in PLA1 loading on the membrane surfaces.

Surface-modified CMOS IC electrochemical sensor array targeting single chromaffin cells for highly parallel amperometry measurements.

Science.gov (United States)

Huang, Meng; Delacruz, Joannalyn B; Ruelas, John C; Rathore, Shailendra S; Lindau, Manfred

2018-01-01

Amperometry is a powerful method to record quantal release events from chromaffin cells and is widely used to assess how specific drugs modify quantal size, kinetics of release, and early fusion pore properties. Surface-modified CMOS-based electrochemical sensor arrays allow simultaneous recordings from multiple cells. A reliable, low-cost technique is presented here for efficient targeting of single cells specifically to the electrode sites. An SU-8 microwell structure is patterned on the chip surface to provide insulation for the circuitry as well as cell trapping at the electrode sites. A shifted electrode design is also incorporated to increase the flexibility of the dimension and shape of the microwells. The sensitivity of the electrodes is validated by a dopamine injection experiment. Microwells with dimensions slightly larger than the cells to be trapped ensure excellent single-cell targeting efficiency, increasing the reliability and efficiency for on-chip single-cell amperometry measurements. The surface-modified device was validated with parallel recordings of live chromaffin cells trapped in the microwells. Rapid amperometric spikes with no diffusional broadening were observed, indicating that the trapped and recorded cells were in very close contact with the electrodes. The live cell recording confirms in a single experiment that spike parameters vary significantly from cell to cell but the large number of cells recorded simultaneously provides the statistical significance.

Development and evaluation of targeting ligands surface modified paclitaxel nanocrystals

Energy Technology Data Exchange (ETDEWEB)

Sohn, Jeong Sun [Division of Undeclared Majors, Chosun University, Gwangju 501-759 (Korea, Republic of); Yoon, Doo-Soo; Sohn, Jun Youn [Department of Bioenvironmental & Chemical Engineering, Chosun College of Science & Technology, Gwangju 501-744 (Korea, Republic of); Park, Jeong-Sook, E-mail: eicosa@cnu.ac.kr [College of Pharmacy and Institute of Drug Research and Development, Chungnam National University, 99 Daehak-ro, Yuseong-gu, Daejeon 34134 (Korea, Republic of); Choi, Jin-Seok, E-mail: c34281@gmail.com [College of Pharmacy and Institute of Drug Research and Development, Chungnam National University, 99 Daehak-ro, Yuseong-gu, Daejeon 34134 (Korea, Republic of)

2017-03-01

To overcome the toxicity of excipient or blank nanoparticles for drug delivery nano-system, the surface modified paclitaxel nanocrystals (PTX-NC) have been developed. PTX-NCs were prepared by nano-precipitation method. The surface of PTX-NCs were modified by grafting with apo-transferrin (Tf) or hyaluronic acid (HA). The physical properties of PTX-NCs were evaluated by field emission scanning electron microscope (FE-SEM), zeta-sizer, zeta-potential, differential scanning calorimetry (DSC) and Fourier transform infrared (FT-IR) spectrometry. In vitro drug release study was performed in phosphate buffered saline (PBS) with or without 0.5% (w/v) Tween 80 for 24 h. Cellular uptake was studied at time intervals of 0.5, 1, and 2 h in MCF-7 cells, and cell growth inhibition study was performed for 24 h using MCF-7 cells (cancer cells), and HaCaT cells (normal cells). Three different types of PTX-NCs with a mean size of 236.0 ± 100.6 nm (PTX-NC), 302.0 ± 152.0 nm (Tf-PTX-NC) and 339 ± 180.6 nm (HA-PTX-NC) were successfully prepared. The drug release profiles showed 29.1%/6.9% (PTX (pure)), 40.7%/23.9% (PTX-NC), 50.5%/25.1% (Tf-PTX-NC) and 46.8/24.8% (HA-PTX-NC) in PBS with/without 0.5% (w/v) Tween 80 for 24 h, respectively. As per the results, the drug release of PTX-NCs showed the faster release as compared to that of PTX (pure). Surface modified PTX-NCs exhibited higher values for cell permeability than unmodified PTX-NC in the cellular uptake study. Surface modified PTX-NCs inhibited the cell growth approximately to 60% in MCF-7 cells, however effect of surface modified PTX-NCs on normal cell line was lower than the PTX-NC and PTX (pure). In conclusion, biological macromolecules (Tf or HA) surface modified PTX-NC enhanced the cellular uptake and the cell growth inhibition. - Highlights: • Surface modified PTX-NCs with HA and Tf are successfully prepared by adsorption method. • Enhanced cellular uptake of modified PTX-NCs compared to unmodified PTX-NC • Improved

Listeria monocytogenes repellence by enzymatically modified PES surfaces

NARCIS (Netherlands)

Veen, van der S.; Nady, N.; Franssen, M.C.R.; Zuilhof, H.; Boom, R.M.; Abee, T.; Schroën, C.G.P.H.

2015-01-01

: The effect of enzyme-catalyzed modification of poly(ethersulfone) (PES) on the adhesion and biofilm formation of two Listeria monocytogenes strains is evaluated under static and dynamic flow conditions. PES has been modified with gallic acid, ferulic acid and 4-hydroxybenzoic acid. The surfaces

Screening for Glycosylphosphatidylinositol-Modified Cell Wall Proteins in Pichia pastoris and Their Recombinant Expression on the Cell Surface

Science.gov (United States)

Zhang, Li; Liang, Shuli; Zhou, Xinying; Jin, Zi; Jiang, Fengchun; Han, Shuangyan; Zheng, Suiping

2013-01-01

Glycosylphosphatidylinositol (GPI)-anchored glycoproteins have various intrinsic functions in yeasts and different uses in vitro. In the present study, the genome of Pichia pastoris GS115 was screened for potential GPI-modified cell wall proteins. Fifty putative GPI-anchored proteins were selected on the basis of (i) the presence of a C-terminal GPI attachment signal sequence, (ii) the presence of an N-terminal signal sequence for secretion, and (iii) the absence of transmembrane domains in mature protein. The predicted GPI-anchored proteins were fused to an alpha-factor secretion signal as a substitute for their own N-terminal signal peptides and tagged with the chimeric reporters FLAG tag and mature Candida antarctica lipase B (CALB). The expression of fusion proteins on the cell surface of P. pastoris GS115 was determined by whole-cell flow cytometry and immunoblotting analysis of the cell wall extracts obtained by β-1,3-glucanase digestion. CALB displayed on the cell surface of P. pastoris GS115 with the predicted GPI-anchored proteins was examined on the basis of potential hydrolysis of p-nitrophenyl butyrate. Finally, 13 proteins were confirmed to be GPI-modified cell wall proteins in P. pastoris GS115, which can be used to display heterologous proteins on the yeast cell surface. PMID:23835174

21 CFR 184.1287 - Enzyme-modified fats.

Science.gov (United States)

2010-04-01

... Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD... that are generally recognized as safe (GRAS). Enzyme-modified milk powder may be prepared with GRAS enzymes from reconstituted milk powder, whole milk, condensed or concentrated whole milk, evaporated milk...

Modified Titanium Surface-Mediated Effects on Human Bone Marrow Stromal Cell Response

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Amol Chaudhari

2013-11-01

Full Text Available Surface modification of titanium implants is used to enhance osseointegration. The study objective was to evaluate five modified titanium surfaces in terms of cytocompatibility and pro-osteogenic/pro-angiogenic properties for human mesenchymal stromal cells: amorphous microporous silica (AMS, bone morphogenetic protein-2 immobilized on AMS (AMS + BMP, bio-active glass (BAG and two titanium coatings with different porosity (T1; T2. Four surfaces served as controls: uncoated Ti (Ti, Ti functionalized with BMP-2 (Ti + BMP, Ti surface with a thickened titanium oxide layer (TiO2 and a tissue culture polystyrene surface (TCPS. The proliferation of eGFP-fLuc (enhanced green fluorescence protein-firefly luciferase transfected cells was tracked non-invasively by fluorescence microscopy and bio-luminescence imaging. The implant surface-mediated effects on cell differentiation potential was tracked by determination of osteogenic and angiogenic parameters [alkaline phosphatase (ALP; osteocalcin (OC; osteoprotegerin (OPG; vascular endothelial growth factor-A (VEGF-A]. Unrestrained cell proliferation was observed on (unfunctionalized Ti and AMS surfaces, whereas BAG and porous titanium coatings T1 and T2 did not support cell proliferation. An important pro-osteogenic and pro-angiogenic potential of the AMS + BMP surface was observed. In contrast, coating the Ti surface with BMP did not affect the osteogenic differentiation of the progenitor cells. A significantly slower BMP-2 release from AMS compared to Ti supports these findings. In the unfunctionalized state, Ti was found to be superior to AMS in terms of OPG and VEGF-A production. AMS is suggested to be a promising implant coating material for bioactive agents delivery.

tRNA modifying enzymes, NSUN2 and METTL1, determine sensitivity to 5-fluorouracil in HeLa cells.

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Mayumi Okamoto

2014-09-01

Full Text Available Nonessential tRNA modifications by methyltransferases are evolutionarily conserved and have been reported to stabilize mature tRNA molecules and prevent rapid tRNA decay (RTD. The tRNA modifying enzymes, NSUN2 and METTL1, are mammalian orthologs of yeast Trm4 and Trm8, which are required for protecting tRNA against RTD. A simultaneous overexpression of NSUN2 and METTL1 is widely observed among human cancers suggesting that targeting of both proteins provides a novel powerful strategy for cancer chemotherapy. Here, we show that combined knockdown of NSUN2 and METTL1 in HeLa cells drastically potentiate sensitivity of cells to 5-fluorouracil (5-FU whereas heat stress of cells revealed no effects. Since NSUN2 and METTL1 are phosphorylated by Aurora-B and Akt, respectively, and their tRNA modifying activities are suppressed by phosphorylation, overexpression of constitutively dephosphorylated forms of both methyltransferases is able to suppress 5-FU sensitivity. Thus, NSUN2 and METTL1 are implicated in 5-FU sensitivity in HeLa cells. Interfering with methylation of tRNAs might provide a promising rationale to improve 5-FU chemotherapy of cancer.

2009 Cellulosomes, Cellulases & Other Carbohydrate Modifying Enzymes GRC

Energy Technology Data Exchange (ETDEWEB)

Gilbert, Harry [Univ. of Newcastle, Callaghan, NSW (Australia)

2009-07-26

The 2009 Gordon Conference on Cellulosomes, Cellulases & Other Carbohydrate Modifying Enzymes will present cutting-edge research on the enzymatic degradation of cellulose and other plant cell wall polysaccharides. The Conference will feature a wide range of topics that includes the enzymology of plant structural degradation, regulation of the degradative apparatus, the mechanism of protein complex assembly, the genomics of cell wall degrading organisms, the structure of the substrate and the industrial application of the process particularly within the biofuel arena. Indeed the deployment of plant cell wall degrading enzymes in biofuel processes will be an important feature of the meeting. It should be emphasized that the 2009 Conference will be expanded to include, in addition to cellulase research, recent advances in other plant cell wall degrading enzymes, and contributions from people working on hemicellulases and pectinases will be particularly welcome. Invited speakers represent a variety of scientific disciplines, including biochemistry, structural biology, genetics and cell biology. The interplay between fundamental research and its industrial exploitation is a particularly important aspect of the meeting, reflecting the appointment of the chair and vice-chair from academia and industry, respectively. The meeting will provide opportunities for junior scientists and graduate students to present their work in poster format and exchange ideas with more established figures in the field. Indeed, some poster presenters will be selected for short talks. The collegial atmosphere of this Conference, with programmed discussion sessions as well as opportunities for informal gatherings in the afternoons and evenings, provides an avenue for scientists from different disciplines to brainstorm and promotes cross-disciplinary collaborations in the various research areas represented. The Conference is likely to be heavily subscribed so we would recommend that you submit

Cytokine Adsorption onto the Modified Carbon Sorbent Surface in vitro in Peritonitis

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T. I. Dolgikh

2009-01-01

Full Text Available Objective: to evaluate the efficiency of cytokine sorption with carbon with a locally aminocaproic acid-modified surface from the plasma of patients with general purulent peritonitis. Materials and methods. The material of the investigation was the plasma obtained during plasmapheresis in 10 patients with acute pancreatitis complicated by pancreonecrosis and general purulent peritonitis, which was used to estimate before and after sorption the content of the cytokines: interleukin (IL-1/8, IL-4, and IL-8 by enzyme immunoassay. The sorption properties of carbon hemosor-bent and aminocaproic acid-modified sorbent were comparatively evaluated. Results. Aminocaproic acid-induced modification of the carbon adsorbent surface with its further polycondensation results in the higher content of superficial functional groups (oxygen- and nitrogen-containing that enhance the hydrophility of the surface and the specific pattern of sorption, thus acting as a means for controlling and regulating the plasma concentration of regulatory proteins, primarily the proinflammatory cytokine IL-1^3, the chemokine IL-8 and the T-helper cell clone cytokine IL-4.

Behaviour of human endothelial cells on surface modified NiTi alloy.

Science.gov (United States)

Plant, Stuart D; Grant, David M; Leach, Lopa

2005-09-01

Intravascular stents are being designed which utilise the shape memory properties of NiTi alloy. Despite the clinical advantages afforded by these stents their application has been limited by concerns about the large nickel ion content of the alloy. In this study, the surface chemistry of NiTi alloy was modified by mechanical polishing and oxidising heat treatments and subsequently characterised using X-ray photon spectroscopy (XPS). The effect of these surfaces on monolayer formation and barrier integrity of human umbilical vein endothelial cells (HUVEC) was then assessed by confocal imaging of the adherens junctional molecule VE-cadherin, perijunctional actin and permeability to 42kDa dextrans. Dichlorofluoroscein assays were used to measure oxidative stress in the cells. XPS analysis of NiTi revealed its surface to be dominated by TiO(2). However, where oxidation had occurred after mechanical polishing or post polishing heat treatments at 300 and 400 degrees C in air, a significant amount of metallic nickel or nickel oxide species (10.5 and 18.5 at%) remained on the surface. Exposure of HUVECs to these surfaces resulted in increased oxidative stress within the cells, loss of VE-cadherin and F-actin and significantly increased paracellular permeability. These pathological phenomena were not found in cells grown on NiTi which had undergone heat treatment at 600 degrees C. At this temperature thickening of the TiO(2) layer had occurred due to diffusion of titanium ions from the bulk of the alloy, displacing nickel ions to sub-surface areas. This resulted in a significant reduction in nickel ions detectable on the sample surface (4.8 at%). This study proposes that the integrity of human endothelial monolayers on NiTi is dependent upon the surface chemistry of the alloy and that this can be manipulated, using simple oxidising heat treatments.

Immobilization of lipases on alkyl silane modified magnetic nanoparticles: effect of alkyl chain length on enzyme activity.

Directory of Open Access Journals (Sweden)

Jiqian Wang

Full Text Available BACKGROUND: Biocatalytic processes often require a full recycling of biocatalysts to optimize economic benefits and minimize waste disposal. Immobilization of biocatalysts onto particulate carriers has been widely explored as an option to meet these requirements. However, surface properties often affect the amount of biocatalysts immobilized, their bioactivity and stability, hampering their wide applications. The aim of this work is to explore how immobilization of lipases onto magnetite nanoparticles affects their biocatalytic performance under carefully controlled surface modification. METHODOLOGY/PRINCIPAL FINDINGS: Magnetite nanoparticles, prepared through a co-precipitation method, were coated with alkyl silanes of different alkyl chain lengths to modulate their surface hydrophobicity. Candida rugosa lipase was then directly immobilized onto the modified nanoparticles through hydrophobic interaction. Enzyme activity was assessed by catalytic hydrolysis of p-nitrophenyl acetate. The activity of immobilized lipases was found to increase with increasing chain length of the alkyl silane. Furthermore, the catalytic activities of lipases immobilized on trimethoxyl octadecyl silane (C18 modified Fe(3O(4 were a factor of 2 or more than the values reported from other surface immobilized systems. After 7 recycles, the activities of the lipases immobilized on C18 modified nanoparticles retained 65%, indicating significant enhancement of stability as well through hydrophobic interaction. Lipase immobilized magnetic nanoparticles facilitated easy separation and recycling with high activity retaining. CONCLUSIONS/SIGNIFICANCE: The activity of immobilized lipases increased with increasing alkyl chain length of the alkyl trimethoxy silanes used in the surface modification of magnetite nanoparticles. Lipase stability was also improved through hydrophobic interaction. Alkyl silane modified magnetite nanoparticles are thus highly attractive carriers for

Immobilization of lipases on alkyl silane modified magnetic nanoparticles: effect of alkyl chain length on enzyme activity.

Science.gov (United States)

Wang, Jiqian; Meng, Gang; Tao, Kai; Feng, Min; Zhao, Xiubo; Li, Zhen; Xu, Hai; Xia, Daohong; Lu, Jian R

2012-01-01

Biocatalytic processes often require a full recycling of biocatalysts to optimize economic benefits and minimize waste disposal. Immobilization of biocatalysts onto particulate carriers has been widely explored as an option to meet these requirements. However, surface properties often affect the amount of biocatalysts immobilized, their bioactivity and stability, hampering their wide applications. The aim of this work is to explore how immobilization of lipases onto magnetite nanoparticles affects their biocatalytic performance under carefully controlled surface modification. Magnetite nanoparticles, prepared through a co-precipitation method, were coated with alkyl silanes of different alkyl chain lengths to modulate their surface hydrophobicity. Candida rugosa lipase was then directly immobilized onto the modified nanoparticles through hydrophobic interaction. Enzyme activity was assessed by catalytic hydrolysis of p-nitrophenyl acetate. The activity of immobilized lipases was found to increase with increasing chain length of the alkyl silane. Furthermore, the catalytic activities of lipases immobilized on trimethoxyl octadecyl silane (C18) modified Fe(3)O(4) were a factor of 2 or more than the values reported from other surface immobilized systems. After 7 recycles, the activities of the lipases immobilized on C18 modified nanoparticles retained 65%, indicating significant enhancement of stability as well through hydrophobic interaction. Lipase immobilized magnetic nanoparticles facilitated easy separation and recycling with high activity retaining. The activity of immobilized lipases increased with increasing alkyl chain length of the alkyl trimethoxy silanes used in the surface modification of magnetite nanoparticles. Lipase stability was also improved through hydrophobic interaction. Alkyl silane modified magnetite nanoparticles are thus highly attractive carriers for enzyme immobilization enabling efficient enzyme recovery and recycling.

Immobilization of glucoamylase on ceramic membrane surfaces modified with a new method of treatment utilizing SPCP-CVD.

Science.gov (United States)

Ida; Matsuyama; Yamamoto

2000-07-01

Glucoamylase, as a model enzyme, was immobilized on a ceramic membrane modified by surface corona discharge induced plasma chemical process-chemical vapor deposition (SPCP-CVD). Characterizations of the immobilized enzyme were then discussed. Three kinds of ceramic membranes with different amounts of amino groups on the surface were prepared utilizing the SPCP-CVD method. Each with 1-time, 3-times and 5-times surface modification treatments and used for supports in glucoamylase immobilization. The amount of immobilized glucoamylase increased with the increase in the number of surface modification treatments and saturated to a certain maximum value estimated by a two-dimensional random packing. The operational stability of the immobilized glucoamylase also increased with the increase in the number of the surface treatment. It was almost the same as the conventional method, while the activity of immobilized enzyme was higher. The results indicated the possibility of designing the performance of the immobilized enzyme by controlling the amount of amino groups. The above results showed that the completely new surface modification method using SPCP was effective in modifying ceramic membranes for enzyme immobilization.

Modified titanium surface with gelatin nano gold composite increases osteoblast cell biocompatibility

International Nuclear Information System (INIS)

Lee, Young-Hee; Bhattarai, Govinda; Aryal, Santosh; Lee, Nan-Hee; Lee, Min-Ho; Kim, Tae-Gun; Jhee, Eun-Chung; Kim, Hak-Yong; Yi, Ho-Keun

2010-01-01

This study examined the gelatin nano gold (GnG) composite for surface modification of titanium in addition to insure biocompatibility on dental implants or biomaterials. The GnG composite was constructed by gelatin and hydrogen tetrachloroaurate in presence of reducing agent, sodium borohydrate (NabH 4 ). The GnG composite was confirmed by UV-VIS spectroscopy and transmission electron microscopy (TEM). A dipping method was used to modify the titanium surface by GnG composite. Surface was characterized by scanning electron microscopy (SEM) and energy dispersive X-ray (EDX). The MC-3T3 E1 cell viability was assessed by trypan blue and the expression of proteins to biocompatibility were analyzed by Western blotting. The GnG composite showed well dispersed character, the strong absorption at 530 nm, roughness, regular crystal and clear C, Na, Cl, P, and Au signals onto titanium. Further, this composite allowed MC-3T3 E1 growth and viability compared to gelatin and pure titanium. It induced ERK activation and the expression of cell adherent molecules, FAK and SPARC, and growth factor, VEGF. However, GnG decreased the level of SAPK/JNK. This shows that GnG composite coated titanium surfaces have a good biocompatibility for osteoblast growth and attachment than in intact by simple and versatile dipping method. Furthermore, it offers good communication between cell and implant surfaces by regulating cell signaling and adherent molecules, which are useful to enhance the biocompatibility of titanium surfaces.

ENZYME RESISTANCE OF GENETICALLY MODIFIED STARCH POTATOES

Directory of Open Access Journals (Sweden)

A. Sh. Mannapova

2015-01-01

Full Text Available Here in this article the justification of expediency of enzyme resistant starch use in therapeutic food products is presented . Enzyme resistant starch is capable to resist to enzymatic hydrolysis in a small intestine of a person, has a low glycemic index, leads to decrease of postprandial concentration of glucose, cholesterol, triglycerides in blood and insulin reaction, to improvement of sensitivity of all organism to insulin, to increase in sense of fulness and to reduction of adjournment of fats. Resistant starch makes bifidogenшс impact on microflora of a intestine of the person, leads to increase of a quantity of lactobacillus and bifidobacterium and to increased production of butyric acid in a large intestine. In this regard the enzyme resistant starch is an important component in food for prevention and curing of human diseases such as diabetes, obesity, colitis, a cancer of large and direct intestine. One method is specified by authors for imitation of starch digestion in a human body. This method is based on the definition of an enzyme resistance of starch in vitro by its hydrolysis to glucose with application of a glucoamylase and digestive enzyme preparation Pancreatin. This method is used in researches of an enzyme resistance of starch, of genetically modified potato, high amylose corn starch Hi-Maize 1043 and HYLON VII (National Starch Food Innovation, USA, amylopectin and amylose. It is shown that the enzyme resistance of the starch emitted from genetically modified potatoes conforms to the enzyme resistance of the high amylose corn starch “Hi-Maize 1043 and HYLON VII starch”, (National Starch Food Innovation, the USA relating to the II type of enzyme resistant starch. It is established that amylopectin doesn't have the enzyme resistant properties. The results of researches are presented. They allow us to make the following conclusion: amylose in comparison with amylopectin possesses higher enzyme resistance and gives to

Immobilized enzymes: understanding enzyme - surface interactions at the molecular level.

Science.gov (United States)

Hoarau, Marie; Badieyan, Somayesadat; Marsh, E Neil G

2017-11-22

Enzymes immobilized on solid supports have important and industrial and medical applications. However, their uses are limited by the significant reductions in activity and stability that often accompany the immobilization process. Here we review recent advances in our understanding of the molecular level interactions between proteins and supporting surfaces that contribute to changes in stability and activity. This understanding has been facilitated by the application of various surface-sensitive spectroscopic techniques that allow the structure and orientation of enzymes at the solid/liquid interface to be probed, often with monolayer sensitivity. An appreciation of the molecular interactions between enzyme and surface support has allowed the surface chemistry and method of enzyme attachement to be fine-tuned such that activity and stability can be greatly enhanced. These advances suggest that a much wider variety of enzymes may eventually be amenable to immobilization as green catalysts.

Specific capture and detection of Staphylococcus aureus with high-affinity modified aptamers to cell surface components.

Science.gov (United States)

Baumstummler, A; Lehmann, D; Janjic, N; Ochsner, U A

2014-10-01

Slow off-rate modified aptamer (SOMAmer) reagents were generated to several Staphylococcus aureus cell surface-associated proteins via SELEX with multiple modified DNA libraries using purified recombinant or native proteins. High-affinity binding agents with sub-nanomolar Kd 's were obtained for staphylococcal protein A (SpA), clumping factors (ClfA, ClfB), fibronectin-binding proteins (FnbA, FnbB) and iron-regulated surface determinants (Isd). Further screening revealed several SOMAmers that specifically bound to Staph. aureus cells from all strains that were tested, but not to other staphylococci or other bacteria. SpA and ClfA SOMAmers proved useful for the selective capture and enrichment of Staph. aureus cells, as shown by culture and PCR, leading to improved limits of detection and efficient removal of PCR inhibitors. Detection of Staph. aureus cells was enhanced by several orders of magnitude when the bacterial cell surface was coated with SOMAmers followed by qPCR of the SOMAmers. Furthermore, fluorescence-labelled SpA SOMAmers demonstrated their utility as direct detection agents in flow cytometry. Significance and impact of the study: Monitoring for microbial contamination of food, water, nonsterile products or the environment is typically based on culture, PCR or antibodies. Aptamers that bind with high specificity and affinity to well-conserved cell surface epitopes represent a promising novel type of reagents to detect bacterial cells without the need for culture or cell lysis, including for the capture and enrichment of bacteria present at low cell densities and for the direct detection via qPCR or fluorescent staining. © 2014 Soma Logic, Inc. published by John Wiley & Sons Ltd On behalf of the society for Applied Microbiology.

Surface modified electrospun nanofibrous scaffolds for nerve tissue engineering

International Nuclear Information System (INIS)

Prabhakaran, Molamma P; Venugopal, J; Chan, Casey K; Ramakrishna, S

2008-01-01

The development of biodegradable polymeric scaffolds with surface properties that dominate interactions between the material and biological environment is of great interest in biomedical applications. In this regard, poly-ε-caprolactone (PCL) nanofibrous scaffolds were fabricated by an electrospinning process and surface modified by a simple plasma treatment process for enhancing the Schwann cell adhesion, proliferation and interactions with nanofibers necessary for nerve tissue formation. The hydrophilicity of surface modified PCL nanofibrous scaffolds (p-PCL) was evaluated by contact angle and x-ray photoelectron spectroscopy studies. Naturally derived polymers such as collagen are frequently used for the fabrication of biocomposite PCL/collagen scaffolds, though the feasibility of procuring large amounts of natural materials for clinical applications remains a concern, along with their cost and mechanical stability. The proliferation of Schwann cells on p-PCL nanofibrous scaffolds showed a 17% increase in cell proliferation compared to those on PCL/collagen nanofibrous scaffolds after 8 days of cell culture. Schwann cells were found to attach and proliferate on surface modified PCL nanofibrous scaffolds expressing bipolar elongations, retaining their normal morphology. The results of our study showed that plasma treated PCL nanofibrous scaffolds are a cost-effective material compared to PCL/collagen scaffolds, and can potentially serve as an ideal tissue engineered scaffold, especially for peripheral nerve regeneration.

Performance enhancement in organic photovoltaic solar cells using iridium (Ir) ultra-thin surface modifier (USM)

Science.gov (United States)

Pandey, Rina; Lim, Ju Won; Kim, Jung Hyuk; Angadi, Basavaraj; Choi, Ji Won; Choi, Won Kook

2018-06-01

In this study, Iridium (Ir) metallic layer as an ultra-thin surface modifier (USM) was deposited on ITO coated glass substrate using radio frequency magnetron sputtering for improving the photo-conversion efficiency of organic photovoltaic cells. Ultra-thin Ir acts as a surface modifier replacing the conventional hole transport layer (HTL) PEDOT:PSS in organic photovoltaic (OPV) cells with two different active layers P3HT:PC60BM and PTB7:PC70BM. The Ir USM (1.0 nm) coated on ITO glass substrate showed transmittance of 84.1% and work function of >5.0 eV, which is higher than that of ITO (4.5-4.7 eV). The OPV cells with Ir USM (1.0 nm) exhibits increased power conversion efficiency of 3.70% (for P3HT:PC60BM active layer) and 7.28% (for PTB7:PC70BM active layer) under 100 mW/cm2 illumination (AM 1.5G) which are higher than those of 3.26% and 6.95% for the same OPV cells but with PEDOT:PSS as HTL instead of Ir USM. The results reveal that the chemically stable Ir USM layer could be used as an alternative material for PEDOT:PSS in organic photovoltaic cells.

Fabrication of endothelial progenitor cell capture surface via DNA aptamer modifying dopamine/polyethyleneimine copolymer film

Energy Technology Data Exchange (ETDEWEB)

Li, Xin; Deng, Jinchuan; Yuan, Shuheng; Wang, Juan; Luo, Rifang; Chen, Si [Key Lab. of Advanced Technology for Materials of Education Ministry, Southwest Jiaotong University, Chengdu 610031 (China); School of Material Science and Engineering, Southwest Jiaotong University, Chengdu 610031 (China); Wang, Jin, E-mail: jinxxwang@263.net [Key Lab. of Advanced Technology for Materials of Education Ministry, Southwest Jiaotong University, Chengdu 610031 (China); Huang, Nan [Key Lab. of Advanced Technology for Materials of Education Ministry, Southwest Jiaotong University, Chengdu 610031 (China); School of Material Science and Engineering, Southwest Jiaotong University, Chengdu 610031 (China)

2016-11-15

Highlights: • The dopamine/PEI film with controlled amine density was successfully prepared. • The DNA aptamer was assembled onto the film via electrostatic incorporation. • The A@DPfilmscanspecificallyandeffectivelycaptureEPCs. • The A@DP film can support the survival of ECs, control the hyperplasia of SMCs. • The dynamic/co-culture models are useful for studying cells competitive adhesion. - Abstract: Endothelial progenitor cells (EPCs) are mainly located in bone marrow and circulate, and play a crucial role in repairmen of injury endothelium. One of the most promising strategies of stents designs were considered to make in-situ endothelialization in vivo via EPC-capture biomolecules on a vascular graft to capture EPCs directly from circulatory blood. In this work, an EPC specific aptamer with a 34 bases single strand DNA sequence was conjugated onto the stent surface via dopamine/polyethyleneimine copolymer film as a platform and linker. The assembled density of DNA aptamer could be regulated by controlling dopamine percentage in this copolymer film. X-ray photoelectron spectroscopy (XPS), water contact angle (WCA) and fluorescence test confirmed the successful immobilization of DNA aptamer. To confirm its biofunctionality and cytocompatibility, the capturing cells ability of the aptamer modified surface and the effects on the growth behavior of human umbilical vein endothelial cells (HUVECs), smooth muscle cells (SMCs) were investigated. The aptamer functionalized sample revealed a good EPC-capture ability, and had a cellular friendly feature for both EPC and EC growth, while not stimulated the hyperplasia of SMCs. And, the co-culture experiment of three types of cells confirmed the specificity capturing of EPCs to aptamer modified surface, rather than ECs and SMCs. These data suggested that this aptamer functionalized surface may have a large potentiality for the application of vascular grafts with targeted endothelialization.

Fabrication of endothelial progenitor cell capture surface via DNA aptamer modifying dopamine/polyethyleneimine copolymer film

International Nuclear Information System (INIS)

Li, Xin; Deng, Jinchuan; Yuan, Shuheng; Wang, Juan; Luo, Rifang; Chen, Si; Wang, Jin; Huang, Nan

2016-01-01

Highlights: • The dopamine/PEI film with controlled amine density was successfully prepared. • The DNA aptamer was assembled onto the film via electrostatic incorporation. • The A@DPfilmscanspecificallyandeffectivelycaptureEPCs. • The A@DP film can support the survival of ECs, control the hyperplasia of SMCs. • The dynamic/co-culture models are useful for studying cells competitive adhesion. - Abstract: Endothelial progenitor cells (EPCs) are mainly located in bone marrow and circulate, and play a crucial role in repairmen of injury endothelium. One of the most promising strategies of stents designs were considered to make in-situ endothelialization in vivo via EPC-capture biomolecules on a vascular graft to capture EPCs directly from circulatory blood. In this work, an EPC specific aptamer with a 34 bases single strand DNA sequence was conjugated onto the stent surface via dopamine/polyethyleneimine copolymer film as a platform and linker. The assembled density of DNA aptamer could be regulated by controlling dopamine percentage in this copolymer film. X-ray photoelectron spectroscopy (XPS), water contact angle (WCA) and fluorescence test confirmed the successful immobilization of DNA aptamer. To confirm its biofunctionality and cytocompatibility, the capturing cells ability of the aptamer modified surface and the effects on the growth behavior of human umbilical vein endothelial cells (HUVECs), smooth muscle cells (SMCs) were investigated. The aptamer functionalized sample revealed a good EPC-capture ability, and had a cellular friendly feature for both EPC and EC growth, while not stimulated the hyperplasia of SMCs. And, the co-culture experiment of three types of cells confirmed the specificity capturing of EPCs to aptamer modified surface, rather than ECs and SMCs. These data suggested that this aptamer functionalized surface may have a large potentiality for the application of vascular grafts with targeted endothelialization.

Surface-modified bacterial nanofibrillar PHB scaffolds for bladder tissue repair.

Science.gov (United States)

Karahaliloğlu, Zeynep; Demirbilek, Murat; Şam, Mesut; Sağlam, Necdet; Mızrak, Alpay Koray; Denkbaş, Emir Baki

2016-01-01

The aim of the study is in vitro investigation of the feasibility of surface-modified bacterial nanofibrous poly [(R)-3-hydroxybutyrate] (PHB) graft for bladder reconstruction. In this study, the surface of electrospun bacterial PHB was modified with PEG- or EDA via radio frequency glow discharge method. After plasma modification, contact angle of EDA-modified PHB scaffolds decreased from 110 ± 1.50 to 23 ± 0.5 degree. Interestingly, less calcium oxalate stone deposition was observed on modified PHB scaffolds compared to that of non-modified group. Results of this study show that surface-modified scaffolds not only inhibited calcium oxalate growth but also enhanced the uroepithelial cell viability and proliferation.

Comparative of fibroblast and osteoblast cells adhesion on surface modified nanofibrous substrates based on polycaprolactone

OpenAIRE

Sharifi, Fereshteh; Irani, Shiva; Zandi, Mojgan; Soleimani, Masoud; Atyabi, Seyed Mohammad

2016-01-01

One of the determinant factors for successful bioengineering is to achieve appropriate nano-topography and three-dimensional substrate. In this research, polycaprolactone (PCL) nano-fibrous mat with different roughness modified with O2 plasma was fabricated via electrospinning. The purpose of this study was to evaluate the effect of plasma modification along with surface nano-topography of mats on the quality of human fibroblast (HDFs) and osteoblast cells (OSTs)-substrate interaction. Surfac...

Immobilization of chlorine dioxide modified cells for uranium absorption

International Nuclear Information System (INIS)

He, Shengbin; Ruan, Binbiao; Zheng, Yueping; Zhou, Xiaobin; Xu, Xiaoping

2014-01-01

There has been a trend towards the use of microorganisms to recover metals from industrial wastewater, for which various methods have been reported to be used to improve microorganism adsorption characteristics such as absorption capacity, tolerance and reusability. In present study, chlorine dioxide(ClO 2 ), a high-efficiency, low toxicity and environment-benign disinfectant, was first reported to be used for microorganism surface modification. The chlorine dioxide modified cells demonstrated a 10.1% higher uranium adsorption capacity than control ones. FTIR analysis indicated that several cell surface groups are involved in the uranium adsorption and cell surface modification. The modified cells were further immobilized on a carboxymethylcellulose (CMC) matrix to improve their reusability. The cell-immobilized adsorbent could be employed either in a high concentration system to move vast UO 2 2+ ions or in a low concentration system to purify UO 2 2+ contaminated water thoroughly, and could be repeatedly used in multiple adsorption-desorption cycles with about 90% adsorption capacity maintained after seven cycles. - Highlights: • Chlorine dioxide was first reported to be used for microorganism surface modification. • The chlorine dioxide modified cells demonstrated a 10.1% higher uranium adsorption capacity than control ones. • The chlorine dioxide modified cells were further immobilized by carboxymethylcellulose to improve their reusability

Ammonia synthesis on Au modified Fe(111) and Ag and Cu modified Fe(100) surfaces

DEFF Research Database (Denmark)

Lytken, Ole; Waltenburg, Hanne Neergaard; Chorkendorff, Ib

2003-01-01

In order to investigate any influence of steps and possible positive effects of making surface alloys the ammonia synthesis has been investigated over Au modified Fe(111) and Ag and Cu modified Fe(100) single crystals in the temperature range 603-773 K, using a system combining ultra-high vacuum...... and a high-pressure cell. Ammonia was synthesized from a stoichiometric (N-2:3H(2)) gas mixture at a pressure of 2 bar. By deposition of small amounts of An, the ammonia production activity of the Fe(1 1 1) surface can be enhanced. More important, for the gold modified surface, the reaction order in ammonia...

Superparamagnetic poly(methyl methacrylate) nanoparticles surface modified with folic acid presenting cell uptake mediated by endocytosis

Energy Technology Data Exchange (ETDEWEB)

Feuser, Paulo Emilio [Federal University of Santa Catarina, Department of Chemical Engineering and Food Engineering (Brazil); Jacques, Amanda Virtuoso [Federal University of Santa Catarina, Department of Clinical Analyses (Brazil); Arévalo, Juan Marcelo Carpio; Rocha, Maria Eliane Merlin [Federal University of Paraná, Department of Biochemistry and Molecular Biology (Brazil); Santos-Silva, Maria Claudia dos [Federal University of Santa Catarina, Department of Clinical Analyses (Brazil); Sayer, Claudia; Araújo, Pedro H. Hermes de, E-mail: pedro.h.araujo@ufsc.br [Federal University of Santa Catarina, Department of Chemical Engineering and Food Engineering (Brazil)

2016-04-15

The encapsulation of superparamagnetic nanoparticles (MNPs) in polymeric nanoparticles (NPs) with modified surfaces can improve targeted delivery and induce cell death by hyperthermia. The goals of this study were to synthesize and characterize surface modified superparamagnetic poly(methyl methacrylate) with folic acid (FA) prepared by miniemulsion polymerization (MNPsPMMA-FA) and to evaluate their in vitro cytotoxicity and cellular uptake in non-tumor cells, murine fibroblast (L929) cells and tumor cells that overexpressed folate receptor (FR) β, and chronic myeloid leukemia cells in blast crisis (K562). Lastly, hemolysis assays were performed on human red blood cells. MNPsPMMA-FA presented an average mean diameter of 135 nm and a saturation magnetization (Ms) value of 37 emu/g of iron oxide, as well as superparamagnetic behavior. The MNPsPMMA-FA did not present cytotoxicity in L929 and K562 cells. Cellular uptake assays showed a higher uptake of MNPsPMMA-FA than MNPsPMMA in K562 cells when incubated at 37 °C. On the other hand, MNPsPMMA-FA showed a low uptake when endocytosis mechanisms were blocked at low temperature (4 °C), suggesting that the MNPsPMMA-FA uptake was mediated by endocytosis. High concentrations of MNPsPMMA-FA showed hemocompatibility when incubated for 24 h in human red blood cells. Therefore, our results suggest that these carrier systems can be an excellent alternative in targeted drug delivery via FR.

Superparamagnetic poly(methyl methacrylate) nanoparticles surface modified with folic acid presenting cell uptake mediated by endocytosis

International Nuclear Information System (INIS)

Feuser, Paulo Emilio; Jacques, Amanda Virtuoso; Arévalo, Juan Marcelo Carpio; Rocha, Maria Eliane Merlin; Santos-Silva, Maria Claudia dos; Sayer, Claudia; Araújo, Pedro H. Hermes de

2016-01-01

The encapsulation of superparamagnetic nanoparticles (MNPs) in polymeric nanoparticles (NPs) with modified surfaces can improve targeted delivery and induce cell death by hyperthermia. The goals of this study were to synthesize and characterize surface modified superparamagnetic poly(methyl methacrylate) with folic acid (FA) prepared by miniemulsion polymerization (MNPsPMMA-FA) and to evaluate their in vitro cytotoxicity and cellular uptake in non-tumor cells, murine fibroblast (L929) cells and tumor cells that overexpressed folate receptor (FR) β, and chronic myeloid leukemia cells in blast crisis (K562). Lastly, hemolysis assays were performed on human red blood cells. MNPsPMMA-FA presented an average mean diameter of 135 nm and a saturation magnetization (Ms) value of 37 emu/g of iron oxide, as well as superparamagnetic behavior. The MNPsPMMA-FA did not present cytotoxicity in L929 and K562 cells. Cellular uptake assays showed a higher uptake of MNPsPMMA-FA than MNPsPMMA in K562 cells when incubated at 37 °C. On the other hand, MNPsPMMA-FA showed a low uptake when endocytosis mechanisms were blocked at low temperature (4 °C), suggesting that the MNPsPMMA-FA uptake was mediated by endocytosis. High concentrations of MNPsPMMA-FA showed hemocompatibility when incubated for 24 h in human red blood cells. Therefore, our results suggest that these carrier systems can be an excellent alternative in targeted drug delivery via FR.

Superparamagnetic poly(methyl methacrylate) nanoparticles surface modified with folic acid presenting cell uptake mediated by endocytosis

Science.gov (United States)

Feuser, Paulo Emilio; Jacques, Amanda Virtuoso; Arévalo, Juan Marcelo Carpio; Rocha, Maria Eliane Merlin; dos Santos-Silva, Maria Claudia; Sayer, Claudia; de Araújo, Pedro H. Hermes

2016-04-01

The encapsulation of superparamagnetic nanoparticles (MNPs) in polymeric nanoparticles (NPs) with modified surfaces can improve targeted delivery and induce cell death by hyperthermia. The goals of this study were to synthesize and characterize surface modified superparamagnetic poly(methyl methacrylate) with folic acid (FA) prepared by miniemulsion polymerization (MNPsPMMA-FA) and to evaluate their in vitro cytotoxicity and cellular uptake in non-tumor cells, murine fibroblast (L929) cells and tumor cells that overexpressed folate receptor (FR) β, and chronic myeloid leukemia cells in blast crisis (K562). Lastly, hemolysis assays were performed on human red blood cells. MNPsPMMA-FA presented an average mean diameter of 135 nm and a saturation magnetization (Ms) value of 37 emu/g of iron oxide, as well as superparamagnetic behavior. The MNPsPMMA-FA did not present cytotoxicity in L929 and K562 cells. Cellular uptake assays showed a higher uptake of MNPsPMMA-FA than MNPsPMMA in K562 cells when incubated at 37 °C. On the other hand, MNPsPMMA-FA showed a low uptake when endocytosis mechanisms were blocked at low temperature (4 °C), suggesting that the MNPsPMMA-FA uptake was mediated by endocytosis. High concentrations of MNPsPMMA-FA showed hemocompatibility when incubated for 24 h in human red blood cells. Therefore, our results suggest that these carrier systems can be an excellent alternative in targeted drug delivery via FR.

Improved mucoadhesion and cell uptake of chitosan and chitosan oligosaccharide surface-modified polymer nanoparticles for mucosal delivery of proteins.

Science.gov (United States)

Dyawanapelly, Sathish; Koli, Uday; Dharamdasani, Vimisha; Jain, Ratnesh; Dandekar, Prajakta

2016-08-01

The main aim of the present study was to compare mucoadhesion and cellular uptake efficiency of chitosan (CS) and chitosan oligosaccharide (COS) surface-modified polymer nanoparticles (NPs) for mucosal delivery of proteins. We have developed poly (D, L-lactide-co-glycolide) (PLGA) NPs, surface-modified COS-PLGA NPs and CS-PLGA NPs, by using double emulsion solvent evaporation method, for encapsulating bovine serum albumin (BSA) as a model protein. Surface modification of NPs was confirmed using physicochemical characterization methods such as particle size and zeta potential, SEM, TEM and FTIR analysis. Both surface-modified PLGA NPs displayed a slow release of protein compared to PLGA NPs. Furthermore, we have explored the mucoadhesive property of COS as a material for modifying the surface of polymeric NPs. During in vitro mucoadhesion test, positively charged COS-PLGA NPs and CS-PLGA NPs exhibited enhanced mucoadhesion, compared to negatively charged PLGA NPs. This interaction was anticipated to improve the cell interaction and uptake of NPs, which is an important requirement for mucosal delivery of proteins. All nanoformulations were found to be safe for cellular delivery when evaluated in A549 cells. Moreover, intracellular uptake behaviour of FITC-BSA loaded NPs was extensively investigated by confocal laser scanning microscopy and flow cytometry. As we hypothesized, positively charged COS-PLGA NPs and CS-PLGA NPs displayed enhanced intracellular uptake compared to negatively charged PLGA NPs. Our results demonstrated that CS- and COS-modified polymer NPs could be promising carriers for proteins, drugs and nucleic acids via nasal, oral, buccal, ocular and vaginal mucosal routes.

Biological Behavior of Osteoblast Cell and Apatite Forming Ability of the Surface Modified Ti Alloys.

Science.gov (United States)

Zhao, Jingming; Hwang, K H; Choi, W S; Shin, S J; Lee, J K

2016-02-01

Titanium as one kind of biomaterials comes in direct contact with the body, making evaluation of biocompatibility an important aspect to biomaterials development. Surface chemistry of titanium plays an important role in osseointegration. Different surface modification alters the surface chemistry and result in different biological response. In this study, three kinds of mixed acid solutions were used to treat Ti specimens to induce Ca-P formation. Following a strong mixed acid activation process, Ca-P coating successfully formed on the Ti surfaces in simulated body fluid. Strong mixed acid increased the roughness of the metal surface, because the porous and rough surface allows better adhesion between Ca-P coatings and substrates. After modification of titanium surface by mixed acidic solution and subsequently H2O2/HCL treatment evaluation of biocompatibility was conducted from hydroxyapatite formation by biomimetic process and cell viability on modified titanium surface. Nano-scale modification of titanium surfaces can alter cellular and tissue responses, which may benefit osseointegration and dental implant therapy. Results from this study indicated that surface treatment methods affect the surface morphology, type of TiO2 layer formed and subsequent apatite deposition and biological responses. The thermo scientific alamarblue cell viability assay reagent is used to quantitatively measure the viability of mammalian cell lines, bacteria and fungi by incorporating a rapid, sensitive and reliable fluorometric/colorimetric growth indicator, without any toxic and side effect to cell line. In addition, mixed acid treatment uses a lower temperature and shorter time period than widely used alkali treatment.

Biofuel cells based on direct enzyme-electrode contacts using PQQ-dependent glucose dehydrogenase/bilirubin oxidase and modified carbon nanotube materials.

Science.gov (United States)

Scherbahn, V; Putze, M T; Dietzel, B; Heinlein, T; Schneider, J J; Lisdat, F

2014-11-15

Two types of carbon nanotube electrodes (1) buckypaper (BP) and (2) vertically aligned carbon nanotubes (vaCNT) have been used for elaboration of glucose/O2 enzymatic fuel cells exploiting direct electron transfer. For the anode pyrroloquinoline quinone dependent glucose dehydrogenase ((PQQ)GDH) has been immobilized on [poly(3-aminobenzoic acid-co-2-methoxyaniline-5-sulfonic acid), PABMSA]-modified electrodes. For the cathode bilirubin oxidase (BOD) has been immobilized on PQQ-modified electrodes. PABMSA and PQQ act as promoter for enzyme bioelectrocatalysis. The voltammetric characterization of each electrode shows current densities in the range of 0.7-1.3 mA/cm(2). The BP-based fuel cell exhibits maximal power density of about 107 µW/cm(2) (at 490 mV). The vaCNT-based fuel cell achieves a maximal power density of 122 µW/cm(2) (at 540 mV). Even after three days and several runs of load a power density over 110 µW/cm(2) is retained with the second system (10mM glucose). Due to a better power exhibition and an enhanced stability of the vaCNT-based fuel cells they have been studied in human serum samples and a maximal power density of 41 µW/cm(2) (390 mV) can be achieved. Copyright © 2014 Elsevier B.V. All rights reserved.

Enhancement of Osteoblastic-Like Cell Activity by Glow Discharge Plasma Surface Modified Hydroxyapatite/β-Tricalcium Phosphate Bone Substitute

Directory of Open Access Journals (Sweden)

Eisner Salamanca

2017-11-01

Full Text Available Glow discharge plasma (GDP treatments of biomaterials, such as hydroxyapatite/β-tricalcium phosphate (HA/β-TCP composites, produce surfaces with fewer contaminants and may facilitate cell attachment and enhance bone regeneration. Thus, in this study we used argon glow discharge plasma (Ar-GDP treatments to modify HA/β-TCP particle surfaces and investigated the physical and chemical properties of the resulting particles (HA/β-TCP + Ar-GDP. The HA/β-TCP particles were treated with GDP for 15 min in argon gas at room temperature under the following conditions: power: 80 W; frequency: 13.56 MHz; pressure: 100 mTorr. Scanning electron microscope (SEM observations showed similar rough surfaces of HA/β-TCP + Ar-GDP HA/β-TCP particles, and energy dispersive spectrometry analyses showed that HA/β-TCP surfaces had more contaminants than HA/β-TCP + Ar-GDP surfaces. Ca/P mole ratios in HA/β-TCP and HA/β-TCP + Ar-GDP were 1.34 and 1.58, respectively. Both biomaterials presented maximal intensities of X-ray diffraction patterns at 27° with 600 a.u. At 25° and 40°, HA/β-TCP + Ar-GDP and HA/β-TCP particles had peaks of 200 a.u., which are similar to XRD intensities of human bone. In subsequent comparisons, MG-63 cell viability and differentiation into osteoblast-like cells were assessed on HA/β-TCP and HA/β-TCP + Ar-GDP surfaces, and Ar-GDP treatments led to improved cell growth and alkaline phosphatase activities. The present data indicate that GDP surface treatment modified HA/β-TCP surfaces by eliminating contaminants, and the resulting graft material enhanced bone regeneration.

Enzyme-catalyzed modification of PES surfaces: Reduction in adsorption of BSA, dextrin and tannin

NARCIS (Netherlands)

Nady, N.; Schroën, C.G.P.H.; Franssen, M.C.R.; Fokkink, R.G.; Mohy Eldin, M.S.; Zuilhof, H.; Boom, R.M.

2012-01-01

Poly(ethersulfone) (PES) can be modified in a flexible manner using mild, environmentally benign components such as 4-hydroxybenzoic acid and gallic acid, which can be attached to the surface via catalysis by the enzyme laccase. This leads to grafting of mostly linear polymeric chains (for

Enzyme detection by microfluidics

DEFF Research Database (Denmark)

2013-01-01

Microfluidic-implemented methods of detecting an enzyme, in particular a DNA-modifying enzyme, are provided, as well as methods for detecting a cell, or a microorganism expressing said enzyme. The enzyme is detected by providing a nucleic acid substrate, which is specifically targeted...... by that enzyme...

Alginate-modifying enzymes: Biological roles and biotechnological uses

Directory of Open Access Journals (Sweden)

Helga eErtesvåg

2015-05-01

Full Text Available Alginate denotes a group of industrially important 1-4-linked biopolymers composed of the C-5-epimers β-D-mannuronic acid (M and α-L-guluronic acid (G. The polysaccharide is manufactured from brown algae where it constitutes the main structural cell wall polymer. The physical properties of a given alginate molecule, e.g. gel-strength, water-binding capacity, viscosity and biocompatibility, are determined by polymer length, the relative amount and distribution of G residues and the acetyl content, all of which are controlled by alginate modifying enzymes. Alginate has also been isolated from some bacteria belonging to the genera Pseudomonas and Azotobacter, and bacterially synthesized alginate may be O-acetylated at O-2 and/or O-3. Initially, alginate is synthesized as polymannuronic acid, and some M residues are subsequently epimerized to G residues. In bacteria a mannuronan C-5-epimerase (AlgG and an alginate acetylase (AlgX are integral parts of the protein complex necessary for alginate polymerisation and export. All alginate-producing bacteria use periplasmic alginate lyases to remove alginate molecules aberrantly released to the periplasm. Alginate lyases are also produced by organisms that utilize alginate as carbon source. Most alginate-producing organisms encode more than one mannuronan C-5 epimerase, each introducing its specific pattern of G residues. Acetylation protects against further epimerization and from most alginate lyases. One enzyme with alginate deacetylase activity from Pseudomonas syringae has been reported. Functional and structural studies reveal that alginate lyases and epimerases have related enzyme mechanisms and catalytic sites. Alginate lyases are now utilized as tools for alginate characterization. Secreted epimerases have been shown to function well in vitro, and have been engineered further in order to obtain enzymes that can provide alginates with new and desired properties for use in medical and

Multijunction Solar Cell Technology for Mars Surface Applications

Science.gov (United States)

Stella, Paul M.; Mardesich, Nick; Ewell, Richard C.; Mueller, Robert L.; Endicter, Scott; Aiken, Daniel; Edmondson, Kenneth; Fetze, Chris

2006-01-01

Solar cells used for Mars surface applications have been commercial space qualified AM0 optimized devices. Due to the Martian atmosphere, these cells are not optimized for the Mars surface and as a result operate at a reduced efficiency. A multi-year program, MOST (Mars Optimized Solar Cell Technology), managed by JPL and funded by NASA Code S, was initiated in 2004, to develop tools to modify commercial AM0 cells for the Mars surface solar spectrum and to fabricate Mars optimized devices for verification. This effort required defining the surface incident spectrum, developing an appropriate laboratory solar simulator measurement capability, and to develop and test commercial cells modified for the Mars surface spectrum. This paper discusses the program, including results for the initial modified cells. Simulated Mars surface measurements of MER cells and Phoenix Lander cells (2007 launch) are provided to characterize the performance loss for those missions. In addition, the performance of the MER rover solar arrays is updated to reflect their more than two (2) year operation.

Co-regulation of histone-modifying enzymes in cancer.

Directory of Open Access Journals (Sweden)

Abul B M M K Islam

Full Text Available Cancer is characterized by aberrant patterns of expression of multiple genes. These major shifts in gene expression are believed to be due to not only genetic but also epigenetic changes. The epigenetic changes are communicated through chemical modifications, including histone modifications. However, it is unclear whether the binding of histone-modifying proteins to genomic regions and the placing of histone modifications efficiently discriminates corresponding genes from the rest of the genes in the human genome. We performed gene expression analysis of histone demethylases (HDMs and histone methyltransferases (HMTs, their target genes and genes with relevant histone modifications in normal and tumor tissues. Surprisingly, this analysis revealed the existence of correlations in the expression levels of different HDMs and HMTs. The observed HDM/HMT gene expression signature was specific to particular normal and cancer cell types and highly correlated with target gene expression and the expression of genes with histone modifications. Notably, we observed that trimethylation at lysine 4 and lysine 27 separated preferentially expressed and underexpressed genes, which was strikingly different in cancer cells compared to normal cells. We conclude that changes in coordinated regulation of enzymes executing histone modifications may underlie global epigenetic changes occurring in cancer.

Probing Chromatin-modifying Enzymes with Chemical Tools

KAUST Repository

Fischle, Wolfgang

2016-02-04

Chromatin is the universal template of genetic information in all eukaryotic organisms. Chemical modifications of the DNA-packaging histone proteins and the DNA bases are crucial signaling events in directing the use and readout of eukaryotic genomes. The enzymes that install and remove these chromatin modifications as well as the proteins that bind these marks govern information that goes beyond the sequence of DNA. Therefore, these so-called epigenetic regulators are intensively studied and represent promising drug targets in modern medicine. We summarize and discuss recent advances in the field of chemical biology that have provided chromatin research with sophisticated tools for investigating the composition, activity, and target sites of chromatin modifying enzymes and reader proteins.

Immobilized enzymes and cells

Energy Technology Data Exchange (ETDEWEB)

Bucke, C; Wiseman, A

1981-04-04

This article reviews the current state of the art of enzyme and cell immobilization and suggests advances which might be made during the 1980's. Current uses of immobilized enzymes include the use of glucoamylase in the production of glucose syrups from starch and glucose isomerase in the production of high fructose corn syrup. Possibilities for future uses of immobilized enzymes and cells include the utilization of whey and the production of ethanol.

Enhanced compatibility of chemically modified titanium surface with periodontal ligament cells

International Nuclear Information System (INIS)

Kado, T.; Hidaka, T.; Aita, H.; Endo, K.; Furuichi, Y.

2012-01-01

Highlights: ► Cell-adhesive molecules were covalently immobilized on a Ti surface. ► Immobilized cell-adhesive molecules maintained native function on the Ti surface. ► Immobilized collagen enhanced adhesion of periodontal ligament cells to the Ti. - Abstract: A simple chemical modification method was developed to immobilize cell-adhesive molecules on a titanium surface to improve its compatibility with human periodontal ligament cells (HPDLCs).The polished titanium disk was immersed in 1% (v/v) p-vinylbenzoic acid solution for 2 h to introduce carboxyl groups onto the surface. After rinsing with distilled deionized water, the titanium disk was dipped into 1.47% 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide solution containing 0.1 mg/ml Gly-Arg-Gly-Asp-Ser (GRGDS), human plasma fibronectin (pFN), or type I collagen from calf skin (Col) to covalently immobilize the cell-adhesive molecules on the titanium surface via formation of peptide bonds. X-ray photoelectron spectroscopy analyses revealed that cell-adhesive molecules were successfully immobilized on the titanium surfaces. The Col-immobilized titanium surface revealed higher values regarding nano rough characteristics than the as-polished titanium surface under scanning probe microscopy. The number of HPDLCs attached to both the pFN- and Col-immobilized titanium surfaces was twice that attached to the as-polished titanium surfaces. The cells were larger with the cellular processes that stretched to a greater extent on the pFN- and Col-immobilized titanium surfaces than on the as-polished titanium surface (p < 0.05). HPDLCs on the Col-immobilized titanium surfaces showed more extensive expression of vinculin at the tips of cell projections and more contiguously along the cell outline than on the as-polished, GRGDS-immobilized and pFN-immobilized titanium surfaces. It was concluded that cell-adhesive molecules successfully immobilized on the titanium surface and improved the compatibility of the surface

Stem cell responses to plasma surface modified electrospun polyurethane scaffolds.

Science.gov (United States)

Zandén, Carl; Hellström Erkenstam, Nina; Padel, Thomas; Wittgenstein, Julia; Liu, Johan; Kuhn, H Georg

2014-07-01

The topographical effects from functional materials on stem cell behavior are currently of interest in tissue engineering and regenerative medicine. Here we investigate the influence of argon, oxygen, and hydrogen plasma surface modification of electrospun polyurethane fibers on human embryonic stem cell (hESC) and rat postnatal neural stem cell (NSC) responses. The plasma gases were found to induce three combinations of fiber surface functionalities and roughness textures. On randomly oriented fibers, plasma treatments lead to substantially increased hESC attachment and proliferation as compared to native fibers. Argon plasma was found to induce the most optimal combination of surface functionality and roughness for cell expansion. Contact guided migration of cells and alignment of cell processes were observed on aligned fibers. Neuronal differentiation around 5% was found for all samples and was not significantly affected by the induced variations of surface functional group distribution or individual fiber topography. In this study the influence of argon, oxygen, and hydrogen plasma surface modification of electrospun polyurethane fibers on human embryonic stem cell and rat postnatal neural stem cell (NSC) responses is studied with the goal of clarifying the potential effects of functional materials on stem cell behavior, a topic of substantial interest in tissue engineering and regenerative medicine. Copyright © 2014 Elsevier Inc. All rights reserved.

Chitosan surface modified electrospun poly(ε-caprolactone)/carbon nanotube composite fibers with enhanced mechanical, cell proliferation and antibacterial properties.

Science.gov (United States)

Wang, Siyu; Li, Yumei; Zhao, Rui; Jin, Toufeng; Zhang, Li; Li, Xiang

2017-11-01

The surface modification is one of the most effective methods to improve the bioactivity and cell affinity effect of electrospun poly(ε-caprolactone) (PCL) fibers. In the present study, chitosan (CS), a cationic polysaccharide, was used to modify the surface of electrospun PCL fibers. To obtain strong interaction between CS and PCL fibers, negatively charged PCL fibers were prepared by the incorporation of acid-treated carbon nanotubes (CNTs) into the fibers. In this way, the positively charged chitosan could be immobilized onto the surface of PCL fibers tightly by the electrostatic attraction. Besides, the incorporation of CNTs could significantly improve the mechanical strength of electrospun PCL fibers even after the CS modification, which guaranteed their usability in practical applications. The CS modification could effectively improve the wettability and bioactivity of electrospun PCL fibers. Cultivation of L929 fibroblast cells on the obtained fibers and the antibacterial activity were both evaluated to discuss the influence of chitosan modification. The results indicated that this modification could enhance the cell proliferation and antibacterial ability in comparison to the non-modified groups. Copyright © 2017 Elsevier B.V. All rights reserved.

Aptamer-conjugated dendrimer-modified quantum dots for glioblastoma cells imaging

International Nuclear Information System (INIS)

Li Zhiming; Huang Peng; He Rong; Bao Chenchen; Cui Daxiang; Zhang Xiaomin; Ren Qiushi

2009-01-01

Targeted quantum dots have shown potential as a platform for development of cancer imaging. Aptamers have recently been demonstrated as ideal candidates for molecular targeting applications. In present work, polyamidoamine dendrimers were used to modify surface of quantum dots and improve their solubility in water solution. Then, dendrimer-modified quantum dots were conjugated with DNA aptamer, GBI-10, can recognize the extracellular matrix protein tenascin-C on the surface of human glioblastoma cells. The dendrimer-modified quantum dots exhibit water-soluble, high quantum yield, and good biocompatibility. Aptamer-conjugated quantum dots can specifically target U251 human glioblastoma cells. High-performance aptamer-conjugated dendrimers modified quantum dot-based nanoprobes have great potential in application such as cancer imaging.

Flow-through 3D biofuel cell anode for NAD+-dependent enzymes

International Nuclear Information System (INIS)

Rincon, Rosalba A.; Lau, Carolin; Garcia, Kristen E.; Atanassov, Plamen

2011-01-01

NAD + -dependent enzymes require the presence of catalysts for cofactor regeneration in order to be employed in enzymatic biofuel cells. Poly-(methylene green) catalysts have proven to help the oxidation reaction of NADH allowing for the use of such enzymes in electrocatalytic oxidation reactions. In this paper we present the development of 3D anode based on NAD + -dependent malate dehydrogenase. The 3D material chosen was reticulated vitreous carbon (RVC) which was modified with poly-(MG) for NADH oxidation and it also accommodated the porous immobilization matrix for MDH consisting of MWCNTs embedded in chitosan; allowing for mass transport of the substrate to the electrode. Scanning electron microscopy was used in order to characterize the poly-(MG)-modified RVC, and electrochemical evaluation of the anode was performed.

Yeast surface displaying glucose oxidase as whole-cell biocatalyst: construction, characterization, and its electrochemical glucose sensing application.

Science.gov (United States)

Wang, Hongwei; Lang, Qiaolin; Li, Liang; Liang, Bo; Tang, Xiangjiang; Kong, Lingrang; Mascini, Marco; Liu, Aihua

2013-06-18

The display of glucose oxidase (GOx) on yeast cell surface using a-agglutinin as an anchor motif was successfully developed. Both the immunochemical analysis and enzymatic assay showed that active GOx was efficiently expressed and translocated on the cell surface. Compared with conventional GOx, the yeast cell surface that displayed GOx (GOx-yeast) demonstrated excellent enzyme properties, such as good stability within a wide pH range (pH 3.5-11.5), good thermostability (retaining over 94.8% enzyme activity at 52 °C and 84.2% enzyme activity at 56 °C), and high d-glucose specificity. In addition, direct electrochemistry was achieved at a GOx-yeast/multiwalled-carbon-nanotube modified electrode, suggesting that the host cell of yeast did not have any adverse effect on the electrocatalytic property of the recombinant GOx. Thus, a novel electrochemical glucose biosensor based on this GOx-yeast was developed. The as-prepared biosensor was linear with the concentration of d-glucose within the range of 0.1-14 mM and a low detection limit of 0.05 mM (signal-to-noise ratio of S/N = 3). Moreover, the as-prepared biosensor is stable, specific, reproducible, simple, and cost-effective, which can be applicable for real sample detection. The proposed strategy to construct robust GOx-yeast may be applied to explore other oxidase-displaying-system-based whole-cell biocatalysts, which can find broad potential application in biosensors, bioenergy, and industrial catalysis.

Osseointegration improvement by plasma electrolytic oxidation of modified titanium alloys surfaces.

Science.gov (United States)

Echeverry-Rendón, Mónica; Galvis, Oscar; Quintero Giraldo, David; Pavón, Juan; López-Lacomba, José Luis; Jiménez-Piqué, Emilio; Anglada, Marc; Robledo, Sara M; Castaño, Juan G; Echeverría, Félix

2015-02-01

Titanium (Ti) is a material frequently used in orthopedic applications, due to its good mechanical properties and high corrosion resistance. However, formation of a non-adherent fibrous tissue between material and bone drastically could affect the osseointegration process and, therefore, the mechanical stability of the implant. Modifications of topography and configuration of the tissue/material interface is one of the mechanisms to improve that process by manipulating parameters such as morphology and roughness. There are different techniques that can be used to modify the titanium surface; plasma electrolytic oxidation (PEO) is one of those alternatives, which consists of obtaining porous anodic coatings by controlling parameters such as voltage, current, anodizing solution and time of the reaction. From all of the above factors, and based on previous studies that demonstrated that bone cells sense substrates features to grow new tissue, in this work commercially pure Ti (c.p Ti) and Ti6Al4V alloy samples were modified at their surface by PEO in different anodizing solutions composed of H2SO4 and H3PO4 mixtures. Treated surfaces were characterized and used as platforms to grow osteoblasts; subsequently, cell behavior parameters like adhesion, proliferation and differentiation were also studied. Although the results showed no significant differences in proliferation, differentiation and cell biological activity, overall results showed an important influence of topography of the modified surfaces compared with polished untreated surfaces. Finally, this study offers an alternative protocol to modify surfaces of Ti and their alloys in a controlled and reproducible way in which biocompatibility of the material is not compromised and osseointegration would be improved.

Investigation of enzyme modified cheese production by two species ...

African Journals Online (AJOL)

ONOS

2010-01-25

Jan 25, 2010 ... African Journal of Biotechnology Vol. 9(4), pp. 508-511, 25 ... were used in this study for production of enzyme modified cheese. The results showed that ... Wheat bran was used as the substrate for cultivation of the molds and ...

An in vitro assay to study the recruitment and substrate specificity of chromatin modifying enzymes

Directory of Open Access Journals (Sweden)

Vermeulen Michiel

2004-01-01

Full Text Available Post-translational modifications of core histones play an important role in regulating fundamental biological processes such as DNA repair, transcription and replication. In this paper, we describe a novel assay that allows sequential targeting of distinct histone modifying enzymes to immobilized nucleosomal templates using recombinant chimeric targeting molecules. The assay can be used to study the histone substrate specificity of chromatin modifying enzymes as well as whether and how certain enzymes affect each other's histone modifying activities. As such the assay can help to understand how a certain histone code is established and interpreted.

Activation of interfacial enzymes at membrane surfaces

DEFF Research Database (Denmark)

Mouritsen, Ole G.; Andresen, Thomas Lars; Halperin, Avi

2006-01-01

A host of water-soluble enzymes are active at membrane surfaces and in association with membranes. Some of these enzymes are involved in signalling and in modification and remodelling of the membranes. A special class of enzymes, the phospholipases, and in particular secretory phospholipase A2 (s...

Novel Enzymes for Targeted Hydrolysis of Algal Cell Walls

DEFF Research Database (Denmark)

Schultz-Johansen, Mikkel

Seaweeds, also known as macroalgae, constitute a rich source of valuable biomolecules which have a potential industrial application in food and pharma products. The use of enzymes can optimize the extraction and separation of these molecules from the seaweed biomass, but most commercial enzymes...... are incapable of breaking the complex polysaccharides found in seaweed cell walls. Therefore, new enzymes are needed for degradation of seaweed biomass. Bacteria that colonize the surfaces of seaweed secrete enzymes that allow them to degrade and utilize seaweed polysaccharides as energy. In addition, sea...... degradation. In addition, three carrageenases were characterised; one as a GH16 κ-carrageenase whereas the other two belong to a new GH16 subfamily of enzymes that degrade furcellaran (κ/β-carrageenan). From metagenome sequence data three putative GH107 fucanases were identified and characterized...

Tissue engineering of bladder using vascular endothelial growth factor gene-modified endothelial progenitor cells.

Science.gov (United States)

Chen, Bai-Song; Xie, Hua; Zhang, Sheng-Li; Geng, Hong-Quan; Zhou, Jun-Mei; Pan, Jun; Chen, Fang

2011-12-01

This study assessed the use of vascular endothelial growth factor (VEGF) gene-modified endothelial progenitor cells (EPCs) seeded onto bladder acellular matrix grafts (BAMGs), to enhance the blood supply in tissue-engineered bladders in a porcine model. Autologous porcine peripheral EPCs were isolated, cultured, expanded, characterized, and modified with the VEGF gene using an adenovirus vector. The expression of VEGF was examined using reverse transcriptase polymerase chain reaction (RT-PCR) and an enzyme-linked immunosorbent assay (ELISA). VEGF gene modified EPCs were seeded onto BAMG and cultured for 3 days before implantation into pigs for bladder tissue engineering. A partial bladder cystectomy was performed in 12 pigs. The experimental group (6 pigs) received VEGF gene-modified EPC-seeded BAMG. The control group (6 pigs) received BAMG without seeded EPCs. The resulting tissue-engineered bladders were subject to a general and histological analysis. Microvessel density (MVD) was assessed using immunohistochemistry. The ex vivo transfection efficiency of EPCs was greater than 60%-70% when concentrated adenovirus was used. The genetically modified cells expressed both VEGF and green fluorescent protein (GFP). Scanning electron microscopy (SEM) and Masson's trichrome staining of cross sections of the cultured cells seeded to BAMG showed cell attachment and proliferation on the surface of the BAMG. Histological examination revealed bladder regeneration in a time-dependent fashion. Significant increases in MVD were observed in the experimental group, in comparison with the control group. VEGF-modified EPCs significantly enhanced neovascularization, compared with BAMG alone. These results indicate that EPCs, combined with VEGF gene therapy, may be a suitable approach for increasing blood supply in the tissue engineering of bladders. Thus, a useful strategy to achieve a tissue-engineered bladder is indicated.

Microplasma jet treatment of bovine serum albumin coatings for controlling enzyme and cell attachmenttype="fn" rid="FN1">

Science.gov (United States)

Szili, Endre J.; Becker, Stefanie; Short, Robert D.; Al-Bataineh, Sameer A.

2017-08-01

We investigated a new approach to control protein and cell attachment inside 96-well polystyrene plates. The wells were first coated with bovine serum albumin (BSA) to inhibit cell and protein attachment. The BSA-coated wells were then treated with a helium microplasma jet for increasing times that resulted in gradual removal of BSA from the surface. It was found that the amount of enzyme and cell attachment could be controlled in the wells where BSA was only partially removed by the microplasma jet. In addition to the surface coverage of BSA, the new surface chemistry induced by the microplasma jet treatment also had an important role in the control of enzyme and cell attachment. In summary, microplasma jet treatment of BSA-coated polystyrene wells is a simple and effective method for controlling enzyme and cell attachment. This might find use for high-throughput screening of new cell culture platforms where control over the level protein, enzyme or cell adherence is needed in order to maintain a specific cell function.

A model for cell wall dissolution in mating yeast cells: polarized secretion and restricted diffusion of cell wall remodeling enzymes induces local dissolution.

Science.gov (United States)

Huberman, Lori B; Murray, Andrew W

2014-01-01

Mating of the budding yeast, Saccharomyces cerevisiae, occurs when two haploid cells of opposite mating types signal using reciprocal pheromones and receptors, grow towards each other, and fuse to form a single diploid cell. To fuse, both cells dissolve their cell walls at the point of contact. This event must be carefully controlled because the osmotic pressure differential between the cytoplasm and extracellular environment causes cells with unprotected plasma membranes to lyse. If the cell wall-degrading enzymes diffuse through the cell wall, their concentration would rise when two cells touched each other, such as when two pheromone-stimulated cells adhere to each other via mating agglutinins. At the surfaces that touch, the enzymes must diffuse laterally through the wall before they can escape into the medium, increasing the time the enzymes spend in the cell wall, and thus raising their concentration at the point of attachment and restricting cell wall dissolution to points where cells touch each other. We tested this hypothesis by studying pheromone treated cells confined between two solid, impermeable surfaces. This confinement increases the frequency of pheromone-induced cell death, and this effect is diminished by reducing the osmotic pressure difference across the cell wall or by deleting putative cell wall glucanases and other genes necessary for efficient cell wall fusion. Our results support the model that pheromone-induced cell death is the result of a contact-driven increase in the local concentration of cell wall remodeling enzymes and suggest that this process plays an important role in regulating cell wall dissolution and fusion in mating cells.

A Model for Cell Wall Dissolution in Mating Yeast Cells: Polarized Secretion and Restricted Diffusion of Cell Wall Remodeling Enzymes Induces Local Dissolution

Science.gov (United States)

Huberman, Lori B.; Murray, Andrew W.

2014-01-01

Mating of the budding yeast, Saccharomyces cerevisiae, occurs when two haploid cells of opposite mating types signal using reciprocal pheromones and receptors, grow towards each other, and fuse to form a single diploid cell. To fuse, both cells dissolve their cell walls at the point of contact. This event must be carefully controlled because the osmotic pressure differential between the cytoplasm and extracellular environment causes cells with unprotected plasma membranes to lyse. If the cell wall-degrading enzymes diffuse through the cell wall, their concentration would rise when two cells touched each other, such as when two pheromone-stimulated cells adhere to each other via mating agglutinins. At the surfaces that touch, the enzymes must diffuse laterally through the wall before they can escape into the medium, increasing the time the enzymes spend in the cell wall, and thus raising their concentration at the point of attachment and restricting cell wall dissolution to points where cells touch each other. We tested this hypothesis by studying pheromone treated cells confined between two solid, impermeable surfaces. This confinement increases the frequency of pheromone-induced cell death, and this effect is diminished by reducing the osmotic pressure difference across the cell wall or by deleting putative cell wall glucanases and other genes necessary for efficient cell wall fusion. Our results support the model that pheromone-induced cell death is the result of a contact-driven increase in the local concentration of cell wall remodeling enzymes and suggest that this process plays an important role in regulating cell wall dissolution and fusion in mating cells. PMID:25329559

[Effects of sintered bone modified with surface mineralization/P24 peptide composite biomaterial on the adhesion, proliferation and osteodifferentiation of MC3T3-E1 cells].

Science.gov (United States)

Li, Jingfeng; Zheng, Qixin; Guo, Xiaodong; Chen, Liaobin

2014-10-01

In the present research, the effects of sintered bone modified with surface mineralization/P24 peptide composite biomaterials on the adhesion, proliferation and osteodifferentiation of MC3T3-E1 cells were investigated. The experiments were divided into three groups due to biomaterials used: Group A (composite materials of sintered bone modified with surface mineralization and P24, a peptide of bone morphogenetic protein-2); Group B (sintered bone modified with surface mineralization) and Group C (sintered bone only). The three groups were observed by scanning electron microscopy (SEM) before the experiments, respectively. Then MC3T3-E1 cells were cultured on the surfaces of the three kinds of material, respectively. The cell adhesion rate was assessed by precipitation method. The proliferative ability of MC3T3-E1 cells were measured with MTT assay. And the ALP staining and measurement of alkaline phosphatase (ALP) activity were performed to assess the differentiation of cells into osteoblasts. The SEM results showed that the materials in the three groups retained the natural pore structure and the pore sizes were in the range between 200-850 μm. The adhesive ratio measurements and MTT assay suggested that adhesion and proliferation of MC3T3-E1 cells in Group A were much higher than those in Group B and Group C (P bone modified with surface mineralization/P24 composite material was confirmed to improve the adhesion rate and proliferation and osteodifferentiation of MC3T3-E1 cells, and maintained their morphology.

An overview of technologies for immobilization of enzymes and surface analysis techniques for immobilized enzymes

Science.gov (United States)

Mohamad, Nur Royhaila; Marzuki, Nur Haziqah Che; Buang, Nor Aziah; Huyop, Fahrul; Wahab, Roswanira Abdul

2015-01-01

The current demands of sustainable green methodologies have increased the use of enzymatic technology in industrial processes. Employment of enzyme as biocatalysts offers the benefits of mild reaction conditions, biodegradability and catalytic efficiency. The harsh conditions of industrial processes, however, increase propensity of enzyme destabilization, shortening their industrial lifespan. Consequently, the technology of enzyme immobilization provides an effective means to circumvent these concerns by enhancing enzyme catalytic properties and also simplify downstream processing and improve operational stability. There are several techniques used to immobilize the enzymes onto supports which range from reversible physical adsorption and ionic linkages, to the irreversible stable covalent bonds. Such techniques produce immobilized enzymes of varying stability due to changes in the surface microenvironment and degree of multipoint attachment. Hence, it is mandatory to obtain information about the structure of the enzyme protein following interaction with the support surface as well as interactions of the enzymes with other proteins. Characterization technologies at the nanoscale level to study enzymes immobilized on surfaces are crucial to obtain valuable qualitative and quantitative information, including morphological visualization of the immobilized enzymes. These technologies are pertinent to assess efficacy of an immobilization technique and development of future enzyme immobilization strategies. PMID:26019635

Flow-through 3D biofuel cell anode for NAD{sup +}-dependent enzymes

Energy Technology Data Exchange (ETDEWEB)

Rincon, Rosalba A.; Lau, Carolin; Garcia, Kristen E. [Department of Chemical and Nuclear Engineering, Center for Emerging Energy Technologies, University of New Mexico, Albuquerque, NM 87131 (United States); Atanassov, Plamen, E-mail: plamen@unm.ed [Department of Chemical and Nuclear Engineering, Center for Emerging Energy Technologies, University of New Mexico, Albuquerque, NM 87131 (United States)

2011-02-01

NAD{sup +}-dependent enzymes require the presence of catalysts for cofactor regeneration in order to be employed in enzymatic biofuel cells. Poly-(methylene green) catalysts have proven to help the oxidation reaction of NADH allowing for the use of such enzymes in electrocatalytic oxidation reactions. In this paper we present the development of 3D anode based on NAD{sup +}-dependent malate dehydrogenase. The 3D material chosen was reticulated vitreous carbon (RVC) which was modified with poly-(MG) for NADH oxidation and it also accommodated the porous immobilization matrix for MDH consisting of MWCNTs embedded in chitosan; allowing for mass transport of the substrate to the electrode. Scanning electron microscopy was used in order to characterize the poly-(MG)-modified RVC, and electrochemical evaluation of the anode was performed.

Cell penetrating peptide-modified poly(lactic-co-glycolic acid) nanoparticles with enhanced cell internalization.

Science.gov (United States)

Steinbach, Jill M; Seo, Young-Eun; Saltzman, W Mark

2016-01-01

The surface modification of nanoparticles (NPs) can enhance the intracellular delivery of drugs, proteins, and genetic agents. Here we studied the effect of different surface ligands, including cell penetrating peptides (CPPs), on the cell binding and internalization of poly(lactic-co-glycolic) (PLGA) NPs. Relative to unmodified NPs, we observed that surface-modified NPs greatly enhanced cell internalization. Using one CPP, MPG (unabbreviated notation), that achieved the highest degree of internalization at both low and high surface modification densities, we evaluated the effect of two different NP surface chemistries on cell internalization. After 2h, avidin-MPG NPs enhanced cellular internalization by 5 to 26-fold relative to DSPE-MPG NP formulations. Yet, despite a 5-fold increase in MPG density on DSPE compared to Avidin NPs, both formulations resulted in similar internalization levels (48 and 64-fold, respectively) after 24h. Regardless of surface modification, all NPs were internalized through an energy-dependent, clathrin-mediated process, and became dispersed throughout the cell. Overall both Avidin- and DSPE-CPP modified NPs significantly increased internalization and offer promising delivery options for applications in which internalization presents challenges to efficacious delivery. Copyright © 2015 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Involvement of epigenetic modifiers in the pathogenesis of testicular dysgenesis and germ cell cancer

DEFF Research Database (Denmark)

Lawaetz, Andreas C.; Almstrup, Kristian

2015-01-01

Testicular germ cell cancer manifests mainly in young adults as a seminoma or non-seminoma. The solid tumors are preceded by the presence of a non-invasive precursor cell, the carcinoma in situ cell (CIS), which shows great similarity to fetal germ cells. It is therefore hypothesized that the CIS...... of epigenetic modifiers with a focus on jumonji C enzymes in the development of testicular dysgenesis and germ cell cancer in men....... cell is a fetal germ cell that has been arrested during development due to testicular dysgenesis. CIS cells retain a fetal and open chromatin structure, and recently several epigenetic modifiers have been suggested to be involved in testicular dysgenesis in mice. We here review the possible involvement...

Surface-modified Y zeolite-filled chitosan membrane for direct methanol fuel cell

Energy Technology Data Exchange (ETDEWEB)

Wu, Hong; Zheng, Bin; Zheng, Xiaohong; Wang, Jingtao; Yuan, Weikang; Jiang, Zhongyi [Key Laboratory for Green Chemical Technology, School of Chemical Engineering and Technology, Tianjin University, Tianjin 300072 (China)

2007-11-15

Hybrid membranes composed of chitosan (CS) as organic matrix and surface-modified Y zeolite as inorganic filler are prepared and their applicability for DMFC is demonstrated by methanol permeability, proton conductivity and swelling property. Y zeolite is modified using silane coupling agents, 3-aminopropyl-triethoxysilane (APTES) and 3-mercaptopropyl-trimethoxysilane (MPTMS), to improve the organic-inorganic interfacial morphology. The mercapto group on MPTMS-modified Y zeolite is further oxidized into sulfonic group. Then, the resultant surface-modified Y zeolites with either aminopropyl groups or sulfonicpropyl groups are mixed with chitosan in acetic acid solution and cast into membranes. The transitional phase generated between chitosan matrix and zeolite filler reduces or even eliminates the nonselective voids commonly exist at the interface. The hybrid membranes exhibit a significant reduction in methanol permeability compared with pure chitosan and Nafion117 membranes, and this reduction extent becomes more pronounced with the increase of methanol concentration. By introducing -SO{sub 3}H groups onto zeolite surface, the conductivity of hybrid membranes is increased up to 2.58 x 10{sup -2} S cm{sup -1}. In terms of the overall selectivity index ({beta} = {sigma}/P), the hybrid membrane is comparable with Nafion117 at low methanol concentration (2 mol L{sup -1}) and much better (three times) at high methanol concentration (12 mol L{sup -1}). (author)

Interaction of human endothelial cells and nickel-titanium materials modified with silicon ions

Energy Technology Data Exchange (ETDEWEB)

Lotkov, Aleksandr I., E-mail: lotkov@ispms.tsc.ru; Kashin, Oleg A., E-mail: okashin@ispms.tsc.ru [Institute of Strength Physics and Materials Science SB RAS, Tomsk, 634055 (Russian Federation); Kudryavtseva, Yuliya A., E-mail: yulia-k1970@mail.ru; Antonova, Larisa V., E-mail: antonova.la@mail.ru; Matveeva, Vera G., E-mail: matveeva-vg@mail.ru; Sergeeva, Evgeniya A., E-mail: sergeewa.ew@yandex.ru [Research Institute for Complex Issues of Cardiovascular Diseases, Kemerovo, 650002 (Russian Federation); Kudryashov, Andrey N., E-mail: kudryashov@angioline.ru [Angioline Interventional Device Ltd, Novosibirsk, 630090 (Russian Federation)

2015-10-27

The paper studies the influence of chemical and phase compositions of NiTi surface layers modified with Si ions by plasma immersion implantation on their interaction with endothelial cells. It is shown that certain technological modes of Si ion implantation enhance the adhesion, proliferation, and viability of endothelial cells. It is found that the Si-modified NiTi surface is capable of stimulating the formation of capillary-like structures in the cell culture.

Self-organization of yeast cells on modified polymer surfaces after dewetting: new perspectives in cellular patterning

Energy Technology Data Exchange (ETDEWEB)

Carnazza, S [Department of Microbiological, Genetic and Molecular Sciences, University of Messina, Messina (Italy); Satriano, S [Department of Chemical Sciences, University of Catania, Catania (Italy); Guglielmino, S [Department of Microbiological, Genetic and Molecular Sciences, University of Messina, Messina (Italy)

2006-08-23

In recent years, biological micro-electro-mechanical systems (commonly referred to as BioMEMS) have found widespread use, becoming increasingly prevalent in diagnostics and therapeutics. Cell-based sensors are nowadays gaining increasing attention, due to cellular built-in natural selectivity and physiologically relevant response to biologically active chemicals. On the other hand, surrogate microbial systems, including yeast models, have become a useful alternative to animal and mammalian cell systems for high-throughput screening for the identification of new pharmacological agents. A main obstacle in biosensor device fabrication is the need for localized geometric confinement of cells, without losing cell viability and sensing capability. Here we illustrate a new approach for cellular patterning using dewetting processes to control cell adhesion and spatial confinement on modified surfaces. By the control of simple system parameters, a rich variety of morphologies, ranging through hexagonal arrays, polygonal networks, bicontinuous structures, and elongated fingers, can be obtained.

Enzyme production in immobilized Trichoderma reesei cells with hydrophobic polymers prepared by radiation polymerization method

International Nuclear Information System (INIS)

Luzhao Xin; Kumakura, Minoru; Kaetsu, Isao

1993-01-01

Trichoderma reesei cells were immobilized on paper covered with hydrophobic monomer, trimethylpropane triacrylate by radiation polymerization. The effect of immobilization condition on enzyme productivity was studied by measuring filter paper and cellobiose activity. The cells were adhered and grew on the surface of the carrier with the polymer giving high enzyme productivity in the immobilized cells in comparison with the free cells. Optimum concentration and volume of the coating monomer for the preparation of the immobilized cells were obtained. (author)

NKT sublineage specification and survival requires the ubiquitin-modifying enzyme TNFAIP3/A20.

Science.gov (United States)

Drennan, Michael B; Govindarajan, Srinath; Verheugen, Eveline; Coquet, Jonathan M; Staal, Jens; McGuire, Conor; Taghon, Tom; Leclercq, Georges; Beyaert, Rudi; van Loo, Geert; Lambrecht, Bart N; Elewaut, Dirk

2016-09-19

Natural killer T (NKT) cells are innate lymphocytes that differentiate into NKT1, NKT2, and NKT17 sublineages during development. However, the signaling events that control NKT sublineage specification and differentiation remain poorly understood. Here, we demonstrate that the ubiquitin-modifying enzyme TNFAIP3/A20, an upstream regulator of T cell receptor (TCR) signaling in T cells, is an essential cell-intrinsic regulator of NKT differentiation. A20 is differentially expressed during NKT cell development, regulates NKT cell maturation, and specifically controls the differentiation and survival of NKT1 and NKT2, but not NKT17, sublineages. Remaining A20-deficient NKT1 and NKT2 thymocytes are hyperactivated in vivo and secrete elevated levels of Th1 and Th2 cytokines after TCR ligation in vitro. Defective NKT development was restored by compound deficiency of MALT1, a key downstream component of TCR signaling in T cells. These findings therefore show that negative regulation of TCR signaling during NKT development controls the differentiation and survival of NKT1 and NKT2 cells. © 2016 Drennan et al.

Design and modelling of enzyme/poly-pyrrole modified electrodes for bio-catalyzed electro-synthesis processes

International Nuclear Information System (INIS)

Gros, Pierre

1996-01-01

This research thesis reports a study which aims at developing, analyzing and integrating an electrode-enzyme interface within an electro-enzymatic reactor to develop electrochemical biosensors. The adopted method comprises a confinement of the enzyme at the electrode surface by means of an electro-formed poly-pyrrole film. The author reports an experimental and theoretical study of the coupling between electrochemical reaction, enzymatic reaction and matter transfer in the polymer in order to better understand the operation of so-modified electrodes. Different parameters have an influence on the reaction rate. A numerical model (validated by experiments) allows the identification of the reaction limiting stages. A new elaboration protocol allows the polymer permeability to be increased. The interface is first applied to the reduction of the NAD coenzyme, and the process is also applied to the production of gluconic acid [fr

Microbial surface displayed enzymes based biofuel cell utilizing degradation products of lignocellulosic biomass for direct electrical energy.

Science.gov (United States)

Fan, Shuqin; Hou, Chuantao; Liang, Bo; Feng, Ruirui; Liu, Aihua

2015-09-01

In this work, a bacterial surface displaying enzyme based two-compartment biofuel cell for the direct electrical energy conversion from degradation products of lignocellulosic biomass is reported. Considering that the main degradation products of the lignocellulose are glucose and xylose, xylose dehydrogenase (XDH) displayed bacteria (XDH-bacteria) and glucose dehydrogenase (GDH) displayed bacteria (GDH-bacteria) were used as anode catalysts in anode chamber with methylene blue as electron transfer mediator. While the cathode chamber was constructed with laccase/multi-walled-carbon nanotube/glassy-carbon-electrode. XDH-bacteria exhibited 1.75 times higher catalytic efficiency than GDH-bacteria. This assembled enzymatic fuel cell exhibited a high open-circuit potential of 0.80 V, acceptable stability and energy conversion efficiency. Moreover, the maximum power density of the cell could reach 53 μW cm(-2) when fueled with degradation products of corn stalk. Thus, this finding holds great potential to directly convert degradation products of biomass into electrical energy. Copyright © 2015 Elsevier Ltd. All rights reserved.

Effect of Surface-Modified Paclitaxel Nanowires on U937 Cells In Vitro: A Novel Drug Delivery Vehicle

Directory of Open Access Journals (Sweden)

Mohamed H. Abumaree

2012-01-01

Full Text Available We have fabricated surface-modified paclitaxel nanowires (SM-PNs with a precise diameter and an average length of 50 μm. The surface of these nanowires is coated with a monolayer of octadecylsiloxane (ODS, which prevents aggregation and enhances dispersivity in aqueous media. This system constitutes a novel drug delivery vehicle based on one-dimensional (1D nanostructures with a large drug to vehicle ratio. We assayed the cytotoxicity of different diameter SM-PNs (200, 80, 35, and 18 nm with U937 cells and compared their activity to microcrystalline paclitaxel. SM-PNs reduced U937 cell proliferation in culture followed by cell death. For the same amount of paclitaxel, different diameter SM-PNs displayed different cytotoxic effect at the same incubation time period. SM-PNs with 35 nm diameters were the most efficient in completely halting cell proliferation following the first 24 hours of treatment, associated with 42% cell death. SM-PNs with 18 nm diameters were least effective. These SM-PNs can be tailored to fit a certain treatment protocol by simply choosing the appropriate diameter.

Assessing cellulolysis in passive treatment systems for mine drainage: a modified enzyme assay.

Science.gov (United States)

McDonald, Corina M; Gould, W Douglas; Lindsay, Matthew B J; Blowes, David W; Ptacek, Carol J; Condon, Peter D

2013-01-01

A modified cellulase enzyme assay was developed to monitor organic matter degradation in passive treatment systems for mine drainage. This fluorogenic substrate method facilitates assessment of exo-(1,4)-β-D-glucanase, endo-(1,4)-β-D-glucanase, and β-glucosidase, which compose an important cellulase enzyme system. The modified method was developed and refined using samples of organic carbon-amended mine tailings from field experiments where sulfate reduction was induced as a strategy for managing water quality. Sample masses (3 g) and the number of replicates ( ≥ 3) were optimized. Matrix interferences within these metal-rich samples were found to be insignificant. Application of this modified cellulase assay method provided insight into the availability and degradation of organic carbon within the amended tailings. Results of this study indicate that cellulase enzyme assays can be applied to passive treatment systems for mine drainage, which commonly contain elevated concentrations of metals. Copyright © by the American Society of Agronomy, Crop Science Society of America, and Soil Science Society of America, Inc.

Influence of the surface structure on the filtration performance of UV-modified PES membranes

DEFF Research Database (Denmark)

Kæselev, Bozena Alicja; Kingshott, P.; Jonsson, Gunnar Eigil

2002-01-01

chemically characterised using X-ray photoelectron spectroscopy (XPS) and time of flight-static secondary ion mass spectrometry (TOF-static SIMS). The filtration performance of irradiated/non-modified and irradiated/modified membranes was examined in a crossflow cell, using a dextran solution. The filtration...... in relation to dextran when compared to membranes modified by AAG and AAP. This work suggests that the structure of the presence of grafted chains seems to be responsible for the observed changes to filtration performance of the modified membrane. Surface analysis supports the claim that the specific surface...

Glycoprotein on cell surfaces

International Nuclear Information System (INIS)

Muramatsu, T.

1975-01-01

There are conjugated polysaccharides in cell membranes and outside of animal cells, and they play important role in the control of cell behavior. In this paper, the studies on the glycoprotein on cell surfaces are reported. It was found that the glycoprotein on cell surfaces have both N-glycoside type and O-glycoside type saccharic chains. Therefore it can be concluded that the basic structure of the saccharic chains in the glycoprotein on cell surfaces is similar to that of blood serum and body fluid. The main glycoprotein in the membranes of red blood corpuscles has been studied most in detail, and it also has both types of saccharic chains. The glycoprotein in liver cell membranes was found to have only the saccharic chains of acid type and to be in different pattern from that in endoplasmic reticula and nuclear membranes, which also has the saccharic chains of neutral type. The structure of the saccharic chains of H-2 antigen, i.e. the peculiar glycoprotein on the surfaces of lymph system cells, has been studied, and it is similar to the saccharic chains of glycoprotein in blood serum. The saccharic chain structures of H-2 antigen and TL antigen are different. TL, H-2 (D), Lna and H-2 (K) are the glycoprotein on cell surfaces, and are independent molecules. The analysis of the saccharic chain patterns on cell surfaces was carried out, and it was shown that the acid type saccharic chains were similar to those of ordinary glycoprotein, because the enzyme of pneumococci hydrolyzed most of the acid type saccharic chains. The change of the saccharic chain patterns of glycoprotein on cell surfaces owing to canceration and multiplication is complex matter. (Kako, I.)

Immobilization of enzymes using non-ionic colloidal liquid aphrons (CLAs): Surface and enzyme effects.

Science.gov (United States)

Ward, Keeran; Xi, Jingshu; Stuckey, David C

2015-12-01

The use of non-ionic colloidal liquid aphrons (CLAs) as a support for enzyme immobilisation was investigated. Formulation required the mixing of an aqueous-surfactant solution with a relatively non-polar solvent-surfactant solution, forming a solvent droplet surrounded by a thin stabilised aqueous film (soapy shell). Studies utilising anionic surfactants have showed increased retention, however, very little have been understood about the forces governing immobilisation. This study seeks to determine the effects of enzyme properties on CLA immobilisation by examining a non-ionic/non-polar solvent system comprised of two non-ionic surfactants, Tween 20 and 80, mineral oil and the enzymes lipase, aprotinin and α-chymotrypsin. From these results it was deduced that hydrophobic interactions strongly governed immobilisation. Confocal Scanning Laser Microscopy (CSLM) revealed that immobilisation was predominantly achieved by surface adsorption attributed to hydrophobic interactions between the enzyme and the CLA surface. Enzyme surface affinity was found to increase when added directly to the formulation (pre-manufacture addition), as opposed to the bulk continuous phase (post-manufacture addition), with α-chymotrypsin and aprotinin being the most perturbed, while lipase was relatively unaffected. The effect of zeta potential on immobilisation showed that enzymes adsorbed better closer to their pI, indicating that charge minimisation was necessary for immobilisation. Finally, the effect of increasing enzyme concentration in the aqueous phase resulted in an increase in adsorption for all enzymes due to cooperativity between protein molecules, with saturation occurring faster at higher adsorption rates. Copyright © 2015 Elsevier B.V. All rights reserved.

Cell wall structure suitable for surface display of proteins in Saccharomyces cerevisiae.

Science.gov (United States)

Matsuoka, Hiroyuki; Hashimoto, Kazuya; Saijo, Aki; Takada, Yuki; Kondo, Akihiko; Ueda, Mitsuyoshi; Ooshima, Hiroshi; Tachibana, Taro; Azuma, Masayuki

2014-02-01

A display system for adding new protein functions to the cell surfaces of microorganisms has been developed, and applications of the system to various fields have been proposed. With the aim of constructing a cell surface environment suitable for protein display in Saccharomyces cerevisiae, the cell surface structures of cell wall mutants were investigated. Four cell wall mutant strains were selected by analyses using a GFP display system via a GPI anchor. β-Glucosidase and endoglucanase II were displayed on the cell surface in the four mutants, and their activities were evaluated. mnn2 deletion strain exhibited the highest activity for both the enzymes. In particular, endoglucanase II activity using carboxymethylcellulose as a substrate in the mutant strain was 1.9-fold higher than that of the wild-type strain. In addition, the activity of endoglucanase II released from the mnn2 deletion strain by Zymolyase 20T treatment was higher than that from the wild-type strain. The results of green fluorescent protein (GFP) and endoglucanase displays suggest that the amounts of enzyme displayed on the cell surface were increased by the mnn2 deletion. The enzyme activity of the mnn2 deletion strain was compared with that of the wild-type strain. The relative value (mnn2 deletion mutant/wild-type strain) of endoglucanase II activity using carboxymethylcellulose as a substrate was higher than that of β-glucosidase activity using p-nitrophenyl-β-glucopyranoside as a substrate, suggesting that the cell surface environment of the mnn2 deletion strain facilitates the binding of high-molecular-weight substrates to the active sites of the displayed enzymes. Copyright © 2014 John Wiley & Sons, Ltd.

Proliferation of Genetically Modified Human Cells on Electrospun Nanofiber Scaffolds

Directory of Open Access Journals (Sweden)

Mandula Borjigin

2012-01-01

Full Text Available Gene editing is a process by which single base mutations can be corrected, in the context of the chromosome, using single-stranded oligodeoxynucleotides (ssODNs. The survival and proliferation of the corrected cells bearing modified genes, however, are impeded by a phenomenon known as reduced proliferation phenotype (RPP; this is a barrier to practical implementation. To overcome the RPP problem, we utilized nanofiber scaffolds as templates on which modified cells were allowed to recover, grow, and expand after gene editing. Here, we present evidence that some HCT116-19, bearing an integrated, mutated enhanced green fluorescent protein (eGFP gene and corrected by gene editing, proliferate on polylysine or fibronectin-coated polycaprolactone (PCL nanofiber scaffolds. In contrast, no cells from the same reaction protocol plated on both regular dish surfaces and polylysine (or fibronectin-coated dish surfaces proliferate. Therefore, growing genetically modified (edited cells on electrospun nanofiber scaffolds promotes the reversal of the RPP and increases the potential of gene editing as an ex vivo gene therapy application.

Effect of Amelogenin Coating of a Nano-Modified Titanium Surface on Bioactivity

Directory of Open Access Journals (Sweden)

Chisato Terada

2018-04-01

Full Text Available The interactions between implants and host tissues depend on several factors. In particular, a growing body of evidence has demonstrated that the surface texture of an implant influences the response of the surrounding cells. The purpose of this study is to develop new implant materials aiming at the regeneration of periodontal tissues as well as hard tissues by coating nano-modified titanium with amelogenin, which is one of the main proteins contained in Emdogain®. We confirmed by quartz crystal microbalance evaluation that amelogenin is easy to adsorb onto the nano-modified titanium surface as a coating. Scanning electron microscopy, scanning probe microscopy, X-ray photoelectron spectroscopy, and Fourier-transform infrared spectroscopy analyses confirmed that amelogenin coated the nano-modified titanium surface following alkali-treatment. In vitro evaluation using rat bone marrow and periodontal ligament cells revealed that the initial adhesion of both cell types and the induction of hard tissue differentiation such as cementum were improved by amelogenin coating. Additionally, the formation of new bone in implanted surrounding tissues was observed in in vivo evaluation using rat femurs. Together, these results suggest that this material may serve as a new implant material with the potential to play a major role in the advancement of clinical dentistry.

Horseradish peroxidase-modified porous silicon for phenol monitoring

Energy Technology Data Exchange (ETDEWEB)

Kermad, A., E-mail: amina_energetique@yahoo.fr [Unité de Recherche Matériaux et Energies Renouvelables (URMER), Département de Physique, Faculté des Sciences, Université Abou Baker Belkaid, B.P. 119, Tlemcen 13000 (Algeria); Sam, S., E-mail: Sabrina.sam@polytechnique.edu [Centre de Recherche en Technologie des Semi-conducteurs pour l’Energétique (CRTSE), 02 Bd. Frantz-Fanon, B.P. 140, Alger-7 merveilles, Algiers (Algeria); Ghellai, N., E-mail: na_ghellai@yahoo.fr [Unité de Recherche Matériaux et Energies Renouvelables (URMER), Département de Physique, Faculté des Sciences, Université Abou Baker Belkaid, B.P. 119, Tlemcen 13000 (Algeria); Khaldi, K., E-mail: Khadidjaphy@yahoo.fr [Unité de Recherche Matériaux et Energies Renouvelables (URMER), Département de Physique, Faculté des Sciences, Université Abou Baker Belkaid, B.P. 119, Tlemcen 13000 (Algeria); Gabouze, N., E-mail: ngabouze@yahoo.fr [Centre de Recherche en Technologie des Semi-conducteurs pour l’Energétique (CRTSE), 02 Bd. Frantz-Fanon, B.P. 140, Alger-7 merveilles, Algiers (Algeria)

2013-11-01

Highlights: • Horseradish peroxidase enzyme (HRP) was covalently immobilized on porous silicon (PSi) surface. • Multistep strategy was used allowing the maintaining of the enzymatic activity of the immobilized enzyme. • Direct electron transfer has occurred between the immobilized enzyme and the surface. • Electrochemical measurements showed a response of HRP-modified PSi toward phenol in the presence of H{sub 2}O{sub 2}. -- Abstract: In this study, horseradish peroxidase enzyme (HRP) was covalently immobilized on porous silicon (PSi) surface using multistep strategy. First, acid terminations were generated on hydrogenated PSi surface by thermal hydrosilylation of undecylenic acid. Then, the carboxyl-terminated monolayer was transformed to active ester (succinimidyl ester) using N-hydroxysuccinimide (NHS) in the presence of the coupling agent N-ethyl-N′-(3-dimethylaminopropyl) carbodiimide (EDC). Subsequently, the enzyme was anchored on the surface via an amidation reaction. The structure of the PSi layers was observed by scanning electron microscopy (SEM). Infrared spectroscopy (FTIR) and contact angle measurements confirmed the efficiency of the modification at each step of the functionalization. Cyclic voltammetry was recorded using the HRP-modified PSi as working electrode. The results show that the enzymatic activity of the immobilized HRP is preserved and in the presence of hydrogen peroxide, the enzyme oxidizes phenolic molecules which were subsequently reduced at the modified-PSi electrode.

Horseradish peroxidase-modified porous silicon for phenol monitoring

International Nuclear Information System (INIS)

Kermad, A.; Sam, S.; Ghellai, N.; Khaldi, K.; Gabouze, N.

2013-01-01

Highlights: • Horseradish peroxidase enzyme (HRP) was covalently immobilized on porous silicon (PSi) surface. • Multistep strategy was used allowing the maintaining of the enzymatic activity of the immobilized enzyme. • Direct electron transfer has occurred between the immobilized enzyme and the surface. • Electrochemical measurements showed a response of HRP-modified PSi toward phenol in the presence of H 2 O 2 . -- Abstract: In this study, horseradish peroxidase enzyme (HRP) was covalently immobilized on porous silicon (PSi) surface using multistep strategy. First, acid terminations were generated on hydrogenated PSi surface by thermal hydrosilylation of undecylenic acid. Then, the carboxyl-terminated monolayer was transformed to active ester (succinimidyl ester) using N-hydroxysuccinimide (NHS) in the presence of the coupling agent N-ethyl-N′-(3-dimethylaminopropyl) carbodiimide (EDC). Subsequently, the enzyme was anchored on the surface via an amidation reaction. The structure of the PSi layers was observed by scanning electron microscopy (SEM). Infrared spectroscopy (FTIR) and contact angle measurements confirmed the efficiency of the modification at each step of the functionalization. Cyclic voltammetry was recorded using the HRP-modified PSi as working electrode. The results show that the enzymatic activity of the immobilized HRP is preserved and in the presence of hydrogen peroxide, the enzyme oxidizes phenolic molecules which were subsequently reduced at the modified-PSi electrode

Glucose oxidase immobilization on different modified surfaces of platinum nanowire for application in glucose detection

International Nuclear Information System (INIS)

Le, Thi Thanh Tuyen; Tran, Phu Duy; Pham, Xuan Tung; Tong, Duy Hien; Dang, Mau Chien

2010-01-01

In this work, the surface of platinum (Pt) nanowires was modified by using several chemicals, including a compound of gelatin gel with SiO 2 , polyvinyl alcohol (PVA) with Prussian blue (PB) mediator and cysteamine self-assembled monolayers (SAM). Then, glucose oxidase (GOD) enzyme was immobilized on the modified surfaces of Pt nanowire electrodes by using techniques of electrochemical adsorption and chemical binding. The GOD immobilized Pt nanowires were used for application in glucose detection by performing a cyclic voltammetry measurement. The detection results showed that GOD was immobilized on all of the tested surfaces and the highest glucose detection sensitivity of 60 μM was obtained when the Pt nanowires were modified by PVA with PB mediator. Moreover, the sensors showed very high current response when the Pt nanowires were modified with the cysteamine SAM. The stability and catalyst activity of GOD are also reported here. For instance, the catalyst activity of GOD retained about 60% of its initial value after it was stored at 4 °C in a 100 mM PBS buffer solution with a pH of 7.2 for a period of 30 days

Glucose oxidase immobilization on different modified surfaces of platinum nanowire for application in glucose detection

Science.gov (United States)

Thanh Tuyen Le, Thi; Duy Tran, Phu; Pham, Xuan Tung; Hien Tong, Duy; Chien Dang, Mau

2010-09-01

In this work, the surface of platinum (Pt) nanowires was modified by using several chemicals, including a compound of gelatin gel with SiO2, polyvinyl alcohol (PVA) with Prussian blue (PB) mediator and cysteamine self-assembled monolayers (SAM). Then, glucose oxidase (GOD) enzyme was immobilized on the modified surfaces of Pt nanowire electrodes by using techniques of electrochemical adsorption and chemical binding. The GOD immobilized Pt nanowires were used for application in glucose detection by performing a cyclic voltammetry measurement. The detection results showed that GOD was immobilized on all of the tested surfaces and the highest glucose detection sensitivity of 60 μM was obtained when the Pt nanowires were modified by PVA with PB mediator. Moreover, the sensors showed very high current response when the Pt nanowires were modified with the cysteamine SAM. The stability and catalyst activity of GOD are also reported here. For instance, the catalyst activity of GOD retained about 60% of its initial value after it was stored at 4 °C in a 100 mM PBS buffer solution with a pH of 7.2 for a period of 30 days.

Effect of functionalized and non-functionalized nanodiamond on the morphology and activities of antioxidant enzymes of lung epithelial cells (A549).

Science.gov (United States)

Solarska-Ściuk, Katarzyna; Gajewska, Agnieszka; Glińska, Sława; Michlewska, Sylwia; Balcerzak, Łucja; Jamrozik, Agnieszka; Skolimowski, Janusz; Burda, Květoslava; Bartosz, Grzegorz

2014-10-05

The development of nanotechnology opens up new ways for biomedical applications of unmodified and modified diamond nanoparticles which are one of the most popular nanomaterials used in biology, biotechnology, medicine, cosmetics and engineering. They have been applied as diagnostic and therapeutic agents because they can be targeted to and localized in cells causing apoptosis and necrosis. The problem of biocompatibility of nanodiamonds at higher concentrations is thus of primary importance. The first step in the modification of DNPs is usually the introduction of hydrogen groups, which can bind other functional groups. The basic method to introduce -OH groups onto nanoparticles is the Fenton reaction. The aim of this study was to compare the effect of unmodified nanodiamond particles and nanoparticles modified by introduction of -OH groups and etoposide onto their surface reaction on human non-small lung cancer cells. A549 cells were incubated with 2-100μg/ml nanopowders and at 0.6-24μg/ml etoposide in the DMEM medium. We observed a decrease of cells viability and generation of reactive oxygen/ nitrogen species in the cells after incubation, estimated by oxidation of H2DCF-DA and DAF-FM-DA. Modified detonation nanoparticles affected also the cellular content of glutathione and activities of main antioxidant enzymes (glutathione peroxidase, glutathione reductase, glutathione S-transferase, superoxide dismutase and catalase). The results of TEM microscopy show changes in cell morphology. These data demonstrate that modified nanoparticles induce oxidative stress in the target cells. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Novel determination of cadmium ions using an enzyme self-assembled monolayer with surface plasmon resonance

International Nuclear Information System (INIS)

May May, Lee; Russell, David A.

2003-01-01

The activity of the enzyme urease is known to be inhibited by the heavy metal cadmium. The binding of cadmium to urease and the consequent changes of the enzyme structure are the basis of the surface plasmon resonance (SPR) biosensing system reported herein. To facilitate the formation of a self-assembled monolayer (SAM) of the urease on gold-coated glass SPR sensor disks, the enzyme has been modified with N-succinimidyl 3-(2-pyridyldithiol) propionate (SPDP). The urease monolayer was exposed to trace levels of cadmium ions and monitored by SPR. From circular dichroism (CD) data, it is believed that the conformation of the active nickel site of the urease changes upon binding of the cadmium ions. It is this change of the enzyme monolayer, measured by SPR, which has been related to the cadmium ion concentration in the range of 0-10 mg l -1 . These data are the first report of a SPR biosensor capable of detecting metal ions

Cellobiose Dehydrogenase Aryl Diazonium Modified Single Walled Carbon Nanotubes: Enhanced Direct Electron Transfer through a Positively Charged Surface

Science.gov (United States)

2011-01-01

One of the challenges in the field of biosensors and biofuel cells is to establish a highly efficient electron transfer rate between the active site of redox enzymes and electrodes to fully access the catalytic potential of the biocatalyst and achieve high current densities. We report on very efficient direct electron transfer (DET) between cellobiose dehydrogenase (CDH) from Phanerochaete sordida (PsCDH) and surface modified single walled carbon nanotubes (SWCNT). Sonicated SWCNTs were adsorbed on the top of glassy carbon electrodes and modified with aryl diazonium salts generated in situ from p-aminobenzoic acid and p-phenylenediamine, thus featuring at acidic pH (3.5 and 4.5) negative or positive surface charges. After adsorption of PsCDH, both electrode types showed excellent long-term stability and very efficient DET. The modified electrode presenting p-aminophenyl groups produced a DET current density of 500 μA cm−2 at 200 mV vs normal hydrogen reference electrode (NHE) in a 5 mM lactose solution buffered at pH 3.5. This is the highest reported DET value so far using a CDH modified electrode and comes close to electrodes using mediated electron transfer. Moreover, the onset of the electrocatalytic current for lactose oxidation started at 70 mV vs NHE, a potential which is 50 mV lower compared to when unmodified SWCNTs were used. This effect potentially reduces the interference by oxidizable matrix components in biosensors and increases the open circuit potential in biofuel cells. The stability of the electrode was greatly increased compared with unmodified but cross-linked SWCNTs electrodes and lost only 15% of the initial current after 50 h of constant potential scanning. PMID:21417322

Fibroblast adhesion and activation onto micro-machined titanium surfaces.

Science.gov (United States)

Guillem-Marti, J; Delgado, L; Godoy-Gallardo, M; Pegueroles, M; Herrero, M; Gil, F J

2013-07-01

Surface modifications performed at the neck of dental implants, in the manner of micro-grooved surfaces, can reduce fibrous tissue encapsulation and prevent bacterial colonization, thereby improving fibrointegration and the formation of a biological seal. However, the applied procedures are technically complex and/or time consuming methods. The aim of this study was to analyse the fibroblast behaviour on modified titanium surfaces obtained, applying a simple and low-cost method. An array of titanium surfaces was obtained using a commercial computerized numerical control lathe, modifying the feed rate and the cutting depth. To elucidate the potential ability of the generated surfaces to activate connective tissue cells, a thorough gene (by real time - qPCR) and protein (by western blot or zymography) expression and cellular response characterization (cell morphology, cell adhesion and cell activation by secreting extracellular matrix (ECM) components and their enzyme regulators) was performed. Micro-grooved surfaces have statistically significant differences in the groove's width (approximately 10, 50 and 100 μm) depending on the applied advancing fixed speed. Field emission scanning electron microscopy images showed that fibroblasts oriented along the generated grooves, but they were only entirely accommodated on the wider grooves (≥50 μm). Micro-grooved surfaces exhibited an earlier cell attachment and activation, as seen by collagen Iα1 and fibronectin deposition and activation of ECM remodelling enzymes, compared with the other surfaces. However, fibroblasts could remain in an activated state on narrower surfaces (fibrotic response. © 2012 John Wiley & Sons A/S.

Adhesive and morphological characteristics of surface chemically modified polytetrafluoroethylene films

International Nuclear Information System (INIS)

Hopp, B.; Kresz, N.; Kokavecz, J.; Smausz, T.; Schieferdecker, H.; Doering, A.; Marti, O.; Bor, Z.

2004-01-01

In the present paper, we report an experimental determination of adhesive and topographic characteristics of chemically modified surface of polytetrafluoroethylene (PTFE) films. The surface chemistry was modified by ArF excimer laser irradiation in presence of triethylene-tetramine photoreagent. The applied laser fluence was varied in the range of 0.4-9 mJ/cm 2 , and the number of laser pulses incident on the same area was 1500. To detect the changes in the adhesive features of the treated Teflon samples, we measured receding contact angle for distilled water and adhesion strength, respectively. It was found that the receding contact angle decreased from 96 deg. to 30-37 deg. and the adhesion strength of two-component epoxy glue to the treated sample surface increased from 0.03 to 9 MPa in the applied laser fluence range. Additionally, it was demonstrated that the adhesion of human cells to the modified Teflon samples is far better than to the untreated ones. The contact mode and pulsed force mode atomic force microscopic investigations of the treated samples demonstrated that the measured effective contact area of the irradiated films does not differ significantly from that of the original films, but the derived adhesion force is stronger on the modified samples than on the untreated ones. Hence, the increased adhesion of the treated Teflon films is caused by the higher surface energy

Voltammetric enzyme sensor for urea using mercaptohydroquinone-modified gold electrode as the base transducer.

Science.gov (United States)

Mizutani, F; Yabuki, S; Sato, Y

1997-01-01

A voltammetric urea-sensing electrode was prepared by combining a lipid-attached urease layer with a 2,5-dihydroxythiophenol-modified gold electrode. A self-assembled monolayer of dihydroxythiophenol was prepared on the gold surface by soaking the electrode into an ethanolic solution containing the modifier. A layer of the lipid-attached enzyme and that of acetyl cellulose overcoat were successively made on the dihydroxythiophenol-modified electrode by applying a dip-coating procedure. The addition of urea in a test solution (10 mM phosphate buffer, pH 7.0) brought about an increase of pH near the urease layer. The pH shift accompanied a negative shift of the anodic peak, which corresponded to the electro-oxidation of dihydroxyphenol moiety to form quinone, on the linear sweep voltammograms for the urease/dihydroxythiophenol electrode. The concentration of urea (0.2-5 mM) could be determined by measuring the electrode current at -0.05 V versus Ag/AgCl from the voltammogram. The electrode was applied to the determination of urea in human urine; the measurement of electrode current at such a low potential provided the urea determination without any electrochemical interference from L-ascorbic acid and uric acid.

Enzyme-modified starch as an oil delivery system for bake-only chicken nuggets.

Science.gov (United States)

Purcell, Sarah; Wang, Ya-Jane; Seo, Han-Seok

2014-05-01

This study investigated the effects of enzyme modification on starch as an effective oil delivery system for bake-only chicken nuggets. Various native starches were hydrolyzed by amyloglucosidase to a hydrolysis degree of 20% to 25% and plated with 50% (w/w, starch dry basis) with canola oil to create a starch-oil matrix. This matrix was then blended into a dry ingredient blend for batter and breader components. Nuggets were prepared by coated with predust, hydrated batter, and breader, and the coated nuggets were steam-baked until fully cooked and then frozen until texture and sensory analyses. The enzyme-modified starches showed a significant decrease in pasting viscosities for all starch types. For textural properties of nuggets, no clear relationship was found between peak force and starch source or amylose content. Sensory attributes related to fried foods (for example, crispness and mouth-coating) did not significantly differ between bake-only nuggets formulated using the enzyme-modified starches and the partially fried and baked ones. The present findings suggest that enzyme-modified starches can deliver sufficient quantity of oil to create sensory attributes similar to those of partially fried chicken nuggets. Further study is needed to optimize the coating formulation of bake-only chicken nugget to become close to the fried one in sensory aspects. The food industry has become increasingly focused on healthier items. Frying imparts several critical and desirable product functionalities, such as developing texture and color, and providing mouth-feel and flavor. The food industry has yet to duplicate all of the unique characteristics of fried chicken nuggets with a baking process. This study investigated the application of enzyme-modified starch as an oil delivery system in bake-only chicken nugget formulation in attempts to provide characteristics of fried items. This information is useful to improve the nutritional value of fried food by eliminating the

Fibrinogen Motif Discriminates Platelet and Cell Capture in Peptide-Modified Gold Micropore Arrays.

Science.gov (United States)

Adamson, Kellie; Spain, Elaine; Prendergast, Una; Moran, Niamh; Forster, Robert J; Keyes, Tia E

2018-01-16

Human blood platelets and SK-N-AS neuroblastoma cancer-cell capture at spontaneously adsorbed monolayers of fibrinogen-binding motifs, GRGDS (generic integrin adhesion), HHLGGAKQAGDV (exclusive to platelet integrin α IIb β 3 ), or octanethiol (adhesion inhibitor) at planar gold and ordered 1.6 μm diameter spherical cap gold cavity arrays were compared. In all cases, arginine/glycine/aspartic acid (RGD) promoted capture, whereas alkanethiol monolayers inhibited adhesion. Conversely only platelets adhered to alanine/glycine/aspartic acid (AGD)-modified surfaces, indicating that the AGD motif is recognized preferentially by the platelet-specific integrin, α IIb β 3 . Microstructuring of the surface effectively eliminated nonspecific platelet/cell adsorption and dramatically enhanced capture compared to RGD/AGD-modified planar surfaces. In all cases, adhesion was reversible. Platelets and cells underwent morphological change on capture, the extent of which depended on the topography of the underlying substrate. This work demonstrates that both the nature of the modified interface and its underlying topography influence the capture of cancer cells and platelets. These insights may be useful in developing cell-based cancer diagnostics as well as in identifying strategies for the disruption of platelet cloaks around circulating tumor cells.

Cell age dependent variations in oxidative protective enzymes

International Nuclear Information System (INIS)

Blakely, E.A.; Chang, P.Y.; Lommel, L.; Tobias, C.A.

1986-01-01

Activity levels of antioxidant enzymes were correlated before and after heavy-ion exposures with cellular radiosensitivity. In preliminary feasibility experiments with human T-1 cells relatively high antioxidant enzyme levels were shown in the unirradiated G 1 phase prior to the normal DNA synthetic phase. Endogenous cellular levels of three antioxidant enzymes were measured at various times in the unirradiated human T-1 cell division cycle. The enzymes measured were: catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GSHPX). Unlike the case in Chinese hamster V79 cells the early data with the synchronized human cell show that in very early G 1 phase (e.g., approximately 1.5 hours after mitotic selection) there are significant peaks in the levels (U/mg cell protein) of both CAT and SOD. Both enzymes show increases as the unirradiated cells progressed from mitosis into G 1 phase while the levels of GSHPX measured in duplicate samples were somewhat more variable than was the case for the other two enzymes. Studies were made in collaboration with the Armed Forces Radiobiology Research Institute

Surface grafting of carboxylic groups onto thermoplastic polyurethanes to reduce cell adhesion

Energy Technology Data Exchange (ETDEWEB)

Alves, P., E-mail: palves@eq.uc.pt [CIEPQPF, Departamento de Engenharia Química, Universidade de Coimbra, Polo II, Pinhal de Marrocos, 3030-790 Coimbra (Portugal); Ferreira, P. [CIEPQPF, Departamento de Engenharia Química, Universidade de Coimbra, Polo II, Pinhal de Marrocos, 3030-790 Coimbra (Portugal); Kaiser, Jean-Pierre [EMPA, St. Gallen, Lerchenfeldstrasse 5, CH-9014 St. Gallen (Switzerland); Salk, Natalie [Mikrofertigung – Micro Engineering, Fraunhofer IFAM, Wiener Strasse 12, D-288359 Bremen (Germany); Bruinink, Arie [EMPA, St. Gallen, Lerchenfeldstrasse 5, CH-9014 St. Gallen (Switzerland); Sousa, Hermínio C. de; Gil, M.H. [CIEPQPF, Departamento de Engenharia Química, Universidade de Coimbra, Polo II, Pinhal de Marrocos, 3030-790 Coimbra (Portugal)

2013-10-15

The interaction of polymers with other materials is an important issue, being their surface properties clearly crucial. For some important polymer applications, their surfaces have to be modified. Surface modification aims to tailor the surface characteristics of a material for a specific application without affecting its bulk properties. Materials can be surface modified by using biological, chemical or physical methods. The aim of this work was to improve the reactivity of the thermoplastic polyurethane (TPU) material (Elastollan{sup ®}) surface and to make its surface cell repellent by grafting carboxylic groups onto its surface. Two TPU materials were studied: a polyether-based TPU and a polyester-based TPU. The grafting efficiency was evaluated by contact angle measurements and by analytical determination of the COOH groups. Scanning electron microscopy (SEM) of the membranes surface was performed as well as cell adhesion tests. It was proved that the surfaces of the TPUs membranes were successfully modified and that cell adhesion was remarkably reduced.

A Disposable Organophosphorus Pesticides Enzyme Biosensor Based on Magnetic Composite Nano-Particles Modified Screen Printed Carbon Electrode

Directory of Open Access Journals (Sweden)

Weigang Wen

2010-01-01

Full Text Available A disposable organophosphorus pesticides (OPs enzyme biosensor based on magnetic composite nanoparticle-modified screen printed carbon electrodes (SPCE has been developed. Firstly, an acetylcholinesterase (AChE-coated Fe3O4/Au (GMP magnetic nanoparticulate (GMP-AChE was synthesized. Then, GMP-AChE was absorbed on the surface of a SPCE modified by carbon nanotubes (CNTs/nano-ZrO2/prussian blue (PB/Nafion (Nf composite membrane by an external magnetic field. Thus, the biosensor (SPCE|CNTs/ZrO2/PB/Nf|GMP-AChE for OPs was fabricated. The surface of the biosensor was characterized by scanning electron micrography (SEM and X-ray fluorescence spectrometery (XRFS and its electrochemical properties were studied by cyclic voltammetry (CV and differential pulse voltammetry (DPV. The degree of inhibition (A% of the AChE by OPs was determined by measuring the reduction current of the PB generated by the AChE-catalyzed hydrolysis of acetylthiocholine (ATCh. In pH = 7.5 KNO3 solution, the A was related linearly to the concentration of dimethoate in the range from 1.0 × 10-3–10 ng•mL-1 with a detection limit of 5.6 × 10-4 ng•mL-1. The recovery rates in Chinese cabbage exhibited a range of 88%–105%. The results were consistent with the standard gas chromatography (GC method. Compared with other enzyme biosensors the proposed biosensor exhibited high sensitivity, good selectivity with disposable, low consumption of sample. In particular its surface can be easily renewed by removal of the magnet. The convenient, fast and sensitive voltammetric measurement opens new opportunities for OPs analysis.

Modification of surface/neuron interfaces for neural cell-type specific responses: a review

International Nuclear Information System (INIS)

Chen, Cen; Kong, Xiangdong; Lee, In-Seop

2016-01-01

Surface/neuron interfaces have played an important role in neural repair including neural prostheses and tissue engineered scaffolds. This comprehensive literature review covers recent studies on the modification of surface/neuron interfaces. These interfaces are identified in cases both where the surfaces of substrates or scaffolds were in direct contact with cells and where the surfaces were modified to facilitate cell adhesion and controlling cell-type specific responses. Different sources of cells for neural repair are described, such as pheochromocytoma neuronal-like cell, neural stem cell (NSC), embryonic stem cell (ESC), mesenchymal stem cell (MSC) and induced pluripotent stem cell (iPS). Commonly modified methods are discussed including patterned surfaces at micro- or nano-scale, surface modification with conducting coatings, and functionalized surfaces with immobilized bioactive molecules. These approaches to control cell-type specific responses have enormous potential implications in neural repair. (paper)

Impacts of papain and neuraminidase enzyme treatment on electrohydrodynamics and IgG-mediated agglutination of type A red blood cells.

Science.gov (United States)

Hyono, Atsushi; Gaboriaud, Fabien; Mazda, Toshio; Takata, Youichi; Ohshima, Hiroyuki; Duval, Jérôme F L

2009-09-15

The stability of native and enzyme-treated human red blood cells of type A (Rh D positive) against agglutination is investigated under conditions where it is mediated by immunoglobuline G (IgG) anti-D antibody binding. The propensity of cells to agglutinate is related to their interphasic (electrokinetic) properties. These properties significantly depend on the concentration of proteolytic papain enzyme and protease-free neuraminidase enzyme that the cells are exposed to. The analysis is based on the interpretation of electrophoretic data of cells by means of the numerical theory for the electrokinetics of soft (bio)particles. A significant reduction of the hydrodynamic permeability of the external soft glycoprotein layer of the cells is reported under the action of papain. This reflects a significant decrease in soft surface layer thickness and a loss in cell surface integrity/rigidity, as confirmed by nanomechanical AFM analysis. Neuraminidase action leads to an important decrease in the interphase charge density by removing sialic acids from the cell soft surface layer. This is accompanied by hydrodynamic softness modulations less significant than those observed for papain-treated cells. On the basis of these electrohydrodynamic characteristics, the overall interaction potential profiles between two native cells and two enzyme-treated cells are derived as a function of the soft surface layer thickness in the Debye-Hückel limit that is valid for cell suspensions under physiological conditions (approximately 0.16 M). The thermodynamic computation of cell suspension stability against IgG-mediated agglutination then reveals that a decrease in the cell surface layer thickness is more favorable than a decrease in interphase charge density for inducing agglutination. This is experimentally confirmed by agglutination data collected for papain- and neuraminidase-treated cells.

Influence of laser surface modifying of polyethylene terephthalate on fibroblast cell adhesion

International Nuclear Information System (INIS)

Mirzadeh, H.; Dadsetan, M.

2003-01-01

Attempts have been made to evaluate the changes in physical and chemical properties of the polyethylene terephthalate (PET) surface due to laser irradiation. These changes have been investigated from viewpoints of microstructuring and its effect on fibroblast cell behavior. The surfaces of PET were irradiated using CO 2 and KrF excimer pulsed laser. The changes were characterized by attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy, scanning electron microscopy (SEM) and contact angle measurements. The data from ATR-FTIR spectra showed that the crystallinity in the surface region decreased due to the CO 2 and excimer laser irradiation. SEM observations showed that specific microstructures were created on the PET surface due to laser irradiation. In order to study biocompatibility and cell behavior, we utilized standard in vitro L929-fibroblast cell culture system. Fibroblast cell adhesion and spreading were significantly correlated to the morphology and wettability of the laser irradiated PET surface

Surface chemical functionalities affect the behavior of human adipose-derived stem cells in vitro

International Nuclear Information System (INIS)

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

2013-01-01

This study examines the effect of surface chemical functionalities on the behavior of human adipose-derived stem cells (hASCs) in vitro. Plasma polymerized films rich in amine (-NH 2 ), carboxyl (-COOH) and methyl (-CH 3 ), were generated on hydroxyapatite (HAp) substrates. The surface chemical functionalities were characterized by X-ray photoelectron spectroscopy (XPS). The ability of different substrates to absorb proteins was evaluated. The results showed that substrates modified with hydrophilic functional group (-COOH and -NH 2 ) can absorb more proteins than these modified with more hydrophobic functional group (-CH 3 ). The behavior of human adipose-derived stem cells (hASCs) cultured on different substrates was investigated in vitro: cell counting kit-8 (CCK-8) analysis was used to characterize cell proliferation, scanning electronic microscopy (SEM) analysis was used to characterize cell morphology and alkaline phosphatase (ALP) activity analysis was used to account for differentiation. The results of this study demonstrated that the -NH 2 modified surfaces encourage osteogenic differentiation; the -COOH modified surfaces promote cell adhesion and spreading and the -CH 3 modified surfaces have the lowest ability to induce osteogenic differentiation. These findings confirmed that the surface chemical states of biomaterials can affect the behavior of hASCs in vitro.

Surface chemical functionalities affect the behavior of human adipose-derived stem cells in vitro

Energy Technology Data Exchange (ETDEWEB)

Liu, Xujie [State key laboratory of new ceramics and fine processing, School of Materials Science and Engineering, Tsinghua University, Beijing 100084 (China); Feng, Qingling, E-mail: biomater@mail.tsinghua.edu.cn [State key laboratory of new ceramics and fine processing, School of Materials Science and Engineering, Tsinghua University, Beijing 100084 (China); Bachhuka, Akash [Mawson Institute, University of South Australia, Mawson Lakes 5095 (Australia); Vasilev, Krasimir [Mawson Institute, University of South Australia, Mawson Lakes 5095 (Australia); School of Advanced Manufacturing, University of South Australia, Mawson Lakes 5095 (Australia)

2013-04-01

This study examines the effect of surface chemical functionalities on the behavior of human adipose-derived stem cells (hASCs) in vitro. Plasma polymerized films rich in amine (-NH{sub 2}), carboxyl (-COOH) and methyl (-CH{sub 3}), were generated on hydroxyapatite (HAp) substrates. The surface chemical functionalities were characterized by X-ray photoelectron spectroscopy (XPS). The ability of different substrates to absorb proteins was evaluated. The results showed that substrates modified with hydrophilic functional group (-COOH and -NH{sub 2}) can absorb more proteins than these modified with more hydrophobic functional group (-CH{sub 3}). The behavior of human adipose-derived stem cells (hASCs) cultured on different substrates was investigated in vitro: cell counting kit-8 (CCK-8) analysis was used to characterize cell proliferation, scanning electronic microscopy (SEM) analysis was used to characterize cell morphology and alkaline phosphatase (ALP) activity analysis was used to account for differentiation. The results of this study demonstrated that the -NH{sub 2} modified surfaces encourage osteogenic differentiation; the -COOH modified surfaces promote cell adhesion and spreading and the -CH{sub 3} modified surfaces have the lowest ability to induce osteogenic differentiation. These findings confirmed that the surface chemical states of biomaterials can affect the behavior of hASCs in vitro.

Immobilization of microbial cells on cellulose-polymer surfaces by radiation polymerization

International Nuclear Information System (INIS)

Kumakura, M.; Kaetsu, I.

1983-01-01

Streptomyces phaeochromogens cells were immobilized on cellulose-polymer surfaces by radiation polymerization using hydrophilic monomers and paper. The enzyme activity of immobilized cell sheets was higher than that of immobilized cell composites obtained by the usual radiation polymerization technique. The enzyme activity of the sheets was affected by monomer concentration, the thickness of paper, and the degree of polymerization of paper. The copolymerization of hydroxyethyl methacrylate and methoxytetraethyleneglycol methacrylate in the sheets led to a further increase of the enzyme activity due to the increase of the hydrophilicity of the polymer matrix. The Michaelis constant of the sheets from low monomer concentration was close to that of intact cells

Glycosylation Helps Cellulase Enzymes Bind to Plant Cell Walls (Fact Sheet)

Energy Technology Data Exchange (ETDEWEB)

2012-06-01

Computer simulations suggest a new strategy to design enhanced enzymes for biofuels production. Large-scale computer simulations predict that the addition of glycosylation on carbohydrate-binding modules can dramatically improve the binding affinity of these protein domains over amino acid mutations alone. These simulations suggest that glycosylation can be used as a protein engineering tool to enhance the activity of cellulase enzymes, which are a key component in the conversion of cellulose to soluble sugars in the production of biofuels. Glycosylation is the covalent attachment of carbohydrate molecules to protein side chains, and is present in many proteins across all kingdoms of life. Moreover, glycosylation is known to serve a wide variety of functions in biological recognition, cell signaling, and metabolism. Cellulase enzymes, which are responsible for deconstructing cellulose found in plant cell walls to glucose, contain glycosylation that when modified can affect enzymatic activity-often in an unpredictable manner. To gain insight into the role of glycosylation on cellulase activity, scientists at the National Renewable Energy Laboratory (NREL) used computer simulation to predict that adding glycosylation on the carbohydrate-binding module of a cellulase enzyme dramatically boosts the binding affinity to cellulose-more than standard protein engineering approaches in which amino acids are mutated. Because it is known that higher binding affinity in cellulases leads to higher activity, this work suggests a new route to designing enhanced enzymes for biofuels production. More generally, this work suggests that tuning glycosylation in cellulase enzymes is a key factor to consider when engineering biochemical conversion processes, and that more work is needed to understand how glycosylation affects cellulase activity at the molecular level.

Effects of synthetic cohesin-containing scaffold protein architecture on binding dockerin-enzyme fusions on the surface of Lactococcus lactis

Directory of Open Access Journals (Sweden)

Wieczorek Andrew S

2012-12-01

Full Text Available Abstract Background The microbial synthesis of fuels, commodity chemicals, and bioactive compounds necessitates the assemblage of multiple enzyme activities to carry out sequential chemical reactions, often via substrate channeling by means of multi-domain or multi-enzyme complexes. Engineering the controlled incorporation of enzymes in recombinant protein complexes is therefore of interest. The cellulosome of Clostridium thermocellum is an extracellular enzyme complex that efficiently hydrolyzes crystalline cellulose. Enzymes interact with protein scaffolds via type 1 dockerin/cohesin interactions, while scaffolds in turn bind surface anchor proteins by means of type 2 dockerin/cohesin interactions, which demonstrate a different binding specificity than their type 1 counterparts. Recombinant chimeric scaffold proteins containing cohesins of different specificity allow binding of multiple enzymes to specific sites within an engineered complex. Results We report the successful display of engineered chimeric scaffold proteins containing both type 1 and type 2 cohesins on the surface of Lactococcus lactis cells. The chimeric scaffold proteins were able to form complexes with the Escherichia coli β-glucuronidase fused to either type 1 or type 2 dockerin, and differences in binding efficiencies were correlated with scaffold architecture. We used E. coli β-galactosidase, also fused to type 1 or type 2 dockerins, to demonstrate the targeted incorporation of two enzymes into the complexes. The simultaneous binding of enzyme pairs each containing a different dockerin resulted in bi-enzymatic complexes tethered to the cell surface. The sequential binding of the two enzymes yielded insights into parameters affecting assembly of the complex such as protein size and position within the scaffold. Conclusions The spatial organization of enzymes into complexes is an important strategy for increasing the efficiency of biochemical pathways. In this study

A radiolabel-release microwell assay for proteolytic enzymes present in cell culture media

International Nuclear Information System (INIS)

Rucklidge, G.J.; Milne, G.

1990-01-01

A modified method for the measurement of proteolytic enzyme activity in cell culture-conditioned media has been developed. Using the release of 3H-labeled peptides from 3H-labeled gelatin the method is performed in microwell plates. The substrate is insolubilized and attached to the wells by glutaraldehyde treatment, thus eliminating the need for a precipitation step at the end of the assay. The assay is sensitive, reproducible, and convenient for small sample volumes. The effect of different protease inhibitors on activity can be assessed rapidly allowing an early characterization of the enzyme. It can also be adapted to microplate spectrophotometric analysis by staining residual substrate with Coomassie blue

Sterol glycosyltransferases--the enzymes that modify sterols.

Science.gov (United States)

Chaturvedi, Pankaj; Misra, Pratibha; Tuli, Rakesh

2011-09-01

Sterols are important components of cell membranes, hormones, signalling molecules and defense-related biotic and abiotic chemicals. Sterol glycosyltransferases (SGTs) are enzymes involved in sterol modifications and play an important role in metabolic plasticity during adaptive responses. The enzymes are classified as a subset of family 1 glycosyltransferases due to the presence of a signature motif in their primary sequence. These enzymes follow a compulsory order sequential mechanism forming a ternary complex. The diverse applications of sterol glycosides, like cytotoxic and apoptotic activity, anticancer activity, medicinal values, anti-stress roles and anti-insect and antibacterial properties, draws attention towards their synthesis mechanisms. Many secondary metabolites are derived from sterol pathways, which are important in defense mechanisms against pathogens. SGTs in plants are involved in changed sensitivity to stress hormones and their agrochemical analogs and changed tolerance to biotic and abiotic stresses. SGTs that glycosylate steroidal hormones, such as brassinosteroids, function as growth and development regulators in plants. In terms of metabolic roles, it can be said that SGTs occupy important position in plant metabolism and may offer future tools for crop improvement.

Cell Adhesion and Proliferation on Sulfonated and Non-Modified Chitosan Films.

Science.gov (United States)

Martínez-Campos, Enrique; Civantos, Ana; Redondo, Juan Alfonso; Guzmán, Rodrigo; Pérez-Perrino, Mónica; Gallardo, Alberto; Ramos, Viviana; Aranaz, Inmaculada

2017-05-01

Three types of chitosan-based films have been prepared and evaluated: a non-modified chitosan film bearing cationizable aliphatic amines and two films made of N-sulfopropyl chitosan derivatives bearing both aliphatic amines and negative sulfonate groups at different ratios. Cell adhesion and proliferation on chitosan films of C2C12 pre-myoblastic cells and B16 cells as tumoral model have been tested. A differential cell behavior has been observed on chitosan films due to their different surface modification. B16 cells have shown lower vinculin expression when cultured on sulfonated chitosan films. This study shows how the interaction among cells and material surface can be modulated by physicochemical characteristics of the biomaterial surface, altering tumoral cell adhesion and proliferation processes.

Comparative of fibroblast and osteoblast cells adhesion on surface modified nanofibrous substrates based on polycaprolactone.

Science.gov (United States)

Sharifi, Fereshteh; Irani, Shiva; Zandi, Mojgan; Soleimani, Masoud; Atyabi, Seyed Mohammad

2016-12-01

One of the determinant factors for successful bioengineering is to achieve appropriate nano-topography and three-dimensional substrate. In this research, polycaprolactone (PCL) nano-fibrous mat with different roughness modified with O 2 plasma was fabricated via electrospinning. The purpose of this study was to evaluate the effect of plasma modification along with surface nano-topography of mats on the quality of human fibroblast (HDFs) and osteoblast cells (OSTs)-substrate interaction. Surface properties were studied using scanning electron microscopy (SEM), atomic force microscopy (AFM), contact angle, Fourier-transformation infrared spectroscopy. We evaluated mechanical properties of fabricated mats by tensile test. The viability and proliferation of HDFs and OSTs on the substrates were followed by 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyltetrazolium bromide (MTT). Mineralization of the substrate was determined by alizarin red staining method and calcium content of OSTs was determined by calcium content kit. Cells morphology was studied by SEM analysis. The results revealed that the plasma-treated electrospun nano-fibrous substrate with higher roughness was an excellent designed substrate. A bioactive topography for stimulating proliferation of HDFs and OSTs is to accelerate the latter's differentiation time. Therefore, the PCL substrate with high density and major nano-topography were considered as a bio-functional and elegant bio-substrate for tissue regeneration applications.

Display of a β-mannanase and a chitosanase on the cell surface of Lactobacillus plantarum towards the development of whole-cell biocatalysts.

Science.gov (United States)

Nguyen, Hoang-Minh; Mathiesen, Geir; Stelzer, Elena Maria; Pham, Mai Lan; Kuczkowska, Katarzyna; Mackenzie, Alasdair; Agger, Jane W; Eijsink, Vincent G H; Yamabhai, Montarop; Peterbauer, Clemens K; Haltrich, Dietmar; Nguyen, Thu-Ha

2016-10-04

Lactobacillus plantarum is considered as a potential cell factory because of its GRAS (generally recognized as safe) status and long history of use in food applications. Its possible applications include in situ delivery of proteins to a host, based on its ability to persist at mucosal surfaces of the human intestine, and the production of food-related enzymes. By displaying different enzymes on the surface of L. plantarum cells these could be used as whole-cell biocatalysts for the production of oligosaccharides. In this present study, we aimed to express and display a mannanase and a chitosanase on the cell surface of L. plantarum. ManB, a mannanase from Bacillus licheniformis DSM13, and CsnA, a chitosanase from Bacillus subtilis ATCC 23857 were fused to different anchoring motifs of L. plantarum for covalent attachment to the cell surface, either via an N-terminal lipoprotein anchor (Lp_1261) or a C-terminal cell wall anchor (Lp_2578), and the resulting fusion proteins were expressed in L. plantarum WCFS1. The localization of the recombinant proteins on the bacterial cell surface was confirmed by flow cytometry and immunofluorescence microscopy. The highest mannanase and chitosanase activities obtained for displaying L. plantarum cells were 890 U and 1360 U g dry cell weight, respectively. In reactions with chitosan and galactomannans, L. plantarum CsnA- and ManB-displaying cells produced chito- and manno-oligosaccharides, respectively, as analyzed by high performance anion exchange chromatography (HPAEC) and mass spectrometry (MS). Surface-displayed ManB is able to break down galactomannan (LBG) into smaller manno-oligosaccharides, which can support growth of L. plantarum. This study shows that mannanolytic and chitinolytic enzymes can be anchored to the cell surface of L. plantarum in active forms. L. plantarum chitosanase- and mannanase-displaying cells should be of interest for the production of potentially 'prebiotic' oligosaccharides. This approach

The influence of the surface chemistry of silver nanoparticles on cell death

International Nuclear Information System (INIS)

Sur, Ilknur; Altunbek, Mine; Kahraman, Mehmet; Culha, Mustafa

2012-01-01

The influence of the surface chemistry of silver nanoparticles (AgNPs) on p53 mediated cell death was evaluated using human dermal fibroblast (HDF) and lung cancer (A549) cells. The citrate reduced AgNPs (C-AgNPs) were modified with either lactose (L-AgNPs) or a 12-base long oligonucleotide (O-AgNPs). Both unmodified and modified AgNPs showed increased concentration and time dependent cytotoxicity and genotoxicity causing an increased p53 up-regulation within 6 h and led to apoptotic or necrotic cell deaths. The C-AgNPs induced more cytotoxicity and cellular DNA damage than the surface modified AgNPs. Modifying the C-AgNPs with lactose or the oligonucleotide reduced both necrotic and apoptotic cell deaths in the HDF cells. The C-AgNPs caused an insignificant necrosis in A549 cells whereas the modified AgNPs caused necrosis and apoptosis in both cell types. Compared to the O-AgNPs, the L-AgNPs triggered more cellular DNA damage, which led to up-regulation of p53 gene inducing apoptosis in A549 cells compared to HDF cells. This suggests that the different surface chemistries of the AgNPs cause different cellular responses that may be important not only for their use in medicine but also for reducing their toxicity. (paper)

Fabrication of tunable microreactor with enzyme modified magnetic nanoparticles for microfluidic electrochemical detection of glucose

Energy Technology Data Exchange (ETDEWEB)

Sheng Jin; Zhang Lei; Lei Jianping [State Key Laboratory of Analytical Chemistry for Life Science, Department of Chemistry, Nanjing University, Nanjing 210093 (China); Ju Huangxian, E-mail: hxju@nju.edu.cn [State Key Laboratory of Analytical Chemistry for Life Science, Department of Chemistry, Nanjing University, Nanjing 210093 (China)

2012-01-04

Highlights: Black-Right-Pointing-Pointer An enzyme microreactor is prepared using an enzyme-nanoparticles packed microchannel. Black-Right-Pointing-Pointer The optimal performance can be obtained by the tunable length of the microreactor. Black-Right-Pointing-Pointer Baseline separation from interferents can be achieved with a microfluidic device. Black-Right-Pointing-Pointer A pretreatment-free determination method for glucose is proposed. - Abstract: A microfluidic device was designed for amperometric determination of glucose by packing enzyme modified magnetic nanoparticles (MNPs) in its microchannel as an enzyme microreactor. Glucose oxidase was covalently attached to the surface of MNPs and localized in the microchannel by the help of an external magnetic field, leading to a tunable packing length. By changing the length of microreactor from 3 to 10 mm, the performance for glucose detection was optimized. The optimal linear range to glucose was from 25 {mu}M to 15 mM with a detection limit of 11 {mu}M at a length of 6 mm. The inter- and intra-day precisions for determination of 1.0 mM glucose were 0.8% and 1.7%, respectively, and the device-to-device reproducibility was 95.6%. The enzyme reactor remained its 81% activity after three-week storage. Due to the advantages of the device and fracture sampling technique, serum samples could be directly sampled through the fracture to achieve baseline separation from ascorbic acid, and proteins in the samples did not interfere with the detection. This work provided a promising way for pretreatment-free determination of glucose with low cost and excellent performance.

Fabrication of tunable microreactor with enzyme modified magnetic nanoparticles for microfluidic electrochemical detection of glucose

International Nuclear Information System (INIS)

Sheng Jin; Zhang Lei; Lei Jianping; Ju Huangxian

2012-01-01

Highlights: ► An enzyme microreactor is prepared using an enzyme-nanoparticles packed microchannel. ► The optimal performance can be obtained by the tunable length of the microreactor. ► Baseline separation from interferents can be achieved with a microfluidic device. ► A pretreatment-free determination method for glucose is proposed. - Abstract: A microfluidic device was designed for amperometric determination of glucose by packing enzyme modified magnetic nanoparticles (MNPs) in its microchannel as an enzyme microreactor. Glucose oxidase was covalently attached to the surface of MNPs and localized in the microchannel by the help of an external magnetic field, leading to a tunable packing length. By changing the length of microreactor from 3 to 10 mm, the performance for glucose detection was optimized. The optimal linear range to glucose was from 25 μM to 15 mM with a detection limit of 11 μM at a length of 6 mm. The inter- and intra-day precisions for determination of 1.0 mM glucose were 0.8% and 1.7%, respectively, and the device-to-device reproducibility was 95.6%. The enzyme reactor remained its 81% activity after three-week storage. Due to the advantages of the device and fracture sampling technique, serum samples could be directly sampled through the fracture to achieve baseline separation from ascorbic acid, and proteins in the samples did not interfere with the detection. This work provided a promising way for pretreatment-free determination of glucose with low cost and excellent performance.

[Treatment of burn surfaces by proteinases: mathematical description of an enzyme distribution].

Science.gov (United States)

Khalili, A S; Domogatskiĭ, S P; Blizniukov, O P; Ruuge, E K

2003-01-01

The process of penetration of a proteolytic enzyme applied to the surface of burn wound into the depth of necrotic tissue was considered. The model approximation describes three factors by a series of mathematical equations: inward-directed enzyme diffusion, counter-flow filtration of interstitial fluid (exudates), and irreversible inactivation of the enzyme by specific inhibitors present in exudates. According to the model, a quasi-stationary distribution of enzymatic activity through the thickness of the necrotic layer is achieved within 3 h and persists as long as the enzyme concentration on the wound surface is constant. The enzyme activity diminishes linearly from the wound surface to the mid-part of the necrotic layer. No enzyme activity is retained in the inner mid-part of the necrotic layer completely protected by the prevalent inhibitor. The ratio of enzyme concentration on the wound surface to inhibitor concentration in the interstitial fluid is the same as the ratio of the depth of active enzyme area to the depth of the inhibitor-protected area through the necrotic layer. The dynamics of accumulation of the active enzyme in the necrotic zone and the rate of enzyme inactivation in the wound by inhibitors were described by formulas applicable for practical purposes.

A Strontium-Modified Titanium Surface Produced by a New Method and Its Biocompatibility In Vitro.

Science.gov (United States)

Liu, Chundong; Zhang, Yanli; Wang, Lichao; Zhang, Xinhua; Chen, Qiuyue; Wu, Buling

2015-01-01

To present a new and effective method of producing titanium surfaces modified with strontium and to investigate the surface characteristics and in vitro biocompatibility of titanium (Ti) surfaces modified with strontium (Sr) for bone implant applications. Sr-modified Ti surfaces were produced by sequential treatments with NaOH, strontium acetate, heat and water. The surface characteristics and the concentration of the Sr ions released from the samples were examined. Cell adhesion, morphology and growth were investigated using osteoblasts isolated from the calvaria of neonatal Sprague-Dawley rats. Expression of osteogenesis-related genes and proteins was examined to assess the effect of the Sr-modified Ti surfaces on osteoblasts. The modified titanium surface had a mesh structure with significantly greater porosity, and approximately5.37±0.35at.% of Sr was incorporated into the surface. The hydrophilicity was enhanced by the incorporation of Sr ions and water treatment. The average amounts of Sr released from the Sr-modified plates subjected to water treatment were slight higher than the plates without water treatment. Sr promoted cellular adhesion, spreading and growth compared with untreated Ti surfaces. The Sr-modified Ti plates also promoted expression of osteogenesis-related genes,and expression of OPN and COL-І by osteoblasts. Ti plates heat treated at 700°C showed increased bioactivity in comparison with those treated at 600°C. Water treatment upregulated the expression of osteogenesis-related genes. These results show that Sr-modification of Ti surfaces may improve bioactivity in vitro. Water treatment has enhanced the response of osteoblasts. The Sr-modified Ti heat-treated at 700°C exhibited better bioactivity compared with that heated at 600°C.

Improving the cell affinity of a poly(D,L-lactide) film modified by grafting collagen via a plasma technique

International Nuclear Information System (INIS)

Zhao Jianhao; Wang Jue; Tu Mei; Luo Binghong; Zhou Changren

2006-01-01

Poly(D,L-lactide) films were surface-modified by grafting collagen via NH 3 plasma to improve cell affinity. The modified films were characterized by IR analysis, contact angle measurement, SEM analysis and collagen quantity determination. It was demonstrated that -NH 2 and collagen were incorporated into the surface of PDLLA films. The hydrophilicity of the PDLLA film increased after NH 3 plasma treatment, but decreased with further collagen modification. More collagen was incorporated into the PDLLA films by a grating method as compared to that with an anchorage treatment. L929 fibroblast cells were used to evaluate the cell affinity of the modified films and control. It was shown that PDLLA films surface-modified by grafting collagen via NH 3 plasma more efficiently enhanced the cells attachment and proliferation than those films modified by collagen anchorage or only NH 3 plasma treatment

Enhanced osteoblast responses to poly ether ether ketone surface modified by water plasma immersion ion implantation.

Science.gov (United States)

Wang, Heying; Lu, Tao; Meng, Fanhao; Zhu, Hongqin; Liu, Xuanyong

2014-05-01

Poly ether ether ketone (PEEK) offers a set of characteristics superior for human implants; however, its application is limited by the bio-inert surface property. In this work, PEEK surface was modified using single step plasma immersion ion implantation (PIII) treatment with a gas mixture of water vapor as a plasma resource and argon as an ionization assistant. Field emission scanning electron microscopy, atomic force microscopy and X-ray photoelectron spectroscopy were used to investigate the microstructure and composition of the modified PEEK surface. The water contact angle and zeta-potential of the surfaces were also measured. Osteoblast precursor cells MC3T3-E1 and rat bone mesenchymal stem cells were cultured on the PEEK samples to evaluate their cytocompatibility. The obtained results show that the hydroxyl groups as well as a "ravined structure" are constructed on water PIII modified PEEK. Compared with pristine PEEK, the water PIII treated PEEK is more favorable for osteoblast adhesion, spreading and proliferation, besides, early osteogenic differentiation indicated by the alkaline phosphatase activity is also up-regulated. Our study illustrates enhanced osteoblast responses to the PEEK surface modified by water PIII, which gives positive information in terms of future biomedical applications. Copyright © 2014 Elsevier B.V. All rights reserved.

Reversible logic gates based on enzyme-biocatalyzed reactions and realized in flow cells: a modular approach.

Science.gov (United States)

Fratto, Brian E; Katz, Evgeny

2015-05-18

Reversible logic gates, such as the double Feynman gate, Toffoli gate and Peres gate, with 3-input/3-output channels are realized using reactions biocatalyzed with enzymes and performed in flow systems. The flow devices are constructed using a modular approach, where each flow cell is modified with one enzyme that biocatalyzes one chemical reaction. The multi-step processes mimicking the reversible logic gates are organized by combining the biocatalytic cells in different networks. This work emphasizes logical but not physical reversibility of the constructed systems. Their advantages and disadvantages are discussed and potential use in biosensing systems, rather than in computing devices, is suggested. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Immobilization of Gibberella fujikuroi cells with carriers modified by radiation polymerization

International Nuclear Information System (INIS)

Lu Zhaoxin; Xie Zhongchuan; Wei Qijiang

1994-01-01

Gibberella fujikuroi cells were immobilized on modified carriers (gauze) by using the radiation polymerization technique. The mycelium was firmly adhered to the surface of fibril covered with hydrophobic polymer, poly (diethylene glycol dimethyl acrylate) and hydrophobic-hydrophilic copolymer poly (diethylene glycol dimethyl acrylate-sodium acrylate), but it was not immobilized onto the polyethylene net, which has a similar network structure to that of the modified carrier. The weight of immobilized cells was affected by covered polymer. Gibberellic acid productivity in immobilized cells was similar to that of free cells, and the immobilized cells was of good stability. A optimum culture condition for gibberellic acid production was at pH 5.5 and under 20 ∼ 30 degree C

Effect of plasma surface functionalization on preosteoblast cells spreading and adhesion on a biomimetic hydroxyapatite layer formed on a titanium surface

International Nuclear Information System (INIS)

Myung, Sung Woon; Ko, Yeong Mu; Kim, Byung Hoon

2013-01-01

This study examined the plasma surface modification of biomimetic hydroxyapatite (HAp) formed on a titanium (Ti) surface as well as its influence on the behavior of preosteoblast cells. Ti substrates pre-treated with a plasma-polymerized thin film rich in carboxyl groups were subjected to a biomimetic process in a simulated body fluid solution to synthesize the HAp. The HAp layer grown on Ti substrate was then coated with two types of plasma polymerized acrylic acid and allyl amine thin film. The different types of Ti substrates were characterized by attenuated total reflection Fourier transform infrared spectroscopy, energy dispersive spectroscopy and X-ray diffraction. HAp with a Ca/P ratio from 1.25 to 1.38 was obtained on the Ti substrate and hydrophilic carboxyl (-COOH) and amine (-NH 2 ) functional groups were introduced to its surface. Scanning electron microscopy was used to observe the surface of the HAp coatings and the morphology of MC3T3-E1 cells. These results showed that the -COOH-modified HAp surfaces promoted the cell spreading synergistically by changing the surface morphology and chemical state.-NH 2 modified HAp had the lowest cell spreading and proliferation compared to HAp and -COOH-modified HAp. These results correspond to fluorescein analysis, which showed many more cell spreading of COOH/HAp/Ti surface compared to HAp and NH 2 modified HAp. A MTT assay was used to evaluate cell proliferation. The results showed that the proliferation of MC3T3-E1 cells increased in the order of COOH/HAp/Ti > HAp/Ti > NH 2 /Ti > Ti, corresponding to the effect of cell spreading for 6 days. The change in morphology and the chemical surface properties of the biomaterial via plasma polymerization can affect the behavior of MC3T3-E1 cells.

Effect of plasma surface functionalization on preosteoblast cells spreading and adhesion on a biomimetic hydroxyapatite layer formed on a titanium surface

Energy Technology Data Exchange (ETDEWEB)

Myung, Sung Woon; Ko, Yeong Mu; Kim, Byung Hoon, E-mail: kim5055@chosun.ac.kr

2013-12-15

This study examined the plasma surface modification of biomimetic hydroxyapatite (HAp) formed on a titanium (Ti) surface as well as its influence on the behavior of preosteoblast cells. Ti substrates pre-treated with a plasma-polymerized thin film rich in carboxyl groups were subjected to a biomimetic process in a simulated body fluid solution to synthesize the HAp. The HAp layer grown on Ti substrate was then coated with two types of plasma polymerized acrylic acid and allyl amine thin film. The different types of Ti substrates were characterized by attenuated total reflection Fourier transform infrared spectroscopy, energy dispersive spectroscopy and X-ray diffraction. HAp with a Ca/P ratio from 1.25 to 1.38 was obtained on the Ti substrate and hydrophilic carboxyl (-COOH) and amine (-NH{sub 2}) functional groups were introduced to its surface. Scanning electron microscopy was used to observe the surface of the HAp coatings and the morphology of MC3T3-E1 cells. These results showed that the -COOH-modified HAp surfaces promoted the cell spreading synergistically by changing the surface morphology and chemical state.-NH{sub 2} modified HAp had the lowest cell spreading and proliferation compared to HAp and -COOH-modified HAp. These results correspond to fluorescein analysis, which showed many more cell spreading of COOH/HAp/Ti surface compared to HAp and NH{sub 2} modified HAp. A MTT assay was used to evaluate cell proliferation. The results showed that the proliferation of MC3T3-E1 cells increased in the order of COOH/HAp/Ti > HAp/Ti > NH{sub 2}/Ti > Ti, corresponding to the effect of cell spreading for 6 days. The change in morphology and the chemical surface properties of the biomaterial via plasma polymerization can affect the behavior of MC3T3-E1 cells.

Effect of Nanosheet Surface Structure of Titanium Alloys on Cell Differentiation

Directory of Open Access Journals (Sweden)

Satoshi Komasa

2014-01-01

Full Text Available Titanium alloys are the most frequently used dental implants partly because of the protective oxide coating that spontaneously forms on their surface. We fabricated titania nanosheet (TNS structures on titanium surfaces by NaOH treatment to improve bone differentiation on titanium alloy implants. The cellular response to TNSs on Ti6Al4V alloy was investigated, and the ability of the modified surfaces to affect osteogenic differentiation of rat bone marrow cells and increase the success rate of titanium implants was evaluated. The nanoscale network structures formed by alkali etching markedly enhanced the functions of cell adhesion and osteogenesis-related gene expression of rat bone marrow cells. Other cell behaviors, such as proliferation, alkaline phosphatase activity, osteocalcin deposition, and mineralization, were also markedly increased in TNS-modified Ti6Al4V. Our results suggest that titanium implants modified with nanostructures promote osteogenic differentiation, which may improve the biointegration of these implants into the alveolar bone.

Biomimetic and enzyme-responsive dynamic hydrogels for studying cell-matrix interactions in pancreatic ductal adenocarcinoma.

Science.gov (United States)

Liu, Hung-Yi; Korc, Murray; Lin, Chien-Chi

2018-04-01

The tumor microenvironment (TME) governs all aspects of cancer progression and in vitro 3D cell culture platforms are increasingly developed to emulate the interactions between components of the stromal tissues and cancer cells. However, conventional cell culture platforms are inadequate in recapitulating the TME, which has complex compositions and dynamically changing matrix mechanics. In this study, we developed a dynamic gelatin-hyaluronic acid hybrid hydrogel system through integrating modular thiol-norbornene photopolymerization and enzyme-triggered on-demand matrix stiffening. In particular, gelatin was dually modified with norbornene and 4-hydroxyphenylacetic acid to render this bioactive protein photo-crosslinkable (through thiol-norbornene gelation) and responsive to tyrosinase-triggered on-demand stiffening (through HPA dimerization). In addition to the modified gelatin that provides basic cell adhesive motifs and protease cleavable sequences, hyaluronic acid (HA), an essential tumor matrix, was modularly and covalently incorporated into the cell-laden gel network. We systematically characterized macromer modification, gel crosslinking, as well as enzyme-triggered stiffening and degradation. We also evaluated the influence of matrix composition and dynamic stiffening on pancreatic ductal adenocarcinoma (PDAC) cell fate in 3D. We found that either HA-containing matrix or a dynamically stiffened microenvironment inhibited PDAC cell growth. Interestingly, these two factors synergistically induced cell phenotypic changes that resembled cell migration and/or invasion in 3D. Additional mRNA expression array analyses revealed changes unique to the presence of HA, to a stiffened microenvironment, or to the combination of both. Finally, we presented immunostaining and mRNA expression data to demonstrate that these irregular PDAC cell phenotypes were a result of matrix-induced epithelial-mesenchymal transition (EMT). Copyright © 2018 Elsevier Ltd. All rights

A Strontium-Modified Titanium Surface Produced by a New Method and Its Biocompatibility In Vitro.

Directory of Open Access Journals (Sweden)

Chundong Liu

Full Text Available To present a new and effective method of producing titanium surfaces modified with strontium and to investigate the surface characteristics and in vitro biocompatibility of titanium (Ti surfaces modified with strontium (Sr for bone implant applications.Sr-modified Ti surfaces were produced by sequential treatments with NaOH, strontium acetate, heat and water. The surface characteristics and the concentration of the Sr ions released from the samples were examined. Cell adhesion, morphology and growth were investigated using osteoblasts isolated from the calvaria of neonatal Sprague-Dawley rats. Expression of osteogenesis-related genes and proteins was examined to assess the effect of the Sr-modified Ti surfaces on osteoblasts.The modified titanium surface had a mesh structure with significantly greater porosity, and approximately5.37±0.35at.% of Sr was incorporated into the surface. The hydrophilicity was enhanced by the incorporation of Sr ions and water treatment. The average amounts of Sr released from the Sr-modified plates subjected to water treatment were slight higher than the plates without water treatment. Sr promoted cellular adhesion, spreading and growth compared with untreated Ti surfaces. The Sr-modified Ti plates also promoted expression of osteogenesis-related genes,and expression of OPN and COL-І by osteoblasts. Ti plates heat treated at 700°C showed increased bioactivity in comparison with those treated at 600°C. Water treatment upregulated the expression of osteogenesis-related genes.These results show that Sr-modification of Ti surfaces may improve bioactivity in vitro. Water treatment has enhanced the response of osteoblasts. The Sr-modified Ti heat-treated at 700°C exhibited better bioactivity compared with that heated at 600°C.

Fc-receptors and surface immunoglobulins in cells of the hairy cell leukemia

International Nuclear Information System (INIS)

Rieber, E.P.; Linke, R.P.; Riethmueller, G.; Heyden, H.W. von; Waller, H.D.

1976-01-01

Using 125 I-labelled aggregated IgG in a quantitative assay a strong expression of Fc-receptors was found on the leukemic cells of a patient with hairy cell leukemia. The Fc-receptor activity on these cells was much higher than that on monocytes and B-lymphocytes from normal blood. Surface immunoglobulins were detected by radioautography using radioactively labelled (Fab') 2 -fragments of monospecific antibodies directed against immunoglobulin heavy chains. Prior to radioautography the cells were stained for the tartrate resistant acid phosphatase. It is found that all cells containing this enzyme bore delta-chains on their surface. On more than 90% of these cells a simultaneous expression of μ-chains was detected. γ-chains could only be demonstrated on cells which were negative for the tartrate resistant acid phosphatase; part of these cells, however, were hairy cells by morphological criteria. (orig.) [de

Fc-receptors and surface immunoglobulins in cells of the hairy cell leukemia

Energy Technology Data Exchange (ETDEWEB)

Rieber, E P; Linke, R P; Riethmueller, G [Tuebingen Univ. (Germany, F.R.). Abt. fuer Experimentelle Chirurgie und Immunologie; Heyden, H.W. von; Waller, H D [Tuebingen Univ. (Germany, F.R.). Abt. Innere Medizin 2

1976-01-01

Using /sup 125/I-labelled aggregated IgG in a quantitative assay a strong expression of Fc-receptors was found on the leukemic cells of a patient with hairy cell leukemia. The Fc-receptor activity on these cells was much higher than that on monocytes and B-lymphocytes from normal blood. Surface immunoglobulins were detected by radioautography using radioactively labelled (Fab')/sub 2/-fragments of monospecific antibodies directed against immunoglobulin heavy chains. Prior to radioautography the cells were stained for the tartrate resistant acid phosphatase. It is found that all cells containing this enzyme bore delta-chains on their surface. On more than 90% of these cells a simultaneous expression of ..mu..-chains was detected. ..gamma..-chains could only be demonstrated on cells which were negative for the tartrate resistant acid phosphatase; part of these cells, however, were hairy cells by morphological criteria.

Radical SAM Enzymes in the Biosynthesis of Ribosomally Synthesized and Post-translationally Modified Peptides (RiPPs

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Alhosna Benjdia

2017-11-01

Full Text Available Ribosomally-synthesized and post-translationally modified peptides (RiPPs are a large and diverse family of natural products. They possess interesting biological properties such as antibiotic or anticancer activities, making them attractive for therapeutic applications. In contrast to polyketides and non-ribosomal peptides, RiPPs derive from ribosomal peptides and are post-translationally modified by diverse enzyme families. Among them, the emerging superfamily of radical SAM enzymes has been shown to play a major role. These enzymes catalyze the formation of a wide range of post-translational modifications some of them having no counterparts in living systems or synthetic chemistry. The investigation of radical SAM enzymes has not only illuminated unprecedented strategies used by living systems to tailor peptides into complex natural products but has also allowed to uncover novel RiPP families. In this review, we summarize the current knowledge on radical SAM enzymes catalyzing RiPP post-translational modifications and discuss their mechanisms and growing importance notably in the context of the human microbiota.

Microarray of neuroblastoma cells on the selectively functionalized nanocrystalline diamond thin film surface

Science.gov (United States)

Park, Young-Sang; Son, Hyeong-Guk; Kim, Dae-Hoon; Oh, Hong-Gi; Lee, Da-Som; Kim, Min-Hye; Lim, Ki-Moo; Song, Kwang-Soup

2016-01-01

Nanocrystalline diamond (NCD) film surfaces were modified with fluorine or oxygen by plasma treatment in an O2 or C3F8 gas environment in order to induce wettability. The oxygenated-NCD (O-NCD) film surface was hydrophilic and the fluorinated-NCD (F-NCD) surface was hydrophobic. The efficiency of early cell adhesion, which is dependent on the wettability of the cell culture plate and necessary for the growth and proliferation of cells, was 89.62 ± 3.92% on the O-NCD film and 7.78 ± 0.77% on the F-NCD film surface after 3 h of cell culture. The wettability of the NCD film surface was artificially modified using a metal mask and plasma treatment to fabricate a micro-pattern. Four types of micro-patterns were fabricated (line, circle, mesh, and word) on the NCD film surface. We precisely arrayed the neuroblastoma cells on the micro-patterned NCD film surfaces by controlling the surface wettability and cell seeding density. The neuroblastoma cells adhered and proliferated along the O-NCD film surface.

Enzymes and other agents that enhance cell wall extensibility

Science.gov (United States)

Cosgrove, D. J.

1999-01-01

Polysaccharides and proteins are secreted to the inner surface of the growing cell wall, where they assemble into a network that is mechanically strong, yet remains extensible until the cells cease growth. This review focuses on the agents that directly or indirectly enhance the extensibility properties of growing walls. The properties of expansins, endoglucanases, and xyloglucan transglycosylases are reviewed and their postulated roles in modulating wall extensibility are evaluated. A summary model for wall extension is presented, in which expansin is a primary agent of wall extension, whereas endoglucanases, xyloglucan endotransglycosylase, and other enzymes that alter wall structure act secondarily to modulate expansin action.

Surface treated fly ash filled modified epoxy composites

Directory of Open Access Journals (Sweden)

Uma Dharmalingam

2015-01-01

Full Text Available Abstract Fly ash, an inorganic alumino silicate has been used as filler in epoxy matrix, but it reduces the mechanical properties due to its poor dispersion and interfacial bonding with the epoxy matrix. To improve its interfacial bonding with epoxy matrix, surface treatment of fly ash was done using surfactant sodium lauryl sulfate and silane coupling agent glycidoxy propyl trimethoxy silane. An attempt is also made to reduce the particle size of fly ash using high pressure pulverizer. To improve fly ash dispersion in epoxy matrix, the epoxy was modified by mixing with amine containing liquid silicone rubber (ACS. The effect of surface treated fly ash with varying filler loadings from 10 to 40% weight on the mechanical, morphological and thermal properties of modified epoxy composites was investigated. The surface treated fly ash was characterized by particle size analyzer and FTIR spectra. Morphological studies of surface treated fly ash filled modified epoxy composites indicate good dispersion of fillers in the modified epoxy matrix and improves its mechanical properties. Impact strength of the surface treated fly ash filled modified epoxy composites show more improvement than unmodified composites.

Antibacterial effect of silver nanofilm modified stainless steel surface

Science.gov (United States)

Fang, F.; Kennedy, J.; Dhillon, M.; Flint, S.

2015-03-01

Bacteria can attach to stainless steel surfaces, resulting in the colonization of the surface known as biofilms. The release of bacteria from biofilms can cause contamination of food such as dairy products in manufacturing plants. This study aimed to modify stainless steel surfaces with silver nanofilms and to examine the antibacterial effectiveness of the modified surface. Ion implantation was applied to produce silver nanofilms on stainless steel surfaces. 35 keV Ag ions were implanted with various fluences of 1 × 1015 to 1 × 1017 ions•cm-2 at room temperature. Representative atomic force microscopy characterizations of the modified stainless steel are presented. Rutherford backscattering spectrometry spectra revealed the implanted atoms were located in the near-surface region. Both unmodified and modified stainless steel coupons were then exposed to two types of bacteria, Pseudomonas fluorescens and Streptococcus thermophilus, to determine the effect of the surface modification on bacterial attachment and biofilm development. The silver modified coupon surface fluoresced red over most of the surface area implying that most bacteria on coupon surface were dead. This study indicates that the silver nanofilm fabricated by the ion implantation method is a promising way of reducing the attachment of bacteria and delay biofilm formation.

Surface modified alginate microcapsules for 3D cell culture

Science.gov (United States)

Chen, Yi-Wen; Kuo, Chiung Wen; Chueh, Di-Yen; Chen, Peilin

2016-06-01

Culture as three dimensional cell aggregates or spheroids can offer an ideal platform for tissue engineering applications and for pharmaceutical screening. Such 3D culture models, however, may suffer from the problems such as immune response and ineffective and cumbersome culture. This paper describes a simple method for producing microcapsules with alginate cores and a thin shell of poly(L-lysine)-graft-poly(ethylene glycol) (PLL-g-PEG) to encapsulate mouse induced pluripotent stem (miPS) cells, generating a non-fouling surface as an effective immunoisolation barrier. We demonstrated the trapping of the alginate microcapsules in a microwell array for the continuous observation and culture of a large number of encapsulated miPS cells in parallel. miPS cells cultured in the microcapsules survived well and proliferated to form a single cell aggregate. Droplet formation of monodisperse microcapsules with controlled size combined with flow cytometry provided an efficient way to quantitatively analyze the growth of encapsulated cells in a high-throughput manner. The simple and cost-effective coating technique employed to produce the core-shell microcapsules could be used in the emerging field of cell therapy. The microwell array would provide a convenient, user friendly and high-throughput platform for long-term cell culture and monitoring.

Micro-Bulges Investigation on Laser Modified Tool Steel Surface

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Fauzun Fazliana

2017-01-01

Full Text Available This paper presents micro-bulges investigation on laser modified tool steel. The aim of this study is to understand the effect of laser irradiance and interaction time on surface morphology configuration. An Nd:YAG laser system with TEM00 pulse processing mode was used to modify the samples. Metallographic study shows samples were analyzed for focal position effect on melted pool size, angle of peaks geometry and laser modified layer depth. Surface morphology were analyzed for surface roughness. Laser modified layer shows depth ranged between 42.22 and 420.12 μm. Angle of peak bulge was found to be increase with increasing peak power. The maximum roughness, Ra, achieved in modified H13 was 21.10 μm. These findings are significant to enhance surface properties of laser modified steel and cast iron for dies and high wear resistance applications.

Composition and microstructure alteration of triticale grain surface after processing by enzymes of cellulase complex

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Elena Kuznetsova

2016-01-01

Full Text Available It is found that the pericarp tissue of grain have considerable strength and stiffness, that has an adverse effect on quality of whole-grain bread. Thereby, there exists the need for preliminary chemical and biochemical processing of durable cell walls before industrial use. Increasingly used in the production of bread finds an artificial hybrid of the traditional grain crops of wheat and rye - triticale, grain which has high nutritional value. The purpose of this research was to evaluate the influence of cellulose complex (Penicillium canescens enzymes on composition and microstructure alteration of triticale grain surface, for grain used in baking. Triticale grain was processed by cellulolytic enzyme preparations with different composition (producer is Penicillium canescens. During experiment it is found that triticale grain processing by enzymes of cellulase complex leads to an increase in the content of water-soluble pentosans by 36.3 - 39.2%. The total amount of low molecular sugars increased by 3.8 - 10.5 %. Studies show that under the influence of enzymes the microstructure of the triticale grain surface is changing. Microphotographs characterizing grain surface structure alteration in dynamic (every 2 hours during 10 hours of substrate hydrolysis are shown. It is found that the depth and direction of destruction process for non-starch polysaccharides of grain integument are determined by the composition of the enzyme complex preparation and duration of exposure. It is found, that xylanase involved in the modification of hemicelluloses fiber having both longitudinal and radial orientation. Hydrolysis of non-starch polysaccharides from grain shells led to increase of antioxidant activity. Ferulic acid was identified in alcoholic extract of triticale grain after enzymatic hydrolysis under the influence of complex preparation containing cellulase, xylanase and β-glucanase. Grain processing by independent enzymes containing in complex

Immobilization of Ag nanoparticles/FGF-2 on a modified titanium implant surface and improved human gingival fibroblasts behavior.

Science.gov (United States)

Ma, Qianli; Mei, Shenglin; Ji, Kun; Zhang, Yumei; Chu, Paul K

2011-08-01

The objective of this study was to form a rapid and firm soft tissue sealing around dental implants that resists bacterial invasion. We present a novel approach to modify Ti surface by immobilizing Ag nanoparticles/FGF-2 compound bioactive factors onto a titania nanotubular surface. The titanium samples were anodized to form vertically organized TiO(2) nanotube arrays and Ag nanoparticles were electrodeposited onto the nanotubular surface, on which FGF-2 was immobilized with repeated lyophilization. A uniform distribution of Ag nanoparticles/FGF-2 was observed on the TiO(2) nanotubular surface. The L929 cell line was used for cytotoxicity assessment. Human gingival fibroblasts (HGFs) were cultured on the modified surface for cytocompatibility determination. The Ag/FGF-2 immobilized samples displayed excellent cytocompatibility, negligible cytotoxicity, and enhanced HGF functions such as cell attachment, proliferation, and ECM-related gene expression. The Ag nanoparticles also exhibit some bioactivity. In conclusion, this modified TiO(2) nanotubular surface has a large potential for use in dental implant abutment. Copyright © 2011 Wiley Periodicals, Inc.

IN VITRO TRANSPLANTATION OF GENETICALLY MODIFIED CELLS TO THE TENDON SURFACE

OpenAIRE

Couvreur, Paulus J. J.; Zhao, Chunfeng; Murphy, Stephen; Amadio, Peter C.

2008-01-01

The objective of this paper was to study in vitro transfection of tendon cells and adherence of transfected cells to different tendon surfaces. Achilles tendon fibroblasts from 2-month-old New Zealand white rabbits were cultured to confluence, after which the cells were transfected by an adenovirus carrying either the β-galactosidase reporter gene or the green fluorescent protein (GFP) gene at multiplicities of infection (MOIs) of 50, 100, or 500. Two days later, the cells were transplanted o...

Relating Nanoscale Accessibility within Plant Cell Walls to Improved Enzyme Hydrolysis Yields in Corn Stover Subjected to Diverse Pretreatments.

Science.gov (United States)

Crowe, Jacob D; Zarger, Rachael A; Hodge, David B

2017-10-04

Simultaneous chemical modification and physical reorganization of plant cell walls via alkaline hydrogen peroxide or liquid hot water pretreatment can alter cell wall structural properties impacting nanoscale porosity. Nanoscale porosity was characterized using solute exclusion to assess accessible pore volumes, water retention value as a proxy for accessible water-cell walls surface area, and solute-induced cell wall swelling to measure cell wall rigidity. Key findings concluded that delignification by alkaline hydrogen peroxide pretreatment decreased cell wall rigidity and that the subsequent cell wall swelling resulted increased nanoscale porosity and improved enzyme binding and hydrolysis compared to limited swelling and increased accessible surface areas observed in liquid hot water pretreated biomass. The volume accessible to a 90 Å dextran probe within the cell wall was found to be correlated to both enzyme binding and glucose hydrolysis yields, indicating cell wall porosity is a key contributor to effective hydrolysis yields.

Osteoblastic differentiating potential of dental pulp stem cells in vitro cultured on a chemically modified microrough titanium surface.

Science.gov (United States)

DE Colli, Marianna; Radunovic, Milena; Zizzari, Vincenzo L; DI Giacomo, Viviana; DI Nisio, Chiara; Piattelli, Adriano; Calvo Guirado, José L; Zavan, Barbara; Cataldi, Amelia; Zara, Susi

2018-03-30

Titanium surface modification is critical for dental implant success. Our aim was to determine surfaces influence on dental pulp stem cells (DPSCs) viability and differentiation. Implants were divided into sandblasted/acid-etched (control) and sandblasted/acid-etched coated with calcium and magnesium ions (CaMg), supplied as composite (test). Proliferation was evaluated by MTT, differentiation checking osteoblastic gene expression, PGE2 secretion and matrix formation, inflammation by Interleukin 6 (IL-6) detection. MTT and IL-6 do not modify on test. A PGE2 increase on test is recorded. BMP2 is higher on test at early experimental points, Osterix and RUNX2 augment later. Alizarin-red S reveals higher matrix production on test. These results suggest that test surface is more osteoinductive, representing a start point for in vivo studies aiming at the construction of more biocompatible dental implants, whose integration and clinical performance are improved and some undesired effects, such as implant stability loss and further surgical procedures, are reduced.

Pool boiling of nanoparticle-modified surface with interlaced wettability

KAUST Repository

Hsu, Chin-Chi; Su, Tsung-Wen; Chen, Ping-Hei

2012-01-01

This study investigated the pool boiling heat transfer under heating surfaces with various interlaced wettability. Nano-silica particles were used as the coating element to vary the interlaced wettability of the surface. The experimental results revealed that when the wettability of a surface is uniform, the critical heat flux increases with the more wettable surface; however, when the wettability of a surface is modified interlacedly, regardless of whether the modified region becomes more hydrophilic or hydrophobic, the critical heat flux is consistently higher than that of the isotropic surface. In addition, this study observed that critical heat flux was higher when the contact angle difference between the plain surface and the modified region was smaller. © 2012 Hsu et al.

Surface modification of parylene-N films for the culture of osteoblast-like cells (MG-63)

Energy Technology Data Exchange (ETDEWEB)

Liaqat, Usman [Graduate Program of Nano Science and Technology, Yonsei University, 50-Yonsei Ro, Seodaemun-gu, Seoul 120-749 (Korea, Republic of); Ko, Hyuk [Department of Materials Science and Engineering, Yonsei University, 50-Yonsei Ro, Seodaemun-gu, Seoul 120-749 (Korea, Republic of); Suh, Hwal [Graduate Program of Nano Science and Technology, Yonsei University, 50-Yonsei Ro, Seodaemun-gu, Seoul 120-749 (Korea, Republic of); Department of Medical Engineering, College of Medicine, Yonsei University, 50-Yonsei Ro, Seodaemun-gu, Seoul, 120-749 (Korea, Republic of); Lee, Misu [Division of Life Sciences, College of Life Science and Bioengineering, Incheon National University, Incheon 406-772 (Korea, Republic of); Pyun, Jae-Chul, E-mail: jcpyun@yonsei.ac.kr [Department of Materials Science and Engineering, Yonsei University, 50-Yonsei Ro, Seodaemun-gu, Seoul 120-749 (Korea, Republic of)

2016-08-15

Highlights: • Osteoblast-like cells (MG-63) was cultured on differently modified surfaces of parylene films. • Proliferation of MG-63 was observed to be far increased on UV-treated parylene-N film. • The influences of UV-treatment were found out on cell viability, proliferation rate and cell cycle. • The influence was estimated to be negligible on the protein synthesis, cell differentiation. • The UV-treated parylene-N was demonstrated to be effectively used for the culture of MG-63. - Abstract: The influence of microenvironments on the culture of osteoblast-like cells (MG-63) has been investigated using parylene films with different surfaces, such as parylene-N film, UV-modified parylene-N film, functional parylene film with amine groups (parylene-A), and UV-modified parylene-A film. In this work, parylene-N film was found to induce dramatic changes in cell adhesion and cell viability before and after UV-treatment with respect to the culture of osteoblast-like cells (MG-63). The influences of such a chemical environment on cell culture were investigated in relation to the cell proliferation (viability and proliferation rate) and the cell physiology (cell cycle, protein synthesis, and differentiation) of cells grown on parylene-N film, UV-modified parylene-N film, parylene-A film, and UV-modified parylene-A film in comparison with cells grown on a polystyrene surface.

Microarray of neuroblastoma cells on the selectively functionalized nanocrystalline diamond thin film surface

International Nuclear Information System (INIS)

Park, Young-Sang; Son, Hyeong-Guk; Kim, Dae-Hoon; Oh, Hong-Gi; Lee, Da-Som; Kim, Min-Hye; Lim, Ki-Moo; Song, Kwang-Soup

2016-01-01

Graphical abstract: - Highlights: • The nanocrystalline diamond (NCD) surface is functionalized with F or O. • The cell adhesion and growth are evaluated on the functionalized NCD surface. • The cell adhesion and growth depend on the wettability of the surface. • Cell patterning was achieved by using of hydrophilic and hydrophobic surfaces. • Neuroblastoma cells were arrayed on the micro-patterned NCD surface. - Abstract: Nanocrystalline diamond (NCD) film surfaces were modified with fluorine or oxygen by plasma treatment in an O_2 or C_3F_8 gas environment in order to induce wettability. The oxygenated-NCD (O-NCD) film surface was hydrophilic and the fluorinated-NCD (F-NCD) surface was hydrophobic. The efficiency of early cell adhesion, which is dependent on the wettability of the cell culture plate and necessary for the growth and proliferation of cells, was 89.62 ± 3.92% on the O-NCD film and 7.78 ± 0.77% on the F-NCD film surface after 3 h of cell culture. The wettability of the NCD film surface was artificially modified using a metal mask and plasma treatment to fabricate a micro-pattern. Four types of micro-patterns were fabricated (line, circle, mesh, and word) on the NCD film surface. We precisely arrayed the neuroblastoma cells on the micro-patterned NCD film surfaces by controlling the surface wettability and cell seeding density. The neuroblastoma cells adhered and proliferated along the O-NCD film surface.

Microarray of neuroblastoma cells on the selectively functionalized nanocrystalline diamond thin film surface

Energy Technology Data Exchange (ETDEWEB)

Park, Young-Sang; Son, Hyeong-Guk; Kim, Dae-Hoon; Oh, Hong-Gi; Lee, Da-Som; Kim, Min-Hye; Lim, Ki-Moo; Song, Kwang-Soup, E-mail: kssong10@kumoh.ac.kr

2016-01-15

Graphical abstract: - Highlights: • The nanocrystalline diamond (NCD) surface is functionalized with F or O. • The cell adhesion and growth are evaluated on the functionalized NCD surface. • The cell adhesion and growth depend on the wettability of the surface. • Cell patterning was achieved by using of hydrophilic and hydrophobic surfaces. • Neuroblastoma cells were arrayed on the micro-patterned NCD surface. - Abstract: Nanocrystalline diamond (NCD) film surfaces were modified with fluorine or oxygen by plasma treatment in an O{sub 2} or C{sub 3}F{sub 8} gas environment in order to induce wettability. The oxygenated-NCD (O-NCD) film surface was hydrophilic and the fluorinated-NCD (F-NCD) surface was hydrophobic. The efficiency of early cell adhesion, which is dependent on the wettability of the cell culture plate and necessary for the growth and proliferation of cells, was 89.62 ± 3.92% on the O-NCD film and 7.78 ± 0.77% on the F-NCD film surface after 3 h of cell culture. The wettability of the NCD film surface was artificially modified using a metal mask and plasma treatment to fabricate a micro-pattern. Four types of micro-patterns were fabricated (line, circle, mesh, and word) on the NCD film surface. We precisely arrayed the neuroblastoma cells on the micro-patterned NCD film surfaces by controlling the surface wettability and cell seeding density. The neuroblastoma cells adhered and proliferated along the O-NCD film surface.

Surface modified nano-hydroxyapatite/poly(lactide acid) composite and its osteocyte compatibility

International Nuclear Information System (INIS)

Diao Huaxin; Si Yunfeng; Zhu Aiping; Ji Lijun; Shi Hongchan

2012-01-01

In this study, melt blending was used to fabricate poly(lactic acid) (PLA)/ hydroxyapatite (HA) nanocomposites. Surface modifying HA nanoparticles (mHA) with dodecyl alcohol through esterification reaction could effectively improve the dispersibility of HA nanoparticles in PLA matrix and the interfacial interactions between PLA and HA nanoparticles, as revealed by field emission scanning electron microscopy (FESEM), rheology analysis, and dynamic mechanical thermal analysis (DMTA). mHA/PLA nanocomposite film demonstrated better cartilage cell attachment, spreading and proliferation than that of PLA and HA/PLA film. The good cytocompatibility could be due to the good dispersibility of the osteoinductive HA nanoparticles, good interfacial interactions between PLA and HA nanoparticles, and balanced hydrophobic/hydrophilic property. This newly developed mHA/PLA nanocomposites may be considered for bone tissue engineering applications. - Highlights: ► Dodecyl alcohol modifies HA nanoparticles via esterification reaction. ► The modified HA results in good dispersibility in PLA matrix. ► The interfacial interactions are improved because of the modified HA. ► The addition of HA and mHA results in good cell affinity and biocompatibility.

Assessing the Nano-Dynamics of the Cell Surface

Energy Technology Data Exchange (ETDEWEB)

Bae, Chil Man [Dept. of Physiology and Biophysics, State University of New York, Buffalo (United States); Park, Ik Keun [Mechanical Engineering, Seoul National University of Technology, Seoul (Korea, Republic of); Bulter, Peter J. [Dept. of Bioengineering, The Pennsylvania State University, University Park (United States)

2012-06-15

It is important to know the mechanism of cell membrane fluctuation because it can be readout for the nanomechanical interaction between cytoskeleton and plasma membrane. Traditional techniques, however, have drawbacks such as probe contact with the cell surface, complicate analysis, and limit spatial and temporal resolution. In this study, we developed a new system for non-contact measurement of nano-scale localized-cell surface dynamics using modified-scanning ion-conductance microscopy. With 2 nm resolution, we determined that endothelial cells have local membrane fluctuations of -20 nm, actin depolymerization causes increase in fluctuation amplitude, and ATP depletion abolishes all membrane fluctuations.

Porous surface modified bioactive bone cement for enhanced bone bonding.

Directory of Open Access Journals (Sweden)

Qiang He

Full Text Available Polymethylmethacrylate bone cement cannot provide an adhesive chemical bonding to form a stable cement-bone interface. Bioactive bone cements show bone bonding ability, but their clinical application is limited because bone resorption is observed after implantation. Porous polymethylmethacrylate can be achieved with the addition of carboxymethylcellulose, alginate and gelatin microparticles to promote bone ingrowth, but the mechanical properties are too low to be used in orthopedic applications. Bone ingrowth into cement could decrease the possibility of bone resorption and promote the formation of a stable interface. However, scarce literature is reported on bioactive bone cements that allow bone ingrowth. In this paper, we reported a porous surface modified bioactive bone cement with desired mechanical properties, which could allow for bone ingrowth.The porous surface modified bioactive bone cement was evaluated to determine its handling characteristics, mechanical properties and behavior in a simulated body fluid. The in vitro cellular responses of the samples were also investigated in terms of cell attachment, proliferation, and osteoblastic differentiation. Furthermore, bone ingrowth was examined in a rabbit femoral condyle defect model by using micro-CT imaging and histological analysis. The strength of the implant-bone interface was also investigated by push-out tests.The modified bone cement with a low content of bioactive fillers resulted in proper handling characteristics and adequate mechanical properties, but slightly affected its bioactivity. Moreover, the degree of attachment, proliferation and osteogenic differentiation of preosteoblast cells was also increased. The results of the push-out test revealed that higher interfacial bonding strength was achieved with the modified bone cement because of the formation of the apatite layer and the osseointegration after implantation in the bony defect.Our findings suggested a new bioactive

Temperature and UV light affect the activity of marine cell-free enzymes

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B. Thomson

2017-09-01

Full Text Available Microbial extracellular enzymatic activity (EEA is the rate-limiting step in the degradation of organic matter in the oceans. These extracellular enzymes exist in two forms: cell-bound, which are attached to the microbial cell wall, and cell-free, which are completely free of the cell. Contrary to previous understanding, cell-free extracellular enzymes make up a substantial proportion of the total marine EEA. Little is known about these abundant cell-free enzymes, including what factors control their activity once they are away from their sites (cells. Experiments were run to assess how cell-free enzymes (excluding microbes respond to ultraviolet radiation (UVR and temperature manipulations, previously suggested as potential control factors for these enzymes. The experiments were done with New Zealand coastal waters and the enzymes studied were alkaline phosphatase (APase, β-glucosidase, (BGase, and leucine aminopeptidase (LAPase. Environmentally relevant UVR (i.e. in situ UVR levels measured at our site reduced cell-free enzyme activities by up to 87 % when compared to controls, likely a consequence of photodegradation. This effect of UVR on cell-free enzymes differed depending on the UVR fraction. Ambient levels of UV radiation were shown to reduce the activity of cell-free enzymes for the first time. Elevated temperatures (15 °C increased the activity of cell-free enzymes by up to 53 % when compared to controls (10 °C, likely by enhancing the catalytic activity of the enzymes. Our results suggest the importance of both UVR and temperature as control mechanisms for cell-free enzymes. Given the projected warming ocean environment and the variable UVR light regime, it is possible that there could be major changes in the cell-free EEA and in the enzymes contribution to organic matter remineralization in the future.

DNA immobilization on polymer-modified Si surface by controlling pH

International Nuclear Information System (INIS)

Demirel, Goekcen Birlik; Caykara, Tuncer

2009-01-01

A novel approach based on polymer-modified Si surface as DNA sensor platforms is presented. The polymer-modified Si surface was prepared by using 3-(methacryloxypropyl)trimethoxysilane [γ-MPS] and poly(acrylamide) [PAAm]. Firstly, a layer of γ-MPS was formed on the hydroxylated silicon surface as a monolayer and then modified with different molecular weight of PAAm to form polymer-modified surface. The polymer-modified Si surface was used for dsDNA immobilization. All steps about formation of layer structure were characterized by ellipsometry, atomic force microscopy (AFM), attenuated total reflectance Fourier transformed infrared (ATR-FTIR), and contact angle (CA) measurements. We found that in this case the amount of dsDNA immobilized onto the surface was dictated by the electrostatic interaction between the substrate surface and the DNA. Our results thus demonstrated that DNA molecules could be immobilized differently onto the polymer-modified support surface via electrostatic interactions.

Plant cell wall extensibility: connecting plant cell growth with cell wall structure, mechanics, and the action of wall-modifying enzymes

Energy Technology Data Exchange (ETDEWEB)

Cosgrove, Daniel J.

2015-11-25

The advent of user-friendly instruments for measuring force/deflection curves of plant surfaces at high spatial resolution has resulted in a recent outpouring of reports of the ‘Young's modulus’ of plant cell walls. The stimulus for these mechanical measurements comes from biomechanical models of morphogenesis of meristems and other tissues, as well as single cells, in which cell wall stress feeds back to regulate microtubule organization, auxin transport, cellulose deposition, and future growth directionality. In this article I review the differences between elastic modulus and wall extensibility in the context of cell growth. Some of the inherent complexities, assumptions, and potential pitfalls in the interpretation of indentation force/deflection curves are discussed. Reported values of elastic moduli from surface indentation measurements appear to be 10- to >1000-fold smaller than realistic tensile elastic moduli in the plane of plant cell walls. Potential reasons for this disparity are discussed, but further work is needed to make sense of the huge range in reported values. The significance of wall stress relaxation for growth is reviewed and connected to recent advances and remaining enigmas in our concepts of how cellulose, hemicellulose, and pectins are assembled to make an extensible cell wall. A comparison of the loosening action of α-expansin and Cel12A endoglucanase is used to illustrate two different ways in which cell walls may be made more extensible and the divergent effects on wall mechanics.

Cell In Situ Zymography: Imaging Enzyme-Substrate Interactions.

Science.gov (United States)

Chhabra, Aastha; Rani, Vibha

2017-01-01

Zymography has long been used for the detection of substrate-specific enzyme activity. In situ zymography (ISZ), an adaptation from the conventional substrate zymography, is a widely employed technique useful for the detection, localization, and estimation of enzyme-substrate interactions in tissues. Here, we describe a protocol to detect 'in position' matrix metalloproteinase (MMP) activity in cells utilizing H9c2 cardiomyoblasts as a model. This technique is primarily adopted from the method used for histological sections and is termed as 'Cell in situ Zymography'. It is a simple, sensitive, and quantifiable methodology to assess the functional activity of an enzyme 'on site/in position' in cell culture.

Surface-modified silk hydrogel containing hydroxyapatite nanoparticle with hyaluronic acid-dopamine conjugate.

Science.gov (United States)

Kim, Hyung Hwan; Park, Jong Bo; Kang, Min Ji; Park, Young Hwan

2014-09-01

Silk fibroin/hydroxyapatite (SF/HAp) composite hydrogels were fabricated in this study, having different HAp contents (0-33 wt%) in SF matrix hydrogel. Surface modification of HAp nanoparticle with hyaluronic acid (HA)-dopamine (DA) conjugate improved a dispersibility of HAp in aqueous SF solution due to its negatively charged surface and therefore, fabrication of the SF composite hydrogel having HAp nanoparticles inside could be possible. Zeta potential of surface-modified HAP was examined by ELS. It demonstrates that surface of HAp was well modified to a negative charge with HA-DA. Morphological structure of SF hydrogel containing surface-modified HAp was examined by FE-SEM for analyzing pore structure of hydrogel and deposition of HAp nanoparticle in SF hydrogel. It was found that HAp nanoparticles were uniformly deposited on the pore wall of SF hydrogel. Structural characteristics of SF/HAp composite hydrogel was performed using X-ray diffraction and FT-IR analysis. It was found that β-sheet crystal conformation of SF was significantly influenced by the HAp content during gelation of a mixture of SF and HAp. As a result of MTT assay, the SF/HAp composite hydrogel showed excellent cell proliferation ability. Therefore, it is expected that SF hydrogel containing HAp nanoparticles has a high potential as bone regeneration scaffold. Copyright © 2014 Elsevier B.V. All rights reserved.

Small and large deformation behaviour of mixtures of xanthan and enzyme modified galactomannans

NARCIS (Netherlands)

Kloek, W.; Luyten, H.; Vliet, van T.

1996-01-01

Small and large deformation properties of aqueous mixtures of xanthan with enzyme modified galactomannans at low ionic strength are discussed in terms of the theory of rubber elasticity and the structure of the galactomannans. The linear deformation region of the gels is small indicating that the

Enzymes in biogenesis of plant cell wall polysaccharides. Enzyme characterization using tracer techniques

International Nuclear Information System (INIS)

Dickinson, D.B.

1975-01-01

Enzymes and metabolic pathways, by which starch and cell wall polysaccharides are formed, were investigated in order to learn how these processes are regulated and to identify the enzymatic regulatory mechanisms involved. Germinating lily pollen was used for studies of cell wall formation, and pollen and maize endosperm for studies of starch biosynthesis. Hexokinase being the first step in conversion of hexoses to starch, wall polysaccharides and respiratory substrates, maize endosperm enzyme was assayed by its conversion of 14 C-hexose to 14 C-hexose-6-P, and rapid separation of the two labelled compounds on anion-exchange paper. This enzyme did not appear to be under tight regulation by feed-back inhibition or activation, nor to be severely inhibited by glucose-6-P or activated by citrate. ADP-glucose pyrophosphorylase and other pyrophosphorylases were assayed radiochemically with 14 C-glucose-1-P (forward direction) or 32-PPsub(i) (reverse direction). They showed that the maize endosperm enzyme was activated by the glycolytic intermediates fructose-6-P and 3-phosphoglycerate, and that low levels of the enzyme were present in the high sucrose-low starch mutant named shrunken-2. Under optimal in-vitro assay conditions, the pollen enzyme reacted four times faster than the observed in-vivo rate of starch accumulation. Biogenesis of plant cell wall polysaccharides requires the conversion of hexose phosphates to various sugar nucleotides and utilization of the latter by the appropriate polysaccharide synthetases. Lily pollen possesses a β-1,3-glucan synthetase which is activated up to six-fold by β-linked oligosaccharides. Hence, the in-vivo activity of this enzyme may be modulated by such effector molecules

Characterization of fatty acid modifying enzyme activity in staphylococcal mastitis isolates and other bacteria

Directory of Open Access Journals (Sweden)

Lu Thea

2012-06-01

Full Text Available Abstract Background Fatty acid modifying enzyme (FAME has been shown to modify free fatty acids to alleviate their bactericidal effect by esterifying fatty acids to cholesterol or alcohols. Although it has been shown in previous studies that FAME is required for Staphylococcus aureus survival in skin abscesses, FAME is poorly studied compared to other virulence factors. FAME activity had also been detected in coagulase-negative staphylococci (CNS. However, FAME activity was only surveyed after a bacterial culture was grown for 24 h. Therefore if FAME activity was earlier in the growth phase, it would not have been detected by the assay and those strains would have been labeled as FAME negative. Results Fifty CNS bovine mastitis isolates and several S. aureus, Escherichia coli, and Streptococcus uberis strains were assayed for FAME activity over 24 h. FAME activity was detected in 54% of CNS and 80% S. aureus strains surveyed but none in E. coli or S. uberis. While some CNS strains produced FAME activity comparable to the lab strain of S. aureus, the pattern of FAME activity varied among strains and across species of staphylococci. All CNS that produced FAME activity also exhibited lipase activity. Lipase activity relative to colony forming units of these CNS decreased over the 24 h growth period. No relationship was observed between somatic cell count in the milk and FAME activity in CNS. Conclusions Some staphylococcal species surveyed produced FAME activity, but E. coli and S. uberis strains did not. All FAME producing CNS exhibited lipase activity which may indicate that both these enzymes work in concert to alter fatty acids in the bacterial environment.

A highly sensitive electrochemical biosensor for catechol using conducting polymer reduced graphene oxide-metal oxide enzyme modified electrode.

Science.gov (United States)

Sethuraman, V; Muthuraja, P; Anandha Raj, J; Manisankar, P

2016-10-15

The fabrication, characterization and analytical performances were investigated for a catechol biosensor, based on the PEDOT-rGO-Fe2O3-PPO composite modified glassy carbon (GC) electrode. The graphene oxide (GO) doped conducting polymer poly (3,4-ethylenedioxythiophene) (PEDOT) was prepared through electrochemical polymerization by potential cycling. Reduction of PEDOT-GO was carried out by amperometric method. Fe2O3 nanoparticles were synthesized in ethanol by hydrothermal method. The mixture of Fe2O3, PPO and glutaraldehyde was casted on the PEDOT-rGO electrode. The surface morphology of the modified electrodes was studied by FE-SEM and AFM. Cyclic voltammetric studies of catechol on the enzyme modified electrode revealed higher reduction peak current. Determination of catechol was carried out successfully by Differential Pulse Voltammetry (DPV) technique. The fabricated biosensor investigated shows a maximum current response at pH 6.5. The catechol biosensor exhibited wide sensing linear range from 4×10(-8) to 6.20×10(-5)M, lower detection limit of 7×10(-9)M, current maxima (Imax) of 92.55µA and Michaelis-Menten (Km) constant of 30.48µM. The activation energy (Ea) of enzyme electrode is 35.93KJmol(-1) at 50°C. There is no interference from d-glucose and l-glutamic acid, ascorbic acid and o-nitrophenol. The PEDOT-rGO-Fe2O3-PPO biosensor was stable for at least 75 days when stored in a buffer at about 4°C. Copyright © 2015 Elsevier B.V. All rights reserved.

Infection Dynamics Vary between Symbiodinium Types and Cell Surface Treatments during Establishment of Endosymbiosis with Coral Larvae

Directory of Open Access Journals (Sweden)

Bette Lynn Willis

2011-07-01

Full Text Available Symbioses between microbes and higher organisms underpin high diversity in many ecosystems, including coral reefs, however mechanisms underlying the early establishment of symbioses remain unclear. Here we examine the roles of Symbiodinium type and cell surface recognition in the establishment of algal endosymbiosis in the reef-building coral, Acropora tenuis. We found 20–70% higher infection success (proportion of larvae infected and five-fold higher Symbiodinium abundance in larvae exposed to ITS-1 type C1 compared to ITS-1 type D in the first 96 h following exposure. The highest abundance of Symbiodinium within larvae occurred when C1-type cells were treated with enzymes that modified the 40–100 kD glycome, including glycoproteins and long chain starch residues. Our finding of declining densities of Symbiodinium C1 through time in the presence of intact cell surface molecules supports a role for cell surface recognition molecules in controlling post-phagocytosis processes, leading to rejection of some Symbiodinium types in early ontogeny. Reductions in the densities of unmodified C1 symbionts after 96 h, in contrast to increases in D symbionts may suggest the early initiation of a winnowing process contributing to the establishment of Symbiodinium D as the dominant type in one-month old juveniles of A. tenuis.

Electrochemical characteristics of Shewanella loihica on carbon nanotubes-modified graphite surfaces

International Nuclear Information System (INIS)

Zhang, Xiaoming; Epifanio, Monica; Marsili, Enrico

2013-01-01

Highlights: • We deposited CNT coatings on graphite electrode by electrophoretic deposition. • CNT coating increased extracellular electron transfer in Shewanella loihica biofilms. • Thick electroactive biofilms hinder the electroactivity of CNT coatings. -- Abstract: High specific surface and electrocatalytic activity of the electrode surface favour extracellular electron transfer from electrochemically active biofilms to polarized electrodes. We coated layer-by-layer carbon nanotubes (CNTs) on graphite electrodes through electrophoretic deposition, thus increasing the electrocatalytic activity. After determining the optimal number of CNT layers through electrochemical methods, we grew Shewanella loihica PV-4 biofilms on the CNT-coated electrodes to quantify the increase in extracellular electron transfer rate compared with unmodified electrodes. Current density on CNT-modified electrodes was 1.7 times higher than that observed on unmodified electrodes after 48 h from inoculation. Rapid microbial cells attachment on CNT-coated electrodes, as determined from scanning electronic microscopy, explained the rapid increase of the current. Also, the CNT reduced the charge transfer resistance of the graphite electrodes, as measured by Electrochemical Impedance Spectroscopy. However, the electrocatalytic activity of the CNT-coated electrode decreased as the biofilm grew thicker and covered the CNT-coating. These result confirmed that surface-modified electrodes improve the electron transfer rate in thin biofilms (<5 μm), but are not feasible for power production in microbial fuel cells, where the biofilm thickness is much higher

Surface cell immobilization within perfluoroalkoxy microchannels

Energy Technology Data Exchange (ETDEWEB)

Stojkovič, Gorazd; Krivec, Matic [Faculty of Chemistry and Chemical Technology, University of Ljubljana, Aškerčeva 5, SI-1000 Ljubljana (Slovenia); Vesel, Alenka [Jožef Stefan Institute, Jamova cesta 39, 1000 Ljubljana (Slovenia); Marinšek, Marjan [Faculty of Chemistry and Chemical Technology, University of Ljubljana, Aškerčeva 5, SI-1000 Ljubljana (Slovenia); Žnidaršič-Plazl, Polona, E-mail: polona.znidarsic@fkkt.uni-lj.si [Faculty of Chemistry and Chemical Technology, University of Ljubljana, Aškerčeva 5, SI-1000 Ljubljana (Slovenia)

2014-11-30

Graphical abstract: - Highlights: • A very efficient approach for immobilization of cells into microreactors is presented. • It is applicable to various materials, including PFA and cyclic olefin (co)polymers. • It was used to immobilize different prokaryotic and eukaryotic microbes. • Cells were immobilized on the surface in high density and showed good stability. • Mechanisms of APTES interactions with target materials are proposed. - Abstract: Perfluoroalkoxy (PFA) is one of the most promising materials for the fabrication of cheap, solvent resistant and reusable microfluidic chips, which have been recently recognized as effective tools for biocatalytic process development. The application of biocatalysts significantly depends on efficient immobilization of enzymes or cells within the reactor enabling long-term biocatalyst use. Functionalization of PFA microchannels by 3-aminopropyltriethoxysilane (ATPES) and glutaraldehyde was used for rapid preparation of microbioreactors with surface-immobilized cells. X-ray photoelectron spectroscopy and scanning electron microscopy were used to accurately monitor individual treatment steps and to select conditions for cell immobilization. The optimized protocol for Saccharomyces cerevisiae immobilization on PFA microchannel walls comprised ethanol surface pretreatment, 4 h contacting with 10% APTES aqueous solution, 10 min treatment with 1% glutaraldehyde and 20 min contacting with cells in deionized water. The same protocol enabled also immobilization of Escherichia coli, Pseudomonas putida and Bacillus subtilis cells on PFA surface in high densities. Furthermore, the developed procedure has been proved to be very efficient also for surface immobilization of tested cells on other materials that are used for microreactor fabrication, including glass, polystyrene, poly (methyl methacrylate), polycarbonate, and two olefin-based polymers, namely Zeonor{sup ®} and Topas{sup ®}.

Surface modification of hydrophobic polymers for improvement of endothelial cell-surface interactions

NARCIS (Netherlands)

Dekker, A.; Dekker, A.; Reitsma, K.; Beugeling, T.; Beugeling, T.; Bantjes, A.; Bantjes, A.; Feijen, Jan; Kirkpatrick, C.J.; van Aken, W.G.

1992-01-01

The aim of this study is to improve the interaction of endothelial cells with polymers used in vascular prostheses. Polytetrafluoroethylene (PTFE; Teflon) films were treated by means of nitrogen and oxygen plasmas. Depending on the plasma exposure time, modified PTFE surfaces showed water-contact

Anaplasma phagocytophilum increases the levels of histone modifying enzymes to inhibit cell apoptosis and facilitate pathogen infection in the tick vector Ixodes scapularis

Czech Academy of Sciences Publication Activity Database

Cabezas-Cruz, A.; Alberdi, P.; Ayllón, N.; Valdés, James J.; Pierce, R.; Villar, M.; de la Fuente, J.

2016-01-01

Roč. 11, č. 4 (2016), s. 303-319 ISSN 1559-2294 EU Projects: European Commission(XE) 278976 - ANTIGONE; European Commission(XE) 316304 - MODBIOLIN Institutional support: RVO:60077344 Keywords : Anaplasma * epigenetic s * histone modifying enzyme * histone * pathogen * tick Subject RIV: GJ - Animal Vermins ; Diseases, Veterinary Medicine Impact factor: 4.394, year: 2016

Interaction of CO2 laser-modified nylon with osteoblast cells in relation to wettability

International Nuclear Information System (INIS)

Waugh, D.G.; Lawrence, J.; Morgan, D.J.; Thomas, C.L.

2009-01-01

It has been amply demonstrated previously that CO 2 lasers hold the ability to surface modify various polymers. In addition, it has been observed that these surface enhancements can augment the biomimetic nature of the laser irradiated materials. This research has employed a CO 2 laser marker to produce trench and hatch topographical patterns with peak heights of around 1 μm on the surface of nylon 6,6. The patterns generated have been analysed using white light interferometry, optical microscopy and X-ray photoelectron spectroscopy was employed to determine the surface oxygen content. Contact angle measurements were used to characterize each sample in terms of wettability. Generally, it was seen that as a result of laser processing the contact angle, surface roughness and surface oxygen content increased whilst the apparent polar and total surface energies decreased. The increase in contact angle and reduction in surface energy components was found to be on account of a mixed intermediate state wetting regime owing to the change in roughness due to the induced topographical patterns. To determine the biomimetic nature of the modified and as-received control samples each one was seeded with 2 x 10 4 cells/ml normal human osteoblast cells and observed after periods of 24 h and 4 days using optical microscopy and SEM to determine mean cell cover densities and variations in cell morphology. In addition, a haemocytometer was used to show that the cell count for the laser patterned samples had increased by up to a factor of 1.5 compared to the as-received control sample after 4 days of incubation. Significantly, it was determined that all laser-induced patterns gave rise to better cell response in comparison to the as-received control sample studied due to increased preferential cell growth on those surfaces with increased surface roughness.

Increasing the Energy Efficiency of Aluminum-Reduction Cells Using Modified Cathodes

Science.gov (United States)

Jianping, Peng; Yang, Song; Yuezhong, Di; Yaowu, Wang; Naixiang, Feng

2017-10-01

A cathode with an inclined surface (5°) and increased bar collector height (230 mm high) was incorporated into two 300-kA industrial aluminum-reduction cells. The voltage of the cells with the modified cathode was reduced by approximately 200 mV when compared with that of a conventional cell with a flat cathode. Through the use of simulations, the reduction in the cell voltage was attributed to the cathode modification (40 mV) and a reduced electrolyte level of 0.5 cm (160 mV). As a result of reduced anode cathode distance (ACD), the ledge toe was extended to the anode shadow by 12 cm. This caused a large inverted horizontal current and a velocity increase. The ledge profile returned to the desired position when the cells were insulated more effectively, and the metal velocity and metal crest in the modified cells were reduced accordingly.

Modified PAMAM dendrimer with 4-carbomethoxypyrrolidone surface groups reveals negligible toxicity against three rodent cell-lines

DEFF Research Database (Denmark)

Janaszewska, Anna; Ciolkowski, Michal; Wróbel, Dominika

2013-01-01

Modification of the surface groups of dendrimers is one of the methods to improve their biocompatibility. This article presents results of experiments related to the toxicity of a modified polyamidoamine (PAMAM) dendrimer of the fourth generation with 4-carbomethoxypyrrolidone surface groups (PAM...

Polymer microfilters with nanostructured surfaces for the culture of circulating cancer cells

International Nuclear Information System (INIS)

Makarova, Olga V.; Adams, Daniel L.; Divan, Ralu; Rosenmann, Daniel; Zhu, Peixuan; Li, Shuhong; Amstutz, Platte; Tang, Cha-Mei

2016-01-01

There is a critical need to improve the accuracy of drug screening and testing through the development of in vitro culture systems that more effectively mimic the in vivo environment. Surface topographical features on the nanoscale level, in short nanotopography, effect the cell growth patterns, and hence affect cell function in culture. We report the preliminary results on the fabrication, and subsequent cellular growth, of nanoscale surface topography on polymer microfilters using cell lines as a precursor to circulating tumor cells (CTCs). To create various nanoscale features on the microfilter surface, we used reactive ion etching (RIE) with and without an etching mask. An anodized aluminum oxide (AAO) membrane fabricated directly on the polymer surface served as an etching mask. Polymer filters with a variety of modified surfaces were used to compare the effects on the culture of cancer cell lines in blank culture wells, with untreated microfilters or with RIE-treated microfilters. We then report the differences of cell shape, phenotype and growth patterns of bladder and glioblastoma cancer cell lines after isolation on the various types of material modifications. Our data suggest that RIE modified polymer filters can isolate model cell lines while retaining ell viability, and that the RIE filter modification allows T24 monolayering cells to proliferate as a structured cluster. - Highlights: • Surface topographical effects the growth patterns and cell function of cancer cells • Nanoscale surface topography on polymer filters for circulating tumor cell culture • Membrane fabricated directly on polymer surfaces utilized for polymer etching • Nanotopography alters cell shape, phenotype and growth patterns of cancer cells • Nanoscale surface topography dictates monolayering or 3D structured cell culture

Polymer microfilters with nanostructured surfaces for the culture of circulating cancer cells

Energy Technology Data Exchange (ETDEWEB)

Makarova, Olga V. [Creatv MicroTech, Inc., 2242 West Harrison St., Chicago 60612, IL (United States); Adams, Daniel L., E-mail: dan@creatvmicrotech.com [Creatv MicroTech, Inc., 1 Deer Park Drive, Monmouth Junction, NJ 08852 (United States); Divan, Ralu; Rosenmann, Daniel [Center for Nanoscale Materials, Argonne National Laboratory, 9700 South Cass Ave., Argonne 60439, IL (United States); Zhu, Peixuan; Li, Shuhong; Amstutz, Platte; Tang, Cha-Mei [Creatv MicroTech, Inc., 11609 Lake Potomac Drive, Potomac 20854, MD (United States)

2016-09-01

There is a critical need to improve the accuracy of drug screening and testing through the development of in vitro culture systems that more effectively mimic the in vivo environment. Surface topographical features on the nanoscale level, in short nanotopography, effect the cell growth patterns, and hence affect cell function in culture. We report the preliminary results on the fabrication, and subsequent cellular growth, of nanoscale surface topography on polymer microfilters using cell lines as a precursor to circulating tumor cells (CTCs). To create various nanoscale features on the microfilter surface, we used reactive ion etching (RIE) with and without an etching mask. An anodized aluminum oxide (AAO) membrane fabricated directly on the polymer surface served as an etching mask. Polymer filters with a variety of modified surfaces were used to compare the effects on the culture of cancer cell lines in blank culture wells, with untreated microfilters or with RIE-treated microfilters. We then report the differences of cell shape, phenotype and growth patterns of bladder and glioblastoma cancer cell lines after isolation on the various types of material modifications. Our data suggest that RIE modified polymer filters can isolate model cell lines while retaining ell viability, and that the RIE filter modification allows T24 monolayering cells to proliferate as a structured cluster. - Highlights: • Surface topographical effects the growth patterns and cell function of cancer cells • Nanoscale surface topography on polymer filters for circulating tumor cell culture • Membrane fabricated directly on polymer surfaces utilized for polymer etching • Nanotopography alters cell shape, phenotype and growth patterns of cancer cells • Nanoscale surface topography dictates monolayering or 3D structured cell culture.

Biomolecular strategies for cell surface engineering

Science.gov (United States)

Wilson, John Tanner

backbone molecular weight, PEG chain length, and grafting ratio, PLL-g-PEG copolymers were rendered cytocompatible and used to initiate and propagate the growth of cell surface-supported PEM films. Planar characterization of this novel class of PEM films indicated that film thickness and composition may be tailored through appropriate control of layer number and copolymer properties. Furthermore, these investigations have helped establish a conceptual framework for the rational design of cell surface-supported thin films, with the objective of translating the diverse biomedical and biotechnological applications of PEM films to cellular interfaces. Important to the development of effective conformal islet coatings is an inherent strategy through which to incorporate bioactive molecules for directing desired biochemical or cellular responses. Towards this end, PLL-g-PEG copolymers functionalized with biotin, azide, and hydrazide moieties were synthesized and used, either alone or in combination, to capture streptavidin-, triphenylphosphine-, and aldehyde-labeled probes, respectively, on the islet surface. Additionally, PEM films assembled using alginate chemically modified to contain aldehyde groups could be used to introduce hydrazide-functionalized molecules to the islet surface. Hence, modified film constituents may be used as modular elements for controlling the chemical composition cell and tissue surfaces. Finally, we report a strategy for tethering thrombomodulin (TM) to the islet surface. Through site-specific, C-terminal biotinylation of TM and optimization of cell surface biotinylation, TM could be integrated with the islet surface. Re-engineering of islet surfaces with TM resulted in an increased catalytic capacity of islets to generate the powerful anti-inflammatory agent, activated protein C (APC), thereby providing a facile strategy for increasing the local concentration of APC at the site of transplantation.

Enzyme electrode configurations : for application in biofuel cells

Energy Technology Data Exchange (ETDEWEB)

Wang Xiaoju

2012-07-01

; their effects on the electrode performance were then investigated. It is proposed that the {eta}-{eta} interaction between the PSS{sup -} and the hydrophobic substrate-binding pocket in the vicinity of the T1 Cu site results in a favorable location of the conducting polymer chain of PEDOT-PSS close to the T1 Cu site and thus facilitates the DET of ThL within this particular architecture. The flavin adenine dinucleotide (FAD)-dependent glucose dehydrogenase from Glomerella cingulata (GcGDH) and cellobiose dehydrogense from Corynascus thermophuilus (CtCDH) have been studied to construct different enzyme electrode configurations as bioanodes towards biofuel cell applications. For GcGDH, six Os-containing polymers, whose redox potentials range across a broad potential window between +15 and +489 mV vs. NHE, were used to 'wire' the GcGDH on the graphite electrodes to catalyze the oxidation of glucose. The ratio of GcGDH:Os-polymer in the overall loading onto the electrode surface significantly affected the catalytic performance of the enzyme electrode on the glucose oxidation. Both the Os-polymer and the GcGDH:Os-polymer ratio were optimized for obtaining the maximum current density; a high current density of 493 {mu}A/cm{sup 2} for 30 mM glucose was produced by a GcGDH/Os c modified electrode. DET type biocatalysis of CtCDH on lactose (and glucose) oxidation was accomplished on Au nanoparticle (AuNP) structured electrode. The haem site in the CtCDH enzyme functions as a 'built-in' mediator for communicating the electron transfer between the FAD site and the AuNP surface. The redox potential of the haem site in CtCDH was determined to be E{sub 1/2} = -122 mV vs. Ag/AgCl/KCl(s) (75 mV vs. NHE). The CtCDH/AuNP/Au bioanode can generate a maximum current response for lactose with I{sub max} = 43.3{+-}1.5 ({mu}A/cm{sup 2}) or for glucose with I{sub max} = 31.2{+-}2.3 ({mu}A/cm{sup 2}). The DET type biocatalysis of CtCDH works most efficiently in a more neutral

Photovoltaic characterization of hybrid solar cells using surface modified TiO2 nanoparticles and poly(3-hexyl)thiophene

International Nuclear Information System (INIS)

Guenes, Serap; Marjanovic, Nenad; Nedeljkovic, Jovan M; Sariciftci, Niyazi Serdar

2008-01-01

We report on the photovoltaic performance of bulk heterojunction solar cells using novel nanoparticles of 6-palmitate ascorbic acid surface modified TiO 2 as an electron acceptor embedded into the donor poly(3-hexyl)thiophene (P3HT) matrix. Devices were fabricated by using P3HT with varying amounts of red TiO 2 nanoparticles (1:1, 1:2, 1:3 w-w ratio). The devices were characterized by measuring current-voltage characteristics under simulated AM 1.5 conditions. Incident photon to current efficiency (IPCE) was spectrally resolved. The nanoscale morphology of such organic/inorganic hybrid blends was also investigated using atomic force microscopy (AFM).

Substrate mediated enzyme prodrug therapy.

Directory of Open Access Journals (Sweden)

Betina Fejerskov

Full Text Available In this report, we detail Substrate Mediated Enzyme Prodrug Therapy (SMEPT as a novel approach in drug delivery which relies on enzyme-functionalized cell culture substrates to achieve a localized conversion of benign prodrug(s into active therapeutics with subsequent delivery to adhering cells or adjacent tissues. For proof-of-concept SMEPT, we use surface adhered micro-structured physical hydrogels based on poly(vinyl alcohol, β-glucuronidase enzyme and glucuronide prodrugs. We demonstrate enzymatic activity mediated by the assembled hydrogel samples and illustrate arms of control over rate of release of model fluorescent cargo. SMEPT was not impaired by adhering cells and afforded facile time - and dose - dependent uptake of the in situ generated fluorescent cargo by hepatic cells, HepG2. With the use of a glucuronide derivative of an anticancer drug, SN-38, SMEPT afforded a decrease in cell viability to a level similar to that achieved using parent drug. Finally, dose response was achieved using SMEPT and administration of judiciously chosen concentration of SN-38 glucuronide prodrug thus revealing external control over drug delivery using drug eluting surface. We believe that this highly adaptable concept will find use in diverse biomedical applications, specifically surface mediated drug delivery and tissue engineering.

Environmentally responsive surface-modified silica nanoparticles for enhanced oil recovery

International Nuclear Information System (INIS)

Behzadi, Abed; Mohammadi, Aliasghar

2016-01-01

Environmentally responsive surface-modified nanoparticles are colloidal nanoparticles coated with, at least, two physicochemically distinct surface groups. Recent advances in the synthesis and production of nanoparticles have enabled the production of environmentally responsive surface-modified nanoparticles with both hydrophilic and hydrophobic surface groups. These nanoparticles act like colloidal surfactants. In this paper, environmentally responsive surface-modified silica nanoparticles are synthesized and used for enhancement of oil recovery. For this purpose, silica nanoparticles are coated with polyethylene glycol chains as hydrophilic agent and propyl chains as hydrophobic agent at various quantities, and their ability to modulate oil–water interface properties and oil recovery is examined. Oil–water interfacial tension and water surface tension are decreased by 50 % in the presence of silica nanoparticles coated with both agents. Measuring oil-drop contact angle on oil-wetted glass slides and carbonate rock sections, after aging in various surface-modified silica nanofluids, indicates that the wettability of various oil-wetted surfaces is modified from strongly oil-wet to water-wet. Flooding nanofluids to glass micro-models and pore-level investigations demonstrate that surface modification of silica nanoparticles, specially, with both hydrophilic and hydrophobic agents improves considerably their performance in increasing oil recovery and wettability alteration.

Environmentally responsive surface-modified silica nanoparticles for enhanced oil recovery

Energy Technology Data Exchange (ETDEWEB)

Behzadi, Abed; Mohammadi, Aliasghar, E-mail: amohammadi@sharif.edu [Sharif University of Technology, Department of Chemical and Petroleum Engineering (Iran, Islamic Republic of)

2016-09-15

Environmentally responsive surface-modified nanoparticles are colloidal nanoparticles coated with, at least, two physicochemically distinct surface groups. Recent advances in the synthesis and production of nanoparticles have enabled the production of environmentally responsive surface-modified nanoparticles with both hydrophilic and hydrophobic surface groups. These nanoparticles act like colloidal surfactants. In this paper, environmentally responsive surface-modified silica nanoparticles are synthesized and used for enhancement of oil recovery. For this purpose, silica nanoparticles are coated with polyethylene glycol chains as hydrophilic agent and propyl chains as hydrophobic agent at various quantities, and their ability to modulate oil–water interface properties and oil recovery is examined. Oil–water interfacial tension and water surface tension are decreased by 50 % in the presence of silica nanoparticles coated with both agents. Measuring oil-drop contact angle on oil-wetted glass slides and carbonate rock sections, after aging in various surface-modified silica nanofluids, indicates that the wettability of various oil-wetted surfaces is modified from strongly oil-wet to water-wet. Flooding nanofluids to glass micro-models and pore-level investigations demonstrate that surface modification of silica nanoparticles, specially, with both hydrophilic and hydrophobic agents improves considerably their performance in increasing oil recovery and wettability alteration.

Cell Wall Remodeling Enzymes Modulate Fungal Cell Wall Elasticity and Osmotic Stress Resistance.

Science.gov (United States)

Ene, Iuliana V; Walker, Louise A; Schiavone, Marion; Lee, Keunsook K; Martin-Yken, Hélène; Dague, Etienne; Gow, Neil A R; Munro, Carol A; Brown, Alistair J P

2015-07-28

The fungal cell wall confers cell morphology and protection against environmental insults. For fungal pathogens, the cell wall is a key immunological modulator and an ideal therapeutic target. Yeast cell walls possess an inner matrix of interlinked β-glucan and chitin that is thought to provide tensile strength and rigidity. Yeast cells remodel their walls over time in response to environmental change, a process controlled by evolutionarily conserved stress (Hog1) and cell integrity (Mkc1, Cek1) signaling pathways. These mitogen-activated protein kinase (MAPK) pathways modulate cell wall gene expression, leading to the construction of a new, modified cell wall. We show that the cell wall is not rigid but elastic, displaying rapid structural realignments that impact survival following osmotic shock. Lactate-grown Candida albicans cells are more resistant to hyperosmotic shock than glucose-grown cells. We show that this elevated resistance is not dependent on Hog1 or Mkc1 signaling and that most cell death occurs within 10 min of osmotic shock. Sudden decreases in cell volume drive rapid increases in cell wall thickness. The elevated stress resistance of lactate-grown cells correlates with reduced cell wall elasticity, reflected in slower changes in cell volume following hyperosmotic shock. The cell wall elasticity of lactate-grown cells is increased by a triple mutation that inactivates the Crh family of cell wall cross-linking enzymes, leading to increased sensitivity to hyperosmotic shock. Overexpressing Crh family members in glucose-grown cells reduces cell wall elasticity, providing partial protection against hyperosmotic shock. These changes correlate with structural realignment of the cell wall and with the ability of cells to withstand osmotic shock. The C. albicans cell wall is the first line of defense against external insults, the site of immune recognition by the host, and an attractive target for antifungal therapy. Its tensile strength is conferred by

Modification of polymer surfaces to enhance enzyme activity and stability

DEFF Research Database (Denmark)

Hoffmann, Christian

Enzyme immobilization is an important concept for the development of improved biocatalytic processes, primarily through facilitated separation procedures. However, enzyme immobilization usually comes at a price of reduced biocatalytic activity. For this reason, different immobilization methods have...... already been developed, combining the same goal to improve enzyme activity, stability and selectivity. Polymer materials have shown, due to their easy processibility and versatile properties, high potential as enzyme support. However, in order to achieve improved enzyme performance, the combination...... on their tailored surface modification in order to obtain improved enzyme-support systems. Firstly, an off-stoichiometric thiol-ene (OSTE) thermosetting material was used for the development of a screening platform allowing the investigation of micro-environmental effects and their impact on the activity...

Atomic diffusion in laser surface modified AISI H13 steel

Science.gov (United States)

Aqida, S. N.; Brabazon, D.; Naher, S.

2013-07-01

This paper presents a laser surface modification process of AISI H13 steel using 0.09 and 0.4 mm of laser spot sizes with an aim to increase surface hardness and investigate elements diffusion in laser modified surface. A Rofin DC-015 diffusion-cooled CO2 slab laser was used to process AISI H13 steel samples. Samples of 10 mm diameter were sectioned to 100 mm length in order to process a predefined circumferential area. The parameters selected for examination were laser peak power, pulse repetition frequency (PRF), and overlap percentage. The hardness properties were tested at 981 mN force. Metallographic study and energy dispersive X-ray spectroscopy (EDXS) were performed to observe presence of elements and their distribution in the sample surface. Maximum hardness achieved in the modified surface was 1017 HV0.1. Change of elements composition in the modified layer region was detected in the laser modified samples. Diffusion possibly occurred for C, Cr, Cu, Ni, and S elements. The potential found for increase in surface hardness represents an important method to sustain tooling life. The EDXS findings signify understanding of processing parameters effect on the modified surface composition.

Engineering of GlcNAc-1-Phosphotransferase for Production of Highly Phosphorylated Lysosomal Enzymes for Enzyme Replacement Therapy.

Science.gov (United States)

Liu, Lin; Lee, Wang-Sik; Doray, Balraj; Kornfeld, Stuart

2017-06-16

Several lysosomal enzymes currently used for enzyme replacement therapy in patients with lysosomal storage diseases contain very low levels of mannose 6-phosphate, limiting their uptake via mannose 6-phosphate receptors on the surface of the deficient cells. These enzymes are produced at high levels by mammalian cells and depend on endogenous GlcNAc-1-phosphotransferase α/β precursor to phosphorylate the mannose residues on their glycan chains. We show that co-expression of an engineered truncated GlcNAc-1-phosphotransferase α/β precursor and the lysosomal enzyme of interest in the producing cells resulted in markedly increased phosphorylation and cellular uptake of the secreted lysosomal enzyme. This method also results in the production of highly phosphorylated acid β-glucocerebrosidase, a lysosomal enzyme that normally has just trace amounts of this modification.

Display of a thermostable lipase on the surface of a solvent-resistant bacterium, Pseudomonas putida GM730, and its applications in whole-cell biocatalysis

Directory of Open Access Journals (Sweden)

Kwon Seok-Joon

2006-04-01

Full Text Available Abstract Background Whole-cell biocatalysis in organic solvents has been widely applied to industrial bioprocesses. In two-phase water-solvent processes, substrate conversion yields and volumetric productivities can be limited by the toxicity of solvents to host cells and by the low mass transfer rates of the substrates from the solvent phase to the whole-cell biocatalysts in water. Results To solve the problem of solvent toxicity, we immobilized a thermostable lipase (TliA from Pseudomonas fluorescens on the cell surface of a solvent-resistant bacterium, Pseudomonas putida GM730. Surface immobilization of enzymes eliminates the mass-transfer limitation imposed by the cell wall and membranes. TliA was successfully immobilized on the surface of P. putida cells using the ice-nucleation protein (INP anchoring motif from Pseudomonas syrinage. The surface location was confirmed by flow cytometry, protease accessibility and whole-cell enzyme activity using a membrane-impermeable substrate. Three hundred and fifty units of whole-cell hydrolytic activity per gram dry cell mass were obtained when the enzyme was immobilized with a shorter INP anchoring motif (INPNC. The surface-immobilized TliA retained full enzyme activity in a two-phase water-isooctane reaction system after incubation at 37°C for 12 h, while the activity of the free form enzyme decreased to 65% of its initial value. Whole cells presenting immobilized TliA were shown to catalyze three representative lipase reactions: hydrolysis of olive oil, synthesis of triacylglycerol and chiral resolution. Conclusion In vivo surface immobilization of enzymes on solvent-resistant bacteria was demonstrated, and appears to be useful for a variety of whole-cell bioconversions in the presence of organic solvents.

Epigenetic Heterogeneity of B-Cell Lymphoma: Chromatin Modifiers

Science.gov (United States)

Hopp, Lydia; Nersisyan, Lilit; Löffler-Wirth, Henry; Arakelyan, Arsen; Binder, Hans

2015-01-01

We systematically studied the expression of more than fifty histone and DNA (de)methylating enzymes in lymphoma and healthy controls. As a main result, we found that the expression levels of nearly all enzymes become markedly disturbed in lymphoma, suggesting deregulation of large parts of the epigenetic machinery. We discuss the effect of DNA promoter methylation and of transcriptional activity in the context of mutated epigenetic modifiers such as EZH2 and MLL2. As another mechanism, we studied the coupling between the energy metabolism and epigenetics via metabolites that act as cofactors of JmjC-type demethylases. Our study results suggest that Burkitt’s lymphoma and diffuse large B-cell Lymphoma differ by an imbalance of repressive and poised promoters, which is governed predominantly by the activity of methyltransferases and the underrepresentation of demethylases in this regulation. The data further suggest that coupling of epigenetics with the energy metabolism can also be an important factor in lymphomagenesis in the absence of direct mutations of genes in metabolic pathways. Understanding of epigenetic deregulation in lymphoma and possibly in cancers in general must go beyond simple schemes using only a few modes of regulation. PMID:26506391

Epigenetic Heterogeneity of B-Cell Lymphoma: Chromatin Modifiers

Directory of Open Access Journals (Sweden)

Lydia Hopp

2015-10-01

Full Text Available We systematically studied the expression of more than fifty histone and DNA (demethylating enzymes in lymphoma and healthy controls. As a main result, we found that the expression levels of nearly all enzymes become markedly disturbed in lymphoma, suggesting deregulation of large parts of the epigenetic machinery. We discuss the effect of DNA promoter methylation and of transcriptional activity in the context of mutated epigenetic modifiers such as EZH2 and MLL2. As another mechanism, we studied the coupling between the energy metabolism and epigenetics via metabolites that act as cofactors of JmjC-type demethylases. Our study results suggest that Burkitt’s lymphoma and diffuse large B-cell Lymphoma differ by an imbalance of repressive and poised promoters, which is governed predominantly by the activity of methyltransferases and the underrepresentation of demethylases in this regulation. The data further suggest that coupling of epigenetics with the energy metabolism can also be an important factor in lymphomagenesis in the absence of direct mutations of genes in metabolic pathways. Understanding of epigenetic deregulation in lymphoma and possibly in cancers in general must go beyond simple schemes using only a few modes of regulation.

Glucose-Driven Fuel Cell Constructed from Enzymes and Filter Paper

Science.gov (United States)

Ge, Jun; Schirhagl, Romana; Zare, Richard N.

2011-01-01

A glucose-driven enzymatic filter-paper fuel cell is described. A strip of filter paper coated with carbon nanotubes and the glucose oxidase enzyme functions as the anode of the enzyme fuel cell. Another strip of filter paper coated with carbon nanotubes and the laccase enzyme functions as the cathode. Between the anode and the cathode, a third…

Peptide-modified PELCL electrospun membranes for regulation of vascular endothelial cells

Energy Technology Data Exchange (ETDEWEB)

Zhou, Fang [School of Materials Science and Engineering, Tianjin Key Laboratory of Composite and Functional Materials, Tianjin University, Tianjin 300072 (China); Jia, Xiaoling [Key Laboratory for Biomechanics and Mechanobiology of Ministry of Education, School of Biological Science and Medical Engineering, Beihang University, Beijing 100083 (China); Yang, Yang [School of Materials Science and Engineering, Tianjin Key Laboratory of Composite and Functional Materials, Tianjin University, Tianjin 300072 (China); Yang, Qingmao; Gao, Chao [Key Laboratory for Biomechanics and Mechanobiology of Ministry of Education, School of Biological Science and Medical Engineering, Beihang University, Beijing 100083 (China); Zhao, Yunhui [School of Materials Science and Engineering, Tianjin Key Laboratory of Composite and Functional Materials, Tianjin University, Tianjin 300072 (China); Fan, Yubo, E-mail: yubofan@buaa.edu.cn [Key Laboratory for Biomechanics and Mechanobiology of Ministry of Education, School of Biological Science and Medical Engineering, Beihang University, Beijing 100083 (China); National Research Center for Rehabilitation Technical Aids, Beijing 100176 (China); Yuan, Xiaoyan, E-mail: yuanxy@tju.edu.cn [School of Materials Science and Engineering, Tianjin Key Laboratory of Composite and Functional Materials, Tianjin University, Tianjin 300072 (China)

2016-11-01

The efficiency of biomaterials used in small vascular repair depends greatly on their ability to interact with vascular endothelial cells (VECs). Rapid endothelialization of the vascular grafts is a promising way to prevent thrombosis and intimal hyperplasia. In this work, modification of electrospun membranes of poly(ethylene glycol)-b-poly(L-lactide-co-ε-caprolactone) (PELCL) by three different peptides for regulation of VECs were studied in order to obtain ideal bioactive biomaterials as small diameter vascular grafts. QK (a mimetic peptide to vascular endothelial growth factor), Arg-Glu-Asp-Val (REDV, a specific adhesive peptide to VECs) and Val-Ala-Pro-Gly (VAPG, a specific adhesive peptide to vascular smooth muscle cells) were investigated. Surface properties of the modified membranes and the response of VECs were verified. It was found that protein adsorption and platelet adhesion were effectively suppressed with the introduction of QK, REDV or VAPG peptides on the PELCL electrospun membranes. Both QK- and REDV-modified electrospun membranes could accelerate the proliferation of VECs in the first 9 days, and the QK-modified electrospun membrane promoted cell proliferation more significantly than the REDV-modified one. The REDV-modified PELCL membrane was the most favorable for VECs adhesion than QK- and VAPG-modified membranes. It was suggested that QK- or REDV-modified PELCL electrospun membranes may have great potential applications in cardiovascular biomaterials for rapid endothelialization in situ. - Highlights: • A series of peptide-modified PELCL electrospun membranes were prepared. • Hemocompatibility of the membranes was greatly improved by the modification. • QK-modified PELCL membrane promoted VECs proliferation more significantly. • REDV-modified PELCL membrane was the most favorable for VEC adhesion.

Peptide-modified PELCL electrospun membranes for regulation of vascular endothelial cells

International Nuclear Information System (INIS)

Zhou, Fang; Jia, Xiaoling; Yang, Yang; Yang, Qingmao; Gao, Chao; Zhao, Yunhui; Fan, Yubo; Yuan, Xiaoyan

2016-01-01

The efficiency of biomaterials used in small vascular repair depends greatly on their ability to interact with vascular endothelial cells (VECs). Rapid endothelialization of the vascular grafts is a promising way to prevent thrombosis and intimal hyperplasia. In this work, modification of electrospun membranes of poly(ethylene glycol)-b-poly(L-lactide-co-ε-caprolactone) (PELCL) by three different peptides for regulation of VECs were studied in order to obtain ideal bioactive biomaterials as small diameter vascular grafts. QK (a mimetic peptide to vascular endothelial growth factor), Arg-Glu-Asp-Val (REDV, a specific adhesive peptide to VECs) and Val-Ala-Pro-Gly (VAPG, a specific adhesive peptide to vascular smooth muscle cells) were investigated. Surface properties of the modified membranes and the response of VECs were verified. It was found that protein adsorption and platelet adhesion were effectively suppressed with the introduction of QK, REDV or VAPG peptides on the PELCL electrospun membranes. Both QK- and REDV-modified electrospun membranes could accelerate the proliferation of VECs in the first 9 days, and the QK-modified electrospun membrane promoted cell proliferation more significantly than the REDV-modified one. The REDV-modified PELCL membrane was the most favorable for VECs adhesion than QK- and VAPG-modified membranes. It was suggested that QK- or REDV-modified PELCL electrospun membranes may have great potential applications in cardiovascular biomaterials for rapid endothelialization in situ. - Highlights: • A series of peptide-modified PELCL electrospun membranes were prepared. • Hemocompatibility of the membranes was greatly improved by the modification. • QK-modified PELCL membrane promoted VECs proliferation more significantly. • REDV-modified PELCL membrane was the most favorable for VEC adhesion.

Surface Modification for Microreactor Fabrication

Directory of Open Access Journals (Sweden)

Wladyslaw Torbicz

2006-04-01

Full Text Available In this paper, methods of surface modification of different supports, i.e. glass andpolymeric beads for enzyme immobilisation are described. The developed method ofenzyme immobilisation is based on Schiff’s base formation between the amino groups onthe enzyme surface and the aldehyde groups on the chemically modified surface of thesupports. The surface of silicon modified by APTS and GOPS with immobilised enzymewas characterised by atomic force microscopy (AFM, time-of-flight secondary ion massspectroscopy (ToF-SIMS and infrared spectroscopy (FTIR. The supports withimmobilised enzyme (urease were also tested in combination with microreactors fabricatedin silicon and Perspex, operating in a flow-through system. For microreactors filled withurease immobilised on glass beads (Sigma and on polymeric beads (PAN, a very high andstable signal (pH change was obtained. The developed method of urease immobilisationcan be stated to be very effective.

Visualization of red-ox proteins on the gold surface using enzymatic polypyrrole formation

International Nuclear Information System (INIS)

Ramanaviciene, A.; Kausaite-Minkstimiene, A.; Voronovic, J.; Ramanavicius, A.; Oztekin, Y.; Carac, G.; German, N.

2011-01-01

We describe a new method for the visualization of the activity of red-ox proteins on a gold interface. Glucose oxidase was selected as a model system. Surfaces were modified by adhesion of glucose oxidase on (a) electrochemically cleaned gold; (b) gold films modified with gold nanoparticles, (c) a gold surface modified with self-assembled monolayer, and (d) covalent immobilization of protein on the gold surface modified with a self-assembled monolayer. The simple optical method for the visualization of enzyme on the surfaces is based on the enzymatic formation of polypyrrole. The activity of the enzyme was quantified via enzymatic formation of polypyrrole, which was detected and investigated by quartz microbalance and amperometric techniques. The experimental data suggest that the enzymatic formation of the polymer may serve as a method to indicate the adhesion of active redox enzyme on such surfaces. (author)

Polymer microfilters with nanostructured surfaces for the culture of circulating cancer cells

Energy Technology Data Exchange (ETDEWEB)

Makarova, Olga V.; Adams, Daniel L.; Divan, Ralu; Rosenmann, Daniel; Zhu, Peixuan; Li, Shuhong; Amstutz, Platte; Tang, Cha-Mei

2016-09-01

There is a critical need to improve the accuracy of drug screening and testing through the development of in vitro culture systems that more effectively mimic the in vivo environment. Surface topographical features on the nanoscale level, in short nanotopography, effect the cell growth patterns, and hence affect cell function in culture. We report the preliminary results on the fabrication, and subsequent cellular growth, of nanoscale surface topography on polymer microfilters using cell lines as a precursor to circulating tumor cells (CTCs). To create various nanoscale features on the microfilter surface, we used reactive ion etching (RIE) with and without an etching mask. An anodized aluminum oxide (AAO) membrane fabricated directly on the polymer surface served as an etching mask. Polymer filters with a variety of modified surfaces were used to compare the effects on the culture of cancer cell lines in blank culture wells, with untreated microfilters or with RIE-treated microfilters. We then report the differences of cell shape, phenotype and growth patterns of bladder and glioblastoma cancer cell lines after isolation on the various types of material modifications. Our data suggest that RIE modified polymer filters can isolate model cell lines while retaining ell viability, and that the RIE filter modification allows T24 monolayering cells to proliferate as a structured cluster. Copyright 2016 The Authors. Published by Elsevier B.V. All rights reserved.

Surface functionalization of a polymeric lipid bilayer for coupling a model biological membrane with molecules, cells, and microstructures.

Science.gov (United States)

Morigaki, Kenichi; Mizutani, Kazuyuki; Saito, Makoto; Okazaki, Takashi; Nakajima, Yoshihiro; Tatsu, Yoshiro; Imaishi, Hiromasa

2013-02-26

We describe a stable and functional model biological membrane based on a polymerized lipid bilayer with a chemically modified surface. A polymerized lipid bilayer was formed from a mixture of two diacetylene-containing phospholipids, 1,2-bis(10,12-tricosadiynoyl)-sn-glycero-3-phosphocholine (DiynePC) and 1,2-bis(10,12-tricosadiynoyl)-sn-glycero-3-phosphoethanolamine (DiynePE). DiynePC formed a stable bilayer structure, whereas the ethanolamine headgroup of DiynePE enabled functional molecules to be grafted onto the membrane surface. Copolymerization of DiynePC and DiynePE resulted in a robust bilayer. Functionalization of the polymeric bilayer provided a route to a robust and biomimetic surface that can be linked with biomolecules, cells, and three-dimensional (3D) microstructures. Biotin and peptides were grafted onto the polymeric bilayer for attaching streptavidin and cultured mammalian cells by molecular recognition, respectively. Nonspecific adsorption of proteins and cells on polymeric bilayers was minimum. DiynePE was also used to attach a microstructure made of an elastomer (polydimethylsiloxan: PDMS) onto the membrane, forming a confined aqueous solution between the two surfaces. The microcompartment enabled us to assay the activity of a membrane-bound enzyme (cyochrome P450). Natural (fluid) lipid bilayers were incorporated together with membrane-bound proteins by lithographically polymerizing DiynePC/DiynePE bilayers. The hybrid membrane of functionalized polymeric bilayers and fluid bilayers offers a novel platform for a wide range of biomedical applications including biosensor, bioassay, cell culture, and cell-based assay.

Immobilization of β-galactosidase from Kluyveromyces lactis onto polymeric membrane surfaces: effect of surface characteristics.

Science.gov (United States)

Güleç, Hacı Ali

2013-04-01

The aim of this study was to investigate the effects of surface characteristics of plain and plasma modified cellulose acetate (CA) membranes on the immobilization yield of β-galactosidases from Kluyveromyces lactis (KLG) and its galacto-oligosaccharide (GOS) yield, respectively. Low pressure plasma treatments involving oxygen plasma activation, plasma polymerization (PlsP) of ethylenediamine (EDA) and PlsP of 2-mercaptoethanol were used to modify plain CA membrane surfaces. KLG enzyme was immobilized onto plain and oxygen plasma treated membrane surfaces by simple adsorption. Oxygen plasma activation increased the hydrophylicity of CA membrane surfaces and it improved the immobilization yield of the enzyme by 42%. KLG enzyme was also immobilized onto CA membrane surfaces through amino groups created by PlsP of EDA via covalent binding. Plasma action at 60W plasma power and 15 min. exposure time improved the amount of membrane bounded enzyme by 3.5-fold. The enrichment of the amount of amino groups via polyethyleneimine (PEI) addition enhanced this increase from 3.5-fold to 4.5-fold. Although high enzyme loading was achived (65-83%), both of the methods dramatically decreased the enzyme activity (11-12%) and GOS yield due to probably negative effects of active amino groups. KLG enzyme was more effectively immobilized onto thiolated CA membrane surface created by PlsP of 2-mercaptoethanol with high immobilization yield (70%) and especially high enzyme activity (46%). Immobilized enzymes on the CA membranes treated by PlsP were successively reutilized for 5-8 cycles at 25°C and enzymatic derivatives retained approximately 75-80% of their initial activites at the end of the reactions. Copyright © 2012 Elsevier B.V. All rights reserved.

Proteasomal targeting and minigene repetition improve cell-surface presentation of a transfected, modified melanoma tumour antigen

DEFF Research Database (Denmark)

Rasmussen, A B; Zocca, M-B; Bonefeld, C M

2004-01-01

Melanoma antigen recognized by T cell 1 (MART-1) is regarded as a candidate peptide for vaccination against malignant melanoma, and it is of importance to develop strategies to improve the vaccine-elicited T-cell activation towards MART-1. T-cell activation is, among other determinants, dependent...... on the density of specific major histocompatibility complex-peptide complexes on the surface of the antigen-presenting cell. In this study, we explored the cell-surface presentation of a substituted MART-1 peptide encoded by transfected minigenes. We investigated the potential of proteasomal targeting compared...... to non-proteasomal targeting of the epitope to increase its cell-surface presentation. Furthermore, we explored the potential of incorporating multiple minigenes instead of one to increase cell-surface presentation. We show that both proteasomal targeting and repetition of the minigene increase cell...

Discovery and Characterization of Enzymes for Degradation of Xyloglucan and Extensin

DEFF Research Database (Denmark)

Feng, Tao; Mikkelsen, Jørn Dalgaard

before the residual polymers are used in the bioethanol production. Therefore, mono-component, substrate-specific enzymes that could selectively degrade or modify plant cell wall components are required. In this PhD study, three enzymes, including two xyloglucan-specific endoglucanases and one...

Surface Plasmon Resonance based sensing of lysozyme in serum on Micrococcus lysodeikticus-modified graphene oxide surfaces.

Science.gov (United States)

Vasilescu, Alina; Gáspár, Szilveszter; Gheorghiu, Mihaela; David, Sorin; Dinca, Valentina; Peteu, Serban; Wang, Qian; Li, Musen; Boukherroub, Rabah; Szunerits, Sabine

2017-03-15

Lysozyme is an enzyme found in biological fluids, which is upregulated in leukemia, renal diseases as well as in a number of inflammatory gastrointestinal diseases. We present here the development of a novel lysozyme sensing concept based on the use of Micrococcus lysodeikticus whole cells adsorbed on graphene oxide (GO)-coated Surface Plasmon Resonance (SPR) interfaces. M. lysodeikticus is a typical enzymatic substrate for lysozyme. Unlike previously reported sensors which are based on the detection of lysozyme through bioaffinity interactions, the bioactivity of lysozyme will be used here for sensing purposes. Upon exposure to lysozyme containing serum, the integrity of the bacterial cell wall is affected and the cells detach from the GO based interfaces, causing a characteristic decrease in the SPR signal. This allows sensing the presence of clinically relevant concentrations of lysozyme in undiluted serum samples. Copyright © 2016 Elsevier B.V. All rights reserved.

Hydrophobicity of electron beam modified surface of hydroxyapatite films

Energy Technology Data Exchange (ETDEWEB)

Gregor, M., E-mail: gregor@fmph.uniba.sk [Department of Experimental Physics, Faculty of Mathematics, Physics and Informatics, Comenius University, 84248 Bratislava (Slovakia); Plecenik, T. [Department of Experimental Physics, Faculty of Mathematics, Physics and Informatics, Comenius University, 84248 Bratislava (Slovakia); Tofail, S.A.M. [Materials & Surface Science Institute, University of Limerick, Limerick (Ireland); Zahoran, M.; Truchly, M. [Department of Experimental Physics, Faculty of Mathematics, Physics and Informatics, Comenius University, 84248 Bratislava (Slovakia); Vargova, M. [Department of Inorganic Chemistry, Faculty of Natural Sciences, Comenius University, 84215 Bratislava (Slovakia); Laffir, F. [Materials & Surface Science Institute, University of Limerick, Limerick (Ireland); Plesch, G. [Department of Inorganic Chemistry, Faculty of Natural Sciences, Comenius University, 84215 Bratislava (Slovakia); Kus, P.; Plecenik, A. [Department of Experimental Physics, Faculty of Mathematics, Physics and Informatics, Comenius University, 84248 Bratislava (Slovakia)

2015-05-15

Highlights: • Surface potential of hydroxyapatite films were modified by focused electron beam. • Micron-sized domains of modified surface potential were created. • Wettability and surface free energy of the irradiated areas was studied. • Possible mechanisms of increased surface hydrophobicity are discussed. - Abstract: Arrays of micron-sized domains of modified surface potential were created on hydroxyapatite films by mid-energy (20 keV) electron beam irradiation available in a laboratory scanning electron microscope. The dosage of electron beam was varied between 10{sup −3} and 10{sup 3} μC/cm{sup 2} to inject charge into the film surface. Contrary to the conventional electrowetting theory, the dosage of injected charge used in creating such microdomains caused a gradual increase of the water contact angle from 57° to 93° due to the elimination of the polar component of the surface free energy. Surface contamination by carbonaceous species can be held only partially responsible for such behavior at lower dosage of electron beam. A transfer of free surface charge to water and an electron beam induced disruption of polar orientation of OH ions have been attributed to be influencial factors in the overall dewetting behavior.

Development of a thiol-ene based screening platform for enzyme immobilization demonstrated using horseradish peroxidase

DEFF Research Database (Denmark)

Hoffmann, Christian; Pinelo, Manuel; Woodley, John

2017-01-01

Efficient immobilization of enzymes on support surfaces requires an exact match between the surface chemistry and the specific enzyme. A successful match would normally be identified through time consuming screening of conventional resins in multiple experiments testing individual immobilization...... strategies. In this study we present a versatile strategy that largely expands the number of possible surface functionalities for enzyme immobilization in a single, generic platform. The combination of many individual surface chemistries and thus immobilization methods in one modular system permits faster...... functionalization by thiol-ene chemistry (TEC) resulted in the formation of a functional monolayer in each well, whereas, polymer surface grafts were introduced through surface chain transfer free radical polymerization (SCT-FRP). Enzyme immobilization on the modified surfaces was evaluated by using a rhodamine...

Radiation-induced heterogeneity of chymotrypsin of mus musculus. On the characterization of structurally and functionally in vitro modified enzyme forms

International Nuclear Information System (INIS)

Amneus, H.

1976-01-01

The distribution of in vitro induced 60 Co-γ (structural heterogeneity of mouse chymotrypsin has been studied in terms of molecular weight, catalytic activity and net charge distribution. It was found that the enzyme stucture, with retained molecular weight, could partly accumulate structural changes subsequently not leading to modification of catalytic properties. Loss of petide fragments (0 < Mw (lt 6000) the enzyme showed native function but also modified as well as total loss of function. Further loss of peptide fragments results in modified function and total loss of function. These results indicate the capability of the enzyme to accumulate in vitro changes partly without a total loss of function. (author)

Photovoltaic characterization of hybrid solar cells using surface modified TiO{sub 2} nanoparticles and poly(3-hexyl)thiophene

Energy Technology Data Exchange (ETDEWEB)

Guenes, Serap [Yildiz Technical University, Faculty of Arts and Science, Department of Physics, Davutpasa Campus, 34220, Esenler, Istanbul (Turkey); Marjanovic, Nenad [Plastic electronic GmbH, Rappetsederweg 28, A-4040 Linz (Austria); Nedeljkovic, Jovan M [Vinca Institute of Nuclear Sciences, PO Box 522, 11001 Belgrade (Serbia); Sariciftci, Niyazi Serdar [Linz Institute for Organic Solar Cells (LIOS), Physical Chemistry, Johannes Kepler University Linz, Altenberger Strasse 69, A-4040, Linz (Austria)], E-mail: sgunes@yildiz.edu.tr

2008-10-22

We report on the photovoltaic performance of bulk heterojunction solar cells using novel nanoparticles of 6-palmitate ascorbic acid surface modified TiO{sub 2} as an electron acceptor embedded into the donor poly(3-hexyl)thiophene (P3HT) matrix. Devices were fabricated by using P3HT with varying amounts of red TiO{sub 2} nanoparticles (1:1, 1:2, 1:3 w-w ratio). The devices were characterized by measuring current-voltage characteristics under simulated AM 1.5 conditions. Incident photon to current efficiency (IPCE) was spectrally resolved. The nanoscale morphology of such organic/inorganic hybrid blends was also investigated using atomic force microscopy (AFM)

Yeast cell surface display for lipase whole cell catalyst and its applications

Energy Technology Data Exchange (ETDEWEB)

Liu, Yun; Zhang, Rui; Lian, Zhongshuai; Wang, Shihui; Wright, Aaron T.

2014-08-01

The cell surface display technique allows for the expression of target proteins or peptides on the microbial cell surface by fusing an appropriate protein as an anchoring motif. Yeast display systems, such as Pichia pastoris, Yarowia lipolytica and Saccharomyces cerevisiae, are ideal, alternative and extensive display systems with the advantage of simple genetic manipulation and post-translational modification of expressed heterologous proteins. Engineered yeasts show high performance characteristics and variant utilizations. Herein, we comprehensively summarize the variant factors affecting lipase whole cell catalyst activity and display efficiency, including the structure and size of target proteins, screening anchor proteins, type and chain length of linkers, and the appropriate matching rules among the above-mentioned display units. Furthermore, we also address novel approaches to enhance stability and activity of recombinant lipases, such as VHb gene co-expression, multi-enzyme co-display technique, and the micro-environmental interference and self-assembly techniques. Finally, we represent the variety of applications of whole cell surface displayed lipases on yeast cells in non-aqueous phases, including synthesis of esters, PUFA enrichment, resolution of chiral drugs, organic synthesis and biofuels. We demonstrate that the lipase surface display technique is a powerful tool for functionalizing yeasts to serve as whole cell catalysts, and increasing interest is providing an impetus for broad application of this technique.

Improvement in Saccharification Yield of Mixed Rumen Enzymes by Identification of Recalcitrant Cell Wall Constituents Using Enzyme Fingerprinting.

Science.gov (United States)

Badhan, Ajay; Wang, Yu-Xi; Gruninger, Robert; Patton, Donald; Powlowski, Justin; Tsang, Adrian; McAllister, Tim A

2015-01-01

Identification of recalcitrant factors that limit digestion of forages and the development of enzymatic approaches that improve hydrolysis could play a key role in improving the efficiency of meat and milk production in ruminants. Enzyme fingerprinting of barley silage fed to heifers and total tract indigestible fibre residue (TIFR) collected from feces was used to identify cell wall components resistant to total tract digestion. Enzyme fingerprinting results identified acetyl xylan esterases as key to the enhanced ruminal digestion. FTIR analysis also suggested cross-link cell wall polymers as principal components of indigested fiber residues in feces. Based on structural information from enzymatic fingerprinting and FTIR, enzyme pretreatment to enhance glucose yield from barley straw and alfalfa hay upon exposure to mixed rumen-enzymes was developed. Prehydrolysis effects of recombinant fungal fibrolytic hydrolases were analyzed using microassay in combination with statistical experimental design. Recombinant hemicellulases and auxiliary enzymes initiated degradation of plant structural polysaccharides upon application and improved the in vitro saccharification of alfalfa and barley straw by mixed rumen enzymes. The validation results showed that microassay in combination with statistical experimental design can be successfully used to predict effective enzyme pretreatments that can enhance plant cell wall digestion by mixed rumen enzymes.

Quantitative kinetics of proteolytic enzymes determined by a surface concentration-based assay using peptide arrays.

Science.gov (United States)

Jung, Se-Hui; Kong, Deok-Hoon; Park, Seoung-Woo; Kim, Young-Myeong; Ha, Kwon-Soo

2012-08-21

Peptide arrays have emerged as a key technology for drug discovery, diagnosis, and cell biology. Despite the promise of these arrays, applications of peptide arrays to quantitative analysis of enzyme kinetics have been limited due to the difficulty in obtaining quantitative information of enzymatic reaction products. In this study, we developed a new approach for the quantitative kinetics analysis of proteases using fluorescence-conjugated peptide arrays, a surface concentration-based assay with solid-phase peptide standards using dry-off measurements, and compared it with an applied concentration-based assay. For fabrication of the peptide arrays, substrate peptides of cMMP-3, caspase-3, caspase-9, and calpain-1 were functionalized with TAMRA and cysteine, and were immobilized onto amine-functionalized arrays using a heterobifunctional linker, N-[γ-maleimidobutyloxy]succinimide ester. The proteolytic activities of the four enzymes were quantitatively analyzed by calculating changes induced by enzymatic reactions in the concentrations of peptides bound to array surfaces. In addition, this assay was successfully applied for calculating the Michaelis constant (K(m,surf)) for the four enzymes. Thus, this new assay has a strong potential for use in the quantitative evaluation of proteases, and for drug discovery through kinetics studies including the determination of K(m) and V(max).

Evaluation of modified stainless steel surfaces targeted to reduce biofilm formation by common milk sporeformers.

Science.gov (United States)

Jindal, Shivali; Anand, Sanjeev; Huang, Kang; Goddard, Julie; Metzger, Lloyd; Amamcharla, Jayendra

2016-12-01

The development of bacterial biofilms on stainless steel (SS) surfaces poses a great threat to the quality of milk and other dairy products as the biofilm-embedded bacteria can survive thermal processing. Established biofilms offer cleaning challenges because they are resistant to most of the regular cleaning protocols. Sporeforming thermoduric organisms entrapped within biofilm matrix can also form heat-resistant spores, and may result in a long-term persistent contamination. The main objective of this study was to evaluate the efficacy of different nonfouling coatings [AMC 18 (Advanced Materials Components Express, Lemont, PA), Dursan (SilcoTek Corporation, Bellefonte, PA), Ni-P-polytetrafluoroethylene (PTFE, Avtec Finishing Systems, New Hope, MN), and Lectrofluor 641 (General Magnaplate Corporation, Linden, NJ)] on SS plate heat exchanger surfaces, to resist the formation of bacterial biofilms. It was hypothesized that modified SS surfaces would promote a lesser amount of deposit buildup and bacterial adhesion as compared with the native SS surface. Vegetative cells of aerobic sporeformers, Geobacillus stearothermophilus (ATCC 15952), Bacillus licheniformis (ATCC 6634), and Bacillus sporothermodurans (DSM 10599), were used to study biofilm development on the modified and native SS surfaces. The adherence of these organisms, though influenced by surface energy and hydrophobicity, exhibited no apparent relation with surface roughness. The Ni-P-PTFE coating exhibited the least bacterial attachment and milk solid deposition, and hence, was the most resistant to biofilm formation. Scanning electron microscopy, which was used to visualize the extent of biofilm formation on modified and native SS surfaces, also revealed lower bacterial attachment on the Ni-P-PTFE as compared with the native SS surface. This study thus provides evidence of reduced biofilm formation on the modified SS surfaces. Copyright © 2016 American Dairy Science Association. Published by Elsevier

Optimizing Immobilized Enzyme Performance in Cell-Free Environments to Produce Liquid Fuels

Energy Technology Data Exchange (ETDEWEB)

Belfort, Georges [Rensselaer Polytechnic Inst., Troy, NY (United States). Dept. of Chemical and Biological Engineering; Grimaldi, Joseph J. [Rensselaer Polytechnic Inst., Troy, NY (United States). Dept. of Chemical and Biological Engineering

2015-01-27

Limitations on biofuel production using cell culture (Escherichia coli, Clostridium, Saccharomyces cerevisiae, brown microalgae, blue-green algae and others) include low product (alcohol) concentrations (≤0.2 vol%) due to feedback inhibition, instability of cells, and lack of economical product recovery processes. To overcome these challenges, an alternate simplified biofuel production scheme was tested based on a cell-free immobilized enzyme system. Using this cell free system, we were able to obtain about 2.6 times higher concentrations of iso-butanol using our non-optimized system as compared with live cell systems. This process involved two steps: (i) converts acid to aldehyde using keto-acid decarboxylase (KdcA), and (ii) produces alcohol from aldehyde using alcohol dehydrogenase (ADH) with a cofactor (NADH) conversion from inexpensive formate using a third enzyme, formate dehydrogenase (FDH). To increase stability and conversion efficiency with easy separations, the first two enzymes were immobilized onto methacrylate resin. Fusion proteins of labile KdcA (fKdcA) were expressed to stabilize the covalently immobilized KdcA. Covalently immobilized ADH exhibited long-term stability and efficient conversion of aldehyde to alcohol over multiple batch cycles without fusions. High conversion rates and low protein leaching were achieved by covalent immobilization of enzymes on methacrylate resin. The complete reaction scheme was demonstrated by immobilizing both ADH and fKdcA and using FDH free in solution. The new system without in situ removal of isobutanol achieved a 55% conversion of ketoisovaleric acid to isobutanol at a concentration of 0.5 % (v/v). Further increases in titer will require continuous removal of the isobutanol using our novel brush membrane system that exhibits a 1.5 fold increase in the separation factor of isobutanol from water versus that obtained for commercial silicone rubber membranes. These bio-inspired brush membranes are based on the

Correction of anemia in uremic mice by genetically modified peritoneal mesothelial cells.

Science.gov (United States)

Einbinder, Tom; Sufaro, Yuval; Yusim, Igor; Byk, Gerardo; Passlick-Deetjen, Jutta; Chaimovitz, Cidio; Douvdevani, Amos

2003-06-01

During peritoneal dialysis, mesothelial cells become detached from the peritoneum and accumulate in the dialysate. Our aim was to evaluate the potential of peritoneal effluent (PF)-derived human peritoneal mesothelial cells (HPMC) as target for gene therapy. We used erythropoietin (EPO) as our target gene. Various extracellular matrixes (ECM) were tested for optimal adhesion and growth of HPMC. The EPO gene was introduced to mouse peritoneal mesothelial cells (MPMC) and HPMC by transfection or retroviral transduction. EPO secretion from PMC was measured by enzyme-linked immunosorbent assay (ELISA) and by the TF-1 cell proliferation assay. We performed intraperitoneal or intramuscular transplantations of the genetically modified cells into regular or 5/6 nephrectomized Balb/c mice and nude mice. Finally, we measured serum EPO and hematocrit levels. ECM-coated plates provided up to sixfold increase in the efficiency of PMC isolation from PF. Gelatin coated dishes (20 microg/cm2) were found optimal for isolation of PF-HPMC. RPR-120535 liposome was found to be best for PMC transduction. In vitro studies showed EPO secretion from modified HPMC over 6 months. Intraperitoneal transplantation aided with collagen matrix was the most effective. EPO, in MPMC transplanted mice, was detected up to 3 weeks (peak at 13 +/- 1 mIU/mL), and anemia of uremic mice was corrected (35.3 +/- 0.9 mIU/mL to 41.9 +/- 1.1 mIU/mL). PF-HPMC can be considered as an appropriate target for gene therapy since these cells can be efficiently isolated, modified, and transplanted. Nevertheless, implantation techniques in the peritoneum should be directed at obtaining longer duration of transgene expression in vivo, and means should be developed for enabling regulated expression of the gene.

Dual-responsive surfaces modified with phenylboronic acid-containing polymer brush to reversibly capture and release cancer cells.

Science.gov (United States)

Liu, Hongliang; Li, Yingying; Sun, Kang; Fan, Junbing; Zhang, Pengchao; Meng, Jingxin; Wang, Shutao; Jiang, Lei

2013-05-22

Artificial stimuli-responsive surfaces that can mimic the dynamic function of living systems have attracted much attention. However, there exist few artificial systems capable of responding to dual- or multistimulation as the natural system does. Herein, we synthesize a pH and glucose dual-responsive surface by grafting poly(acrylamidophenylboronic acid) (polyAAPBA) brush from aligned silicon nanowire (SiNW) array. The as-prepared surface can reversibly capture and release targeted cancer cells by precisely controlling pH and glucose concentration, exhibiting dual-responsive AND logic. In the presence of 70 mM glucose, the surface is pH responsive, which can vary from a cell-adhesive state to a cell-repulsive state by changing the pH from 6.8 to 7.8. While keeping the pH at 7.8, the surface becomes glucose responsive--capturing cells in the absence of glucose and releasing cells by adding 70 mM glucose. Through simultaneously changing the pH and glucose concentration from pH 6.8/0 mM glucose to pH 7.8/70 mM glucose, the surface is dual responsive with the capability to switch between cell capture and release for at least 5 cycles. The cell capture and release process on this dual-responsive surface is noninvasive with cell viability higher than 95%. Moreover, topographical interaction between the aligned SiNW array and cell protrusions greatly amplifies the responsiveness and accelerates the response rate of the dual-responsive surface between cell capture and release. The responsive mechanism of the dual-responsive surface is systematically studied using a quartz crystal microbalance, which shows that the competitive binding between polyAAPBA/sialic acid and polyAAPBA/glucose contributes to the dual response. Such dual-responsive surface can significantly impact biomedical and biological applications including cell-based diagnostics, in vivo drug delivery, etc.

Characterization of electrochemically modified polycrystalline platinum surfaces

Energy Technology Data Exchange (ETDEWEB)

Krebs, L.C.; Ishida, Takanobu.

1991-12-01

The characterization of electrochemically modified polycrystalline platinum surfaces has been accomplished through the use of four major electrochemical techniques. These were chronoamperometry, chronopotentiommetry, cyclic voltammetry, and linear sweep voltammetry. A systematic study on the under-potential deposition of several transition metals has been performed. The most interesting of these were: Ag, Cu, Cd, and Pb. It was determined, by subjecting the platinum electrode surface to a single potential scan between {minus}0.24 and +1.25 V{sub SCE} while stirring the solution, that the electrocatalytic activity would be regenerated. As a consequence of this study, a much simpler method for producing ultra high purity water from acidic permanganate has been developed. This method results in water that surpasses the water produced by pyrocatalytic distillation. It has also been seen that the wettability of polycrystalline platinum surfaces is greatly dependent on the quantity of oxide present. Oxide-free platinum is hydrophobic and gives a contact angle in the range of 55 to 62 degrees. We have also modified polycrystalline platinum surface with the electrically conducting polymer poly-{rho}-phenylene. This polymer is very stable in dilute sulfuric acid solutions, even under applied oxidative potentials. It is also highly resistant to electrochemical hydrogenation. The wettability of the polymer modified platinum surface is severely dependent on the choice of supporting electrolyte chosen for the electrochemical polymerization. Tetraethylammonium tetrafluoroborate produces a film that is as hydrophobic as Teflon, whereas tetraethylammonium perchlorate produces a film that is more hydrophilic than oxide-free platinum.

Characterization of electrochemically modified polycrystalline platinum surfaces

Energy Technology Data Exchange (ETDEWEB)

Krebs, Leonard C. [State Univ. of New York (SUNY), Stony Brook, NY (United States); Ishida, Takanobu [State Univ. of New York (SUNY), Stony Brook, NY (United States)

1991-12-01

The characterization of electrochemically modified polycrystalline platinum surfaces has been accomplished through the use of four major electrochemical techniques. These were chronoamperometry, chronopotentiommetry, cyclic voltammetry, and linear sweep voltammetry. A systematic study on the under-potential deposition of several transition metals has been performed. The most interesting of these were: Ag, Cu, Cd, and Pb. It was determined, by subjecting the platinum electrode surface to a single potential scan between -0.24 and +1.25 V_SCE while stirring the solution, that the electrocatalytic activity would be regenerated. As a consequence of this study, a much simpler method for producing ultra high purity water from acidic permanganate has been developed. This method results in water that surpasses the water produced by pyrocatalytic distillation. It has also been seen that the wettability of polycrystalline platinum surfaces is greatly dependent on the quantity of oxide present. Oxide-free platinum is hydrophobic and gives a contact angle in the range of 55 to 62 degrees. We have also modified polycrystalline platinum surface with the electrically conducting polymer poly-ρ-phenylene. This polymer is very stable in dilute sulfuric acid solutions, even under applied oxidative potentials. It is also highly resistant to electrochemical hydrogenation. The wettability of the polymer modified platinum surface is severely dependent on the choice of supporting electrolyte chosen for the electrochemical polymerization. Tetraethylammonium tetrafluoroborate produces a film that is as hydrophobic as Teflon, whereas tetraethylammonium perchlorate produces a film that is more hydrophilic than oxide-free platinum.

Improving cytoactive of endothelial cell by introducing fibronectin to the surface of poly L-Lactic acid fiber mats via dopamine

Energy Technology Data Exchange (ETDEWEB)

Yang, Wufeng; Zhang, Xiazhi; Wu, Keke; Liu, Xiaoyan; Jiao, Yanpeng, E-mail: tjiaoyp@jnu.edu.cn; Zhou, Changren

2016-12-01

A simple but straightforward approach was reported to prepare fiber mats modified with fibronectin (Fn) protein for endothelial cells activity study. Based on the self-polymerization and strong adhesion feature of dopamine, poly L-Lactic acid (PLLA) fibers mat was modified via simply immersing them into dopamine solution for 16 h. Subsequently, Fn was immobilized onto the fiber mats surface by the coupling reactive polydopamine (PDA) layer and Fn. Attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy and X-ray photoelectron spectroscopy (XPS) were used to determine the chemical compositions of fiber mats surface, which confirmed the successful immobilization of PDA and Fn molecules on the fiber surface. Scanning electronic microscopy (SEM) was used to observe the surface morphology changes after modification with PDA and Fn. The data of water contact angle showed that the hydrophilicity of the fiber mats was improved after surface modification. The data of in vitro cell culture proved that the PDA and Fn modified surface significantly enhanced the adhesion, proliferation and cell activity of endothelial cells on the fiber mats. And the release of tumor necrosis factor-α (TNF-α) by endothelial cells on the modified surface was suppressed compared to that on culture plate and PLLA film at 2 and 4 days, while the secretion of interleukin-1β (IL-1β) was increased compared to that on culture plate and PLLA film at 2 days. - Highlights: • Fibronectin (Fn) was grafted on PLLA fiber surface mediated by polydopamine coating. • Fn modified PLLA fiber enhanced the adhesion, proliferation of endothelial cells. • Fn and polydopamine modified PLLA fiber could adjust the release of inflammatory factor.

An amperometric uric acid biosensor based on Bis[sulfosuccinimidyl] suberate crosslinker/3-aminopropyltriethoxysilane surface modified ITO glass electrode

International Nuclear Information System (INIS)

Ahuja, Tarushee; Rajesh; Kumar, Devendra; Tanwar, Vinod Kumar; Sharma, Vikash; Singh, Nahar; Biradar, Ashok M.

2010-01-01

A label free, amperometric uric acid biosensor is described by immobilizing enzyme uricase through a self assembled monolayer (SAM) of 3-aminopropyltriethoxysilane (APTES) using a crosslinker, Bis[sulfosuccinimidyl]suberate (BS 3 ) on an indium-tin-oxide (ITO) coated glass plate. The biosensor (uricase/BS 3 /APTES/ITO) was characterized by, scanning electron microscopy (SEM), atomic force microscopy (AFM) and electrochemical techniques. Chronoamperometric response was measured as a function of uric acid concentration in aqueous solution (pH 7.4). The biosensor shows a linear response over a concentration range of 0.05 to 0.58 mM with a sensitivity of 39.35 μA mM -1 . The response time is 50 s reaching to a 95% steady state current value and about 90% of enzyme activity is retained for about 7 weeks. These results indicate an efficient binding of enzyme with the crosslinker over the surface of APTES modified ITO glass plates, which leads to an improved sensitivity and shelf life of the biosensor.

Modified gold surfaces by 6-(ferrocenyl)hexanethiol/dendrimer/gold nanoparticles as a platform for the mediated biosensing applications

Energy Technology Data Exchange (ETDEWEB)

Karadag, Murat; Geyik, Caner; Demirkol, Dilek Odaci [Ege University, Faculty of Science, Biochemistry Department, 35100 Bornova-Izmir (Turkey); Ertas, F. Nil [Ege University, Faculty of Science, Chemistry Department, 35100, Bornova-Izmir (Turkey); Timur, Suna, E-mail: suna.timur@ege.edu.tr [Ege University, Faculty of Science, Biochemistry Department, 35100 Bornova-Izmir (Turkey)

2013-03-01

An electrochemical biosensor mediated by using 6-(Ferrocenyl) hexanethiol (FcSH) was fabricated by construction of gold nanoparticles (AuNPs) on the surface of polyamidoamine dendrimer (PAMAM) modified gold electrode. Glucose oxidase (GOx) was used as a model enzyme and was immobilized onto the gold surface forming a self assembled monolayer via FcSH and cysteamine. Cyclic voltammetry and amperometry were used for the characterization of electrochemical response towards glucose substrate. Following the optimization of medium pH, enzyme loading, AuNP and FcSH amount, the linear range for the glucose was studied and found as 1.0 to 5.0 mM with the detection limit (LOD) of 0.6 mM according to S/N = 3. Finally, the proposed Au/AuNP/(FcSH + Cyst)/PAMAM/GOx biosensor was successfully applied for the glucose analysis in beverages, and the results were compared with those obtained by HPLC. Highlights: Black-Right-Pointing-Pointer Immobilized mediator in SAM layer and dendrimeric structure to expand surface area. Black-Right-Pointing-Pointer Au nanoparticles for enhanced electron transfer. Black-Right-Pointing-Pointer Satisfactory Limit of Detection with 0.6 mM.

From mannan to bioethanol: cell surface co-display of β-mannanase and β-mannosidase on yeast Saccharomyces cerevisiae.

Science.gov (United States)

Ishii, Jun; Okazaki, Fumiyoshi; Djohan, Apridah Cameliawati; Hara, Kiyotaka Y; Asai-Nakashima, Nanami; Teramura, Hiroshi; Andriani, Ade; Tominaga, Masahiro; Wakai, Satoshi; Kahar, Prihardi; Yopi; Prasetya, Bambang; Ogino, Chiaki; Kondo, Akihiko

2016-01-01

Mannans represent the largest hemicellulosic fraction in softwoods and also serve as carbohydrate stores in various plants. However, the utilization of mannans as sustainable resources has been less advanced in sustainable biofuel development. Based on a yeast cell surface-display technology that enables the immobilization of multiple enzymes on the yeast cell walls, we constructed a recombinant Saccharomyces cerevisiae strain that co-displays β-mannanase and β-mannosidase; this strain is expected to facilitate ethanol fermentation using mannan as a biomass source. Parental yeast S. cerevisiae assimilated mannose and glucose as monomeric sugars, producing ethanol from mannose. We constructed yeast strains that express tethered β-mannanase and β-mannosidase; co-display of the two enzymes on the cell surface was confirmed by immunofluorescence staining and enzyme activity assays. The constructed yeast cells successfully hydrolyzed 1,4-β-d-mannan and produced ethanol by assimilating the resulting mannose without external addition of enzymes. Furthermore, the constructed strain produced ethanol from 1,4-β-d-mannan continually during the third batch of repeated fermentation. Additionally, the constructed strain produced ethanol from ivory nut mannan; ethanol yield was improved by NaOH pretreatment of the substrate. We successfully displayed β-mannanase and β-mannosidase on the yeast cell surface. Our results clearly demonstrate the utility of the strain co-displaying β-mannanase and β-mannosidase for ethanol fermentation from mannan biomass. Thus, co-tethering β-mannanase and β-mannosidase on the yeast cell surface provides a powerful platform technology for yeast fermentation toward the production of bioethanol and other biochemicals from lignocellulosic materials containing mannan components.

In vitro cytotoxicity and phototoxicity of surface-modified gold nanoparticles associated with neutral red as a potential drug delivery system in phototherapy

Energy Technology Data Exchange (ETDEWEB)

Verissimo, Tanira V. [School of Health Sciences, University of Brasilia, Brasilia (Brazil); Laboratory of Photochemistry and Nanobiotechnology, Faculty of Ceilandia, University of Brasilia, Brasilia (Brazil); Santos, Naiara T. [School of Health Sciences, University of Brasilia, Brasilia (Brazil); Silva, Jaqueline R.; Azevedo, Ricardo B. [Institute of Biological Sciences, University of Brasilia, Brasilia (Brazil); Gomes, Anderson J., E-mail: ajgomes@unb.br [School of Health Sciences, University of Brasilia, Brasilia (Brazil); Laboratory of Photochemistry and Nanobiotechnology, Faculty of Ceilandia, University of Brasilia, Brasilia (Brazil); Lunardi, Claure N., E-mail: clunardi@unb.br [School of Health Sciences, University of Brasilia, Brasilia (Brazil); Laboratory of Photochemistry and Nanobiotechnology, Faculty of Ceilandia, University of Brasilia, Brasilia (Brazil)

2016-08-01

The surface of gold nanoparticles (AuNP) was modified, improving their interaction with neutral red (NR), by using sodium thioglycolate (TGA) as a covering agent. The resulting NR-AuNPTGA system was evaluated as a potential drug delivery system for photodynamic therapy (PDT). The associations of NR with the gold nanoparticles were evaluated using UV-vis spectrometry and measurement of their zeta potential and size distribution. The toxicity and phototoxicity of NR, AuNPTGA and NR-AuNPTGA were evaluated in NIH-3T3 fibroblast and 4T1 tumor cell lines. The compounds NR and NR-AuNPTGA induced toxicity in 4T1 tumor cells and NIH-3T3 fibroblasts under visible light irradiation. Modification of the surface of AuNP with TGA prevented nanoparticle aggregation and allowed greater association with NR molecules than for naked AuNP. The photosensitizer (PS) characteristics were not affected by its association with the modified surface of the gold nanoparticles, leading to a reduction of cell viability in both cell lines assayed. This NR-AuNPTGA system is a promising drug delivery system for photodynamic cancer therapy. - Highlights: • Modified gold nanoparticle (AuNP) by sodium thioglicolate (TGA) prevents aggregation. • Neutral red (NR) adsorbed on the surface of modified gold nanoparticles (AuNPTGA). • AuNPTGA is suitable as a platform to deliver the NR under irradiation process. • Photodamage of 90% was achieved by NR added to AuNPTGA in 4T1 and NIH-3T3 cells.

Sorption of nonpolar aromatic contaminants by chlorosilane surface modified natural minerals.

Science.gov (United States)

Huttenloch, P; Roehl, K E; Czurda, K

2001-11-01

The efficacy of the surface modification of natural diatomite and zeolite material by chlorosilanes is demonstrated. Chlorosilanes used were trimethylchlorosilane (TMSCI), tert-butyldimethylchlorosilane (TBDMSCI), dimethyloctadecylchlorosilane (DMODSCI), and diphenyldichlorosilane (DPDSCI) possessing different headgroups and chemical properties. Silanol groups of the diatomite and zeolite were modified by chemical reaction with the chlorosilanes resulting in a stable covalent attachment of the organosilanes to the mineral surface. The alteration of surface properties of the modified material was proved by measurements of water adsorption capacity, total organic carbon (TOC) content, and thermoanalytical data. The surface modified material showed great stability even when exposed to extremes in ionic strength, pH, and to pure organic solvents. Sorption of toluene, o-xylene, and naphthalene from water was greatly enhanced by the surface modification compared to the untreated materials which showed no measurable sorption of these compounds. The enhanced sorption was dependent on the organic carbon content as well as on chemical characteristics of the chlorosilanes used. Batch sorption experiments showed that the phenyl headgroups of DPDSCI have the best affinity for aromatic compounds. Removal from an aqueous solution of 10 mg/L of naphthalene, o-xylene, and toluene was 71%, 60%, and 30% for surface modified diatomite and 51%, 30%, and 16% for modified clinoptilolite, respectively. Sorption data were well described by the Freundlich isotherm equation, which indicated physical adsorption onto the lipophilic surface rather than partitioning into the surface organic phase. The chlorosilane modified materials have an apparent potential for application in environmental technologies such as permeable reactive barriers (PRB) or wastewater treatment.

Hydrolytic enzymes in the central vacuole of plant cells.

Science.gov (United States)

Boller, T; Kende, H

1979-06-01

The hydrolase content of vacuoles isolated from protoplasts of suspension-cultured tobacco cells, of tulip petals, and of pineapple leaves, and the sedimentation behavior of tobacco tonoplasts were studied. Three precautions were found to be important for the analysis of vacuolar hydrolases and of the tonoplast. (a) Purification of protoplasts in a Ficoll gradient was necessary to remove cell debris which contained contaminating hydrolases adsorbed from the fungal cell-wall-degrading enzyme preparation. (b) Hydrolase activities in the homogenates of the intact cells or the tissue used and of the purified protoplasts had to be compared to verify the absence of contaminating hydrolases in the protoplast preparation. (c) Vacuoles obtained from the protoplasts by an osmotic shock had to be purified from the lysate in a Ficoll gradient. Since the density of the central vacuole approximates that of the protoplasts, about a 10% contamination of the vacuolar preparation by surviving protoplasts could not be eliminated and had to be taken into account when the distribution of enzymes and of radioactivity was calculated.THE INTRACELLULAR ACTIVITIES OF THE FOLLOWING ACID HYDROLASES WERE PRIMARILY LOCALIZED IN THE VACUOLE OF TOBACCO CELLS: alpha-mannosidase, beta-N-acetylglucosaminidase, beta-fructosidase, nuclease, phosphatase, phosphodiesterase. A similar composition of acid hydrolases was found in vacuoles obtained from protoplasts of tulip petals. Proteinase, a hydrolase with low activity in tobacco cells and tulip petals and therefore difficult to localize unequivocally, was found to be vacuolar in pineapple leaves, a tissue containing high levels of this enzyme. Our data support the hypothesis that the central vacuole of higher plant cells has an enzyme composition analogous to that of the animal lysosome.None of the vacuolar enzymes investigated was found to be bound to the tonoplast. When vacuoles were isolated from cells labeled with radioactive choline, the vacuolar

The effect of fluoride surface modification of ceramic TiO{sub 2} on the surface properties and biological response of osteoblastic cells in vitro

Energy Technology Data Exchange (ETDEWEB)

Tiainen, H; Knychala, J; Lyngstadaas, S P; Haugen, H J [Department of Biomaterials, Institute for Clinical Dentistry, University of Oslo, PO Box 1109 Blindern, NO-0317 Oslo (Norway); Monjo, M [Department of Fundamental Biology and Health Sciences, Research Institute on Health Sciences (IUNICS), University of the Balearic Islands, Cra. de Valldemossa, km 7.5, 07122 Palma de Mallorca (Spain); Nilsen, O [Department of Chemistry, University of Oslo, PO Box 1033 Blindern, NO-0315 Oslo (Norway); Ellingsen, J E, E-mail: h.j.haugen@odont.uio.no [Oral Research Laboratory, Institute for Clinical Dentistry, University of Oslo, PO Box 1109 Blindern, NO-0317 Oslo (Norway)

2011-08-15

This study investigates the effect of fluoride surface modification on the surface properties of polycrystalline ceramic TiO{sub 2} and the biological response of murine osteoblast cells to fluoride-modified TiO{sub 2} in vitro. Fluoride concentrations up to 9 at.% were detected and the fluoride was found to bind to the surface in a ligand exchange reaction between surface hydroxyl groups and the fluoride anions from the HF. No significant changes in the surface topography were detected. In vitro experiments were performed in order to evaluate the biological response of the MC3T3-E1 cells to the fluoride-modified ceramic TiO{sub 2} surfaces. No difference in the lactate dehydrogenase (LDH) activity was seen in comparison to unmodified samples, apart from the highest fluoride concentration ({approx}9 at.%) which was found to be more toxic to the cells. Real-time PCR analysis showed no conclusive evidence for the fluoride-induced promotion of osteoblast differentiation as no significant increase in the collagen-1, osteocalcin, or BMP-2 mRNA levels was detected on the fluoride-modified ceramic TiO{sub 2} surfaces apart from one group, which showed an elevated osteocalcin level and higher number of cells. Since the observed grain boundary corrosion is also anticipated to reduce the mechanical properties of ceramic TiO{sub 2}, this surface modification method may not be an ideal method for improving the osteogenic response of ceramic TiO{sub 2} scaffolds.

Cysteine modified polyaniline films improve biocompatibility for two cell lines

Energy Technology Data Exchange (ETDEWEB)

Yslas, Edith I., E-mail: eyslas@exa.unrc.edu.ar [Departamento de Biología Molecular, Universidad Nacional de Río Cuarto, Agencia Postal Nro3, X580BYA Río Cuarto (Argentina); Cavallo, Pablo; Acevedo, Diego F.; Barbero, César A. [Departamento de Química, Universidad Nacional de Río Cuarto, Agencia Postal Nro3, X580BYA Río Cuarto (Argentina); Rivarola, Viviana A. [Departamento de Biología Molecular, Universidad Nacional de Río Cuarto, Agencia Postal Nro3, X580BYA Río Cuarto (Argentina)

2015-06-01

This work focuses on one of the most exciting application areas of conjugated conducting polymers, which is cell culture and tissue engineering. To improve the biocompatibility of conducting polymers we present an easy method that involves the modification of the polymer backbone using L-cysteine. In this publication, we show the synthesis of polyaniline (PANI) films supported onto Polyethylene terephthalate (PET) films, and modified using cysteine (PANI-Cys) in order to generate a biocompatible substrate for cell culture. The PANI-Cys films are characterized by Fourier Transform infrared and UV–visible spectroscopy. The changes in the hydrophilicity of the polymer films after and before the modification were tested using contact angle measurements. After modification the contact angle changes from 86° ± 1 to 90° ± 1, suggesting a more hydrophylic surface. The adhesion properties of LM2 and HaCaT cell lines on the surface of PANI-Cys films in comparison with tissue culture plastic (TCP) are studied. The PANI-Cys film shows better biocompatibility than PANI film for both cell lines. The cell morphologies on the TCP and PANI-Cys film were examined by florescence and Atomic Force Microscopy (AFM). Microscopic observations show normal cellular behavior when PANI-Cys is used as a substrate of both cell lines (HaCaT and LM2) as when they are cultured on TCP. The ability of these PANI-Cys films to support cell attachment and growth indicates their potential use as biocompatible surfaces and in tissue engineering. - Highlights: • A new surface PANI-Cys was produced on films of polyethylene terephthalate. • The relationship between surface characteristics and biocompatibility is analyzed. • The PANI-Cys film presents good biocompatibility for two cell lines.

Modified stainless steel for high performance and stable anode in microbial fuel cells

International Nuclear Information System (INIS)

Peng, Xinwen; Chen, Shuiliang; Liu, Lang; Zheng, Suqi; Li, Ming

2016-01-01

Graphical abstract: A high performance and stable anode was prepared for microbial fuel cells by surface modification of stainless steel mesh including steps of acid etching, binder-free carbon black (CB) coating and the low-temperature heat treatment below 400 °C. The modified anode could deliver a stable and high current density of 1.91 mA cm −2 . - Highlights: • A high-performance anode for MFC is prepared by surface modification of SSM. • The modified SSM could generate a high current density of up to 1.91 mA cm −2 . • The formation of Fe 3 O 4 layer enhanced the interaction between the CB and SSM. • The modified SSM was stable under the potential of +0.2 V (vs. Ag/AgCl). • The modified SSM was an ideal anode for upscaling applications of MFCs. - Abstract: The surface modification of the stainless steel mesh (SSM) was conducted by acid etching, binder-free carbon black (CB) coating and the low-temperature heat treatment below 400 °C to improve the microbial bioelectrocatalytic activity for use as high-performance anode in microbial fuel cells. The modified SSM, such as SSM/CB-400, could generate a high current density of up to 1.91 mA cm −2 , which was nearly three orders of magnitude higher than the untreated SSM electrode (0.0025 mA cm −2 ). Moreover, it was stable and recovered the equal current density after removal of the formed biofilms. Surface characterization results demonstrate that the performance improvement was attributed to the CB/Fe 3 O 4 composite layer formed onto the surface of the SSM, which protected the biofilms from being poisoned by the Cr component in the SSM and ensured a rapid electron transfer from biofilms to the SSM surface. The CB/Fe 3 O 4 composite layer showed excellent corrosion-resistant under the oxidizing potential of + 0.2 V (vs. Ag/AgCl). Rising the heating temperature to 500 °C, the SSM-500 and SSM/CB-500 electrodes suffered from corrosion due to the formation of α-Fe 2 O 3 crystals.

Preparation and antifouling properties of 2-(meth-acryloyloxy)ethyl cholinephosphate based polymers modified surface with different molecular architectures by ATRP.

Science.gov (United States)

Jiang, Yuchen; Su, Yuling; Zhao, Lili; Meng, Fancui; Wang, Quanxin; Ding, Chunmei; Luo, Jianbin; Li, Jianshu

2017-08-01

Choline phosphate (CP) containing polymers modified surfaces have been shown good resist to the adhesion of proteins while prompt the attaching of mammalian cells due to the dipole pairing between the CP groups of the polymer and the phosphorylcholine (PC) groups on the cell membrane. However, the antifouling activities of CP modified surface against microbes have not been investigated at present. In addition, CP containing polymers modified surface with different molecular architectures has not been prepared and studied. To this end, glass slides surface modified with two different 2-(meth-acryloyloxy)ethyl cholinephosphate (MCP) containing polymer (PMCP) structures, i.e. brush-like (Glass-PMCP) and bottle brush-like (Glass-PHEMA-g-PMCP) architectures, were prepared in this work by surface-initiated atom transfer radical polymerization (SI-ATRP). The surface physichemical and antifouling properties of the prepared surfaces were characterized and studied. The Glass-PMCP shows improved antifouling properties against proteins and bacteria as compared to pristine glass slides (Glass-OH) and glass slides grafted with poly(2-hydroxyethyl methacrylate) (Glass-PHEMA). Notably, a synergetic fouling resistant properties of PHEMA and PMCP is presented for Glass-PHEMA-g-PMCP, which shows superior antifouling activities over Glass-PHEMA and Glass-PMCP. Furthermore, glass slides containing PMCP, i.e. Glass-PMCP and Glas-PHEMA-g-PMCP, decrease platelet adhesion and prevent their activation significantly. Therefore, the combination of antifouling PHEMA and PMCP into one system holds potential for prevention of bacterial fouling and biomaterial-centered infections. Copyright © 2017 Elsevier B.V. All rights reserved.

PNIPAAM modified mesoporous hydroxyapatite for sustained osteogenic drug release and promoting cell attachment

Energy Technology Data Exchange (ETDEWEB)

Wu, Tao [The State Key Laboratory Breeding Base of Basic Science of Stomatology (Hubei-MOST) & Key Laboratory of Oral Biomedicine Ministry of Education, School & Hospital of Stomatology, Wuhan University, 237 Luoyu Road, Wuhan 430079 (China); Tan, Lei [Key Laboratory of Biomedical Polymers of Ministry of Education, College of Chemistry and Molecular Science, Wuhan University, Wuhan 430072 (China); Cheng, Ning; Yan, Qi; Zhang, Yu-Feng [The State Key Laboratory Breeding Base of Basic Science of Stomatology (Hubei-MOST) & Key Laboratory of Oral Biomedicine Ministry of Education, School & Hospital of Stomatology, Wuhan University, 237 Luoyu Road, Wuhan 430079 (China); Liu, Chuan-Jun, E-mail: cjliu@whu.edu.cn [Key Laboratory of Biomedical Polymers of Ministry of Education, College of Chemistry and Molecular Science, Wuhan University, Wuhan 430072 (China); Shi, Bin, E-mail: shibin_dentist@126.com [The State Key Laboratory Breeding Base of Basic Science of Stomatology (Hubei-MOST) & Key Laboratory of Oral Biomedicine Ministry of Education, School & Hospital of Stomatology, Wuhan University, 237 Luoyu Road, Wuhan 430079 (China)

2016-05-01

This work presented a sustained release system of simvastatin (SIM) based on the mesoporous hydroxyapatite (MHA) capped with poly(N-isopropylacrylamide) (PNIPAAM). The MHA was prepared by using cetyltrimethylammonium bromide (CTAB) as a template and the modified PNIPAAM layer on the surface of MHA was fabricated through surface-initiated atom transfer radical polymerization (SI-ATRP). The SIM loaded MHA-PNIPAAM showed a sustained release of SIM at 37 °C over 16 days. The bone marrow mesenchymal stem cell (BMSC) proliferation was assessed by cell counting kit-8 (CCK-8) assay, and the osteogenic differentiation was evaluated by alkaline phosphatase (ALP) activity and Alizarin Red staining. The release profile showed that the release of SIM from MHA-SIM-PNIPAAM lasted 16 days and the cumulative amount of released SIM was almost seven-fold than MHA-SIM. Besides, SIM loaded MHA-PNIPAAM exhibited better performance on cell proliferation, ALP activity, and calcium deposition than pure MHA due to the sustained release of SIM. The quantity of ALP in MHA-SIM-PNIPAAM group was more than two fold than pure MHA group at 7 days. Compared to pure MHA, better BMSC attachment on PNIPAAM modified MHA was observed using fluorescent microscopy, indicating the better biocompatibility of MHA-PNIPAAM. - Highlights: • PNIPAAM modified mesoporous hydroxyapatite (MHA) was fabricated by SI-ATRP. • SIM loaded MHA-PNIPAAM continually released SIM in effect concentration for 16 days. • MHA-SIM-PNIPAAM behaved well on cell proliferation, ALP activity and calcium deposition.

Pathogenicity and cell wall-degrading enzyme activities of some ...

African Journals Online (AJOL)

Dr. J. T. Ekanem

2005-12-17

Dec 17, 2005 ... be attributed to the activities of these cell wall degrading enzymes. Keywords: Cowpea ... bacteria have long been known to produce enzymes capable of ... Inoculated seeds were sown in small plastic pots filled with steam- ...

Enzymatic biofuel cell based on electrodes modified with lipid liquid-crystalline cubic phases

Science.gov (United States)

Nazaruk, Ewa; Smoliński, Sławomir; Swatko-Ossor, Marta; Ginalska, Grażyna; Fiedurek, Jan; Rogalski, Jerzy; Bilewicz, Renata

Two glassy carbon electrodes modified with enzymes embedded in lyotropic liquid-crystalline cubic phase were used for the biofuel cell construction. The monoolein liquid-crystalline film allowed to avoid separators in the biofuel cell. Glucose and oxygen as fuels, and glucose oxidase and laccase as anode and cathode biocatalysts, respectively were used. The biofuel cell parameters were examined in McIlvaine buffer, pH 7 solution containing 15 mM of glucose and saturated with dioxygen. A series of mediators were tested taking into account their formal potentials, stability in the cubic phase and efficiency of mediation. Most stable was the biofuel cell based on tetrathiafulvalene (TTF) and 2,2‧-azino-bis(3-ethylbenzothiazoline-6-sulfonate) (ABTS) as anode and cathode mediators, respectively. The open-circuit voltage was equal to 450 ± 40 mV. The power densities and current densities were measured for all the systems studied.

Surface modification for interaction study with bacteria and preosteoblast cells

Science.gov (United States)

Song, Qing

Surface modification plays a pivotal role in bioengineering. Polymer coatings can provide biocompatibility and biofunctionalities to biomaterials through surface modification. In this dissertation, initiated chemical vapor deposition (iCVD) was utilized to coat two-dimensional (2D) and three-dimensional (3D) substrates with differently charged polyelectrolytes in order to generate antimicrobial and osteocompatible biomaterials. ICVD is a modified CVD technique that enables surface modification in an all-dry condition without substrate damage and solvent contamination. The free-radical polymerization allows the vinyl polymers to conformally coat on various micro- and nano-structured substrates and maintains the delicate structure of the functional groups. The vapor deposition of polycations provided antimicrobial activity to planar and porous substrates through destroying the negatively charged bacterial membrane and brought about high contact-killing efficiency (99.99%) against Gram-positive Bacillus subtilis and Gram-negative Escherichia coli. Additionally, the polyampholytes synthesized by iCVD exhibited excellent antifouling performance against the adhesion of Gram-positive Listeria innocua and Gram-negative E. coli in phosphate buffered saline (PBS). Their antifouling activities were attributed to the electrostatic interaction and hydration layers that served as physical and energetic barriers to prevent bacterial adhesion. The contact-killing and antifouling polymers synthesized by iCVD can be applied to surface modification of food processing equipment and medical devices with the aim of reducing foodborne diseases and medical infections. Moreover, the charged polyelectrolyte modified 2D polystyrene surfaces displayed good osteocompatibility and enhanced osteogenesis of preosteoblast cells than the un-modified polystyrene surface. In order to promote osteoinduction of hydroxyapatite (HA) scaffolds, bioinspired polymer-controlled mineralization was conducted

Interactions between endothelial progenitor cells (EPC) and titanium implant surfaces.

Science.gov (United States)

Ziebart, Thomas; Schnell, Anne; Walter, Christian; Kämmerer, Peer W; Pabst, Andreas; Lehmann, Karl M; Ziebart, Johanna; Klein, Marc O; Al-Nawas, Bilal

2013-01-01

Endothelial cells play an important role in peri-implant angiogenesis during early bone formation. Therefore, interactions between endothelial progenitor cells (EPCs) and titanium dental implant surfaces are of crucial interest. The aim of our in vitro study was to investigate the reactions of EPCs in contact with different commercially available implant surfaces. EPCs from buffy coats were isolated by Ficoll density gradient separation. After cell differentiation, EPC were cultured for a period of 7 days on different titanium surfaces. The test surfaces varied in roughness and hydrophilicity: acid-etched (A), sand-blasted-blasted and acid-etched (SLA), hydrophilic A (modA), and hydrophilic SLA (modSLA). Plastic and fibronectin-coated plastic surfaces served as controls. Cell numbers and morphology were analyzed by confocal laser scanning microscopy. Secretion of vascular endothelial growth factor (VEGF)-A was measured by enzyme-linked immunosorbent assay and expressions of iNOS and eNOS were investigated by real-time polymerase chain reaction. Cell numbers were higher in the control groups compared to the cells of titanium surfaces. Initially, hydrophilic titanium surfaces (modA and modSLA) showed lower cell numbers than hydrophobic surfaces (A and SLA). After 7 days smoother surfaces (A and modA) showed increased cell numbers compared to rougher surfaces (SLA and modSLA). Cell morphology of A, modA, and control surfaces was characterized by a multitude of pseudopodia and planar cell soma architecture. SLA and modSLA promoted small and plump cell soma with little quantity of pseudopodia. The lowest VEGF level was measured on A, the highest on modSLA. The highest eNOS and iNOS expressions were found on modA surfaces. The results of this study demonstrate that biological behaviors of EPCs can be influenced by different surfaces. The modSLA surface promotes an undifferentiated phenotype of EPCs that has the ability to secrete growth factors in great quantities. In

Cell adhesion and growth on ultrananocrystalline diamond and diamond-like carbon films after different surface modifications

Energy Technology Data Exchange (ETDEWEB)

Miksovsky, J. [Institute of Nanostructure Technologies and Analytics, Center for Interdisciplinary Nanostructure Science and Technology, University of Kassel (Germany); Institute of Physics ASCR, Prague (Czech Republic); Czech Technical University in Prague, Faculty of Biomedical Engineering, Kladno (Czech Republic); Voss, A. [Institute of Nanostructure Technologies and Analytics, Center for Interdisciplinary Nanostructure Science and Technology, University of Kassel (Germany); Kozarova, R. [Institute of Molecular Biology, Bulgarian Academy of Sciences, Sofia (Bulgaria); Kocourek, T.; Pisarik, P. [Institute of Physics ASCR, Prague (Czech Republic); Czech Technical University in Prague, Faculty of Biomedical Engineering, Kladno (Czech Republic); Ceccone, G. [Unit Nanobiosciences, European Commission Joint Research Centre, Ispra (Italy); Kulisch, W. [Institute of Nanostructure Technologies and Analytics, Center for Interdisciplinary Nanostructure Science and Technology, University of Kassel (Germany); Jelinek, M. [Institute of Physics ASCR, Prague (Czech Republic); Czech Technical University in Prague, Faculty of Biomedical Engineering, Kladno (Czech Republic); Apostolova, M.D. [Institute of Molecular Biology, Bulgarian Academy of Sciences, Sofia (Bulgaria); Reithmaier, J.P. [Institute of Nanostructure Technologies and Analytics, Center for Interdisciplinary Nanostructure Science and Technology, University of Kassel (Germany); Popov, C., E-mail: popov@ina.uni-kassel.de [Institute of Nanostructure Technologies and Analytics, Center for Interdisciplinary Nanostructure Science and Technology, University of Kassel (Germany)

2014-04-01

Graphical abstract: - Highlights: • UNCD and DLC films were modified by UV/O{sub 3} treatments, O{sub 2} or NH{sub 3}-containing plasmas. • Surface composition, wettability and surface energy change upon modifications. • Higher efficiency of UNCD modifications was observed. • Cell attachment and growth were influenced by the surface termination and roughness. - Abstract: Diamond and diamond-like carbon (DLC) films possess a set of excellent physical and chemical properties which together with a high biocompatibility make them attractive candidates for a number of medical and biotechnological applications. In the current work thin ultrananocrystalline diamond (UNCD) and DLC films were comparatively investigated with respect to cell attachment and proliferation after different surface modifications. The UNCD films were prepared by microwave plasma enhanced chemical vapor deposition, the DLC films by pulsed laser deposition (PLD). The films were comprehensively characterized with respect to their basic properties, e.g. crystallinity, morphology, chemical bonding nature, etc. Afterwards the UNCD and DLC films were modified applying O{sub 2} or NH{sub 3}/N{sub 2} plasmas and UV/O{sub 3} treatments to alter their surface termination. The surface composition of as-grown and modified samples was studied by X-ray photoelectron spectroscopy (XPS). Furthermore the films were characterized by contact angle measurements with water, formamide, 1-decanol and diiodomethane; from the results obtained the surface energy with its dispersive and polar components was calculated. The adhesion and proliferation of MG63 osteosarcoma cells on the different UNCD and DLC samples were assessed by measurement of the cell attachment efficiency and MTT assays. The determined cell densities were compared and correlated with the surface properties of as-deposited and modified UNCD and DLC films.

Genetic and proteomic evidences support the localization of yeast enolase in the cell surface

DEFF Research Database (Denmark)

López-Villar, Elena; Monteoliva, Lucía; Larsen, Martin Røssel

2006-01-01

Although enolase, other glycolytic enzymes, and a variety of cytoplasmic proteins lacking an N-terminal secretion signal have been widely described as located at the cell surface in yeast and in mammalian cells, their presence in this external location is still controversial. Here, we report that...

Enzyme-Gelatin Electrochemical Biosensors: Scaling Down

Directory of Open Access Journals (Sweden)

Hendrik A. Heering

2012-03-01

Full Text Available In this article we investigate the possibility of scaling down enzyme-gelatin modified electrodes by spin coating the enzyme-gelatin layer. Special attention is given to the electrochemical behavior of the selected enzymes inside the gelatin matrix. A glassy carbon electrode was used as a substrate to immobilize, in the first instance, horse heart cytochrome c (HHC in a gelatin matrix. Both a drop dried and a spin coated layer was prepared. On scaling down, a transition from diffusion controlled reactions towards adsorption controlled reactions is observed. Compared to a drop dried electrode, a spin coated electrode showed a more stable electrochemical behavior. Next to HHC, we also incorporated catalase in a spin coated gelatin matrix immobilized on a glassy carbon electrode. By spincoating, highly uniform sub micrometer layers of biocompatible matrices can be constructed. A full electrochemical study and characterization of the modified surfaces has been carried out. It was clear that in the case of catalase, gluteraldehyde addition was needed to prevent leaking of the catalase from the gelatin matrix.

Improved cellular response of ion modified poly(lactic acid-co-glycolic acid) substrates for mouse fibroblast cells

International Nuclear Information System (INIS)

Adhikari, Ananta Raj; Geranpayeh, Tanya; Chu, Wei Kan; Otteson, Deborah C.

2016-01-01

In this report, the effects of argon (Ar) ion irradiation on poly(lactic acid-co-glycolic acid) (PLGA) substrates on biocompatibility were studied. PLGA scaffold substrates were prepared by spin coating glass surfaces with PLGA dissolved in anhydrous chloroform. Previously, we showed that surface modifications of PLGA films using ion irradiation modulate the inherent hydrophobicity of PLGA surface. Here we show that with increasing ion dose (1 × 10 12 to 1 × 10 14 ions/cm 2 ), hydrophobicity and surface roughness decreased. Biocompatibility for NIH3T3 mouse fibroblast cells was increased by argon irradiation of PLGA substrates. On unirradiated PLGA films, fibroblasts had a longer doubling time and cell densities were 52% lower than controls after 48 h in vitro. Argon irradiated PLGA substrates supported growth rates similar to control. Despite differences in cell cycle kinetics, there was no detectible cytotoxicity observed on any substrate. This demonstrates that argon ion irradiation can be used to tune the surface microstructure and generate substrates that are more compatible for the cell growth and proliferation. - Highlights: • Argon irradiation modifies surface chemistry and increases hydrophilicity of poly(lactic-glycolic) acid (PLGA) films. • Both native and irradiated PLGA films were not cytotoxic for mouse fibroblasts. • Fibroblast proliferation increased on PLGA substrates modified with higher doses of Argon irradiation. • Surface modification with Argon irradiation increases biocompatibility of PLGA films.

In situ immobilization of proteins and RGD peptide on polyurethane surfaces via poly(ethylene oxide) coupling polymers for human endothelial cell growth.

Science.gov (United States)

Wang, Dong-an; Ji, Jian; Sun, Yong-hong; Shen, Jia-cong; Feng, Lin-xian; Elisseeff, Jennifer H

2002-01-01

A "CBABC"-type pentablock coupling polymer, mesylMPEO, was designed and synthesized to promote human endothelial cell growth on the surfaces of polyurethane biomaterials. The polymer was composed of a central 4,4'-methylenediphenyl diisocyanate (MDI) coupling unit and poly(ethylene oxide) (PEO) spacer arms with methanesulfonyl (mesyl) end groups pendent on both ends. As the presurface modifying additive (pre-SMA), the mesylMPEO was noncovalently introduced onto the poly(ether urethane) (PEU) surfaces by dip coating, upon which the protein/peptide factors (gelatin, albumin, and arginine-glycine-aspartic acid tripeptide [RGD]) were covalently immobilized in situ by cleavage of the original mesyl end groups. The pre-SMA synthesis and PEU surface modification were characterized using nuclear magnetic resonance spectroscopy ((1)H NMR), attenuated total reflection infrared spectroscopy (ATR-FTIR), and X-ray photoelectron spectroscopy (XPS). Human umbilical vein endothelial cells (HUVEC) were harvested manually by collagenase digestion and seeded on the modified PEU surfaces. Cell adhesion ratios (CAR) and cell proliferation ratios (CPR) were measured using flow cytometry, and the individual cell viability (ICV) was determined by MTT assay. The cell morphologies were investigated by optical inverted microscopy (OIM) and scanning electrical microscopy (SEM). The gelatin- and RGD-modified surfaces were HUVEC-compatible and promoted HUVEC growth. The albumin-modified surfaces were compatible but inhibited cell adhesion. The results also indicated that, for HUVEC in vitro cultivation, the cell adhesion stage was of particular importance and had a significant impact on the cell responses to the modified surfaces.

Correspondence between radioactive and functional methods in the quality control of DNA restriction and modifying enzymes

DEFF Research Database (Denmark)

Trujillo, L E; Pupo, E; Miranda, F

1996-01-01

We evaluated the use of two radiolabeled lambda DNA/Hpa II substrates to detect 5'-->3', 3'-->5' single and double stranded DNA dependent exonuclease and phosphatase activities found as contaminants in restriction and modifying enzyme preparations. Looking for the meaning of the radioactive assay...

METABOLIC MAPPING BY ENZYME HISTOCHEMISTRY IN LIVING ANIMALS, TISSUES AND CELLS

NARCIS (Netherlands)

van Noorden, C. J. F.

2009-01-01

Imaging of reporter molecules such as fluorescent proteins in intact animals, tissue and cells has become an indispensable tool in cell biology Imaging activity of enzymes, which is called metabolic mapping, provides information on subcellular localisation in combination with function of the enzymes

Host cell capable of producing enzymes useful for degradation of lignocellulosic material

Energy Technology Data Exchange (ETDEWEB)

Los, Alrik Pieter; Sagt, Cornelis Maria Jacobus; Schoonneveld-Bergmans, Margot Elisabeth Francoise; Damveld, Robbertus Antonius

2017-08-22

The invention relates to a host cell comprising at least four different heterologous polynucleotides chosen from the group of polynucleotides encoding cellulases, hemicellulases and pectinases, wherein the host cell is capable of producing the at least four different enzymes chosen from the group of cellulases, hemicellulases and pectinases, wherein the host cell is a filamentous fungus and is capable of secretion of the at least four different enzymes. This host cell can suitably be used for the production of an enzyme composition that can be used in a process for the saccharification of cellulosic material.

Host cell capable of producing enzymes useful for degradation of lignocellulosic material

Science.gov (United States)

Los, Alrik Pieter; Sagt, Cornelis Maria Jacobus; Schooneveld-Bergmans, Margot Elisabeth Francoise; Damveld, Robbertus Antonius

2015-08-18

The invention relates to a host cell comprising at least four different heterologous polynucleotides chosen from the group of polynucleotides encoding cellulases, hemicellulases and pectinases, wherein the host cell is capable of producing the at least four different enzymes chosen from the group of cellulases, hemicellulases and pectinases, wherein the host cell is a filamentous fungus and is capable of secretion of the at least four different enzymes. This host cell can suitably be used for the production of an enzyme composition that can be used in a process for the saccharification of cellulosic material.

Human procollagen type I surface-modified PHB-based non-woven textile scaffolds for cell growth: preparation and short-term biological tests

International Nuclear Information System (INIS)

Kawalec, Michał; Sobota, Michał; Kurcok, Piotr; Sitkowska, Anna; Sieroń, Aleksander L; Komar, Patrycja

2014-01-01

3D fine porous structures obtained by electrospinning a poly[(R,S)-3-hydroxybutyrate] (aPHB)/ poly[(R)-3-hydroxybutyrate] (PHB) (85/15 w/w) blend were successfully modified with human procollagen type I by simple immersion of the polyester scaffold in an aqueous solution of the protein. Effective modification of the scaffold with human procollagen I was confirmed by an immunodetection test, which revealed the presence of the procollagen type I as an outer layer even on inner structures of the porous matrixes. Biological tests of 3D fabrics made of the PHB blend provide support for the adhesion and proliferation of human fibroblasts, while their modification with procollagen type I increased the biocompatibility of the final scaffolds significantly, as shown by the notable increase in the number of attached cells during the early hours of their incubation. Based on these findings, human procollagen type I surface-modified aPHB/PHB scaffolds should be considered a promising material in regenerative medicine. (paper)

Watch Out for the “Living Dead”: Cell-Free Enzymes and Their Fate

Directory of Open Access Journals (Sweden)

Federico Baltar

2018-01-01

Full Text Available Microbes are the engines driving biogeochemical cycles. Microbial extracellular enzymatic activities (EEAs are the “gatekeepers” of the carbon cycle. The total EEA is the sum of cell-bound (i.e., cell-attached, and dissolved (i.e., cell-free enzyme activities. Cell-free enzymes make up a substantial proportion (up to 100% of the total marine EEA. Although we are learning more about how microbial diversity and function (including total EEA will be affected by environmental changes, little is known about what factors control the importance of the abundant cell-free enzymes. Since cell-attached EEAs are linked to the cell, their fate will likely be linked to the factors controlling the cell’s fate. In contrast, cell-free enzymes belong to a kind of “living dead” realm because they are not attached to a living cell but still are able to perform their function away from the cell; and as such, the factors controlling their activity and fate might differ from those affecting cell-attached enzymes. This article aims to place cell-free EEA into the wider context of hydrolysis of organic matter, deal with recent studies assessing what controls the production, activity and lifetime of cell-free EEA, and what their fate might be in response to environmental stressors. This perspective article advocates the need to go “beyond the living things,” studying the response of cells/organisms to different stressors, but also to study cell-free enzymes, in order to fully constrain the future and evolution of marine biogeochemical cycles.

Watch Out for the “Living Dead”: Cell-Free Enzymes and Their Fate

Science.gov (United States)

Baltar, Federico

2018-01-01

Microbes are the engines driving biogeochemical cycles. Microbial extracellular enzymatic activities (EEAs) are the “gatekeepers” of the carbon cycle. The total EEA is the sum of cell-bound (i.e., cell-attached), and dissolved (i.e., cell-free) enzyme activities. Cell-free enzymes make up a substantial proportion (up to 100%) of the total marine EEA. Although we are learning more about how microbial diversity and function (including total EEA) will be affected by environmental changes, little is known about what factors control the importance of the abundant cell-free enzymes. Since cell-attached EEAs are linked to the cell, their fate will likely be linked to the factors controlling the cell’s fate. In contrast, cell-free enzymes belong to a kind of “living dead” realm because they are not attached to a living cell but still are able to perform their function away from the cell; and as such, the factors controlling their activity and fate might differ from those affecting cell-attached enzymes. This article aims to place cell-free EEA into the wider context of hydrolysis of organic matter, deal with recent studies assessing what controls the production, activity and lifetime of cell-free EEA, and what their fate might be in response to environmental stressors. This perspective article advocates the need to go “beyond the living things,” studying the response of cells/organisms to different stressors, but also to study cell-free enzymes, in order to fully constrain the future and evolution of marine biogeochemical cycles. PMID:29354095

Proteolytic Enzymes Clustered in Specialized Plasma-Membrane Domains Drive Endothelial Cells' Migration.

Directory of Open Access Journals (Sweden)

Monica Salamone

Full Text Available In vitro cultured endothelial cells forming a continuous monolayer establish stable cell-cell contacts and acquire a "resting" phenotype; on the other hand, when growing in sparse conditions these cells acquire a migratory phenotype and invade the empty area of the culture. Culturing cells in different conditions, we compared expression and clustering of proteolytic enzymes in cells having migratory versus stationary behavior. In order to observe resting and migrating cells in the same microscopic field, a continuous cell monolayer was wounded. Increased expression of proteolytic enzymes was evident in cell membranes of migrating cells especially at sprouting sites and in shed membrane vesicles. Gelatin zymography and western blotting analyses confirmed that in migrating cells, expression of membrane-bound and of vesicle-associated proteolytic enzymes are increased. The enzymes concerned include MMP-2, MMP-9, MT1-MMP, seprase, DPP4 (DiPeptidyl Peptidase 4 and uPA. Shed membrane vesicles were shown to exert degradative activity on ECM components and produce substrates facilitating cell migration. Vesicles shed by migrating cells degraded ECM components at an increased rate; as a result their effect on cell migration was amplified. Inhibiting either Matrix Metallo Proteases (MMPs or Serine Integral Membrane Peptidases (SIMPs caused a decrease in the stimulatory effect of vesicles, inhibiting the spontaneous migratory activity of cells; a similar result was also obtained when a monoclonal antibody acting on DPP4 was tested. We conclude that proteolytic enzymes have a synergistic stimulatory effect on cell migration and that their clustering probably facilitates the proteolytic activation cascades needed to produce maximal degradative activity on cell substrates during the angiogenic process.

Proteolytic Enzymes Clustered in Specialized Plasma-Membrane Domains Drive Endothelial Cells' Migration.

Science.gov (United States)

Salamone, Monica; Carfì Pavia, Francesco; Ghersi, Giulio

2016-01-01

In vitro cultured endothelial cells forming a continuous monolayer establish stable cell-cell contacts and acquire a "resting" phenotype; on the other hand, when growing in sparse conditions these cells acquire a migratory phenotype and invade the empty area of the culture. Culturing cells in different conditions, we compared expression and clustering of proteolytic enzymes in cells having migratory versus stationary behavior. In order to observe resting and migrating cells in the same microscopic field, a continuous cell monolayer was wounded. Increased expression of proteolytic enzymes was evident in cell membranes of migrating cells especially at sprouting sites and in shed membrane vesicles. Gelatin zymography and western blotting analyses confirmed that in migrating cells, expression of membrane-bound and of vesicle-associated proteolytic enzymes are increased. The enzymes concerned include MMP-2, MMP-9, MT1-MMP, seprase, DPP4 (DiPeptidyl Peptidase 4) and uPA. Shed membrane vesicles were shown to exert degradative activity on ECM components and produce substrates facilitating cell migration. Vesicles shed by migrating cells degraded ECM components at an increased rate; as a result their effect on cell migration was amplified. Inhibiting either Matrix Metallo Proteases (MMPs) or Serine Integral Membrane Peptidases (SIMPs) caused a decrease in the stimulatory effect of vesicles, inhibiting the spontaneous migratory activity of cells; a similar result was also obtained when a monoclonal antibody acting on DPP4 was tested. We conclude that proteolytic enzymes have a synergistic stimulatory effect on cell migration and that their clustering probably facilitates the proteolytic activation cascades needed to produce maximal degradative activity on cell substrates during the angiogenic process.

The development, characterization, and application of biomimetic nanoscale enzyme immobilization

Science.gov (United States)

Haase, Nicholas R.

The utilization of enzymes is of interest for applications such as biosensors and biofuel cells. Immobilizing enzymes provides a means to develop these applications. Previous immobilization efforts have been accomplished by exposing surfaces on which silica-forming molecules are present to solutions containing an enzyme and a silica precursor. This approach leads to the enzyme being entrapped in a matrix three orders of magnitude larger than the enzyme itself, resulting in low retention of enzyme activity. The research herein introduces a method for the immobilization of enzymes during the layer-by-layer buildup of Si-O and Ti-O coatings which are nanoscale in thickness. This approach is an application of a peptide-induced mineral deposition method developed in the Sandhage and Kroger groups, and it involves the alternating exposure of a surface to solutions containing the peptide protamine and then an aqueous precursor solution of silicon- or titanium-oxide at near-neutral pH. A method has been developed that enables in situ immobilization of enzymes in the protamine/mineral oxide coatings. Depending on the layer and mineral (silica or titania) within which the enzyme is incorporated, the resulting multilayer biocatalytic hybrid materials retain 20 -- 100% of the enzyme activity. Analyses of kinetic properties of the immobilized enzyme, coupled with characterization of physical properties of the mineral-bearing layers (thickness, porosity, pore size distribution), indicates that the catalytic activities of the enzymes immobilized in the different layers are largely determined by substrate diffusion. The enzyme was also found to be substantially stabilized against heat-induced denaturation and largely protected from proteolytic attack. These functional coatings are then developed for use as antimicrobial materials. Glucose oxidase, which catalyzes production of the cytotoxic agent hydrogen peroxide, was immobilized with silver nanoparticles, can release

Designing and Producing Modified, New-to-Nature Peptides with Antimicrobial Activity by Use of a Combination of Various Lantibiotic Modification Enzymes

NARCIS (Netherlands)

van Heel, Auke J.; Mu, Dongdong; Montalban-Lopez, Manuel; Hendriks, Djoke; Kuipers, Oscar P.

Lanthipeptides are peptides that contain several post-translationally modified amino acid residues and commonly show considerable antimicrobial activity. After translation, the amino acid residues of these peptides are modified by a distinct set of modification enzymes. This process results in

CELLISA: reporter cell-based immunization and screening of hybridomas specific for cell surface antigens.

Science.gov (United States)

Chen, Peter; Mesci, Aruz; Carlyle, James R

2011-01-01

Monoclonal antibodies (mAbs) specific for cell surface antigens are an invaluable tool to study immune receptor expression and function. Here, we outline a generalized reporter cell-based approach to the generation and high-throughput screening of mAbs specific for cell surface antigens. Termed CELLISA, this technology hinges upon the capture of hybridoma supernatants in mAb arrays that facilitate ligation of an antigen of interest displayed on BWZ reporter cells in the form of a CD3ζ-fusion chimeric antigen receptor (zCAR); in turn, specific mAb-mediated cross-linking of zCAR on BWZ cells results in the production of β-galactosidase enzyme (β-gal), which can be assayed colorimetrically. Importantly, the BWZ reporter cells bearing the zCAR of interest may be used for immunization as well as screening. In addition, serial immunizations employing additional zCAR- or native antigen-bearing cell lines can be used to increase the frequency of the desired antigen-specific hybridomas. Finally, the use of a cohort of epitope-tagged zCAR (e.g., zCAR(FLAG)) variants allows visualization of the cell surface antigen prior to immunization, and coimmunization using these variants can be used to enhance the immunogenicity of the target antigen. Employing the CELLISA strategy, we herein describe the generation of mAb directed against an uncharacterized natural killer cell receptor protein.

Facile direct electron transfer in glucose oxidase modified electrodes

International Nuclear Information System (INIS)

Wang Dan; Chen Liwei

2009-01-01

Glucose oxidase (GOx) is widely used in the glucose biosensor industry. However, mediatorless direct electron transfer (DET) from GOx to electrode surfaces is very slow. Recently, mediatorless DET has been reported via the incorporation of nanomaterials such as carbon nanotubes and nanoparticles in the modification of electrodes. Here we report GOx electrodes showing DET without the need for any nanomaterials. The enzyme after immobilization with poly-L-lysine (PLL) and Nafion retains the biocatalytic activities and oxidizes glucose efficiently. The amperometric response of Nafion-PLL-GOx modified electrode is linearly proportional to the concentration of glucose up to 10 mM with a sensitivity of 0.75 μA/mM at a low detection potential (-0.460 V vs. Ag/AgCl). The methodology developed in this study will have impact on glucose biosensors and biofuel cells and may potentially simplify enzyme immobilization in other biosensing systems.

The ultrasound technology for modifying enzyme activity

Directory of Open Access Journals (Sweden)

Meliza Lindsay

2016-06-01

Full Text Available Enzymes are protein complexes compounds widely studied and used due to their ability to catalyze reactions. The food processing mainly aims the inactivation of enzymes due to various undesirable effects. However, there are many processes that can be optimized by its catalytic activity. In this context, different technologies have been applied both to inactivate or to improve the enzymes efficiency. The Ultrasound technology emerges as an alternative mainly applied to achieve the enzyme inactivation. On the contrary, very few investigations show the ability of this technology under certain conditions to achieve the opposite effect (i.e. increase the catalytic activity of enzymes. The objective of this study was to correlate the ultrasonic energy delivered to the sample (J/mL with the residual enzymatic activity and explain the possible mechanisms which results in the enzymatic activation/inactivation complex behavior. The activity of POD in coconut water was evaluated as a model. The enzymatic activity initially increased, followed by reduction with a trend to enzyme inactivation. This complex behavior is directly related to the applied ultrasonic energy and their direct mechanical effects on the product, as well as the effect in the enzymatic infinite intermediate states and its structural conformation changes. The obtained results are useful for both academic and industrial perspectives.

Evaluation of cell behavior on modified polypropylene with swift heavy ion irradiation

International Nuclear Information System (INIS)

Arbeitman, Claudia R.; Ibañez, Irene L.; García Bermúdez, Gerardo; Durán, Hebe; Grosso, Mariela F. del; Salguero, Noelia; Mazzei, Rubén

2012-01-01

Ion beam irradiation is a well known means to change the physico-chemical properties of polymers, and induced bio and citocompatibility in controlled conditions and in selected areas of surface. However, the enhancement of cell adhesion on a modified substrate does not mean that the surface is adequate for functional cells. The purpose of the present work is to study proliferation, changes in cytoskeleton and cell morphology on substrates as a function of irradiation parameters. We irradiated polypropylene with sulfur (S) ion-beam at energies of 110 MeV with fluences between 1 × 10 6 and 2 × 10 10 ions cm −2 . NIH 3T3 cells were cultured on each sample. Cell morphology was observed using phase contrast microscopy and cytoskeleton proteins with fluorescence microscopy. The analysis show different cellular responses as a functions of irradiation parameter, strongly suggests that different underlying substratum can result in distinct types of cytoskeleton reorganization.

Evaluation of cell behavior on modified polypropylene with swift heavy ion irradiation

Energy Technology Data Exchange (ETDEWEB)

Arbeitman, Claudia R., E-mail: arbeitman@tandar.cnea.gov.ar [Gerencia de Investigacion y Aplicaciones, TANDAR-CNEA, Av. Gral. Paz 1499, 1650 San Martin, Bs. As. (Argentina); CONICET, Av. Rivadavia 1917, C1033AAJ, CABA (Argentina); Ibanez, Irene L. [CONICET, Av. Rivadavia 1917, C1033AAJ, CABA (Argentina); Garcia Bermudez, Gerardo [Gerencia de Investigacion y Aplicaciones, TANDAR-CNEA, Av. Gral. Paz 1499, 1650 San Martin, Bs. As. (Argentina); CONICET, Av. Rivadavia 1917, C1033AAJ, CABA (Argentina); Duran, Hebe [CONICET, Av. Rivadavia 1917, C1033AAJ, CABA (Argentina); Gerencia de Desarrollo Tecnologico y Proyectos Especiales, TANDAR-CNEA (Argentina); Grosso, Mariela F. del [Gerencia de Investigacion y Aplicaciones, TANDAR-CNEA, Av. Gral. Paz 1499, 1650 San Martin, Bs. As. (Argentina); CONICET, Av. Rivadavia 1917, C1033AAJ, CABA (Argentina); Salguero, Noelia [Gerencia de Investigacion y Aplicaciones, TANDAR-CNEA, Av. Gral. Paz 1499, 1650 San Martin, Bs. As. (Argentina); Mazzei, Ruben [U.A. Tecnologicas y Agropecuarias, CNEA (Argentina)

2012-02-15

Ion beam irradiation is a well known means to change the physico-chemical properties of polymers, and induced bio and citocompatibility in controlled conditions and in selected areas of surface. However, the enhancement of cell adhesion on a modified substrate does not mean that the surface is adequate for functional cells. The purpose of the present work is to study proliferation, changes in cytoskeleton and cell morphology on substrates as a function of irradiation parameters. We irradiated polypropylene with sulfur (S) ion-beam at energies of 110 MeV with fluences between 1 Multiplication-Sign 10{sup 6} and 2 Multiplication-Sign 10{sup 10} ions cm{sup -2}. NIH 3T3 cells were cultured on each sample. Cell morphology was observed using phase contrast microscopy and cytoskeleton proteins with fluorescence microscopy. The analysis show different cellular responses as a functions of irradiation parameter, strongly suggests that different underlying substratum can result in distinct types of cytoskeleton reorganization.

Extraction of Active Enzymes from "Hard-to-Break-Cells"

DEFF Research Database (Denmark)

Ottaviani, Alessio; Tesauro, Cinzia; Fjelstrup, S

We present the utilization of a rolling circle amplification (RCA) based assay to investigate the extraction efficiency of active enzymes from a class of “hard-to-break” cells, yeast Saccaramyces cerevisiae. Current analyses of microorganisms, such as pathogenic bacteria, parasites or particular...... life stages of microorganisms (e.g. spores from bacteria or fungi) is hampered by the lack of efficient lysis protocols that preserve the activity and integrity of the cellular content. Presented herein is a flexible scheme to screen lysis protocols for active enzyme extraction. We also report a gentle...... yet effective approach for extraction of active enzymes by entrapping cells in microdroplets. Combined effort of optimized extraction protocols and effective analytical approaches is expected to generate impact in future disease diagnosis and environmental safety....

Proteomic analyses for profiling regulated proteins/enzymes by Fucus vesiculosus fucoidan in B16 melanoma cells: A combination of enzyme kinetics functional study.

Science.gov (United States)

Wang, Zhi-Jiang; Zheng, Li; Yang, Jun-Mo; Kang, Yani; Park, Yong-Doo

2018-06-01

Fucoidans are complex sulfated polysaccharides that have a wide range of biological activities. Previously, we reported the various effects of Fucus vesiculosus fucoidan on tyrosinase and B16 melanoma cells. In this study, to identify fucoidan-targeted proteins in B16 melanoma cells, we performed a proteomics study and integrated enzyme kinetics. We detected 19 candidate proteins dysregulated by fucoidan treatment. Among the probed proteins, the enzyme kinetics of two candidate enzymes, namely lactate dehydrogenase (LDH) as an upregulated protein and superoxide dismutase (SOD) as a downregulated enzyme, were determined. The enzyme kinetics results showed that Fucus vesiculosus fucoidan significantly inhibited LDH catalytic function while it did not affect SOD activity even at a high dose, while only slightly decreased activity (up to 10%) at a low dose. Based on our previous and present observations, fucoidan could inhibit B16 melanoma cells growth via regulating proteins/enzymes expression levels such as LDH and SOD known as cell survival biomarkers. Interestingly, both expression level and enzyme catalytic activity of LDH were regulated by fucoidan, which could directly induce the apoptotic effect on B16 melanoma cells along with SOD downregulation. This study highlights how combining proteomics with enzyme kinetics can yield valuable insights into fucoidan targets. Copyright © 2018 Elsevier B.V. All rights reserved.

Surface localization of the nuclear receptor CAR in influenza A virus-infected cells

International Nuclear Information System (INIS)

Takahashi, Tadanobu; Moriyama, Yusuke; Ikari, Akira; Sugatani, Junko; Suzuki, Takashi; Miwa, Masao

2008-01-01

Constitutive active/androstane receptor CAR is a member of the nuclear receptors which regulate transcription of xenobiotic metabolism enzymes. CAR is usually localized in the cytosol and nucleus. Here, we found that CAR was localized at the cell surface of influenza A virus (IAV)-infected cells. Additionally, we demonstrated that expression of a viral envelope glycoprotein, either hemagglutinin (HA) or neuraminidase (NA), but not viral nucleoprotein (NP), was responsible for this localization. This report is the first demonstration of CAR at the surface of tissue culture cells, and suggests that CAR may exert the IAV infection mechanism

In vitro mesenchymal stem cell response to a CO{sub 2} laser modified polymeric material

Energy Technology Data Exchange (ETDEWEB)

Waugh, D.G., E-mail: d.waugh@chester.ac.uk [Laser Engineering and Manufacturing Research Centre, Faculty of Science and Engineering, University of Chester, Chester CH1 4BJ (United Kingdom); Hussain, I. [School of Life Sciences, Brayford Pool, University of Lincoln, Lincoln LN6 7TS (United Kingdom); Lawrence, J.; Smith, G.C. [Laser Engineering and Manufacturing Research Centre, Faculty of Science and Engineering, University of Chester, Chester CH1 4BJ (United Kingdom); Cosgrove, D. [School of Life Sciences, Brayford Pool, University of Lincoln, Lincoln LN6 7TS (United Kingdom); Toccaceli, C. [Laser Engineering and Manufacturing Research Centre, Faculty of Science and Engineering, University of Chester, Chester CH1 4BJ (United Kingdom)

2016-10-01

With an ageing world population it is becoming significantly apparent that there is a need to produce implants and platforms to manipulate stem cell growth on a pharmaceutical scale. This is needed to meet the socio-economic demands of many countries worldwide. This paper details one of the first ever studies in to the manipulation of stem cell growth on CO{sub 2} laser surface treated nylon 6,6 highlighting its potential as an inexpensive platform to manipulate stem cell growth on a pharmaceutical scale. Through CO{sub 2} laser surface treatment discrete changes to the surfaces were made. That is, the surface roughness of the nylon 6,6 was increased by up to 4.3 μm, the contact angle was modulated by up to 5° and the surface oxygen content increased by up to 1 atom %. Following mesenchymal stem cell growth on the laser treated samples, it was identified that CO{sub 2} laser surface treatment gave rise to an enhanced response with an increase in viable cell count of up to 60,000 cells/ml when compared to the as-received sample. The effect of surface parameters modified by the CO{sub 2} laser surface treatment on the mesenchymal stem cell response is also discussed along with potential trends that could be identified to govern the mesenchymal stem cell response.

The role of surface charge on the uptake and biocompatibility of hydroxyapatite nanoparticles with osteoblast cells

Energy Technology Data Exchange (ETDEWEB)

Chen Liang; Mccrate, Joseph M; Li Hao [Department of Mechanical and Aerospace Engineering, University of Missouri, Columbia, MO 65211 (United States); Lee, James C-M, E-mail: liha@missouri.edu [Department of Biological Engineering, University of Missouri, Columbia, MO 65211 (United States)

2011-03-11

The objective of this study is to evaluate the effect of hydroxyapatite (HAP) nanoparticles with different surface charges on the cellular uptake behavior and in vitro cell viability and proliferation of MC3T3-E1 cell lines (osteoblast). The nanoparticles' surface charge was varied by surface modification with two carboxylic acids: 12-aminododecanoic acid (positive) and dodecanedioic acid (negative). The untreated HAP nanoparticles and dodecanoic acid modified HAP nanoparticles (neutral) were used as the control. X-ray diffraction (XRD) revealed that surface modifications by the three carboxylic acids did not change the crystal structure of HAP nanoparticles; Fourier transform infrared spectroscopy (FT-IR) confirmed the adsorption and binding of the carboxylic acids on the HAP nanoparticles' surfaces; and zeta potential measurement confirmed that the chemicals successfully modified the surface charge of HAP nanoparticles in water based solution. Transmission electron microscopy (TEM) images showed that positively charged, negatively charged and untreated HAP nanoparticles, with similar size and shape, all penetrated into the cells and cells had more uptake of HAP nanoparticles with positive charge compared to those with negative charge, which might be attributed to the attractive or repulsive interaction between the negatively charged cell membrane and positively/negatively charged HAP nanoparticles. The neutral HAP nanoparticles could not penetrate the cell membrane due to their larger size. MTT assay and LDH assay results indicated that as compared with the polystyrene control, greater cell viability and cell proliferation were measured on MC3T3-E1 cells treated with the three kinds of HAP nanoparticles (neutral, positive, and untreated), among which positively charged HAP nanoparticles showed the strongest improvement for cell viability and cell proliferation. In summary, the surface charge of HAP nanoparticles can be modified to influence the cellular

The role of surface charge on the uptake and biocompatibility of hydroxyapatite nanoparticles with osteoblast cells

International Nuclear Information System (INIS)

Chen Liang; Mccrate, Joseph M; Li Hao; Lee, James C-M

2011-01-01

The objective of this study is to evaluate the effect of hydroxyapatite (HAP) nanoparticles with different surface charges on the cellular uptake behavior and in vitro cell viability and proliferation of MC3T3-E1 cell lines (osteoblast). The nanoparticles' surface charge was varied by surface modification with two carboxylic acids: 12-aminododecanoic acid (positive) and dodecanedioic acid (negative). The untreated HAP nanoparticles and dodecanoic acid modified HAP nanoparticles (neutral) were used as the control. X-ray diffraction (XRD) revealed that surface modifications by the three carboxylic acids did not change the crystal structure of HAP nanoparticles; Fourier transform infrared spectroscopy (FT-IR) confirmed the adsorption and binding of the carboxylic acids on the HAP nanoparticles' surfaces; and zeta potential measurement confirmed that the chemicals successfully modified the surface charge of HAP nanoparticles in water based solution. Transmission electron microscopy (TEM) images showed that positively charged, negatively charged and untreated HAP nanoparticles, with similar size and shape, all penetrated into the cells and cells had more uptake of HAP nanoparticles with positive charge compared to those with negative charge, which might be attributed to the attractive or repulsive interaction between the negatively charged cell membrane and positively/negatively charged HAP nanoparticles. The neutral HAP nanoparticles could not penetrate the cell membrane due to their larger size. MTT assay and LDH assay results indicated that as compared with the polystyrene control, greater cell viability and cell proliferation were measured on MC3T3-E1 cells treated with the three kinds of HAP nanoparticles (neutral, positive, and untreated), among which positively charged HAP nanoparticles showed the strongest improvement for cell viability and cell proliferation. In summary, the surface charge of HAP nanoparticles can be modified to influence the cellular uptake of

Covalent attachment of cell-adhesive peptide Gly-Arg-Gly-Asp (GRGD) to poly(etheretherketone) surface by tailored silanization layers technique

International Nuclear Information System (INIS)

Zheng, Yanyan; Xiong, Chengdong; Li, Xiaoyu; Zhang, Lifang

2014-01-01

Highlights: • The carbonyl groups on PEEK surface were effectively reduced to hydroxyl groups using sodium borohydride. • Silanization layers technique was employed to immobilize the cell-adhesive peptide Gly-Arg-Gly-Asp (GRGD) on hydroxylation-pretreated PEEK sheet surface by covalent chemical attachment. • XPS, surface profiler and water contact angle measurements proved the presence of GRGD on PEEK surface. • Osteoblast-like cells (MC3T3-E1) attachment and proliferation were improved effectively on GRGD-modified PEEK surface. - Abstract: Poly(etheretherketone) (PEEK) is a rigid semicrystalline polymer that combines excellent mechanical properties, broad chemical resistance and bone-like stiffness and is widely used in biomedical fields. However, PEEK is naturally bioinert, leading to limited biomedical applications, especially when a direct bone-implant osteointegration is desired. In this study, a three-step reaction procedure was employed to immobilize the cell-adhesive peptide Gly-Arg-Gly-Asp (GRGD) on the surface of PEEK sheet by covalent chemical attachment to favor cell adhesion and proliferation. First, hydroxylation-pretreated PEEK surfaces were silanized with 7-Oct-1-enyltrichlorosilane (OETS) in dry cyclohexane, resulting in a silanization layer with terminal ethenyl. Second, the terminal ethylenic double bonds of the silanization layer on PEEK surface were converted to carboxyl groups through acidic potassium manganate oxidation. Finally, GRGD was covalently attached by carbodiimide mediated condensation between the carboxyl on PEEK surface and amine presents in GRGD. X-ray photoelectron spectroscopy (XPS), attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy, surface profiler and water contact angle measurements were applied to characterize the modified surfaces. The effect of cells attachment and proliferation on each specimen was investigated. Pre-osteoblast cells (MC3T3-E1) attachment, spreading and proliferation

Covalent attachment of cell-adhesive peptide Gly-Arg-Gly-Asp (GRGD) to poly(etheretherketone) surface by tailored silanization layers technique

Energy Technology Data Exchange (ETDEWEB)

Zheng, Yanyan [Chengdu Institute of Organic Chemistry, Chinese Academy of Sciences, Chengdu 610041 (China); University of Chinese Academy of Sciences, Beijing 100049 (China); Xiong, Chengdong [Chengdu Institute of Organic Chemistry, Chinese Academy of Sciences, Chengdu 610041 (China); Li, Xiaoyu [State Key Laboratory of Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu 610041 (China); Zhang, Lifang, E-mail: zhanglfcioc@163.com [Chengdu Institute of Organic Chemistry, Chinese Academy of Sciences, Chengdu 610041 (China)

2014-11-30

Highlights: • The carbonyl groups on PEEK surface were effectively reduced to hydroxyl groups using sodium borohydride. • Silanization layers technique was employed to immobilize the cell-adhesive peptide Gly-Arg-Gly-Asp (GRGD) on hydroxylation-pretreated PEEK sheet surface by covalent chemical attachment. • XPS, surface profiler and water contact angle measurements proved the presence of GRGD on PEEK surface. • Osteoblast-like cells (MC3T3-E1) attachment and proliferation were improved effectively on GRGD-modified PEEK surface. - Abstract: Poly(etheretherketone) (PEEK) is a rigid semicrystalline polymer that combines excellent mechanical properties, broad chemical resistance and bone-like stiffness and is widely used in biomedical fields. However, PEEK is naturally bioinert, leading to limited biomedical applications, especially when a direct bone-implant osteointegration is desired. In this study, a three-step reaction procedure was employed to immobilize the cell-adhesive peptide Gly-Arg-Gly-Asp (GRGD) on the surface of PEEK sheet by covalent chemical attachment to favor cell adhesion and proliferation. First, hydroxylation-pretreated PEEK surfaces were silanized with 7-Oct-1-enyltrichlorosilane (OETS) in dry cyclohexane, resulting in a silanization layer with terminal ethenyl. Second, the terminal ethylenic double bonds of the silanization layer on PEEK surface were converted to carboxyl groups through acidic potassium manganate oxidation. Finally, GRGD was covalently attached by carbodiimide mediated condensation between the carboxyl on PEEK surface and amine presents in GRGD. X-ray photoelectron spectroscopy (XPS), attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy, surface profiler and water contact angle measurements were applied to characterize the modified surfaces. The effect of cells attachment and proliferation on each specimen was investigated. Pre-osteoblast cells (MC3T3-E1) attachment, spreading and proliferation

Modifying the red cell surface: towards an ABO-universal blood supply

DEFF Research Database (Denmark)

Olsson, Martin L; Clausen, Henrik

2007-01-01

with these novel enzymes resulting in ECO RBCs typing as O can now be achieved with low enzyme protein consumption, short incubation times and at neutral pH. Presently, clinical trials evaluating safety and efficacy of ECO RBCs are ongoing. Here, we review the status of the ECO technology, its impact and potential...... for introduction into clinical component preparation laboratories....

Improved cellular response of ion modified poly(lactic acid-co-glycolic acid) substrates for mouse fibroblast cells

Energy Technology Data Exchange (ETDEWEB)

Adhikari, Ananta Raj, E-mail: aa8381@gmail.com [Department of Sciences, Wentworth Institute of Technology, Boston MA 02115 (United States); Geranpayeh, Tanya [Department of Biology and Biochemistry, University of Houston, Houston, TX 77204 (United States); Chu, Wei Kan [Texas Center for Superconductivity, University of Houston, Houston, TX 77204 (United States); Department of Physics, University of Houston, Houston, TX 77204 (United States); Otteson, Deborah C. [Department of Biology and Biochemistry, University of Houston, Houston, TX 77204 (United States); Department of Basic and Vision Sciences, College of Optometry, University of Houston, Houston, TX 77204 (United States)

2016-03-01

In this report, the effects of argon (Ar) ion irradiation on poly(lactic acid-co-glycolic acid) (PLGA) substrates on biocompatibility were studied. PLGA scaffold substrates were prepared by spin coating glass surfaces with PLGA dissolved in anhydrous chloroform. Previously, we showed that surface modifications of PLGA films using ion irradiation modulate the inherent hydrophobicity of PLGA surface. Here we show that with increasing ion dose (1 × 10{sup 12} to 1 × 10{sup 14} ions/cm{sup 2}), hydrophobicity and surface roughness decreased. Biocompatibility for NIH3T3 mouse fibroblast cells was increased by argon irradiation of PLGA substrates. On unirradiated PLGA films, fibroblasts had a longer doubling time and cell densities were 52% lower than controls after 48 h in vitro. Argon irradiated PLGA substrates supported growth rates similar to control. Despite differences in cell cycle kinetics, there was no detectible cytotoxicity observed on any substrate. This demonstrates that argon ion irradiation can be used to tune the surface microstructure and generate substrates that are more compatible for the cell growth and proliferation. - Highlights: • Argon irradiation modifies surface chemistry and increases hydrophilicity of poly(lactic-glycolic) acid (PLGA) films. • Both native and irradiated PLGA films were not cytotoxic for mouse fibroblasts. • Fibroblast proliferation increased on PLGA substrates modified with higher doses of Argon irradiation. • Surface modification with Argon irradiation increases biocompatibility of PLGA films.

Surface modifications of magnesium alloys for biomedical applications.

Science.gov (United States)

Yang, Jingxin; Cui, Fuzhai; Lee, In Seop

2011-07-01

In recent years, research on magnesium (Mg) alloys had increased significantly for hard tissue replacement and stent application due to their outstanding advantages. Firstly, Mg alloys have mechanical properties similar to bone which avoid stress shielding. Secondly, they are biocompatible essential to the human metabolism as a factor for many enzymes. In addition, main degradation product Mg is an essential trace element for human enzymes. The most important reason is they are perfectly biodegradable in the body fluid. However, extremely high degradation rate, resulting in too rapid loss of mechanical strength in chloride containing environments limits their applications. Engineered artificial biomaterials with appropriate mechanical properties, surface chemistry, and surface topography are in a great demand. As the interaction between the cells and tissues with biomaterials at the tissue--implant interface is a surface phenomenon; surface properties play a major role in determining both the biological response to implants and the material response to the physiological condition. Therefore, the ability to modify the surface properties while preserve the bulk properties is important, and surface modification to form a hard, biocompatible and corrosion resistant modified layer have always been an interesting topic in biomaterials field. In this article, attempts are made to give an overview of the current research and development status of surface modification technologies of Mg alloys for biomedical materials research. Further, the advantages/disadvantages of the different methods and with regard to the most promising method for Mg alloys are discussed. Finally, the scientific challenges are proposed based on own research and the work of other scientists.

Recent advances in yeast cell-surface display technologies for waste biorefineries.

Science.gov (United States)

Liu, Zhuo; Ho, Shih-Hsin; Hasunuma, Tomohisa; Chang, Jo-Shu; Ren, Nan-Qi; Kondo, Akihiko

2016-09-01

Waste biorefinery aims to maximize the output of value-added products from various artificial/agricultural wastes by using integrated bioprocesses. To make waste biorefinery economically feasible, it is thus necessary to develop a low-cost, environment-friendly technique to perform simultaneous biodegradation and bioconversion of waste materials. Cell-surface display engineering is a novel, cost-effective technique that can auto-immobilize proteins on the cell exterior of microorganisms, and has been applied for use with waste biofinery. Through tethering different enzymes (e.g., cellulase, lipase, and protease) or metal-binding peptides on cell surfaces, various yeast strains can effectively produce biofuels and biochemicals from sugar/protein-rich waste materials, catalyze waste oils into biodiesels, or retrieve heavy metals from wastewater. This review critically summarizes recent applications of yeast cell-surface display on various types of waste biorefineries, highlighting its potential and future challenges with regard to commercializing this technology. Copyright © 2016 Elsevier Ltd. All rights reserved.

The ultrasound technology for modifying enzyme activity

Directory of Open Access Journals (Sweden)

Meliza Lindsay Rojas

2016-01-01

Full Text Available Enzymes are protein complexes compounds widely studied and used due to their ability to catalyze reactions. The food processing mainly a ims the inactivation of enzymes due to various undesirable effects. However, there are many processes that can be optimized by its catalytic activity. In this context, different technologies have been applied both to inactivate or to improve the enzymes ef ficiency. The Ultrasound technology emerges as an alternative mainly applied to achieve the enzyme inactivation. On the contrary, very few investigations show the ability of this technology under certain conditions to achieve the opposite effect (i.e. increase the catalytic activity of enzymes. The objective of this study was to correlate the ultrasonic energy delivered to the sample (J/mL with the residual enzymatic activity and explain the possible mechanisms which results in the enzymatic activation/in activation complex behavior. The activity of POD in coconut water was evaluated as a model. The enzymatic activity initially increased, followed by reduction with a trend to enzyme inactivation. This complex behavior is directly related to the applied ultr asonic energy and their direct mechanical effects on the product, as well as the effect in the enzymatic infinite intermediate states and its structural conformation changes. The obtained results are useful for both academic and industrial perspectives.

Photocatalysis of Modified Transition Metal Oxide Surfaces

Energy Technology Data Exchange (ETDEWEB)

Batzill, Matthias [Univ. of South Florida, Tampa, FL (United States). Dept. of Physics

2018-02-28

The goal of this project has been to establish a cause-effect relationship for photocatalytic activity variations of different structures of the same material; and furthermore gain fundamental understanding on modification of photocatalysts by compositional or surface modifications. The reasoning is that gaining atomic scale understanding of how surface and bulk modifications alter the photo reactivity will lead to design principles for next generation photocatalysts. As a prototypical photocatalyst the research focused on TiO₂ synthesized in well-defined single crystalline form to enable fundamental characterizations.We have obtained results in the following areas: (a) Preparation of epitaxial anatase TiO₂ samples by pulsed laser deposition. (b) Comparison of hydrogen diffusion on different crystallographic surface. (c) Determining the stability of the TiO₂(011)-2x1 reconstruction upon interactions with adsorbates. (d) Characterization of adsorption and (thermal and photo) reaction of molecules with nitro-endgroups, (e) Exploring the possibility of modifying planar model photocatalyst surfaces with graphene to enable fundamental studies on reported enhanced photocatalytic activities of graphene modified transition metal oxides, (f) gained fundamental understanding on the role of crystallographic polymorphs of the same material for their photocatalytic activities.

Metabolic signature of breast cancer cell line MCF-7: profiling of modified nucleosides via LC-IT MS coupling

Directory of Open Access Journals (Sweden)

Gleiter Christoph H

2007-11-01

Full Text Available Abstract Background Cancer, like other diseases accompanied by strong metabolic disorders, shows characteristic effects on cell turnover rate, activity of modifying enzymes and DNA/RNA modifications, resulting also in elevated amounts of excreted modified nucleosides. For a better understanding of the impaired RNA metabolism in breast cancer cells, we screened these metabolites in the cell culture supernatants of the breast cancer cell line MCF-7 and compared it to the human mammary epithelial cells MCF-10A. The nucleosides were isolated and analyzed via 2D-chromatographic techniques: In the first dimension by cis-diol specific boronate affinity extraction and subsequently by reversed phase chromatography coupled to an ion trap mass spectrometer. Results Besides the determination of ribonucleosides, additional compounds with cis-diol structure, deriving from cross-linked biochemical pathways, like purine-, histidine- and polyamine metabolism were detected. In total, 36 metabolites were identified by comparison of fragmentation patterns and retention time. Relation to the internal standard isoguanosine yielded normalized area ratios for each identified compound and enabled a semi-quantitative metabolic signature of both analyzed cell lines. 13 of the identified 26 modified ribonucleosides were elevated in the cell culture supernatants of MCF-7 cells, with 5-methyluridine, N2,N2,7-trimethylguanosine, N6-methyl-N6-threonylcarbamoyladenosine and 3-(3-aminocarboxypropyl-uridine showing the most significant differences. 1-ribosylimidazole-4-acetic acid, a histamine metabolite, was solely found in the supernatants of MCF-10A cells, whereas 1-ribosyl-4-carboxamido-5-aminoimidazole and S-adenosylmethionine occurred only in supernatants of MCF-7 cells. Conclusion The obtained results are discussed against the background of pathological changes in cell metabolism, resulting in new perspectives for modified nucleosides and related metabolites as possible

Surface-Enhanced Raman Scattering Nanoparticles as Optical Labels for Imaging Cell Surface Proteins

Science.gov (United States)

MacLaughlin, Christina M.

Assaying the expression of cell surface proteins has widespread application for characterizing cell type, developmental stage, and monitoring disease transformation. Immunophenotyping is conducted by treating cells with labelled targeting moieties that have high affinity for relevant surface protein(s). The sensitivity and specificity of immunophenotyping is defined by the choice of contrast agent and therefore, the number of resolvable signals that can be used to simultaneously label cells. Narrow band width surface-enhanced Raman scattering (SERS) nanoparticles are proposed as optical labels for multiplexed immunophenotying. Two types of surface coatings were investigated to passivate the gold nanoparticles, incorporate SERS functionality, and to facilitate attachment of targeting antibodies. Thiolated poly(ethylene glycol) forms dative bonds with the gold surface and is compatible with multiple physisorbed Raman-active reporter molecules. Ternary lipid bilayers are used to encapsulate the gold nanoparticles particles, and incorporate three different classes of Raman reporters. TEM, UV-Visible absorbance spectroscopy, DLS, and electrophoretic light scattering were used characterize the particle coating. Colourimetric protein assay, and secondary antibody labelling were used to quantify the antibody conjugation. Three different in vitromodels were used to investigate the binding efficacy and specificity of SERS labels for their biomarker targets. Primary human CLL cells, LY10 B lymphoma, and A549 adenocarcinoma lines were targeted. Dark field imaging was used to visualize the colocalization of SERS labels with cells, and evidence of receptor clustering was obtained based on colour shifts of the particles' Rayleigh scattering. Widefield, and spatially-resolved Raman spectra were used to detect labels singly, and in combination from labelled cells. Fluorescence flow cytometry was used to test the particles' binding specificity, and SERS from labelled cells was also

Metabolic enzymes: key modulators of functionality in cancer stem-like cells.

Science.gov (United States)

Dong, Bo-Wen; Qin, Guang-Ming; Luo, Yan; Mao, Jian-Shan

2017-02-21

Cancer Stem-like Cells (CSCs) are a subpopulation of cancer cells with self-renewal capacity and are important for the initiation, progression and recurrence of cancer diseases. The metabolic profile of CSCs is consistent with their stem-like properties. Studies have indicated that enzymes, the main regulators of cellular metabolism, dictate functionalities of CSCs in both catalysis-dependent and catalysis-independent manners. This paper reviews diverse studies of metabolic enzymes, and describes the effects of these enzymes on metabolic adaptation, gene transcription and signal transduction, in CSCs.

Culture conditions affect photoreactivating enzyme levels in human fibroblasts

International Nuclear Information System (INIS)

Sutherland, B.M.; Oliver, R.

1976-01-01

Photoreactivation of pyrimidine dimers occured under the experimental conditions given in this study, but has not been observed under conditions used by others. Three possible differences were tested in experimental procedures including dimer separation and analysis methods, illumination conditions and cell culture techniques. The methods in this study of dimer separation and analysis indeed measure cis-syn pyrimidine dimers and give results in quantitative agreement with the methods of others. It was found that white light pre-illumination of fibroblasts from the xeroderma pigmentosum line XP12BE or of normal cells does not affect the cellular capacity for dimer photoreactivation. However, the cell culture conditions can affect photoreactivating enzyme levels, and thus cellular dimer photoreactivation capacity. Cells grown in Eagle's minimal essential medium (supplemented with 15% fetal bovine serum) contain very low levels of photoreactivating enzyme and cannot photoreactivate dimers in their DNA; but companion cultures maintained in Dulbecco's modified Eagle's minimal medium do contain photoreactivating enzyme and can reactivate photoreactive cellular dimers

Study of wettability of calcite surfaces using oil-brine-enzyme systems for enhanced oil recovery applications

DEFF Research Database (Denmark)

Khusainova, Alsu; Nielsen, Sidsel Marie; Pedersen, Hanne Høst

2015-01-01

and adhesion behaviour tests. Comparative studies with a surfactant, protein, purified enzyme, enzyme stabiliser using n-decane (as a model for the oil) have also been carried out in order to verify experimental results. The enzymes that have the highest effect on the wettability have been identified. Those...... action has been found to be replacement of oil at the solid surface by the enzyme. Other mechanisms (modification of the surface tension or catalytic modification of hydrocarbons resulting in reducing the oil viscosity) have shown to be much less pronounced from the measurements reported here....

Cell adhesive ability of a biological foam ceramic with surface modification

International Nuclear Information System (INIS)

Zhang Yong; Li Xiaoyu; Feng Fan; Lin Yunfeng; Liao Yunmao; Tian, Weidong; Liu Lei

2008-01-01

Biological foam ceramic is a promising material for tissue engineering scaffold because of its biocompatibility, biodegradation and adequate pores measured from micrometer to nanometers. The aim of this study was to evaluate the adhesion and proliferation of adipose-derived stromal cells (ADSCs) on the biological foam ceramic coated with fibronectin. ADSCs were harvested from SD rats and passaged three times prior to seeding onto biological foam surface modified with fibronectin (50 μg/ml). Scaffold without surface modification served as control. To characterize cellular attachment, cells were incubated on the scaffold for 1 h and 3 h and then the cells attached onto the scaffold were counted. The difference of proliferation was appraised using MTT assay at day 1, 3, 5 and 7 before the cells reached confluence. After 7 days of culture, scanning electron microscope (SEM) was chosen to assess cell morphology and attachment of ADSCs on the biological foam ceramic. Attachment of ADSCs on the biological foam ceramic surface modified with fibronectin at 1 h or 3 h was substantially greater than that in control. MTT assay revealed that ADSCs proliferation tendency of the experimental group was nearly parallel to that of control. SEM view showed that ADSCs in the experimental groups connected more tightly and excreted more collagen than that in control. The coating of fibronectin could improve the cell adhesive ability of biological foam ceramics without evident effect on proliferation

Enzyme activity assay of glycoprotein enzymes based on a boronate affinity molecularly imprinted 96-well microplate.

Science.gov (United States)

Bi, Xiaodong; Liu, Zhen

2014-12-16

Enzyme activity assay is an important method in clinical diagnostics. However, conventional enzyme activity assay suffers from apparent interference from the sample matrix. Herein, we present a new format of enzyme activity assay that can effectively eliminate the effects of the sample matrix. The key is a 96-well microplate modified with molecularly imprinted polymer (MIP) prepared according to a newly proposed method called boronate affinity-based oriented surface imprinting. Alkaline phosphatase (ALP), a glycoprotein enzyme that has been routinely used as an indicator for several diseases in clinical tests, was taken as a representative target enzyme. The prepared MIP exhibited strong affinity toward the template enzyme (with a dissociation constant of 10(-10) M) as well as superb tolerance for interference. Thus, the enzyme molecules in a complicated sample matrix could be specifically captured and cleaned up for enzyme activity assay, which eliminated the interference from the sample matrix. On the other hand, because the boronate affinity MIP could well retain the enzymatic activity of glycoprotein enzymes, the enzyme captured by the MIP was directly used for activity assay. Thus, additional assay time and possible enzyme or activity loss due to an enzyme release step required by other methods were avoided. Assay of ALP in human serum was successfully demonstrated, suggesting a promising prospect of the proposed method in real-world applications.

Dynamic moisture sorption characteristics of enzyme-resistant recrystallized cassava starch.

Science.gov (United States)

Mutungi, Christopher; Schuldt, Stefan; Onyango, Calvin; Schneider, Yvonne; Jaros, Doris; Rohm, Harald

2011-03-14

The interaction of moisture with enzyme-resistant recrystallized starch, prepared by heat-moisture treatment of debranched acid-modified or debranched non-acid-modified cassava starch, was investigated in comparison with the native granules. Crystallinities of the powdered products were estimated by X-ray diffraction. Moisture sorption was determined using dynamic vapor sorption analyzer and data fitted to various models. Percent crystallinities of native starch (NS), non-acid-modified recrystallized starch (NAMRS), and acid-modified recrystallized starch (AMRS) were 39.7, 51.9, and 56.1%, respectively. In a(w) below 0.8, sorption decreased in the order NS > NAMRS > AMRS in line with increasing sample crystallinities but did not follow this crystallinity dependence at higher a(w) because of condensation and polymer dissolution effects. Adsorbed moisture became internally absorbed in NS but not in NAMRS and AMRS, which might explain the high resistance of the recrystallized starches to digestion because enzyme and starch cannot approach each other over fairly sufficient surface at the molecular level.

Surface noble metal modified PdM/C (M = Ru, Pt, Au) as anode catalysts for direct ethanol fuel cells

International Nuclear Information System (INIS)

Mao, Han; Huang, Tao; Yu, Aishui

2016-01-01

In this article, we studied the surface noble metal modification on Pd nanoparticles, other than the homogeneous or core-shell structure. The surface modification will lead to the uneven constitution within the nanoparticles and thus more obvious optimization effect toward the catalyst brought by the lattice deformation. The surface of the as-prepared Pd nanoparticles was modified with Ru, Pt or Au by a moderate and green approach, respectively. XPS results confirm the interactive electron effects between Pd and the modified noble metal. Electrochemical measurements show that the surface noble metal modified catalysts not only show higher catalytic activity, but also better stability and durability. The PdM/C catalysts all exhibit good dispersion and very little agglomeration after long-term potential cycles toward ethanol oxidation. With only 10% metallic atomic ratio of Au, PdAu/C catalyst shows extraordinary catalytic activity and stability, the peak current reaches 1700 mA mg"−"1 Pd, about 2.5 times that of Pd/C. Moreover, the PdAu/C maintains 40% of the catalytic activity after 4500 potential cycles. - Highlights: • Pd-based catalysts with complicated exposed facets. • Much enhanced electrocatalytic activity and stability with about 10% noble metal M (M = Ru, Pt, Au) on Pd nanoparticles. • The outstanding electrocatalytic performance of PdAu/C towards ethanol oxidation after the Au modification.

Surface noble metal modified PdM/C (M = Ru, Pt, Au) as anode catalysts for direct ethanol fuel cells

Energy Technology Data Exchange (ETDEWEB)

Mao, Han; Huang, Tao, E-mail: huangt@fudan.edu.cn; Yu, Aishui, E-mail: asyu@fudan.edu.cn

2016-08-15

In this article, we studied the surface noble metal modification on Pd nanoparticles, other than the homogeneous or core-shell structure. The surface modification will lead to the uneven constitution within the nanoparticles and thus more obvious optimization effect toward the catalyst brought by the lattice deformation. The surface of the as-prepared Pd nanoparticles was modified with Ru, Pt or Au by a moderate and green approach, respectively. XPS results confirm the interactive electron effects between Pd and the modified noble metal. Electrochemical measurements show that the surface noble metal modified catalysts not only show higher catalytic activity, but also better stability and durability. The PdM/C catalysts all exhibit good dispersion and very little agglomeration after long-term potential cycles toward ethanol oxidation. With only 10% metallic atomic ratio of Au, PdAu/C catalyst shows extraordinary catalytic activity and stability, the peak current reaches 1700 mA mg{sup −1} Pd, about 2.5 times that of Pd/C. Moreover, the PdAu/C maintains 40% of the catalytic activity after 4500 potential cycles. - Highlights: • Pd-based catalysts with complicated exposed facets. • Much enhanced electrocatalytic activity and stability with about 10% noble metal M (M = Ru, Pt, Au) on Pd nanoparticles. • The outstanding electrocatalytic performance of PdAu/C towards ethanol oxidation after the Au modification.

Studies on a photoreactivating enzyme from Drosophila melanogaster cultured cells

International Nuclear Information System (INIS)

Beck, L.A.

1982-01-01

A photoreactivating enzyme was purified from Schneider's Line No. 2 Drosophila melanogaster cultured cells. DEAE cellulose chromatography with high potassium phosphate buffer conditions was used to separate nucleic acids from the protein component of the crude cell extract. The protein pass-through fraction from DEAE cellulose was chromatographed on phosphocellulose followed by hydroxylapatite, using linear potassium phosphate gradients to elute the enzyme. Gel filtration chromatography on Sephacryl S-200 resulted in a 4500-fold purification of the enzyme with a final recovery of 4%. The enzyme has an apparent gel filtration molecular weight of 32,900 (+/- 1350 daltons) and an isoelectric pH of 4.9. Optimum ionic strength for activity is 0.17 at pH 6.5 in potassium phosphate buffer. The action spectrum for photoreactivation in Drosophila has an optimum at 365 nm with a response to wavelengths in the range of 313 to 465 nm. Drosophila photoreactivating enzyme contains an essential RNA that is necessary for activity in vitro. The ability of the enzyme to photoreactivate dimers in vitro is abolished by treatment of the enzyme with ribonucleases, or by disruption of the enzyme-RNA complex by electrophoresis or adsorption to DEAE cellulose. The essential RNA is heterogeneous in size but contains a 10-12 base region that may interact with the active site of the enzyme, and thus is protected from degradation by contaminating RNase activities during purification. The RNA is thought to stabilize the photoreactivating enzyme by maintaining the enzyme in the proper configuration for binding to dimer-containing DNA. It is not known whether this RNA is essential for in vivo photoreactivation

The plant cell wall in the feeding sites of cyst nematodes.

Science.gov (United States)

Bohlmann, Holger; Sobczak, Miroslaw

2014-01-01

Plant parasitic cyst nematodes (genera Heterodera and Globodera) are serious pests for many crops. They enter the host roots as migratory second stage juveniles (J2) and migrate intracellularly toward the vascular cylinder using their stylet and a set of cell wall degrading enzymes produced in the pharyngeal glands. They select an initial syncytial cell (ISC) within the vascular cylinder or inner cortex layers to induce the formation of a multicellular feeding site called a syncytium, which is the only source of nutrients for the parasite during its entire life. A syncytium can consist of more than hundred cells whose protoplasts are fused together through local cell wall dissolutions. While the nematode produces a cocktail of cell wall degrading and modifying enzymes during migration through the root, the cell wall degradations occurring during syncytium development are due to the plants own cell wall modifying and degrading proteins. The outer syncytial cell wall thickens to withstand the increasing osmotic pressure inside the syncytium. Furthermore, pronounced cell wall ingrowths can be formed on the outer syncytial wall at the interface with xylem vessels. They increase the surface of the symplast-apoplast interface, thus enhancing nutrient uptake into the syncytium. Processes of cell wall degradation, synthesis and modification in the syncytium are facilitated by a variety of plant proteins and enzymes including expansins, glucanases, pectate lyases and cellulose synthases, which are produced inside the syncytium or in cells surrounding the syncytium.

The plant cell wall in the feeding sites of cyst nematodes

Directory of Open Access Journals (Sweden)

Holger eBohlmann

2014-03-01

Full Text Available Plant parasitic cyst nematodes (genera Heterodera and Globodera are serious pests for many crops. They enter the host roots as migratory second stage juveniles (J2 and migrate intracellularly towards the vascular cylinder using their stylet and a set of cell wall degrading enzymes produced in the pharyngeal glands. They select an initial syncytial cell (ISC within the vascular cylinder or inner cortex layers to induce the formation of a multicellular feeding site called a syncytium, which is the only source of nutrients for the parasite during its entire life. A syncytium can consist of more than hundred cells whose protoplasts are fused together through local cell wall dissolutions. While the nematode produces a cocktail of cell wall degrading and modifying enzymes during migration through the root, the cell wall degradations occurring during syncytium development are due to the plants own cell wall modifying and degrading proteins. The outer syncytial cell wall thickens to withstand the increasing osmotic pressure inside the syncytium. Furthermore, pronounced cell wall ingrowths can be formed on the outer syncytial wall at the interface with xylem vessels. They increase the surface of the symplast-apoplast interface, thus enhancing nutrient uptake into the syncytium. Processes of cell wall degradation, synthesis and modification in the syncytium are facilitated by a variety of plant proteins and enzymes including expansins, glucanases, pectate lyases and cellulose synthases, which are produced inside the syncytium or in cells surrounding the syncytium.

Clinical efficacy of gene-modified stem cells in adenosine deaminase-deficient immunodeficiency.

Science.gov (United States)

Shaw, Kit L; Garabedian, Elizabeth; Mishra, Suparna; Barman, Provaboti; Davila, Alejandra; Carbonaro, Denise; Shupien, Sally; Silvin, Christopher; Geiger, Sabine; Nowicki, Barbara; Smogorzewska, E Monika; Brown, Berkley; Wang, Xiaoyan; de Oliveira, Satiro; Choi, Yeong; Ikeda, Alan; Terrazas, Dayna; Fu, Pei-Yu; Yu, Allen; Fernandez, Beatriz Campo; Cooper, Aaron R; Engel, Barbara; Podsakoff, Greg; Balamurugan, Arumugam; Anderson, Stacie; Muul, Linda; Jagadeesh, G Jayashree; Kapoor, Neena; Tse, John; Moore, Theodore B; Purdy, Ken; Rishi, Radha; Mohan, Kathey; Skoda-Smith, Suzanne; Buchbinder, David; Abraham, Roshini S; Scharenberg, Andrew; Yang, Otto O; Cornetta, Kenneth; Gjertson, David; Hershfield, Michael; Sokolic, Rob; Candotti, Fabio; Kohn, Donald B

2017-05-01

Autologous hematopoietic stem cell transplantation (HSCT) of gene-modified cells is an alternative to enzyme replacement therapy (ERT) and allogeneic HSCT that has shown clinical benefit for adenosine deaminase-deficient (ADA-deficient) SCID when combined with reduced intensity conditioning (RIC) and ERT cessation. Clinical safety and therapeutic efficacy were evaluated in a phase II study. Ten subjects with confirmed ADA-deficient SCID and no available matched sibling or family donor were enrolled between 2009 and 2012 and received transplantation with autologous hematopoietic CD34+ cells that were modified with the human ADA cDNA (MND-ADA) γ-retroviral vector after conditioning with busulfan (90 mg/m2) and ERT cessation. Subjects were followed from 33 to 84 months at the time of data analysis. Safety of the procedure was assessed by recording the number of adverse events. Efficacy was assessed by measuring engraftment of gene-modified hematopoietic stem/progenitor cells, ADA gene expression, and immune reconstitution. With the exception of the oldest subject (15 years old at enrollment), all subjects remained off ERT with normalized peripheral blood mononuclear cell (PBMC) ADA activity, improved lymphocyte numbers, and normal proliferative responses to mitogens. Three of nine subjects were able to discontinue intravenous immunoglobulin replacement therapy. The MND-ADA vector was persistently detected in PBMCs (vector copy number [VCN] = 0.1-2.6) and granulocytes (VCN = 0.01-0.3) through the most recent visits at the time of this writing. No patient has developed a leukoproliferative disorder or other vector-related clinical complication since transplant. These results demonstrate clinical therapeutic efficacy from gene therapy for ADA-deficient SCID, with an excellent clinical safety profile. ClinicalTrials.gov NCT00794508. Food and Drug Administration Office of Orphan Product Development award, RO1 FD003005; NHLBI awards, PO1 HL73104 and Z01 HG000122; UCLA

Clinical efficacy of gene-modified stem cells in adenosine deaminase–deficient immunodeficiency

Science.gov (United States)

Shaw, Kit L.; Garabedian, Elizabeth; Mishra, Suparna; Barman, Provaboti; Davila, Alejandra; Carbonaro, Denise; Shupien, Sally; Silvin, Christopher; Geiger, Sabine; Nowicki, Barbara; Smogorzewska, E. Monika; Brown, Berkley; Wang, Xiaoyan; de Oliveira, Satiro; Choi, Yeong; Ikeda, Alan; Terrazas, Dayna; Fu, Pei-Yu; Yu, Allen; Fernandez, Beatriz Campo; Cooper, Aaron R.; Engel, Barbara; Podsakoff, Greg; Balamurugan, Arumugam; Anderson, Stacie; Muul, Linda; Jagadeesh, G. Jayashree; Kapoor, Neena; Tse, John; Moore, Theodore B.; Purdy, Ken; Rishi, Radha; Mohan, Kathey; Skoda-Smith, Suzanne; Buchbinder, David; Abraham, Roshini S.; Scharenberg, Andrew; Yang, Otto O.; Cornetta, Kenneth; Gjertson, David; Hershfield, Michael; Sokolic, Rob; Candotti, Fabio

2017-01-01

BACKGROUND. Autologous hematopoietic stem cell transplantation (HSCT) of gene-modified cells is an alternative to enzyme replacement therapy (ERT) and allogeneic HSCT that has shown clinical benefit for adenosine deaminase–deficient (ADA-deficient) SCID when combined with reduced intensity conditioning (RIC) and ERT cessation. Clinical safety and therapeutic efficacy were evaluated in a phase II study. METHODS. Ten subjects with confirmed ADA-deficient SCID and no available matched sibling or family donor were enrolled between 2009 and 2012 and received transplantation with autologous hematopoietic CD34+ cells that were modified with the human ADA cDNA (MND-ADA) γ-retroviral vector after conditioning with busulfan (90 mg/m2) and ERT cessation. Subjects were followed from 33 to 84 months at the time of data analysis. Safety of the procedure was assessed by recording the number of adverse events. Efficacy was assessed by measuring engraftment of gene-modified hematopoietic stem/progenitor cells, ADA gene expression, and immune reconstitution. RESULTS. With the exception of the oldest subject (15 years old at enrollment), all subjects remained off ERT with normalized peripheral blood mononuclear cell (PBMC) ADA activity, improved lymphocyte numbers, and normal proliferative responses to mitogens. Three of nine subjects were able to discontinue intravenous immunoglobulin replacement therapy. The MND-ADA vector was persistently detected in PBMCs (vector copy number [VCN] = 0.1–2.6) and granulocytes (VCN = 0.01–0.3) through the most recent visits at the time of this writing. No patient has developed a leukoproliferative disorder or other vector-related clinical complication since transplant. CONCLUSION. These results demonstrate clinical therapeutic efficacy from gene therapy for ADA-deficient SCID, with an excellent clinical safety profile. TRIAL REGISTRATION. ClinicalTrials.gov NCT00794508. FUNDING. Food and Drug Administration Office of Orphan Product

Covalent immobilization of invertase on PAMAM-dendrimer modified superparamagnetic iron oxide nanoparticles

International Nuclear Information System (INIS)

Uzun, K.; Cevik, E.; Senel, M.; Soezeri, H.; Baykal, A.; Abasiyanik, M. F.; Toprak, M. S.

2010-01-01

In this study, polyamidoamine (PAMAM) dendrimer was synthesized on the surface of superparamagnetite nanoparticles to enhance invertase immobilization. The amount of immobilized enzyme on the surface-hyperbranched magnetite nanoparticle was up to 2.5 times (i.e., 250%) as much as that of magnetite nanoparticle modified with only amino silane. Maximum reaction rate (V max ) and Michaelis-Menten constant (K m ) were determined for the free and immobilized enzymes. Various characteristics of immobilized invertase such as; the temperature activity, thermal stability, operational stability, and storage stability were evaluated and results revealed that stability of the enzyme is improved upon immobilization.

Cellular redox homeostasis in endothelial cells treated with nonmodified and Fenton-modified nanodiamond powders.

Science.gov (United States)

Solarska-Ściuk, K; Gajewska, A; Skolimowski, J; Gajek, A; Bartosz, G

2014-01-01

Diamond nanoparticles find numerous applications in pharmacy, medicine, cosmetics, and biotechnology. However, possible adverse cellular effects of diamond nanoparticle cells have been reported, which may limit their use. The aim of this study was to compare the effect of nonmodified diamond nanoparticles (D) and diamond nanoparticles modified by the Fenton reaction (D+OH) on human umbilical cord endothelial cells (HUVEC-ST). We found that both D and D+OH show time- and concentration-dependent cytotoxicity, inducing apoptosis and necrosis of HUVEC-ST. Interaction with D and D+OH also induced changes in the production of reactive oxygen and nitrogen species and changes in the level of glutathione and activities of antioxidant enzymes in the cells. These data demonstrate that diamond nanoparticles may induce oxidative stress in human endothelial cells, which contributes to their cytotoxic effects seen at higher concentrations of D and D+OH. © 2014 International Union of Biochemistry and Molecular Biology, Inc.

Bone induction by surface-double-modified true bone ceramics in vitro and in vivo

International Nuclear Information System (INIS)

Li, Jingfeng; Chen, Liaobin; Deng, Yu; Zheng, Qixin; Guo, Xiaodong; Zou, Zhenwei; Liu, Yudong; Lan, Shenghui

2013-01-01

True bone ceramic (TBC), obtained by twice sintering fresh bovine cancellous bone at high temperatures, is an osteoconductive and bioactive bone substitute material that exhibits excellent biocompatibility with hard tissue. The authors have previously synthesized a novel BMP-2-related peptide, P24, and found that it could enhance the osteoblastic differentiation of cells. The objective of the present study was to construct a double-modified TBC via mineralization into simulated body fluid and P24 incorporation for enhanced bone formation. In vitro experiments revealed that surface mineralization-modified (SMM) TBC scaffolds demonstrated efficiency for sustained release of P24. The P24/SMM-TBC composite exhibited increased osteogenic activity by cell adhesion rate determination, MTT assay, alkaline phosphatase staining, and calcium nodule staining with alizarin red compared with SMM-TBC and TBC. In vivo studies showed that the P24/SMM-TBC composite scaffold promoted significant bone defect repair, in marked contrast to stand-alone SMM-TBC and TBC, based on the results of radiographic evaluation and histological examination. These findings indicate that SMM-TBC is a good scaffold for the controlled release of P24 and that the P24/SMM-TBC composite could improve the adhesion, proliferation and differentiation of cells and repair bone defects. The double-modified P24/SMM-TBC composite biomaterial shows potential for clinical application in bone tissue engineering. (paper)

Cell in situ zymography: an in vitro cytotechnology for localization of enzyme activity in cell culture.

Science.gov (United States)

Chhabra, Aastha; Jaiswal, Astha; Malhotra, Umang; Kohli, Shrey; Rani, Vibha

2012-09-01

In situ zymography is a unique technique for detection and localization of enzyme-substrate interactions majorly in histological sections. Substrate with quenched fluorogenic molecule is incorporated in gel over which tissue sections are mounted and then incubated in buffer. The enzymatic activity is observed in the form of fluorescent signal. With the advancements in the field of biological research, use of in vitro cell culture has become very popular and holds great significance in multiple fields including inflammation, cancer, stem cell biology and the still emerging 3-D cell cultures. The information on analysis of enzymatic activity in cell lines is inadequate presently. We propose a single-step methodology that is simple, sensitive, cost-effective, and functional to perform and study the 'in position' activity of enzyme on substrate for in vitro cell cultures. Quantification of enzymatic activity to carry out comparative studies on cells has also been illustrated. This technique can be applied to a variety of enzyme classes including proteases, amylases, xylanases, and cellulases in cell cultures.

[Treatment of surface burns with proteolytic enzymes: mathematic description of lysis kinetics].

Science.gov (United States)

Domogatskaia, A S; Domogatskiĭ, S P; Ruuge, E K

2003-01-01

The lysis of necrotic tissue by a proteolytic enzyme applied to the surface of a burn wound was studied. A mathematical model was proposed, which describes changes in the thickness of necrotic tissue as a function of the proteolytic activity of the enzyme. The model takes into account the inward-directed diffusion of the enzyme, the counterflow of interstitial fluid (exudates) containing specific inhibitors, and the extracellular matrix proteolysis. It was shown in terms of the quasi-stationary approach that the thickness of the necrotic tissue layer decreases exponentially with time; i.e., the lysis slows down as the thickness of the necrotic tissue layer decreases. The dependence of the characteristic time of this decrease on enzyme concentration was obtained. It was shown that, at high enzyme concentrations (more than 5 mg/ml), the entire time of lysis (after the establishment of quasi-stationary equilibrium) is inversely proportional to the concentration of the enzyme.

A modified stanton number for heat transfer through fabric surface

Directory of Open Access Journals (Sweden)

Zhang Shen-Zhong

2015-01-01

Full Text Available The Stanton number was originally proposed for describing heat transfer through a smooth surface. A modified one is suggested in this paper to take into account non-smooth surface or fractal surface. The emphasis is put on the heat transfer through fabrics.

Improvement of in vitro corrosion and cytocompatibility of biodegradable Fe surface modified by Zn ion implantation

Science.gov (United States)

Wang, Henan; Zheng, Yang; Li, Yan; Jiang, Chengbao

2017-05-01

Pure Fe was surface-modified by Zn ion implantation to improve the biodegradable behavior and cytocompatibility. Surface topography, chemical composition, corrosion resistance and cytocompatibility were investigated. Atomic force microscopy, auger electron spectroscopy and X-ray photoelectron spectroscopy results showed that Zn was implanted into the surface of pure Fe in the depth of 40-60 nm and Fe2O3/ZnO oxides were formed on the outmost surface. Electrochemical measurements and immersion tests revealed an improved degradable behavior for the Zn-implanted Fe samples. An approximately 12% reduction in the corrosion potential (Ecorr) and a 10-fold increase in the corrosion current density (icorr) were obtained after Zn ion implantation with a moderate incident ion dose, which was attributed to the enhanced pitting corrosion. The surface free energy of pure Fe was decreased by Zn ion implantation. The results of direct cell culture indicated that the short-term (4 h) cytocompatibility of MC3T3-E1 cells was promoted by the implanted Zn on the surface.

Fluorogenic Substrates for Visualizing Acidic Organelle Enzyme Activities.

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Fiona Karen Harlan

Full Text Available Lysosomes are acidic cytoplasmic organelles that are present in all nucleated mammalian cells and are involved in a variety of cellular processes including repair of the plasma membrane, defense against pathogens, cholesterol homeostasis, bone remodeling, metabolism, apoptosis and cell signaling. Defects in lysosomal enzyme activity have been associated with a variety of neurological diseases including Parkinson's Disease, Lysosomal Storage Diseases, Alzheimer's disease and Huntington's disease. Fluorogenic lysosomal staining probes were synthesized for labeling lysosomes and other acidic organelles in a live-cell format and were shown to be capable of monitoring lysosomal metabolic activity. The new targeted substrates were prepared from fluorescent dyes having a low pKa value for optimum fluorescence at the lower physiological pH found in lysosomes. They were modified to contain targeting groups to direct their accumulation in lysosomes as well as enzyme-cleavable functions for monitoring specific enzyme activities using a live-cell staining format. Application to the staining of cells derived from blood and skin samples of patients with Metachromatic Leukodystrophy, Krabbe and Gaucher Diseases as well as healthy human fibroblast and leukocyte control cells exhibited localization to the lysosome when compared with known lysosomal stain LysoTracker® Red DND-99 as well as with anti-LAMP1 Antibody staining. When cell metabolism was inhibited with chloroquine, staining with an esterase substrate was reduced, demonstrating that the substrates can be used to measure cell metabolism. When applied to diseased cells, the intensity of staining was reflective of lysosomal enzyme levels found in diseased cells. Substrates specific to the enzyme deficiencies in Gaucher or Krabbe disease patient cell lines exhibited reduced staining compared to that in non-diseased cells. The new lysosome-targeted fluorogenic substrates should be useful for research

Mitochondrial thiol modification by a targeted electrophile inhibits metabolism in breast adenocarcinoma cells by inhibiting enzyme activity and protein levels

Directory of Open Access Journals (Sweden)

M. Ryan Smith

2016-08-01

Full Text Available Many cancer cells follow an aberrant metabolic program to maintain energy for rapid cell proliferation. Metabolic reprogramming often involves the upregulation of glutaminolysis to generate reducing equivalents for the electron transport chain and amino acids for protein synthesis. Critical enzymes involved in metabolism possess a reactive thiolate group, which can be modified by certain oxidants. In the current study, we show that modification of mitochondrial protein thiols by a model compound, iodobutyl triphenylphosphonium (IBTP, decreased mitochondrial metabolism and ATP in MDA-MB 231 (MB231 breast adenocarcinoma cells up to 6 days after an initial 24 h treatment. Mitochondrial thiol modification also depressed oxygen consumption rates (OCR in a dose-dependent manner to a greater extent than a non-thiol modifying analog, suggesting that thiol reactivity is an important factor in the inhibition of cancer cell metabolism. In non-tumorigenic MCF-10A cells, IBTP also decreased OCR; however the extracellular acidification rate was significantly increased at all but the highest concentration (10 µM of IBTP indicating that thiol modification can have significantly different effects on bioenergetics in tumorigenic versus non-tumorigenic cells. ATP and other adenonucleotide levels were also decreased by thiol modification up to 6 days post-treatment, indicating a decreased overall energetic state in MB231 cells. Cellular proliferation of MB231 cells was also inhibited up to 6 days post-treatment with little change to cell viability. Targeted metabolomic analyses revealed that thiol modification caused depletion of both Krebs cycle and glutaminolysis intermediates. Further experiments revealed that the activity of the Krebs cycle enzyme, aconitase, was attenuated in response to thiol modification. Additionally, the inhibition of glutaminolysis corresponded to decreased glutaminase C (GAC protein levels, although other protein levels were

Hydrolytic enzymes in the central vacuole of plant cells

International Nuclear Information System (INIS)

Boller, T.; Kende, H.

1979-01-01

The hydrolase content of vacuoles isolated from protoplasts of suspension-cultured tobacco cells, of tulip petals, and pineapple leaves, and the sedimentation behavior of tobacco tonoplasts were studied. Three precautions were found to be important for the analysis of vacuolar hydrolases and of the tonoplast: (a) purification of protoplasts in a Ficoll gradient was necessary to remove cell debris which contained contaminating hydrolases adsorbed from the fungal cell-wall-degrading enzyme preparation; (b) hydrolase activities in the homogenates of the intact cells or the tissue used and of the purified protoplasts had to be compared to verify the absence of contaminating hydrolases in the protoplast preparation; and (c) vacuoles obtained from the protoplasts by an osmotic shock had to be purified from the lysate in a Ficoll gradient. Since the density of the central vacuole approximates that of the protoplasts, about a 10% contamination of the vacuolar preparation by surviving protoplasts could not be eliminated. The intracellular activities of the following acid hydrolases were primarily localized in the vacuole of tobacco cells: α-mannosidase, β-N-acetylglucosaminidase, β-fructosidase, nuclease, phosphatase, phosphodiesterase. A similar composition of acid hydrolases was found in vacuoles obtained from protoplasts of tulip petals. Proteinase, a hydrolase with low activity in tobacco cells and tulip petals was found to be vacuolar in pineapple leaves, a tissue containing high levels of this enzyme. None of the vacuolar enzymes investigated ws found to be bound to the tonoplast. When vacuoles were isolated from cells labeled with radioactive choline, the vacuolar membrane was found to contain radioactivity. On sucrose gradients, the label incorporated into tonoplasts banded around a density of 1.10 grams per cubic centimeter

Characterization of Modified and Polymer Coated Alumina Surfaces by Infrared Spectroscopy

Directory of Open Access Journals (Sweden)

Ashraf Yehia El-Naggar

2013-01-01

Full Text Available The prepared, modified, and coated alumina surfaces were characterized by infrared spectroscopy (FTIR to investigate the surface properties of the individual and double modified samples. FTIR helps in reporting the changes occurred in hydroxyl groups as well as the structure changes as a result of thermal treating, hydrothermal treating, silylation treating, alkali metal treating, coating, and bonding with polymer. FTIR spectroscopy represents the strength and abundance of surface acidic OH which determine the adsorption properties of polar and nonpolar sorbents. Generally, all treated samples exhibit decrease of OH groups compared with those of parent ones producing alumina surfaces of different adsorptive powers.

Uniform surface-to-line integral reduction of physical optics for curved surfaces by modified edge representation with higher-order correction

Science.gov (United States)

Lyu, Pengfei; Ando, Makoto

2017-09-01

The modified edge representation is one of the equivalent edge currents approximation methods for calculating the physical optics surface radiation integrals in diffraction analysis. The Stokes' theorem is used in the derivation of the modified edge representation from the physical optics for the planar scatterer case, which implies that the surface integral is rigorously reduced into the line integral of the modified edge representation equivalent edge currents, defined in terms of the local shape of the edge. On the contrary, for curved surfaces, the results of radiation integrals depend upon the global shape of the scatterer. The physical optics surface integral consists of two components, from the inner stationary phase point and the edge. The modified edge representation is defined independently from the orientation of the actual edge, and therefore, it could be available not only at the edge but also at the arbitrary points on the scatterer except the stationary phase point where the modified edge representation equivalent edge currents becomes infinite. If stationary phase point exists inside the illuminated region, the physical optics surface integration is reduced into two kinds of the modified edge representation line integrations, along the edge and infinitesimally small integration around the inner stationary phase point, the former and the latter give the diffraction and reflection components, respectively. The accuracy of the latter has been discussed for the curved surfaces and published. This paper focuses on the errors of the former and discusses its correction. It has been numerically observed that the modified edge representation works well for the physical optics diffraction in flat and concave surfaces; errors appear especially for the observer near the reflection shadow boundary if the frequency is low for the convex scatterer. This paper gives the explicit expression of the higher-order correction for the modified edge representation.

A model of extracellular enzymes in free-living microbes: which strategy pays off?

Science.gov (United States)

Traving, Sachia J; Thygesen, Uffe H; Riemann, Lasse; Stedmon, Colin A

2015-11-01

An initial modeling approach was applied to analyze how a single, nonmotile, free-living, heterotrophic bacterial cell may optimize the deployment of its extracellular enzymes. Free-living cells live in a dilute and complex substrate field, and to gain enough substrate, their extracellular enzymes must be utilized efficiently. The model revealed that surface-attached and free enzymes generate unique enzyme and substrate fields, and each deployment strategy has distinctive advantages. For a solitary cell, surface-attached enzymes are suggested to be the most cost-efficient strategy. This strategy entails potential substrates being reduced to very low concentrations. Free enzymes, on the other hand, generate a radically different substrate field, which suggests significant benefits for the strategy if free cells engage in social foraging or experience high substrate concentrations. Swimming has a slight positive effect for the attached-enzyme strategy, while the effect is negative for the free-enzyme strategy. The results of this study suggest that specific dissolved organic compounds in the ocean likely persist below a threshold concentration impervious to biological utilization. This could help explain the persistence and apparent refractory state of oceanic dissolved organic matter (DOM). Microbial extracellular enzyme strategies, therefore, have important implications for larger-scale processes, such as shaping the role of DOM in ocean carbon sequestration. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Facile modification of gelatin-based microcarriers with multiporous surface and proliferative growth factors delivery to enhance cell growth

Energy Technology Data Exchange (ETDEWEB)

Huang Sha [Department of Oral Histology and Pathology, School of Stomatology, Fourth Military Medical University, Xi' an 710032 (China); Research and Development Center for Tissue Engineering, Fourth Military Medical University, Xi' an 710032 (China); Wang Yijuan [Key Laboratory for Macromolecular Science of Shaanxi Province, Shaanxi Normal University, Xi' an 710062 (China); Deng, Tianzheng [Research and Development Center for Tissue Engineering, Fourth Military Medical University, Xi' an 710032 (China); Department of Oral and Maxillofacial Surgery, School of Stomatology, Fourth Military Medical University, Xi' an 710032 (China); Jin Fang [Department of Orthodontics, School of Stomatology, Fourth Military Medical University, Xi' an, 710032 (China); Liu Shouxin [Key Laboratory for Macromolecular Science of Shaanxi Province, Shaanxi Normal University, Xi' an 710062 (China); Zhang Yongjie [Department of Oral Histology and Pathology, School of Stomatology, Fourth Military Medical University, Xi' an 710032 (China); Research and Development Center for Tissue Engineering, Fourth Military Medical University, Xi' an 710032 (China); Feng Feng [Research and Development Center for Tissue Engineering, Fourth Military Medical University, Xi' an 710032 (China); Department of Dermatology, Tangdu Hospital, Fourth Military Medical University, Xi' an 710038 (China); Jin Yan [Department of Oral Histology and Pathology, School of Stomatology, Fourth Military Medical University, Xi' an 710032 (China); Research and Development Center for Tissue Engineering, Fourth Military Medical University, Xi' an 710032 (China)], E-mail: yanjin@fmmu.edu.cn

2008-07-28

The design of microcarriers plays an important role in the success of cell expansion. The present article provides a facile approach to modify the gelatin-based particles and investigates the feasibility of their acting as microcarriers for cell attachment and growth. Gelatin particles (150-320 {mu}m) were modified by cryogenic treatment and lyophilization to develop the surface with the features of multiporous morphology and were incorporated with proliferative growth factors (bFGF) by adsorption during the post-preparation, which enables them to serve as microcarriers for cells amplification, together with the advantages of larger cell-surface contact area and capability of promoting cell propagation. The microstructure and release assay of the modified microcarriers demonstrated that the pores on surface were uniform and bFGF was released in a controlled manner. Through in vitro fibroblast culture, these features resulted in a prominent increase in the cell attachment rate and cell growth rate relative to the conditions without modification. Although the scanning electron microscopy and optical microscopy analysis results indicated that cells attached, spread, and proliferated on all the microcarriers, cell growth clearly showed a significant correlation with the multiporous structure of microcarriers, in particular on bFGF combined ones. These results validate our previous assumption that the facile modification could improve cell growth on the gelatin-based microcarriers obviously and the novel microcarriers may be a promising candidate in tissue engineering.

Facile modification of gelatin-based microcarriers with multiporous surface and proliferative growth factors delivery to enhance cell growth

International Nuclear Information System (INIS)

Huang Sha; Wang Yijuan; Deng, Tianzheng; Jin Fang; Liu Shouxin; Zhang Yongjie; Feng Feng; Jin Yan

2008-01-01

The design of microcarriers plays an important role in the success of cell expansion. The present article provides a facile approach to modify the gelatin-based particles and investigates the feasibility of their acting as microcarriers for cell attachment and growth. Gelatin particles (150-320 μm) were modified by cryogenic treatment and lyophilization to develop the surface with the features of multiporous morphology and were incorporated with proliferative growth factors (bFGF) by adsorption during the post-preparation, which enables them to serve as microcarriers for cells amplification, together with the advantages of larger cell-surface contact area and capability of promoting cell propagation. The microstructure and release assay of the modified microcarriers demonstrated that the pores on surface were uniform and bFGF was released in a controlled manner. Through in vitro fibroblast culture, these features resulted in a prominent increase in the cell attachment rate and cell growth rate relative to the conditions without modification. Although the scanning electron microscopy and optical microscopy analysis results indicated that cells attached, spread, and proliferated on all the microcarriers, cell growth clearly showed a significant correlation with the multiporous structure of microcarriers, in particular on bFGF combined ones. These results validate our previous assumption that the facile modification could improve cell growth on the gelatin-based microcarriers obviously and the novel microcarriers may be a promising candidate in tissue engineering

Enhancing anticoagulation and endothelial cell proliferation of titanium surface by sequential immobilization of poly(ethylene glycol) and collagen

International Nuclear Information System (INIS)

Pan, Chang-Jiang; Hou, Yan-Hua; Ding, Hong-Yan; Dong, Yun-Xiao

2013-01-01

In the present study, poly(ethylene glycol) (PEG) and collagen I were sequentially immobilized on the titanium surface to simultaneously improve the anticoagulation and endothelial cell proliferation. Attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR) and X-ray photoelectron spectroscopy analysis confirmed that PEG and collagen I were successfully immobilized on the titanium surface. Water contact angle results suggested the excellent hydrophilic surface after the immobilization. The anticoagulation experiments demonstrated that the immobilized PEG and collagen I on the titanium surface could not only obviously prevent platelet adhesion and aggregation but also prolong activated partial thromboplastin time (APTT), leading to the improved blood compatibility. Furthermore, immobilization of collagen to the end of PEG chain did not abate the anticoagulation. As compared to those on the pristine and PEG-modified titanium surfaces, endothelial cells exhibited improved proliferative profiles on the surface modified by the sequential immobilization of PEG and collagen in terms of CCK-8 assay, implying that the modified titanium may promote endothelialization without abating the blood compatibility. Our method may be used to modify the surface of blood-contacting biomaterials such as titanium to promote endothelialization and improve the anticoagulation, it may be helpful for development of the biomedical devices such as coronary stents, where endothelializaton and excellent anticoagulation are required.

Wettability Control of Gold Surfaces Modified with Benzenethiol Derivatives: Water Contact Angle and Thermal Stability.

Science.gov (United States)

Tatara, Shingo; Kuzumoto, Yasutaka; Kitamura, Masatoshi

2016-04-01

The water wettability of Au surfaces has been controlled using various benzenethiol derivatives including 4-methylbenzenethiol, pentafluorobenzenethiol, 4-flubrobenzenethiol, 4-methoxy-benzenethiol, 4-nitrobenzenethiol, and 4-hydroxybenzenethiol. The water contact angle of the Au surface modified with the benzenethiol derivative was found to vary in the wide range of 30.9° to 88.3°. The contact angle of the modified Au films annealed was also measured in order to investigate their thermal stability. The change in the contact angle indicated that the modified surface is stable at temperatures below about 400 K. Meanwhile, the activation energy of desorption from the modified surface was estimated from the change in the contact angle. The modified Au surface was also examined using X-ray photoelectron spectroscopy.

Enhanced adherence of mouse fibroblast and vascular cells to plasma modified polyethylene

Energy Technology Data Exchange (ETDEWEB)

Reznickova, Alena, E-mail: alena.reznickova@vscht.cz [Department of Solid State Engineering, Institute of Chemical Technology Prague, 166 28 Prague 6 (Czech Republic); Novotna, Zdenka, E-mail: zdenka1.novotna@vscht.cz [Department of Solid State Engineering, Institute of Chemical Technology Prague, 166 28 Prague 6 (Czech Republic); Kolska, Zdenka [Faculty of Science, J.E. Purkyně University, 400 96 Usti nad Labem (Czech Republic); Kasalkova, Nikola Slepickova [Department of Solid State Engineering, Institute of Chemical Technology Prague, 166 28 Prague 6 (Czech Republic); Rimpelova, Silvie [Department of Biochemistry and Microbiology, Institute of Chemical Technology Prague, 166 28 Prague 6 (Czech Republic); Svorcik, Vaclav [Department of Solid State Engineering, Institute of Chemical Technology Prague, 166 28 Prague 6 (Czech Republic)

2015-07-01

Since the last decade, tissue engineering has shown a sensational promise in providing more viable alternatives to surgical procedures for harvested tissues, implants and prostheses. Biomedical polymers, such as low-density polyethylene (LDPE), high-density polyethylene (HDPE) and ultra-high molecular weight polyethylene (UHMWPE), were activated by Ar plasma discharge. Degradation of polymer chains was examined by determination of the thickness of ablated layer. The amount of an ablated polymer layer was measured by gravimetry. Contact angle, measured by goniometry, was studied as a function of plasma exposure and post-exposure aging times. Chemical structure of modified polymers was characterized by angle resolved X-ray photoelectron spectroscopy. Surface chemistry and polarity of the samples were investigated by electrokinetic analysis. Changes in surface morphology were followed using atomic force microscopy. Cytocompatibility of plasma activated polyethylene foils was studied using two distinct model cell lines; VSMCs (vascular smooth muscle cells) as a model for vascular graft testing and connective tissue cells L929 (mouse fibroblasts) approved for standardized material cytotoxicity testing. Specifically, the cell number, morphology, and metabolic activity of the adhered and proliferated cells on the polyethylene matrices were studied in vitro. It was found that the plasma treatment caused ablation of the polymers, resulting in dramatic changes in their surface morphology and roughness. ARXPS and electrokinetic measurements revealed oxidation of the polymer surface. It was found that plasma activation has a positive effect on the adhesion and proliferation of VSMCs and L929 cells. - Highlights: • Plasma activation of LDPE, HDPE and UHMWPE • Study of surface properties by several techniques: ARXPS, AFM, zeta-potential, and goniometry • Investigation of adhesion and spreading of vascular smooth muscle cells (VSMCs) and mouse fibroblasts (L929)

H3PO4 treated surface modified CuS counter electrodes with high electrocatalytic activity for enhancing photovoltaic performance of quantum dot-sensitized solar cells

Science.gov (United States)

Panthakkal Abdul Muthalif, Mohammed; Sunesh, Chozhidakath Damodharan; Choe, Youngson

2018-05-01

Herein we report a simple synthetic strategy to prepare highly efficient and surface modified CuS counter electrodes (CEs) for quantum dot-sensitized solar cells (QDSSCs) in the presence of phosphoric acid (H3PO4) using the chemical bath deposition method. This is the first report of successful treatment of H3PO4 on the surface of CuS CEs for designing a high-performance QDSSCs with improved photovoltaic properties. After optimization, the 4 ml H3PO4 treated CuS CE-based QDSSC exhibits excellent photovoltaic performance with a conversion efficiency (η) of 4.20% (Voc = 0.592 V, Jsc = 13.35 mA cm-2, FF = 0.532) under one full-sun illumination (100 mW cm-2, AM 1.5 G).

Genetic Modifiers of Sickle Cell Disease

Science.gov (United States)

Steinberg, Martin H.; Sebastiani, Paola

2015-01-01

Sickle cell anemia is associated with unusual clinical heterogeneity for a Mendelian disorder. Fetal hemoglobin concentration and coincident ∝ thalassemia, both which directly affect the sickle erythrocyte, are the major modulators of the phenotype of disease. Understanding the genetics underlying the heritable subphenotypes of sickle cell anemia would be prognostically useful, could inform personalized therapeutics, and might help the discovery of new “druggable” pathophysiologic targets. Genotype-phenotype association studies have been used to identify novel genetic modifiers. In the future, whole genome sequencing with its promise of discovering hitherto unsuspected variants could add to our understanding of the genetic modifiers of this disease. PMID:22641398

Mucopolysaccharidosis enzyme production by bone marrow and dental pulp derived human mesenchymal stem cells.

Science.gov (United States)

Jackson, Matilda; Derrick Roberts, Ainslie; Martin, Ellenore; Rout-Pitt, Nathan; Gronthos, Stan; Byers, Sharon

2015-04-01

Mucopolysaccharidoses (MPS) are inherited metabolic disorders that arise from a complete loss or a reduction in one of eleven specific lysosomal enzymes. MPS children display pathology in multiple cell types leading to tissue and organ failure and early death. Mesenchymal stem cells (MSCs) give rise to many of the cell types affected in MPS, including those that are refractory to current treatment protocols such as hematopoietic stem cell (HSC) based therapy. In this study we compared multiple MPS enzyme production by bone marrow derived (hBM) and dental pulp derived (hDP) MSCs to enzyme production by HSCs. hBM MSCs produce significantly higher levels of MPS I, II, IIIA, IVA, VI and VII enzyme than HSCs, while hDP MSCs produce significantly higher levels of MPS I, IIIA, IVA, VI and VII enzymes. Higher transfection efficiency was observed in MSCs (89%) compared to HSCs (23%) using a lentiviral vector. Over-expression of four different lysosomal enzymes resulted in up to 9303-fold and up to 5559-fold greater levels in MSC cell layer and media respectively. Stable, persistent transduction of MSCs and sustained over-expression of MPS VII enzyme was observed in vitro. Transduction of MSCs did not affect the ability of the cells to differentiate down osteogenic, adipogenic or chondrogenic lineages, but did partially delay differentiation down the non-mesodermal neurogenic lineage. Copyright © 2015 Elsevier Inc. All rights reserved.

A model of extracellular enzymes in free-living microbes: Which strategy pays off?

DEFF Research Database (Denmark)

Traving, Sachia J; Thygesen, Uffe Høgsbro; Riemann, Lasse

2015-01-01

a slight positive effect for the attached-enzyme strategy, while the effect is negative for the free-enzyme strategy. The results of this study suggest that specific dissolved organic compounds in the ocean likely persist below a threshold concentration impervious to biological utilization. This could help...... must be utilized efficiently. The model revealed that surface-attached and free enzymes generate unique enzyme and substrate fields, and each deployment strategy has distinctive advantages. For a solitary cell, surface-attached enzymes are suggested to be the most cost-efficient strategy. This strategy...

Diffusion of MMPs on the surface of collagen fibrils: the mobile cell surface-collagen substratum interface.

Directory of Open Access Journals (Sweden)

Ivan E Collier

Full Text Available Remodeling of the extracellular matrix catalyzed by MMPs is central to morphogenetic phenomena during development and wound healing as well as in numerous pathologic conditions such as fibrosis and cancer. We have previously demonstrated that secreted MMP-2 is tethered to the cell surface and activated by MT1-MMP/TIMP-2-dependent mechanism. The resulting cell-surface collagenolytic complex (MT1-MMP(2/TIMP-2/MMP-2 can initiate (MT1-MMP and complete (MMP-2 degradation of an underlying collagen fibril. The following question remained: What is the mechanism of substrate recognition involving the two structures of relatively restricted mobility, the cell surface enzymatic complex and a collagen fibril embedded in the ECM? Here we demonstrate that all the components of the complex are capable of processive movement on a surface of the collagen fibril. The mechanism of MT1-MMP movement is a biased diffusion with the bias component dependent on the proteolysis of its substrate, not adenosine triphosphate (ATP hydrolysis. It is similar to that of the MMP-1 Brownian ratchet we described earlier. In addition, both MMP-2 and MMP-9 as well as their respective complexes with TIMP-1 and -2 are capable of Brownian diffusion on the surface of native collagen fibrils without noticeable dissociation while the dimerization of MMP-9 renders the enzyme immobile. Most instructive is the finding that the inactivation of the enzymatic activity of MT1-MMP has a detectable negative effect on the cell force developed in miniaturized 3D tissue constructs. We propose that the collagenolytic complex (MT1-MMP(2/TIMP-2/MMP-2 represents a Mobile Cell Surface-Collagen Substratum Interface. The biological implications of MT1-MMP acting as a molecular ratchet tethered to the cell surface in complex with MMP-2 suggest a new mechanism for the role of spatially regulated peri-cellular proteolysis in cell-matrix interactions.

Effects of a hybrid micro/nanorod topography-modified titanium implant on adhesion and osteogenic differentiation in rat bone marrow mesenchymal stem cells.

Science.gov (United States)

Zhang, Wenjie; Li, Zihui; Huang, Qingfeng; Xu, Ling; Li, Jinhua; Jin, Yuqin; Wang, Guifang; Liu, Xuanyong; Jiang, Xinquan

2013-01-01

Various methods have been used to modify titanium implant surfaces with the aim of achieving better osseointegration. In this study, we fabricated a clustered nanorod structure on an acid-etched, microstructured titanium plate surface using hydrogen peroxide. We also evaluated biofunctionalization of the hybrid micro/nanorod topography on rat bone marrow mesenchymal stem cells. Scanning electron microscopy and x-ray diffraction were used to investigate the surface topography and phase composition of the modified titanium plate. Rat bone marrow mesenchymal stem cells were cultured and seeded on the plate. The adhesion ability of the cells was then assayed by cell counting at one, 4, and 24 hours after cell seeding, and expression of adhesion-related protein integrin β1 was detected by immunofluorescence. In addition, a polymerase chain reaction assay, alkaline phosphatase and Alizarin Red S staining assays, and osteopontin and osteocalcin immunofluorescence analyses were used to evaluate the osteogenic differentiation behavior of the cells. The hybrid micro/nanoscale texture formed on the titanium surface enhanced the initial adhesion activity of the rat bone marrow mesenchymal stem cells. Importantly, the hierarchical structure promoted osteogenic differentiation of these cells. This study suggests that a hybrid micro/nanorod topography on a titanium surface fabricated by treatment with hydrogen peroxide followed by acid etching might facilitate osseointegration of a titanium implant in vivo.

Controlled immobilisation of active enzymes on the cowpea mosaic virus capsid

Science.gov (United States)

Aljabali, Alaa A. A.; Barclay, J. Elaine; Steinmetz, Nicole F.; Lomonossoff, George P.; Evans, David J.

2012-08-01

Immobilisation of horseradish peroxidase (HRP) and glucose oxidase (GOX) via covalent attachment of modified enzyme carbohydrate to the exterior of the cowpea mosaic virus (CPMV) capsid gave high retention of enzymatic activity. The number of enzymes bound per virus was determined to be about eleven for HRP and 2-3 for GOX. This illustrates that relatively large biomacromolecules can be readily coupled to the virus surface using simple conjugation strategies. Virus-biomacromolecule hybrids have great potential for uses in catalysis, diagnostic assays or biosensors.Immobilisation of horseradish peroxidase (HRP) and glucose oxidase (GOX) via covalent attachment of modified enzyme carbohydrate to the exterior of the cowpea mosaic virus (CPMV) capsid gave high retention of enzymatic activity. The number of enzymes bound per virus was determined to be about eleven for HRP and 2-3 for GOX. This illustrates that relatively large biomacromolecules can be readily coupled to the virus surface using simple conjugation strategies. Virus-biomacromolecule hybrids have great potential for uses in catalysis, diagnostic assays or biosensors. Electronic supplementary information (ESI) available: Alternative conjugation strategies, agarose gel electrophoresis of CPMV and CPMV-HRP conjugates, UV-vis spectrum of HRP-ADHCPMV, agarose gel electrophoresis of GOX-ADHCPMV particles and corresponding TEM image, calibration curves for HRP-ADHCPMV and GOX-ADHCPMV, DLS data for GOX-ADHCPMV are made available. See DOI: 10.1039/c2nr31485a

High endothelin-converting enzyme-1 expression independently predicts poor survival of patients with esophageal squamous cell carcinoma.

Science.gov (United States)

Wu, Ching-Fang; Lee, Ching-Tai; Kuo, Yao-Hung; Chen, Tzu-Haw; Chang, Chi-Yang; Chang, I-Wei; Wang, Wen-Lun

2017-09-01

Patients with esophageal squamous cell carcinoma have poor survival and high recurrence rate, thus an effective prognostic biomarker is needed. Endothelin-converting enzyme-1 is responsible for biosynthesis of endothelin-1, which promotes growth and invasion of human cancers. The role of endothelin-converting enzyme-1 in esophageal squamous cell carcinoma is still unknown. Therefore, this study investigated the significance of endothelin-converting enzyme-1 expression in esophageal squamous cell carcinoma clinically. We enrolled patients with esophageal squamous cell carcinoma who provided pretreated tumor tissues. Tumor endothelin-converting enzyme-1 expression was evaluated by immunohistochemistry and was defined as either low or high expression. Then we evaluated whether tumor endothelin-converting enzyme-1 expression had any association with clinicopathological findings or predicted survival of patients with esophageal squamous cell carcinoma. Overall, 54 of 99 patients with esophageal squamous cell carcinoma had high tumor endothelin-converting enzyme-1 expression, which was significantly associated with lymph node metastasis ( p = 0.04). In addition, tumor endothelin-converting enzyme-1 expression independently predicted survival of patients with esophageal squamous cell carcinoma, and the 5-year survival was poorer in patients with high tumor endothelin-converting enzyme-1 expression ( p = 0.016). Among patients with locally advanced and potentially resectable esophageal squamous cell carcinoma (stage II and III), 5-year survival was poorer with high tumor endothelin-converting enzyme-1 expression ( p = 0.003). High tumor endothelin-converting enzyme-1 expression also significantly predicted poorer survival of patients in this population. In patients with esophageal squamous cell carcinoma, high tumor endothelin-converting enzyme-1 expression might indicate high tumor invasive property. Therefore, tumor endothelin-converting enzyme-1 expression

Evaluation of Antibacterial Activity of Titanium Surface Modified by PVD/PACVD Process.

Science.gov (United States)

Ji, Min-Kyung; Lee, Min-Joo; Park, Sang-Won; Lee, Kwangmin; Yun, Kwi-Dug; Kim, Hyun-Seung; Oh, Gye-Jeong; Kim, Ji-Hyun; Lim, Hyun-Pil

2016-02-01

The aim of this study was to evaluate the response of Streptococcus mutans (S. mutans) via crystal violet staining assay on titanium surface modified by physical vapor deposition/plasma assisted chemical vapor deposition process. Specimens were divided into the following three groups: polished titanium (control group), titanium modified by DC magnetron sputtering (group TiN-Ti), and titanium modified by plasma nitriding (group N-Ti). Surface characteristics of specimens were observed by using nanosurface 3D optical profiler and field emission scanning electron microscope. Group TiN-Ti showed TiN layer of 1.2 microm in thickness. Group N-Ti was identified as plasma nitriding with X-ray photoelectron spectroscopy. Roughness average (Ra) of all specimens had values 0.05). Within the process condition of this study, modified titanium surfaces by DC magnetron sputtering and plasma nitriding did not influence the adhesion of S. mutans.

Surface characterization and free thyroid hormones response of chemically modified poly(ethylene terephthalate) blood collection tubes

Science.gov (United States)

Jalali Dil, Ebrahim; Kim, Samuel C.; Saffar, Amir; Ajji, Abdellah; Zare, Richard N.; Sattayapiwat, Annie; Esguerra, Vanessa; Bowen, Raffick A. R.

2018-06-01

The surface chemistry and surface energy of chemically modified polyethylene terephthalate (PET) blood collection tubes (BCTs) were studied and the results showed a significant increase in hydrophilicity and polarity of modified PET surface. The surface modification created nanometer-sized, needle-like asperities through molecular segregation at the surface. The surface dynamics of the modified PET was examined by tracking its surface properties over a 280-day period. The results showed surface rearrangement toward a surface with lower surface energy and fewer nanometer-sized asperities. Thromboelastography (TEG) was used to evaluate and compare the thrombogenicity of the inner walls of various types of BCTs. The TEG tracings and data from various types of BCTs demonstrated differences in the reactionand coagulation times but not in clot strength. The performance of the modified tubes in free triiodothyronine (FT3) and free thyroxine (FT4) hormone tests was examined, and it was found that the interference of modified PET tubes was negligible compared to that of commercially available PET BCTs.

Protein-modified nanocrystalline diamond thin films for biosensor applications.

Science.gov (United States)

Härtl, Andreas; Schmich, Evelyn; Garrido, Jose A; Hernando, Jorge; Catharino, Silvia C R; Walter, Stefan; Feulner, Peter; Kromka, Alexander; Steinmüller, Doris; Stutzmann, Martin

2004-10-01

Diamond exhibits several special properties, for example good biocompatibility and a large electrochemical potential window, that make it particularly suitable for biofunctionalization and biosensing. Here we show that proteins can be attached covalently to nanocrystalline diamond thin films. Moreover, we show that, although the biomolecules are immobilized at the surface, they are still fully functional and active. Hydrogen-terminated nanocrystalline diamond films were modified by using a photochemical process to generate a surface layer of amino groups, to which proteins were covalently attached. We used green fluorescent protein to reveal the successful coupling directly. After functionalization of nanocrystalline diamond electrodes with the enzyme catalase, a direct electron transfer between the enzyme's redox centre and the diamond electrode was detected. Moreover, the modified electrode was found to be sensitive to hydrogen peroxide. Because of its dual role as a substrate for biofunctionalization and as an electrode, nanocrystalline diamond is a very promising candidate for future biosensor applications.

Effects of commercial pectolytic and cellulolytic enzyme preparations on the apple cell wall.

Science.gov (United States)

Dongowski, G; Sembries, S

2001-09-01

The action of three different commercial enzyme combinations on apple cell wall material has been examined in a model system under conditions of mash and pomace treatment by using an alcohol-insoluble substance prepared from apples. A part of the total dietary fiber, for example, galacturonan (pectin), appeared in the soluble fraction after enzymatic mash treatment. The soluble fraction increased intensely during pomace treatment. Furthermore, enzyme actions caused a change in the water-binding capacity of residues as well as changes in the monosaccharide composition and in the molecular weight distribution of saccharides in filtrates (soluble parts). The extent of decomposition of cell wall material and the increase of soluble oligomeric and/or polymeric dietary fiber components are caused by both the composition (pectinases, cellulases, and hemicellulases) and the activities of the enzyme preparations. The model experiments allow an insight into the reactions occurring during enzyme action on the plant cell wall, for example, during apple juice production using pectolytic and cellulolytic enzyme preparations.

Electrochemical Glucose Biosensor Based on Glucose Oxidase Displayed on Yeast Surface.

Science.gov (United States)

Wang, Hongwei; Lang, Qiaolin; Liang, Bo; Liu, Aihua

2015-01-01

The conventional enzyme-based biosensor requires chemical or physical immobilization of purified enzymes on electrode surface, which often results in loss of enzyme activity and/or fractions immobilized over time. It is also costly. A major advantage of yeast surface display is that it enables the direct utilization of whole cell catalysts with eukaryote-produced proteins being displayed on the cell surface, providing an economic alternative to traditional production of purified enzymes. Herein, we describe the details of the display of glucose oxidase (GOx) on yeast cell surface and its application in the development of electrochemical glucose sensor. In order to achieve a direct electrochemistry of GOx, the entire cell catalyst (yeast-GOx) was immobilized together with multiwalled carbon nanotubes on the electrode, which allowed sensitive and selective glucose detection.

Applications of Enzymes in Oil and Oilseed Processing

DEFF Research Database (Denmark)

Xu, Xuebing

Enzymes, through the last 20-30 years research and development, have been widely explored for the uses in oil and oilseed processing. Following the conventional processing technology from oilseeds, the oil can be produced through pressing or solvent extraction. The crude oil is then refined to meet...... edible requirements. The oil can be also modified to meet functional or even nutritional needs. In each of those steps, enzymes have been used in industry successfully. For the oil processing stage, enzymes have been used to destroy the cell structure so that makes the oil release easier, where...... conventionally high temperature conditioning or cooking is necessary. The good story in industry is the fish oil and olive oil processing. Good quality and higher oil yield have been achieved through the use of enzymes in the processing stages. For the refining stage, the use of enzymes for degumming has...

Nedd8 processing enzymes in Schizosaccharomyces pombe

DEFF Research Database (Denmark)

O'Donoghue, Jean; Bech-Otschir, Dawadschargal; Larsen, Ida

2013-01-01

Conjugation of the ubiquitin-like modifier Nedd8 to cullins is critical for the function of SCF-type ubiquitin ligases and thus facilitates ubiquitin conjugation and ultimately degradation of SCF substrates, including several cell cycle regulators. Like ubiquitin, Nedd8 is produced as a precursor...... that must first be processed before it becomes active. In Saccharomyces cerevisiae this is carried out exclusively by the enzyme Yuh1....

Phospholipase D specific for the phosphatidylinositol anchor of cell-surface proteins is abundant in plasma

International Nuclear Information System (INIS)

Low, M.G.; Prasad, A.R.S.

1988-01-01

An enzyme activity capable of degrading the glycosyl-phosphatidylinositol membrane anchor of cell-surface proteins has previously been reported in a number of mammalian tissues. The experiments reported here demonstrate that this anchor-degrading activity is also abundant in mammalian plasma. The activity was inhibited by EGTA or 1,10-phenanthroline. It was capable of removing the anchor from alkaline phosphatase, 5'-nucleotidase, and variant surface glycoprotein but had little or no activity toward phosphatidylinositol or phosphatidylcholine. Phosphatidic acid was the only 3 H-labeled product when this enzyme hydrolyzed [ 3 H]myristate-labeled variant surface glycoprotein. It could be distinguished from the Ca 2 =-dependent inositol phospholipid-specific phospholipase C activity in several rat tissues on the basis of its molecular size and its sensitivity to 1,10-phenanthroline. The data therefore suggest that this activity is due to a phospholipase D with specificity for glycosylphosphatidylinositol structures. Although the precise physiological function of this anchor-specific phospholipase D remains to be determined, these findings indicate that it could play an important role in regulating the expression and release of cell-surface proteins in vivo

Reconfigurable modified surface layers using plasma capillaries around the neutral inclusion regime

Energy Technology Data Exchange (ETDEWEB)

Varault, S. [ONERA—The French Aerospace Lab 2, Avenue Edouard Belin, BP4025, 31055 Toulouse Cedex (France); Universite Paul Sabatier—CNRS-Laplace 118, Route de Narbonne, F-31062 Toulouse Cedex 9 (France); Gabard, B. [ONERA—The French Aerospace Lab 2, Avenue Edouard Belin, BP4025, 31055 Toulouse Cedex (France); STAE—4, Rue Emile Monso, BP84234, 31030 Toulouse Cedex 4 (France); Crépin, T.; Bolioli, S. [ONERA—The French Aerospace Lab 2, Avenue Edouard Belin, BP4025, 31055 Toulouse Cedex (France); Sokoloff, J. [Universite Paul Sabatier—CNRS-Laplace 118, Route de Narbonne, F-31062 Toulouse Cedex 9 (France)

2014-02-28

We show both theoretically and experimentally reconfigurable properties achieved by plasma inclusions placed in modified surface layers generally used to tailor the transmission and beaming properties of electromagnetic bandgap based waveguiding structures. A proper parametrization of the plasma capillaries allows to reach the neutral inclusion regime, where the inclusions appear to be electromagnetically transparent, letting the surface mode characteristics unaltered. Varying the electron density of the plasma inclusions provoques small perturbations around this peculiar regime, and we observe significant modifications of the transmission/beaming properties. This offers a way to dynamically select the enhanced transmission frequency or to modify the radiation pattern of the structure, depending on whether the modified surface layer is placed at the entrance/exit of the waveguide.

Reconfigurable modified surface layers using plasma capillaries around the neutral inclusion regime

International Nuclear Information System (INIS)

Varault, S.; Gabard, B.; Crépin, T.; Bolioli, S.; Sokoloff, J.

2014-01-01

We show both theoretically and experimentally reconfigurable properties achieved by plasma inclusions placed in modified surface layers generally used to tailor the transmission and beaming properties of electromagnetic bandgap based waveguiding structures. A proper parametrization of the plasma capillaries allows to reach the neutral inclusion regime, where the inclusions appear to be electromagnetically transparent, letting the surface mode characteristics unaltered. Varying the electron density of the plasma inclusions provoques small perturbations around this peculiar regime, and we observe significant modifications of the transmission/beaming properties. This offers a way to dynamically select the enhanced transmission frequency or to modify the radiation pattern of the structure, depending on whether the modified surface layer is placed at the entrance/exit of the waveguide

Water-soluble nanoconjugates of quantum dot-chitosan-antibody for in vitro detection of cancer cells based on “enzyme-free” fluoroimmunoassay

Energy Technology Data Exchange (ETDEWEB)

Mansur, Herman S., E-mail: hmansur@demet.ufmg.br [Center of Nanoscience, Nanotechnology, and Innovation-CeNano" 2I, Department of Metallurgical and Materials Engineering, Federal University of Minas Gerais, UFMG, Av. Presidente Antônio Carlos, 6627, Belo Horizonte, MG 31.270-901 (Brazil); Mansur, Alexandra A.P. [Center of Nanoscience, Nanotechnology, and Innovation-CeNano" 2I, Department of Metallurgical and Materials Engineering, Federal University of Minas Gerais, UFMG, Av. Presidente Antônio Carlos, 6627, Belo Horizonte, MG 31.270-901 (Brazil); Soriano-Araújo, Amanda [Center of Nanoscience, Nanotechnology, and Innovation-CeNano" 2I, Department of Metallurgical and Materials Engineering, Federal University of Minas Gerais, UFMG, Av. Presidente Antônio Carlos, 6627, Belo Horizonte, MG 31.270-901 (Brazil); Department of Preventive Veterinary Medicine, Veterinary School, UFMG (Brazil); Lobato, Zélia I.P. [Department of Preventive Veterinary Medicine, Veterinary School, UFMG (Brazil); Carvalho, Sandhra M. de [Center of Nanoscience, Nanotechnology, and Innovation-CeNano" 2I, Department of Metallurgical and Materials Engineering, Federal University of Minas Gerais, UFMG, Av. Presidente Antônio Carlos, 6627, Belo Horizonte, MG 31.270-901 (Brazil); Department of Physiology and Biophysics, ICB, UFMG (Brazil); Leite, Maria de Fatima [Department of Physiology and Biophysics, ICB, UFMG (Brazil)

2015-07-01

Cancer remains one of the world's most devastating diseases with millions of fatalities and new cases every year. In this work, we attempted to develop a facile “enzyme-free” fluoroimmunoassay based on the novel nanoconjugates composed of CdS quantum dots (QDs) as the fluorescent inorganic core and an antibody-modified polysaccharide as the organic shell, modeling their possible application for the in vitro diagnosis of non-Hodgkin lymphoma (NHL) cancer. Chitosan was conjugated with an anti-CD20 polyclonal antibody (pAbCD20) by the formation of covalent amide bonds. In the sequence, these chitosan-antibody conjugates were utilized as direct ligands for the surface biofunctionalization of CdS QDs (CdS/chitosan-pAbCD20) using a single-step colloidal process in aqueous medium at room temperature. The most relevant physico-chemical properties of these nanoconjugates were assessed by morphological and spectroscopic techniques. The results indicated that CdS nanocrystals were produced with an average diameter of 2.5 nm and with cubic zinc blende crystalline nanostructure. The CdS-immunoconjugates (CdS/chitosan-pAbCD20) presented colloidal hydrodynamic diameter (H{sub D}) of 15.0 ± 1.2 nm. In addition, the results evidenced that the “enzyme-free” QD-linked immunosorbent assay (QLISA) was effective for the in vitro detection against the antigen CD20 (aCD20) based on fluorescent behavior of the CdS nanoconjugates. Moreover, the CdS-immunoconjugates were successfully used for fluorescence bioimaging of NHL cancer cells. Finally, the cell viability results using different cell cultures based on LDH, MTT and Resazurin bio-assays have demonstrated no cytotoxicity of the new CdS-chitosan bioconjugates relative to the standard controls. Thus, CdS conjugates may offer a promising platform for the future development of in vitro and in vivo applications for the detection and diagnosis of NHL cancer cells. - Highlights: • CdS quantum dots (QDs) were prepared using

Construction of Hydrophobic Wood Surface and Mechanical Property of Wood Cell Wall on Nanoscale Modified by Dimethyldichlorosilane

Science.gov (United States)

Yang, Rui; Wang, Siqun; Zhou, Dingguo; Zhang, Jie; Lan, Ping; Jia, Chong

2018-01-01

Dimethyldichlorosilane was used to improve the hydrophobicity of wood surface. The water contact angle of the treated wood surface increased from 85° to 143°, which indicated increased hydrophobicity. The nanomechanical properties of the wood cell wall were evaluated using a nanoindentation test to analyse the hydrophobic mechanism on the nano scale. The elastic modulus of the cell wall was significantly affected by the concentration but the influence of treatment time is insignificant. The hardness of the cell wall for treated samples was significantly affected by both treatment time and concentration. The interaction between treatment time and concentration was extremely significant for the elastic modulus of the wood cell wall.

Controlling cell adhesion via replication of laser micro/nano-textured surfaces on polymers

Energy Technology Data Exchange (ETDEWEB)

Koufaki, Niki; Ranella, Anthi; Barberoglou, Marios; Psycharakis, Stylianos; Fotakis, Costas; Stratakis, Emmanuel [Institute of Electronic Structure and Laser (IESL), Foundation for Research and Technology-Hellas (FORTH), 711 10, Heraklion, Crete (Greece); Aifantis, Katerina E, E-mail: stratak@iesl.forth.gr [Lab of Mechanics and Materials, Aristotle University of Thessaloniki, Thessaloniki (Greece)

2011-12-15

The aim of this study is to investigate cell adhesion and viability on highly rough polymeric surfaces with gradient roughness ratios and wettabilities prepared by microreplication of laser micro/nano-textured Si surfaces. Negative replicas on polydimethylsiloxane as well as positive ones on a photocurable (organically modified ceramic) and a biodegradable (poly(lactide-co-glycolide)) polymer have been successfully reproduced. The final culture substrates comprised from forests of micron-sized conical spikes exhibiting a range of roughness ratios and wettabilities, was achieved by changing the laser fluence used to fabricate the original template surfaces. Cell culture experiments were performed with the fibroblast NIH/3T3 and PC12 neuronal cell lines in order to investigate how these surfaces are capable of modulating different types of cellular responses including, viability, adhesion and morphology. The results showed a preferential adhesion of both cell types on the microstructured surfaces compared to the unstructured ones. In particular, the fibroblast NIH/3T3 cells show optimal adhesion for small roughness ratios, independent of the surface wettability and polymer type, indicating a non-monotonic dependence of cell adhesion on surface energy. In contrast, the PC12 cells were observed to adhere well to the patterned surfaces independent of the roughness ratio and wettability. These experimental findings are correlated with micromechanical measurements performed on the unstructured and replicated surfaces and discussed on the basis of previous observations describing the relation of cell response to surface energy and rigidity.

Controlling cell adhesion via replication of laser micro/nano-textured surfaces on polymers

International Nuclear Information System (INIS)

Koufaki, Niki; Ranella, Anthi; Barberoglou, Marios; Psycharakis, Stylianos; Fotakis, Costas; Stratakis, Emmanuel; Aifantis, Katerina E

2011-01-01

The aim of this study is to investigate cell adhesion and viability on highly rough polymeric surfaces with gradient roughness ratios and wettabilities prepared by microreplication of laser micro/nano-textured Si surfaces. Negative replicas on polydimethylsiloxane as well as positive ones on a photocurable (organically modified ceramic) and a biodegradable (poly(lactide-co-glycolide)) polymer have been successfully reproduced. The final culture substrates comprised from forests of micron-sized conical spikes exhibiting a range of roughness ratios and wettabilities, was achieved by changing the laser fluence used to fabricate the original template surfaces. Cell culture experiments were performed with the fibroblast NIH/3T3 and PC12 neuronal cell lines in order to investigate how these surfaces are capable of modulating different types of cellular responses including, viability, adhesion and morphology. The results showed a preferential adhesion of both cell types on the microstructured surfaces compared to the unstructured ones. In particular, the fibroblast NIH/3T3 cells show optimal adhesion for small roughness ratios, independent of the surface wettability and polymer type, indicating a non-monotonic dependence of cell adhesion on surface energy. In contrast, the PC12 cells were observed to adhere well to the patterned surfaces independent of the roughness ratio and wettability. These experimental findings are correlated with micromechanical measurements performed on the unstructured and replicated surfaces and discussed on the basis of previous observations describing the relation of cell response to surface energy and rigidity.

21 CFR 864.7100 - Red blood cell enzyme assay.

Science.gov (United States)

2010-04-01

... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Red blood cell enzyme assay. 864.7100 Section 864.7100 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7100 Red blood cell...

Immobilization of the Enzyme Glucose Oxidase on Both Bulk and Porous SiO2 Surfaces

Directory of Open Access Journals (Sweden)

Fulvia Sinatra

2008-09-01

Full Text Available Silicon dioxide surfaces, both bulk and porous, were used to anchor the enzyme glucose oxidase. The immobilization protocol was optimized and the samples characterized using X-ray Photoelectron Spectroscopy, Energy Dispersive X-rays coupled to scanning electron microscopy and enzymatic activity measurements. We show that a uniform layer was obtained by activating the oxide before immobilization. X-ray Photoelectron Spectroscopy measurements carried out on bulk oxide showed that the silicon substrate signal was fully screened after the enzyme deposition showing the absence of uncovered surface regions. The enzyme presence was detected monitoring both the C 1s and N 1s signals. Finally, enzymatic activity measurements confirmed that the glucose oxidase activity was preserved after immobilization and maintained after three months of shelf life if the sample was properly stored. The importance of using porous silicon oxide to maximize the surface area was also evidenced.

T-style keratoprosthesis based on surface-modified poly (2-hydroxyethyl methacrylate) hydrogel for cornea repairs

International Nuclear Information System (INIS)

Xiang, Jun; Sun, Jianguo; Hong, Jiaxu; Wang, Wentao; Wei, Anji; Le, Qihua; Xu, Jianjiang

2015-01-01

Corneal disease is a common cause of blindness, and keratoplasty is considered as an effective treatment method. However, there is a severe shortage of donor corneas worldwide. This paper presents a novel T-style design of a keratoprosthesis and its preparation methods, in which a mechanically and structurally effective artificial cornea is made based on a poly(2-hydroxyethyl methacrylate) hydrogel. The porous skirt was modified with hyaluronic acid and cationized gelatin, and the bottom of the optical column was coated with poly(ethylene glycol). The physical properties of the T-style Kpro were analyzed using ultraviolet and visible spectrophotometry and electron scanning microscopy. The surface chemical properties were characterized using Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy. The surface modification in the spongy skirt promoted cell adhesion and produced a firm bond between the corneal tissue and the implant device, while the surface modification in the optic column resisted cell adhesion and prevented retroprosthetic membrane formation. Through improved surgical techniques, the novel T-style keratoprosthesis provides enough mechanical stability to facilitate long-term biointegration with the host environment. In vivo implantation experiments showed that the T-style keratoprosthesis is a promising cornea alternative for patients with severe limbal stem cell deficiency and corneal opacity. - Highlights: • T-style keratoprosthesis was designed and prepared based on a PHEMA hydrogel. • Selective surface modifications effectively regulated cells' selective adhesion. • T-style keratoprosthesis provides enough mechanical stability to facilitate long-term biointegration with host tissues

T-style keratoprosthesis based on surface-modified poly (2-hydroxyethyl methacrylate) hydrogel for cornea repairs

Energy Technology Data Exchange (ETDEWEB)

Xiang, Jun [Department of Ophthalmology, Eye & ENT Hospital, Shanghai Medical College, Fudan University (China); Key Laboratory of Myopia, Ministry of Health, Fudan University (China); Shanghai Key Laboratory of Visual Impairment and Restoration, Fudan University (China); Sun, Jianguo [Research Center, Eye & ENT Hospital, Shanghai Medical College, Fudan University (China); Key Laboratory of Myopia, Ministry of Health, Fudan University (China); Shanghai Key Laboratory of Visual Impairment and Restoration, Fudan University (China); State Key Laboratory of Molecular Engineering of Polymers, Fudan University (China); Hong, Jiaxu [Department of Ophthalmology, Eye & ENT Hospital, Shanghai Medical College, Fudan University (China); Key Laboratory of Myopia, Ministry of Health, Fudan University (China); Shanghai Key Laboratory of Visual Impairment and Restoration, Fudan University (China); Wang, Wentao [Research Center, Eye & ENT Hospital, Shanghai Medical College, Fudan University (China); Key Laboratory of Myopia, Ministry of Health, Fudan University (China); Shanghai Key Laboratory of Visual Impairment and Restoration, Fudan University (China); Wei, Anji [Department of Ophthalmology, Eye & ENT Hospital, Shanghai Medical College, Fudan University (China); Key Laboratory of Myopia, Ministry of Health, Fudan University (China); Shanghai Key Laboratory of Visual Impairment and Restoration, Fudan University (China); Le, Qihua [Research Center, Eye & ENT Hospital, Shanghai Medical College, Fudan University (China); Key Laboratory of Myopia, Ministry of Health, Fudan University (China); Shanghai Key Laboratory of Visual Impairment and Restoration, Fudan University (China); Xu, Jianjiang, E-mail: jianjiang-xu@163.com [Department of Ophthalmology, Eye & ENT Hospital, Shanghai Medical College, Fudan University (China); Key Laboratory of Myopia, Ministry of Health, Fudan University (China); Shanghai Key Laboratory of Visual Impairment and Restoration, Fudan University (China)

2015-05-01

Corneal disease is a common cause of blindness, and keratoplasty is considered as an effective treatment method. However, there is a severe shortage of donor corneas worldwide. This paper presents a novel T-style design of a keratoprosthesis and its preparation methods, in which a mechanically and structurally effective artificial cornea is made based on a poly(2-hydroxyethyl methacrylate) hydrogel. The porous skirt was modified with hyaluronic acid and cationized gelatin, and the bottom of the optical column was coated with poly(ethylene glycol). The physical properties of the T-style Kpro were analyzed using ultraviolet and visible spectrophotometry and electron scanning microscopy. The surface chemical properties were characterized using Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy. The surface modification in the spongy skirt promoted cell adhesion and produced a firm bond between the corneal tissue and the implant device, while the surface modification in the optic column resisted cell adhesion and prevented retroprosthetic membrane formation. Through improved surgical techniques, the novel T-style keratoprosthesis provides enough mechanical stability to facilitate long-term biointegration with the host environment. In vivo implantation experiments showed that the T-style keratoprosthesis is a promising cornea alternative for patients with severe limbal stem cell deficiency and corneal opacity. - Highlights: • T-style keratoprosthesis was designed and prepared based on a PHEMA hydrogel. • Selective surface modifications effectively regulated cells' selective adhesion. • T-style keratoprosthesis provides enough mechanical stability to facilitate long-term biointegration with host tissues.

High Stability Pentacene Transistors Using Polymeric Dielectric Surface Modifier.

Science.gov (United States)