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Sample records for vitro glutathionylation potential

  1. Susceptibility of isolated myofibrils to in vitro glutathionylation: Potential relevance to muscle functions.

    Science.gov (United States)

    Passarelli, Chiara; Di Venere, Almerinda; Piroddi, Nicoletta; Pastore, Anna; Scellini, Beatrice; Tesi, Chiara; Petrini, Stefania; Sale, Patrizio; Bertini, Enrico; Poggesi, Corrado; Piemonte, Fiorella

    2010-02-01

    In this study we investigated the molecular mechanism of glutathionylation on isolated human cardiac myofibrils using several pro-glutathionylating agents. Total glutathionylated proteins appeared significantly enhanced with all the pro-oxidants used. The increase was completely reversed by the addition of a reducing agent, demonstrating that glutathione binding occurs by a disulfide and that the process is reversible. A sensitive target of glutathionylation was alpha-actin, showing a different reactivity to the several pro-glutathionylating agents by ELISA. Noteworthy, myosin although highly sensitive to the in vitro glutathionylation does not represent the primary glutathionylation target in isolated myofibrils. Light scattering measurements of the glutathionylated alpha-actin showed a slower polymerisation compared to the non-glutathionylated protein and force development was depressed after glutathionylation, when the myofibrils were mounted in a force recording apparatus. Interestingly, confocal laser scanning microscopy of cardiac cryosections indicated, for the first time, the constitutive glutathionylation of alpha-cardiac actin in human heart. Due to the critical location of alpha-actin in the contractile machinery and to its susceptibility to the oxidative modifications, glutathionylation may represent a mechanism for modulating sarcomere assembly and muscle functionality under patho-physiological conditions in vivo. 2009 Wiley-Liss, Inc.

  2. Protein glutathionylation in health and disease.

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    Ghezzi, Pietro

    2013-05-01

    It is now recognized that protein cysteines exist not only as free thiols or intramolecular disulfides, that help maintain the 3D structure of proteins, but can also undergo different types of oxidation, one of which is glutathionylation, or the formation of mixed disulfides with glutathione (GSH). We will discuss how proteins can undergo glutathionylation and how this can affect the protein characteristics/function. Glutathionylation is reversible and de-glutathionylation can be catalysed by protein thiol-disulfide oxidoreductases. Genetic modification of the expression of these enzymes, particularly glutaredoxin, using overexpression, knockout mice or siRNA, is becoming an important tool to study the role of protein glutathionylation. While in the past this post-translational modification was mainly known in the context of oxidative stress, measurement of glutathionylated proteins in patients is pointing out a potential importance if this modification in pathogenesis and could identify new biomarkers. We also wanted to point out the main findings in the role of glutathionylation in diseases and drug action. We identify two major open problems in the field, namely the complexity of the mechanisms responsible for glutathionylation and de-glutathionylation, as well as what makes a protein susceptible to glutathionylation. This review underlines the peculiarities of this post-translational modification and their biological role. This article is part of a Special Issue entitled Cellular functions of glutathione. Copyright © 2013 Elsevier B.V. All rights reserved.

  3. Glutathionylation of the Aquaporin-2 Water Channel

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    Tamma, Grazia; Ranieri, Marianna; Di Mise, Annarita; Centrone, Mariangela; Svelto, Maria; Valenti, Giovanna

    2014-01-01

    Aquaporin-2 (AQP2) is the vasopressin-regulated water channel that controls renal water reabsorption and urine concentration. AQP2 undergoes different regulated post-translational modifications, including phosphorylation and ubiquitylation, which are fundamental for controlling AQP2 cellular localization, stability, and function. The relationship between AQP2 and S-glutathionylation is of potential interest because reactive oxygen species (ROS), produced under renal failure or nephrotoxic drugs, may influence renal function as well as the expression and the activity of different transporters and channels, including aquaporins. Here, we show for the first time that AQP2 is subjected to S-glutathionylation in kidney and in HEK-293 cells stably expressing AQP2. S-Glutathionylation is a redox-dependent post-translational modification controlling several signal transduction pathways and displaying an acute effect on free cytosolic calcium concentration. Interestingly, we found that in fresh kidney slices, the increased AQP2 S-glutathionylation correlated with tert-butyl hydroperoxide-induced ROS generation. Moreover, we also found that cells expressing wild-type human calcium-sensing receptor (hCaSR-wt) and its gain of function (hCaSR-R990G; hCaSR-N124K) had a significant decrease in AQP2 S-glutathionylation secondary to reduced ROS levels and reduced basal intracellular calcium concentration compared with mock cells. Together, these new findings provide fundamental insight into cell biological aspects of AQP2 function and may be relevant to better understand and explain pathological states characterized by an oxidative stress and AQP2-dependent water reabsorption disturbs. PMID:25112872

  4. Histone h3 glutathionylation in proliferating mammalian cells destabilizes nucleosomal structure.

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    García-Giménez, José Luis; Òlaso, Gloria; Hake, Sandra B; Bönisch, Clemens; Wiedemann, Sonja M; Markovic, Jelena; Dasí, Francisco; Gimeno, Amparo; Pérez-Quilis, Carme; Palacios, Oscar; Capdevila, Mercè; Viña, José; Pallardó, Federico V

    2013-10-20

    Here we report that chromatin, the complex and dynamic eukaryotic DNA packaging structure, is able to sense cellular redox changes. Histone H3, the only nucleosomal protein that possesses cysteine(s), can be modified by glutathione (GSH). Using Biotin labeled glutathione ethyl ester (BioGEE) treatment of nucleosomes in vitro, we show that GSH, the most abundant antioxidant in mammals, binds to histone H3. BioGEE treatment of NIH3T3 cells indicates that glutathionylation of H3 is maximal in fast proliferating cells, correlating well with enhanced levels of H3 glutathionylation in different tumor cell lines. Furthermore, glutathionylation of H3 in vivo decreases in livers from aged SAMP8 and C57BL/6J mice. We demonstrate biochemically and by mass spectrometry that histone variants H3.2/H3.3 are glutathionylated on their cysteine residue 110. Furthermore, circular dichroism, thermal denaturation of reconstituted nucleosomes, and molecular modeling indicate that glutathionylation of histone H3 produces structural changes affecting nucleosomal stability. We characterize the implications of histone H3 glutathionylation in cell physiology and the modulation of core histone proteins structure affected by this modification. Histone H3 senses cellular redox changes through glutathionylation of Cys, which increases during cell proliferation and decreases during aging. Glutathionylation of histone H3 affects nucleosome stability structure leading to a more open chromatin structure.

  5. Cadmium-induced glutathionylation of actin occurs through a ROS-independent mechanism: Implications for cytoskeletal integrity

    Energy Technology Data Exchange (ETDEWEB)

    Choong, Grace; Liu, Ying; Xiao, Weiqun; Templeton, Douglas M., E-mail: doug.templeton@utoronto.ca

    2013-10-15

    Cadmium disrupts the actin cytoskeleton in rat mesangial cells, and we have previously shown that this involves a complex interplay involving activation of kinase signaling, protein translocation, and disruption of focal adhesions. Here we investigate the role that glutathionylation of actin plays in Cd{sup 2+}-associated cytoskeletal reorganization. Low concentrations of Cd{sup 2+} (0.5–2 μM) caused an increase in actin glutathionylation by 6 h, whereas at higher concentrations glutathionylation remained at basal levels. Although oxidation with diamide increased glutathionylation, reactive oxygen species (ROS) were not involved in the Cd{sup 2+}-dependent effect, as only Cd{sup 2+} concentrations above 2 μM were sufficient to increase ROS. However, low [Cd{sup 2+}] increased total glutathione levels without affecting the ratio of reduced/oxidized glutathione, and inhibition of glutathione synthesis suppressed actin glutathionylation. Cadmium increased the activity of the enzyme glutaredoxin, which influences the equilibrium between glutathionylated and deglutathionylated proteins and thus may influence levels of glutathionylated actin. Together these observations show that cadmium-dependent effects on actin glutathionylation are affected by glutathione metabolism and not by direct effects of ROS on thiol chemistry. In vitro polymerization assays with glutathionylated actin show a decreased rate of polymerization. In contrast, immunofluorescence of cytoskeletal structure in intact cells suggests that increases in actin glutathionylation accompanying increased glutathione levels occurring under low Cd{sup 2+} exposure are protective in vivo, with cytoskeletal disruption ensuing only when higher Cd{sup 2+} concentrations increase ROS levels and prevent an increase in actin–glutathione conjugates. - Highlights: • Cadmium disrupts the actin cytoskeleton in mesangial cells. • Cadmium induces glutathionylation of actin at low concentrations.

  6. S-glutathionylation reactions in mitochondrial function and disease

    Directory of Open Access Journals (Sweden)

    Ryan J. Mailloux

    2014-11-01

    Full Text Available Mitochondria are highly efficient energy-transforming organelles that convert energy stored in carbon bonds into the universal energy currency ATP. The production of ATP by mitochondria is dependent on oxidation of nutrients and coupling of exergonic electron transfer reactions to the genesis of transmembrane electrochemical potential of protons. Electrons can also prematurely spin-off from the respiratory complexes and univalently reduce di-oxygen to generate ROS, an important signaling molecule that can be toxic at high concentrations. Production of ATP and ROS are intimately linked by the respiratory chain and the genesis of one or the other inherently depends on the metabolic state of mitochondria. Various control mechanisms converge on mitochondria to adjust ATP and ROS output in response to changing cellular demands. One control mechanism that has gained a high amount of attention recently is S-glutathionylation, a redox sensitive covalent modification that involves formation of a disulfide bridge between glutathione and an available protein cysteine thiol. A number of S-glutathionylation targets have been identified in mitochondria. It has also been established that S-glutathionylation reactions in mitochondria are mediated by the thiol oxidoreductase glutaredoxin-2 (Grx2. In the following review, emerging knowledge on S-glutathionylation reactions and its importance in modulation mitochondrial ATP and ROS production will be discussed. Major focus will be placed on Complex I of the respiratory chain since 1 it is a target for reversible S-glutathionylation by Grx2 and 2 deregulation of Complex I S-glutathionylation is associated with development of various disease states particularly heart disease. Other mitochondrial enzymes and how their S-glutathionylation profile is affected in different disease states will also be discussed.

  7. Proteomic Identification and Quantification of S-glutathionylation in Mouse Macrophages Using Resin-Assisted Enrichment and Isobaric Labeling

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    Su, Dian; Gaffrey, Matthew J.; Guo, Jia; Hatchell, Kayla E.; Chu, Rosalie K.; Clauss, Therese RW; Aldrich, Joshua T.; Wu, Si; Purvine, Samuel O.; Camp, David G.; Smith, Richard D.; Thrall, Brian D.; Qian, Weijun

    2014-02-11

    Protein S-glutathionylation (SSG) is an important regulatory posttranslational modification of protein cysteine (Cys) thiol redox switches, yet the role of specific cysteine residues as targets of modification is poorly understood. We report a novel quantitative mass spectrometry (MS)-based proteomic method for site-specific identification and quantification of S-glutathionylation across different conditions. Briefly, this approach consists of initial blocking of free thiols by alkylation, selective reduction of glutathionylated thiols and enrichment using thiol affinity resins, followed by on-resin tryptic digestion and isobaric labeling with iTRAQ (isobaric tags for relative and absolute quantitation) for MS-based identification and quantification. The overall approach was validated by application to RAW 264.7 mouse macrophages treated with different doses of diamide to induce glutathionylation. A total of 1071 Cys-sites from 690 proteins were identified in response to diamide treatment, with ~90% of the sites displaying >2-fold increases in SSG-modification compared to controls.. This approach was extended to identify potential SSG modified Cys-sites in response to H2O2, an endogenous oxidant produced by activated macrophages and many pathophysiological stimuli. The results revealed 364 Cys-sites from 265 proteins that were sensitive to S-glutathionylation in response to H2O2 treatment. These proteins covered a range of molecular types and molecular functions with free radical scavenging, and cell death and survival included as the most significantly enriched functional categories. Overall the results demonstrate that our approach is effective for site-specific identification and quantification of S-glutathionylated proteins. The analytical strategy also provides a unique approach to determining the major pathways and cell processes most susceptible to glutathionylation at a proteome-wide scale.

  8. S-Glutathionylation of estrogen receptor alpha affects dendritic cell function.

    Science.gov (United States)

    Zhang, Jie; Ye, Zhi-Wei; Chen, Wei; Manevich, Yefim; Mehrotra, Shikhar; Ball, Lauren E; Janssen-Heininger, Yvonne M; Tew, Kenneth D; Townsend, Danyelle M

    2018-01-26

    Glutathione S-transferase P (GSTP) is a thiolase that catalyzes the addition of glutathione (GSH) to receptive cysteines in target proteins, producing an S-glutathionylated residue. Accordingly, previous studies have reported that S-glutathionylation is constitutively decreased in cells from mice lacking GSTP ( Gstp1/p2 -/- ). Here, we found that bone marrow-derived dendritic cells (BMDDC) from Gstp1/p2 -/- mice have proliferation rates that are greater than those in their WT counterparts ( Gstp1/p2 +/+ ). Moreover, Gstp1/p2 -/- BMDCC had increased reactive oxygen species (ROS) levels and decreased GSH:glutathione disulfide (GSSG) ratios. Estrogen receptor alpha (ERα) is linked to myeloproliferation and differentiation, and we observed that its steady-state levels are elevated in Gstp1/p2 -/- BMDDC, indicating a link between GSTP and ERα activities. BMDDC differentiated by granulocyte-macrophage colony-stimulating factor had elevated ERα levels, which were more pronounced in Gstp1/p2 -/- than WT mice. When stimulated with lipopolysaccharide for maturation, Gstp1/p2 -/- BMDDC exhibited augmented endocytosis, maturation rate, cytokine secretion, and T-cell activation; heightened glucose uptake and glycolysis; increased Akt signaling (in the mTOR pathway); and decreased AMPK-mediated phosphorylation of proteins. Of note, GSTP formed a complex with ERα, stimulating ERα S-glutathionylation at cysteines 221, 245, 417, and 447, altering ERα's binding affinity for estradiol and reduced overall binding potential (receptor density and affinity) threefold. Moreover, in Gstp1/p2 -/- BMDDC, ERα S-glutathionylation was constitutively decreased. Taken together, these findings suggest that GSTP-mediated S-glutathionylation of ERα controls BMDDC differentiation and affects metabolic function in dendritic cells. Copyright © 2018, The American Society for Biochemistry and Molecular Biology.

  9. Modulation of the specific glutathionylation of mitochondrial proteins in the yeast Saccharomyces cerevisiae under basal and stress conditions.

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    Gergondey, Rachel; Garcia, Camille; Marchand, Christophe H; Lemaire, Stephane D; Camadro, Jean-Michel; Auchère, Françoise

    2017-03-15

    The potential biological consequences of oxidative stress and changes in glutathione levels include the oxidation of susceptible protein thiols and reversible covalent binding of glutathione to the -SH groups of proteins by S-glutathionylation. Mitochondria are central to the response to oxidative stress and redox signaling. It is therefore crucial to explore the adaptive response to changes in thiol-dependent redox status in these organelles. We optimized the purification protocol of glutathionylated proteins in the yeast Saccharomyces cerevisiae and present a detailed proteomic analysis of the targets of protein glutathionylation in cells undergoing constitutive metabolism and after exposure to various stress conditions. This work establishes the physiological importance of the glutathionylation process in S. cerevisiae under basal conditions and provides evidence for an atypical and unexpected cellular distribution of the process between the cytosol and mitochondria. In addition, our data indicate that each oxidative condition (diamide, GSSG, H2O2, or the presence of iron) elicits an adaptive metabolic response affecting specific mitochondrial metabolic pathways, mainly involved in the energetic maintenance of the cells. The correlation of protein modifications with intracellular glutathione levels suggests that protein deglutathionylation may play a role in protecting mitochondria from oxidative stress. This work provides further insights into the diversity of proteins undergoing glutathionylation and the role of this post-translational modification as a regulatory process in the adaptive response of the cell. © 2017 The Author(s); published by Portland Press Limited on behalf of the Biochemical Society.

  10. Prediction of S-glutathionylation sites based on protein sequences.

    Directory of Open Access Journals (Sweden)

    Chenglei Sun

    Full Text Available S-glutathionylation, the reversible formation of mixed disulfides between glutathione(GSH and cysteine residues in proteins, is a specific form of post-translational modification that plays important roles in various biological processes, including signal transduction, redox homeostasis, and metabolism inside cells. Experimentally identifying S-glutathionylation sites is labor-intensive and time consuming, whereas bioinformatics methods provide an alternative way to this problem by predicting S-glutathionylation sites in silico. The bioinformatics approaches give not only candidate sites for further experimental verification but also bio-chemical insights into the mechanism of S-glutathionylation. In this paper, we firstly collect experimentally determined S-glutathionylated proteins and their corresponding modification sites from the literature, and then propose a new method for predicting S-glutathionylation sites by employing machine learning methods based on protein sequence data. Promising results are obtained by our method with an AUC (area under ROC curve score of 0.879 in 5-fold cross-validation, which demonstrates the predictive power of our proposed method. The datasets used in this work are available at http://csb.shu.edu.cn/SGDB.

  11. Increased TRPC5 glutathionylation contributes to striatal neuron loss in Huntington's disease.

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    Hong, Chansik; Seo, Hyemyung; Kwak, Misun; Jeon, Jeha; Jang, Jihoon; Jeong, Eui Man; Myeong, Jongyun; Hwang, Yu Jin; Ha, Kotdaji; Kang, Min Jueng; Lee, Kyu Pil; Yi, Eugene C; Kim, In-Gyu; Jeon, Ju-Hong; Ryu, Hoon; So, Insuk

    2015-10-01

    Aberrant glutathione or Ca(2+) homeostasis due to oxidative stress is associated with the pathogenesis of neurodegenerative disorders. The Ca(2+)-permeable transient receptor potential cation (TRPC) channel is predominantly expressed in the brain, which is sensitive to oxidative stress. However, the role of the TRPC channel in neurodegeneration is not known. Here, we report a mechanism of TRPC5 activation by oxidants and the effect of glutathionylated TRPC5 on striatal neurons in Huntington's disease. Intracellular oxidized glutathione leads to TRPC5 activation via TRPC5 S-glutathionylation at Cys176/Cys178 residues. The oxidized glutathione-activated TRPC5-like current results in a sustained increase in cytosolic Ca(2+), activated calmodulin-dependent protein kinase and the calpain-caspase pathway, ultimately inducing striatal neuronal cell death. We observed an abnormal glutathione pool indicative of an oxidized state in the striatum of Huntington's disease transgenic (YAC128) mice. Increased levels of endogenous TRPC5 S-glutathionylation were observed in the striatum in both transgenic mice and patients with Huntington's disease. Both knockdown and inhibition of TRPC5 significantly attenuated oxidation-induced striatal neuronal cell death. Moreover, a TRPC5 blocker improved rearing behaviour in Huntington's disease transgenic mice and motor behavioural symptoms in littermate control mice by increasing striatal neuron survival. Notably, low levels of TRPC1 increased the formation of TRPC5 homotetramer, a highly Ca(2+)-permeable channel, and stimulated Ca(2+)-dependent apoptosis in Huntington's disease cells (STHdh(Q111/111)). Taken together, these novel findings indicate that increased TRPC5 S-glutathionylation by oxidative stress and decreased TRPC1 expression contribute to neuronal damage in the striatum and may underlie neurodegeneration in Huntington's disease. © The Author (2015). Published by Oxford University Press on behalf of the Guarantors of Brain

  12. Increased TRPC5 glutathionylation contributes to striatal neuron loss in Huntington’s disease

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    Hong, Chansik; Seo, Hyemyung; Kwak, Misun; Jeon, Jeha; Jang, Jihoon; Jeong, Eui Man; Myeong, Jongyun; Hwang, Yu Jin; Ha, Kotdaji; Kang, Min Jueng; Lee, Kyu Pil; Yi, Eugene C.; Kim, In-Gyu; Jeon, Ju-Hong

    2015-01-01

    Aberrant glutathione or Ca2+ homeostasis due to oxidative stress is associated with the pathogenesis of neurodegenerative disorders. The Ca2+-permeable transient receptor potential cation (TRPC) channel is predominantly expressed in the brain, which is sensitive to oxidative stress. However, the role of the TRPC channel in neurodegeneration is not known. Here, we report a mechanism of TRPC5 activation by oxidants and the effect of glutathionylated TRPC5 on striatal neurons in Huntington’s disease. Intracellular oxidized glutathione leads to TRPC5 activation via TRPC5 S-glutathionylation at Cys176/Cys178 residues. The oxidized glutathione-activated TRPC5-like current results in a sustained increase in cytosolic Ca2+, activated calmodulin-dependent protein kinase and the calpain-caspase pathway, ultimately inducing striatal neuronal cell death. We observed an abnormal glutathione pool indicative of an oxidized state in the striatum of Huntington’s disease transgenic (YAC128) mice. Increased levels of endogenous TRPC5 S-glutathionylation were observed in the striatum in both transgenic mice and patients with Huntington’s disease. Both knockdown and inhibition of TRPC5 significantly attenuated oxidation-induced striatal neuronal cell death. Moreover, a TRPC5 blocker improved rearing behaviour in Huntington’s disease transgenic mice and motor behavioural symptoms in littermate control mice by increasing striatal neuron survival. Notably, low levels of TRPC1 increased the formation of TRPC5 homotetramer, a highly Ca2+-permeable channel, and stimulated Ca2+-dependent apoptosis in Huntington’s disease cells (STHdhQ111/111). Taken together, these novel findings indicate that increased TRPC5 S-glutathionylation by oxidative stress and decreased TRPC1 expression contribute to neuronal damage in the striatum and may underlie neurodegeneration in Huntington’s disease. PMID:26133660

  13. S-Glutathionylation and Redox Protein Signaling in Drug Addiction.

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    Womersley, Jacqueline S; Uys, Joachim D

    2016-01-01

    Drug addiction is a chronic relapsing disorder that comes at a high cost to individuals and society. Therefore understanding the mechanisms by which drugs exert their effects is of prime importance. Drugs of abuse increase the production of reactive oxygen and nitrogen species resulting in oxidative stress. This change in redox homeostasis increases the conjugation of glutathione to protein cysteine residues; a process called S-glutathionylation. Although traditionally regarded as a protective mechanism against irreversible protein oxidation, accumulated evidence suggests a more nuanced role for S-glutathionylation, namely as a mediator in redox-sensitive protein signaling. The reversible modification of protein thiols leading to alteration in function under different physiologic/pathologic conditions provides a mechanism whereby change in redox status can be translated into a functional response. As such, S-glutathionylation represents an understudied means of post-translational protein modification that may be important in the mechanisms underlying drug addiction. This review will discuss the evidence for S-glutathionylation as a redox-sensing mechanism and how this may be involved in the response to drug-induced oxidative stress. The function of S-glutathionylated proteins involved in neurotransmission, dendritic spine structure, and drug-induced behavioral outputs will be reviewed with specific reference to alcohol, cocaine, and heroin. Copyright © 2016. Published by Elsevier Inc.

  14. Identification of o-quinone/quinone methide metabolites of quercetin in a cellular in vitro system

    NARCIS (Netherlands)

    Awad, H.M.; Boersma, M.G.; Boeren, S.; Woude, van der H.; Zanden, van J.; Bladeren, van P.J.; Vervoort, J.; Rietjens, I.M.C.M.

    2002-01-01

    Formation of quercetin quinone/quinone methide metabolites, reflected by formation of the glutathionyl quercetin adducts as authentic metabolites, was investigated in an in vitro cell model (B16F-10 melanoma cells). Results of the present study clearly indicate the formation of glutathionyl

  15. Short Communication In vitro evaluation of antagonistic potential ...

    African Journals Online (AJOL)

    Short Communication In vitro evaluation of antagonistic potential activity and assay of culture filtrates of Trichoderma harzianum and Trichoderma viride against coffee wilt pathogen ( Gibberella xylarioides )

  16. Positive Regulation of Interleukin-1β Bioactivity by Physiological ROS-Mediated Cysteine S-Glutathionylation

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    Xue Zhang

    2017-07-01

    Full Text Available Reactive oxygen species (ROS-induced cysteine S-glutathionylation is an important posttranslational modification (PTM that controls a wide range of intracellular protein activities. However, whether physiological ROS can modulate the function of extracellular components via S-glutathionylation is unknown. Using a screening approach, we identified ROS-mediated cysteine S-glutathionylation on several extracellular cytokines. Glutathionylation of the highly conserved Cys-188 in IL-1β positively regulates its bioactivity by preventing its ROS-induced irreversible oxidation, including sulfinic acid and sulfonic acid formation. We show this mechanism protects IL-1β from deactivation by ROS in an in vivo system of irradiation-induced bone marrow (BM injury. Glutaredoxin 1 (Grx1, an enzyme that catalyzes deglutathionylation, was present and active in the extracellular space in serum and the BM, physiologically regulating IL-1β glutathionylation and bioactivity. Collectively, we identify cysteine S-glutathionylation as a cytokine regulatory mechanism that could be a therapeutic target in the treatment of various infectious and inflammatory diseases.

  17. GSHSite: exploiting an iteratively statistical method to identify s-glutathionylation sites with substrate specificity.

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    Yi-Ju Chen

    Full Text Available S-glutathionylation, the covalent attachment of a glutathione (GSH to the sulfur atom of cysteine, is a selective and reversible protein post-translational modification (PTM that regulates protein activity, localization, and stability. Despite its implication in the regulation of protein functions and cell signaling, the substrate specificity of cysteine S-glutathionylation remains unknown. Based on a total of 1783 experimentally identified S-glutathionylation sites from mouse macrophages, this work presents an informatics investigation on S-glutathionylation sites including structural factors such as the flanking amino acids composition and the accessible surface area (ASA. TwoSampleLogo presents that positively charged amino acids flanking the S-glutathionylated cysteine may influence the formation of S-glutathionylation in closed three-dimensional environment. A statistical method is further applied to iteratively detect the conserved substrate motifs with statistical significance. Support vector machine (SVM is then applied to generate predictive model considering the substrate motifs. According to five-fold cross-validation, the SVMs trained with substrate motifs could achieve an enhanced sensitivity, specificity, and accuracy, and provides a promising performance in an independent test set. The effectiveness of the proposed method is demonstrated by the correct identification of previously reported S-glutathionylation sites of mouse thioredoxin (TXN and human protein tyrosine phosphatase 1b (PTP1B. Finally, the constructed models are adopted to implement an effective web-based tool, named GSHSite (http://csb.cse.yzu.edu.tw/GSHSite/, for identifying uncharacterized GSH substrate sites on the protein sequences.

  18. Increased oxidation-related glutathionylation and carbonic anhydrase activity in endometriosis.

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    Andrisani, Alessandra; Donà, Gabriella; Brunati, Anna Maria; Clari, Giulio; Armanini, Decio; Ragazzi, Eugenio; Ambrosini, Guido; Bordin, Luciana

    2014-06-01

    permeability and adhesion molecule expression, thus contributing to ongoing inflammation. Due to their main cellular functions--delivery of O2 from lung to tissue and removal of CO2 from tissue to lung--red blood cells (RBC) are exposed to oxidative stress. Carbon dioxide in tissue capillaries diffuses into red cells, where it is rapidly hydrated by the action of cytosolic carbonic anhydrase. Analysis of the oxidation status of endometriotic RBC membranes showed a high content of glutathionylated proteins, indicating pre-existing oxidation-related alterations. The increase in glutathionylated proteins was correlated to increased carbonic anhydrase activity in endometriotic RBC compared with healthy controls. Carbonic anhydrase is a family of metalloenzymes involved in many physiological processes such as acid-base homeostasis, respiration, carbon dioxide and ion transport, and bone resorption, and in the regulation of ureagenesis, gluconeogenesis, lipogenesis and tumourigenesis. Due to the potential implication of carbonic anhydrase activation in many pathologies, such as glaucoma, hypertension, obesity and infections, carbonic anhydrase activity should be closely monitored in endometriosis to prevent possible complications and/or worsening of related conditions. Copyright © 2014 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.

  19. S-glutathionylation of cryptic cysteines enhances titin elasticity by blocking protein folding

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    Alegre-Cebollada, Jorge; Kosuri, Pallav; Giganti, David; Eckels, Edward; Rivas-Pardo, Jaime Andrés; Hamdani, Nazha; Warren, Chad M.; Solaro, R. John; Linke, Wolfgang A.; Fernández, Julio M.

    2014-01-01

    The giant elastic protein titin is a determinant factor in how much blood fills the left ventricle during diastole, and thus in the etiology of heart disease. Titin has been identified as a target of S-glutathionylation, an end product of the nitric oxide signaling cascade that increases cardiac muscle elasticity. However, it is unknown how S-glutathionylation may regulate the elasticity of titin and cardiac tissue. Here we show that mechanical unfolding of titin immunoglobulin (Ig) domains exposes buried cysteine residues, which then can be S-glutathionylated. S-glutathionylation of cryptic cysteines greatly decreases the mechanical stability of the parent Ig domain as well as its ability to fold. Both effects favor a more extensible state of titin. Furthermore we demonstrate that S-glutathionylation of cryptic cysteines in titin mediates mechano-chemical modulation of the elasticity of human cardiomyocytes. We propose that posttranslational modification of cryptic residues is a general mechanism to regulate tissue elasticity. PMID:24630725

  20. Identification of DUOX1-dependent redox signaling through protein S-glutathionylation in airway epithelial cells☆

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    Hristova, Milena; Veith, Carmen; Habibovic, Aida; Lam, Ying-Wai; Deng, Bin; Geiszt, Miklos; Janssen-Heininger, Yvonne M.W.; van der Vliet, Albert

    2014-01-01

    The NADPH oxidase homolog dual oxidase 1 (DUOX1) plays an important role in innate airway epithelial responses to infection or injury, but the precise molecular mechanisms are incompletely understood and the cellular redox-sensitive targets for DUOX1-derived H2O2 have not been identified. The aim of the present study was to survey the involvement of DUOX1 in cellular redox signaling by protein S-glutathionylation, a major mode of reversible redox signaling. Using human airway epithelial H292 cells and stable transfection with DUOX1-targeted shRNA as well as primary tracheal epithelial cells from either wild-type or DUOX1-deficient mice, DUOX1 was found to be critical in ATP-stimulated transient production of H2O2 and increased protein S-glutathionylation. Using cell pre-labeling with biotin-tagged GSH and analysis of avidin-purified proteins by global proteomics, 61 S-glutathionylated proteins were identified in ATP-stimulated cells compared to 19 in untreated cells. Based on a previously established role of DUOX1 in cell migration, various redox-sensitive proteins with established roles in cytoskeletal dynamics and/or cell migration were evaluated for S-glutathionylation, indicating a critical role for DUOX1 in ATP-stimulated S-glutathionylation of β-actin, peroxiredoxin 1, the non-receptor tyrosine kinase Src, and MAPK phosphatase 1. Overall, our studies demonstrate the importance of DUOX1 in epithelial redox signaling through reversible S-glutathionylation of a range of proteins, including proteins involved in cytoskeletal regulation and MAPK signaling pathways involved in cell migration. PMID:24624333

  1. Identification of DUOX1-dependent redox signaling through protein S-glutathionylation in airway epithelial cells

    Directory of Open Access Journals (Sweden)

    Milena Hristova

    2014-01-01

    Full Text Available The NADPH oxidase homolog dual oxidase 1 (DUOX1 plays an important role in innate airway epithelial responses to infection or injury, but the precise molecular mechanisms are incompletely understood and the cellular redox-sensitive targets for DUOX1-derived H2O2 have not been identified. The aim of the present study was to survey the involvement of DUOX1 in cellular redox signaling by protein S-glutathionylation, a major mode of reversible redox signaling. Using human airway epithelial H292 cells and stable transfection with DUOX1-targeted shRNA as well as primary tracheal epithelial cells from either wild-type or DUOX1-deficient mice, DUOX1 was found to be critical in ATP-stimulated transient production of H2O2 and increased protein S-glutathionylation. Using cell pre-labeling with biotin-tagged GSH and analysis of avidin-purified proteins by global proteomics, 61 S-glutathionylated proteins were identified in ATP-stimulated cells compared to 19 in untreated cells. Based on a previously established role of DUOX1 in cell migration, various redox-sensitive proteins with established roles in cytoskeletal dynamics and/or cell migration were evaluated for S-glutathionylation, indicating a critical role for DUOX1 in ATP-stimulated S-glutathionylation of β-actin, peroxiredoxin 1, the non-receptor tyrosine kinase Src, and MAPK phosphatase 1. Overall, our studies demonstrate the importance of DUOX1 in epithelial redox signaling through reversible S-glutathionylation of a range of proteins, including proteins involved in cytoskeletal regulation and MAPK signaling pathways involved in cell migration.

  2. In vitro control of Alternaria citri using antifungal potentials of ...

    African Journals Online (AJOL)

    In vitro control of Alternaria citri using antifungal potentials of Trichoderma species. Asma Murtaza, Shazia Shafique, Tehmina Anjum, Sobiya Shafique. Abstract. The antifungal potential of five species of Trichoderma viz., Trichoderma viride, Trichoderma aureoviride, Trichoderma reesei, Trichoderma koningii and ...

  3. Glutathionylation of Yersinia pestis LcrV and Its Effects on Plague Pathogenesis

    Directory of Open Access Journals (Sweden)

    Anthony Mitchell

    2017-05-01

    Full Text Available Glutathionylation, the formation of reversible mixed disulfides between glutathione and protein cysteine residues, is a posttranslational modification previously observed for intracellular proteins of bacteria. Here we show that Yersinia pestis LcrV, a secreted protein capping the type III secretion machine, is glutathionylated at Cys273 and that this modification promotes association with host ribosomal protein S3 (RPS3, moderates Y. pestis type III effector transport and killing of macrophages, and enhances bubonic plague pathogenesis in mice and rats. Secreted LcrV was purified and analyzed by mass spectrometry to reveal glutathionylation, a modification that is abolished by the codon substitution Cys273Ala in lcrV. Moreover, the lcrVC273A mutation enhanced the survival of animals in models of bubonic plague. Investigating the molecular mechanism responsible for these virulence attributes, we identified macrophage RPS3 as a ligand of LcrV, an association that is perturbed by the Cys273Ala substitution. Furthermore, macrophages infected by the lcrVC273A variant displayed accelerated apoptotic death and diminished proinflammatory cytokine release. Deletion of gshB, which encodes glutathione synthetase of Y. pestis, resulted in undetectable levels of intracellular glutathione, and we used a Y. pestis ΔgshB mutant to characterize the biochemical pathway of LcrV glutathionylation, establishing that LcrV is modified after its transport to the type III needle via disulfide bond formation with extracellular oxidized glutathione.

  4. Glutathione controls the redox state of the mitochondrial carnitine/acylcarnitine carrier Cys residues by glutathionylation.

    Science.gov (United States)

    Giangregorio, Nicola; Palmieri, Ferdinando; Indiveri, Cesare

    2013-11-01

    The mitochondrial carnitine/acylcarnitine carrier (CAC) is essential for cell metabolism since it catalyzes the transport of acylcarnitines into mitochondria allowing the β-oxidation of fatty acids. CAC functional and structural properties have been characterized. Cys residues which could form disulfides suggest the involvement of CAC in redox switches. The effect of GSH and GSSG on the [(3)H]-carnitine/carnitine antiport catalyzed by the CAC in proteoliposomes has been studied. The Cys residues involved in the redox switch have been identified by site-directed mutagenesis. Glutathionylated CAC has been assessed by glutathionyl-protein specific antibody. GSH led to increase of transport activity of the CAC extracted from liver mitochondria. A similar effect was observed on the recombinant CAC. The presence of glutaredoxin-1 (Grx1) accelerated the GSH activation of the recombinant CAC. The effect was more evident at 37°C. GSSG led to transport inhibition which was reversed by dithioerythritol (DTE). The effects of GSH and GSSG were studied on CAC Cys-mutants. CAC lacking C136 and C155 was insensitive to both reagents. Mutants containing these two Cys responded as the wild-type. Anti-glutathionyl antibody revealed the formation of glutathionylated CAC. CAC is redox-sensitive and it is regulated by the GSH/GSSG couple. C136 and C155 are responsible for the regulation which occurs through glutathionylation. CAC is sensitive to the redox state of the cell switching between oxidized and reduced forms in response to variation of GSSG and GSH concentrations. © 2013.

  5. Phytochemicals and in vitro antioxidant potentials of defatted ...

    African Journals Online (AJOL)

    Plant-based dietary components and additives are known to protect cells from deleterious effect of reactive oxygen species (ROS). Proximate, phytochemical and antioxidant potentials of methanolic extract of defatted Holarrhena floribunda (G.Don) leaves were assessed using in vitro systems such as, 1,1 ...

  6. Androgenic potential and anther in vitro culture of Lagenaria ...

    African Journals Online (AJOL)

    Androgenic potential and anther in vitro culture of Lagenaria siceraria (Molina) Standl an edible-seed cucurbit. Kouakou Laurent Kouakou, Tra Serge Doubi, Tra Serge Koffi, Kouadio Ignace Kouassi, Tanoh Hilaire Kouakou, Jean-Pierre Baudoin, Irié Arsène Zoro Bi ...

  7. Phytochemicals and in vitro antioxidant potentials of defatted ...

    African Journals Online (AJOL)

    ONOS

    2010-01-18

    Jan 18, 2010 ... Plant-based dietary components and additives are known to protect cells from deleterious effect of reactive oxygen species (ROS). Proximate, phytochemical and antioxidant potentials of methanolic extract of defatted Holarrhena floribunda (G.Don) leaves were assessed using in vitro systems such as,.

  8. Evaluating in-vitro regeneration potential for high betacarotene ...

    African Journals Online (AJOL)

    In-vitro regeneration potential for three high beta-carotene (pro-vitamin A) cassava varieties - UMUCASS 36, UMUCASS 37 and UMUCASS 38, and a control variety TMS 60444 were evaluated and optimized as a preliminary step towards the introgression of more nutritional and agronomic traits. Somatic embryos ...

  9. The effect of exercise and beta2-adrenergic stimulation on glutathionylation and function of the Na,K-ATPase in human skeletal muscle

    DEFF Research Database (Denmark)

    Juel, Carsten; Hostrup, Morten; Bangsbo, Jens

    2015-01-01

    subunits are further glutathionylated by exercise and beta2-adrenergic stimulation. Our data suggest that glutathionylation contributes to the complex regulation of Na,K-ATPase function in human skeletal muscle. Glutathionylation of the Na,K-ATPase may explain reductions in maximal Na,K-ATPase activity......Potassium and sodium displacements across the skeletal muscle membrane during exercise may cause fatigue and are in part controlled by the Na,K-ATPase. Regulation of the Na,K-ATPase is therefore important for muscle functioning. We investigated the effect of oxidative stress (glutathionylation...

  10. In vitro dentine remineralization with a potential salivary phosphoprotein homologue.

    Science.gov (United States)

    Romero, Maria Jacinta Rosario H; Nakashima, Syozi; Nikaido, Toru; Sadr, Alireza; Tagami, Junji

    2016-08-01

    Advantages of introducing a salivary phosphoprotein homologue under standardized in vitro conditions to simulate the mineral-stabilizing properties of saliva have been proposed. This study longitudinally investigates the effects of casein, incorporated as a potential salivary phosphoprotein homologue in artificial saliva (AS) solutions with/without fluoride (F) on in vitro dentine lesion remineralization. Thin sections of bovine root dentine were demineralized and allocated randomly into 6 groups (n=18) having equivalent mineral loss (ΔZ) after transverse microradiography (TMR). The specimens were remineralized using AS solutions containing casein 0μg/ml, F 0ppm (C0-F0); casein 0μg/ml, F 1ppm (C0-F1); casein 10μg/ml, F 0ppm (C10-F0); casein 10μg/ml, F 1ppm (C10-F1); casein 100μg/ml, F 0ppm (C100-F0) or casein 100μg/ml, F 1ppm (C100-F1) for 28days with TMR taken every 7 days. Surface mineral precipitation, evident in group C0-F1, was apparently inhibited in groups with casein incorporation. Repeated measures ANOVA with Bonferroni correction revealed higher ΔZ for non-F and non-casein groups than for their counterparts (p<0.001). Subsequent multiple comparisons showed that mineral gain was higher (p<0.001) with 10μg/ml casein than with 100μg/ml when F was present in the earlier stages of remineralization, with both groups achieving almost complete remineralization after 28 days. Casein is a potential salivary phosphoprotein homologue that could be employed for in vitro dentine remineralization studies. Concentration related effects may be clinically significant and thus must be further examined. Copyright © 2016 Elsevier Ltd. All rights reserved.

  11. Aloe arborescens Polysaccharides: In Vitro Immunomodulation and Potential Cytotoxic Activity.

    Science.gov (United States)

    Nazeam, Jilan A; Gad, Haidy A; Esmat, Ahmed; El-Hefnawy, Hala M; Singab, Abdel-Naser B

    2017-05-01

    Different polysaccharides were isolated from the leaves of Aloe arborescens using the gradient power of hydrogen followed by antitumor and immunomodulatory assay. The total polysaccharide content of different fractions, water-soluble polysaccharide (WAP), acid-soluble polysaccharide (ACP), and alkaline-soluble polysaccharide (ALP), was estimated using a phenol-sulfuric acid spectrophotometric method. WAP possessed a higher content of mannose and glucose than either ACP or ALP. In vitro antitumor activity was investigated in three different cancer cell lines, and in vitro immunomodulatory potential was assessed through phagocytosis and lymphocyte transformation assay. The results showed that WAP and ALP exhibited the most significant cytotoxicity against HepG2 human liver cancer cells, with IC 50 values of 26.14 and 21.46 μg/mL, respectively. In contrast, ALP was able to enhance lymphocyte transformation, whereas WAP had the most potent phagocytic activity. Molecular weight, total sugar and uronic acid content, Fourier transform-infrared analysis, and linkage type of bioactive polysaccharides were investigated. These findings revealed that the potential antitumor activity of the natural agents WAP and ALP was through an immunomodulation mechanism, which verifies the use of the plant as adjuvant supplement for cancer patients suffering immunosuppression during chemotherapy.

  12. In vitro studies on the cytotoxic potential of surface sealants.

    Science.gov (United States)

    Zingler, S; Matthei, B; Kohl, A; Saure, D; Ludwig, B; Diercke, K; Lux, C J; Erber, R

    2015-01-01

    The objective of this in vitro study was an initial screening of the cytotoxic potential of widely used smooth enamel surface sealants. A total of 20 products were allocated to four groups based on their chemical composition: (1) filled resin-based sealants, (2) unfilled resin-based sealants, (3) a resin-modified, glass ionomer-based sealant, and (4) silicone-based sealants. All materials were applied to human enamel slices both in accordance with manufacturers' instructions and in additional experiments applying 50% undercuring and 50% overcuring. An agar overlay assay was then used to test the specimens following ISO 10933. The cytotoxic potential of each material was interpreted based on a reaction index that summarized the decolorization and lysis scores obtained. The cytotoxic potential decreased as follows: unfilled resin-based sealants > filled resin-based sealants > resin-modified, glass ionomer-based sealant > silicone-based sealants. In 75% of the resin-based products, deliberate undercuring was associated with more extensive decolorization zones, leading to higher rates of cytotoxic potential in two of those products. Overcuring, by contrast, was associated with a tendency for smaller decolorization zones in 50% of the resin-based products. Surface sealants derived from resin monomers exhibited cytotoxic potential in the agar overlay assay. There is also evidence of a possible association with curing, as undercuring can increase the cytotoxic potential, whereas normal curing (as per manufacturers' instructions) or overcuring may help minimize such effects. More research into the biological implications of these materials is needed, especially with regard to their potential impact on the adjacent gingiva.

  13. Potential in vitro antimicrobial efficacy of Holigarna arnottiana (Hook F).

    Science.gov (United States)

    Manilal, Aseer; Idhayadhulla, Akbar

    2014-01-01

    To explore the in vitro antimicrobial potential of Holigarna arnottiana (H. arnottiana) against human and shrimp pathogenic bacteria and use GC-MS analysis to elucidate its antimicrobial principles. In the present study, organic extract of H. arnottiana was examined for in vitro antimicrobial potency against five clinical human pathogens, seven species of human type culture pathogens, six pathogenic Vibrio strains isolated from moribund tiger shrimp (Penaeus monodon) and seven type cultures (Microbial Type Culture Collection, MTCC) of prominent shrimp pathogens. The extraction of H. arnottiana with ethyl acetate yielded bioactive crude extract that efficiently repressed the growth of all tested pathogens. Among the pathogens tested, shrimp pathogens were the most susceptible organisms while clinical pathogens were found to be a little resistant. The chemical constituents of the H. arnottiana were analysed by GC-MS which revealed the presence of major compounds such as 3,7,11,15-tetramethyl-2-hexadecen-1-o1 (42.1%), 1-lodo-2-methylundecane (34.5%) and squalene (11.1%) which might have a functional role in the chemical defence against microbial invasion. Based on the finding it could be inferred that H. arnottiana would be a reliable source for developing shrimp and human bio-therapeutics in future. Copyright © 2014 Asian Pacific Tropical Biomedical Magazine. Published by Elsevier B.V. All rights reserved.

  14. Evaluation of In Vitro Antioxidant Potential of Cordia retusa.

    Science.gov (United States)

    Amudha, Murugesan; Rani, Shanmugam

    2016-01-01

    The present study was carried out to investigate the antioxidant potential, total flavonoid and phenolic content in extracts of aerial parts of Cordia retua (Vahl.) Masam. The samples such as ethyl acetate and ethanol extracts were tested using six in vitro models such as 2,2-diphenyl-1-picrylhydrazyl, nitric oxide radical, iron chelating, hydroxyl radical, superoxide radical scavenging activity and total antioxidant activity to evaluate the in vitro antioxidant potential of C. retusa by spectrophotometrically. Total flavonoid and phenolic content in samples were estimated using aluminum chloride colorimetric and Folin-Ciocalteu method. The results were analyzed statistically by the regression method. Half maximal inhibitory concentration (IC50) of the ethanol extract was found to be 596 μg/ml for DPPH, 597 μg/ml for nitric oxide radical, 554 μg/ml for iron chelating, 580 μg/ml for hydroxyl radical, 562 μg/ml for superoxide radical and 566 μg/ml for total antioxidant capacity. Furthermore, the total flavonoid content and total phenolic content of the ethanol extract were found to be 2.71 mg gallic acid equivalent per gram of extract and 1.86 mg quercetin equivalent per gram of extract, respectively. In all the testing, a significant correlation existed between concentrations of the extract and percentage inhibition of free radicals. The results of the present comprehensive analysis demonstrated that C. retusa possess potent antioxidant activity, high flavonoid and phenolic content. The antioxidant property may be related to the polyphenols and flavonoids present in the extract. These results clearly indicated that C. retusa is effective against free radical mediated diseases as a natural antioxidant.

  15. Glutathionylation of Yersinia pestis LcrV and Its Effects on Plague Pathogenesis.

    Science.gov (United States)

    Mitchell, Anthony; Tam, Christina; Elli, Derek; Charlton, Thomas; Osei-Owusu, Patrick; Fazlollahi, Farbod; Faull, Kym F; Schneewind, Olaf

    2017-05-16

    Glutathionylation, the formation of reversible mixed disulfides between glutathione and protein cysteine residues, is a posttranslational modification previously observed for intracellular proteins of bacteria. Here we show that Yersinia pestis LcrV, a secreted protein capping the type III secretion machine, is glutathionylated at Cys273 and that this modification promotes association with host ribosomal protein S3 (RPS3), moderates Y. pestis type III effector transport and killing of macrophages, and enhances bubonic plague pathogenesis in mice and rats. Secreted LcrV was purified and analyzed by mass spectrometry to reveal glutathionylation, a modification that is abolished by the codon substitution Cys273Ala in lcrV Moreover, the lcrVC273A mutation enhanced the survival of animals in models of bubonic plague. Investigating the molecular mechanism responsible for these virulence attributes, we identified macrophage RPS3 as a ligand of LcrV, an association that is perturbed by the Cys273Ala substitution. Furthermore, macrophages infected by the lcrVC273A variant displayed accelerated apoptotic death and diminished proinflammatory cytokine release. Deletion of gshB, which encodes glutathione synthetase of Y. pestis, resulted in undetectable levels of intracellular glutathione, and we used a Y. pestis ΔgshB mutant to characterize the biochemical pathway of LcrV glutathionylation, establishing that LcrV is modified after its transport to the type III needle via disulfide bond formation with extracellular oxidized glutathione.IMPORTANCEYersinia pestis, the causative agent of plague, has killed large segments of the human population; however, the molecular bases for the extraordinary virulence attributes of this pathogen are not well understood. We show here that LcrV, the cap protein of bacterial type III secretion needles, is modified by host glutathione and that this modification contributes to the high virulence of Y. pestis in mouse and rat models for bubonic

  16. Quantitation of protein S-glutathionylation by liquid chromatograph-tandem mass spectrometry: Correction for contaminating glutathione and glutathione disulfide

    Science.gov (United States)

    Protein S-glutathionylation is a posttranslational modification that links oxidative stimuli to reversible changes in cellular function. Protein-glutathione mixed disulfides (PSSG) are commonly quantified by the reduction of the disulfide and detection of the resultant glutathione species. This met...

  17. Disulfiram attenuates osteoclast differentiation in vitro: a potential antiresorptive agent.

    Directory of Open Access Journals (Sweden)

    Hua Ying

    Full Text Available Disulfiram (DSF, a cysteine modifying compound, has long been clinically employed for the treatment of alcohol addiction. Mechanistically, DSF acts as a modulator of MAPK and NF-κB pathways signaling pathways. While these pathways are crucial for osteoclast (OC differentiation, the potential influence of DSF on OC formation and function has not been directly assessed. Here, we explore the pharmacological effects of DSF on OC differentiation, activity and the modulation of osteoclastogenic signaling cascades. We first analyzed cytotoxicity of DSF on bone marrow monocytes isolated from C57BL/6J mice. Upon the establishment of optimal dosage, we conducted osteoclastogenesis and bone resorption assays in the presence or absence of DSF treatment. Luciferase assays in RAW264.7 cells were used to examine the effects of DSF on major transcription factors activation. Western blot, reverse transcription polymerase chain reaction, intracellular acidification and proton influx assays were employed to further dissect the underlying mechanism. DSF treatment dose-dependently inhibited both mouse and human osteoclastogenesis, especially at early stages of differentiation. This inhibition correlated with a decrease in the expression of key osteoclastic marker genes including CtsK, TRAP, DC-STAMP and Atp6v0d2 as well as a reduction in bone resorption in vitro. Suppression of OC differentiation was found to be due, at least in part, to the blockade of several key receptor activators of nuclear factor kappa-B ligand (RANKL-signaling pathways including ERK, NF-κB and NFATc1. On the other hand, DSF failed to suppress intracellular acidification and proton influx in mouse and human osteoclasts using acridine orange quenching and microsome-based proton transport assays. Our findings indicate that DSF attenuates OC differentiation via the collective suppression of several key RANKL-mediated signaling cascades, thus making it an attractive agent for the treatment of OC

  18. An in vitro investigation of the erosive potential of smoothies.

    Science.gov (United States)

    Blacker, S M; Chadwick, R G

    2013-02-01

    Recent health promotion campaigns have encouraged the public to consume at least five portions of fruit and vegetables per day. Many see consuming fruit smoothies as a way of achieving this. To ascertain the potential or otherwise for fruit smoothies to bring about dental erosion. Laboratory study.Method This was an in vitro investigation in which five varieties of shop bought fruit smoothies, including a 'thickie' were investigated, with respect to their initial pH, titratable acidity and effect upon exposure to the surface microhardness and profile of extracted human teeth. In addition their performance was compared to negative (Volvic(™) water) and positive (orange juice) control drinks as well as a homemade smoothie, based upon the recipe of one of the commercially bought drinks, from which ingredient omissions were made. The majority of the drinks investigated had a baseline pH below the critical pH of enamel (5.5) and required comparable volumes of 0.1M NaOH to raise their pH to neutrality as the positive control. Only two drinks (Volvic(™) still mineral water, the negative control, and the yoghurt, vanilla bean and honey 'thickie') displayed a higher pH, though to neutralise the thickie, a lesser quantity of alkali addition was required. The immersion of the tooth samples in the drinks brought about reductions in their surface hardness (expressed as a percentage change of median hardness) but these were only significant (p benefits of their consumption, their consumption should be confined to mealtimes.

  19. Studies on in vitro evaluation of antidiabetic potentials of ...

    African Journals Online (AJOL)

    At present, the prevalence of Diabetes has increased worldwide and predicted to increase to greater extent in future generations. Among various therapeutic approaches implemented to prevent diabetes is to regulate the blood glucose levels by various mechanisms. This is being assessed by in vitro antidiabetic assays ...

  20. In – Vitro Propagation and Antimycotic Potential of Extracts and ...

    African Journals Online (AJOL)

    In spite of the therapeutic importance of Aristolochia bracteolata Linn. in Nigerian ethnomedicine, it is largely collected from the wild. Owing to the acclaimed potency of the plant and the difficulty in treating candidiasis, the anticandidal activity and in vitro propagation of the plant were investigated. Phytochemical screening ...

  1. In vitro antimicrobial potential of organic solvent extracts of novel ...

    African Journals Online (AJOL)

    SAM

    In vitro screening of antibacterial and antifungal activities of hexane, chloroform, ethyl acetate, methanol and water extracts of selected promising actinomycetes strains were studied towards Gram- positive, Gram-negative bacteria, dermatophytes and opportunistic pathogens. Crude antimicrobial metabolites were extracted ...

  2. In vitro antimicrobial potential of organic solvent extracts of novel ...

    African Journals Online (AJOL)

    In vitro screening of antibacterial and antifungal activities of hexane, chloroform, ethyl acetate, methanol and water extracts of selected promising actinomycetes strains were studied towards Gram-positive, Gram-negative bacteria, dermatophytes and opportunistic pathogens. Crude antimicrobial metabolites were extracted ...

  3. In vitro interactions between Armillaria species and potential biocontrol fungi

    Directory of Open Access Journals (Sweden)

    Keča Nenad

    2009-01-01

    Full Text Available Interaction between Armillaria species and seven other fungi were tested in vitro. Tree antagonistic (Trichoderma viride, Trichotecium roseum and Penicillium sp. and four decaying (Hypholoma fasciculare¸ Hypholoma capnoides, Phlebiopsis gigantea, and Pleurotus ostreatus fungi were chosen for this study. The best results were noted for Trichoderma viride, because fungus was able to kill both mycelia and rhizomorphs of Armillaria species, while Hypholoma spp. inhibited both growth of Armillaria colonies and rhizomorph production.

  4. In vitro investigation of Debaryomyces hansenii strains for potential probiotic properties

    DEFF Research Database (Denmark)

    Ochangco, Honeylet Sabas; Gamero, Amparo; Smith, Ida Mosbech

    2016-01-01

    In this study, 23 Debaryomyces hansenii strains, isolated from cheese and fish gut, were investigated in vitro for potential probiotic properties i.e. (1) survival under in vitro GI (gastrointestinal) conditions with different oxygen levels, (2) adhesion to Caco-2 intestinal epithelial cells and ...

  5. Protein thiol oxidation and formation of S-glutathionylated cyclophilin A in cells exposed to chloramines and hypochlorous acid.

    Science.gov (United States)

    Stacey, Melissa M; Cuddihy, Sarah L; Hampton, Mark B; Winterbourn, Christine C

    2012-11-01

    Neutrophil oxidants, including the myeloperoxidase products, HOCl and chloramines, have been linked to endothelial dysfunction in inflammatory diseases such as atherosclerosis. As they react preferentially with sulfur centers, thiol proteins are likely to be cellular targets. Our objectives were to establish whether there is selective protein oxidation in vascular endothelial cells treated with HOCl or chloramines, and to identify sensitive proteins. Cells were treated with HOCl, glycine chloramine and monochloramine, reversibly oxidized cysteines were labeled and separated by 1D or 2D SDS-PAGE, and proteins were characterized by mass spectrometry. Selective protein oxidation was observed, with chloramines and HOCl causing more changes than H(2)O(2). Cyclophilin A was one of the most sensitive targets, particularly with glycine chloramine. Cyclophilin A was also oxidized in Jurkat T cells where its identity was confirmed using a knockout cell line. The product was a mixed disulfide with glutathione, with glutathionylation at Cys-161. Glyceraldehyde-3-phosphate dehydrogenase, peroxiredoxins and cofilin were also highly sensitive to HOCl/chloramines. Cyclophilins are becoming recognized as redox regulatory proteins, and glutathionylation is an important mechanism for redox regulation. Cells lacking Cyclophilin A showed more glutathionylation of other proteins than wild-type cells, suggesting that cyclophilin-regulated deglutathionylation could contribute to redox changes in HOCl/chloramine-exposed cells. Copyright © 2012 Elsevier Inc. All rights reserved.

  6. Glutathionylation of the L-type Ca2+ Channel in Oxidative Stress-Induced Pathology of the Heart

    Directory of Open Access Journals (Sweden)

    Victoria P. A. Johnstone

    2014-10-01

    Full Text Available There is mounting evidence to suggest that protein glutathionylation is a key process contributing to the development of pathology. Glutathionylation occurs as a result of posttranslational modification of a protein and involves the addition of a glutathione moiety at cysteine residues. Such modification can occur on a number of proteins, and exerts a variety of functional consequences. The L-type Ca2+ channel has been identified as a glutathionylation target that participates in the development of cardiac pathology. Ca2+ influx via the L-type Ca2+ channel increases production of mitochondrial reactive oxygen species (ROS in cardiomyocytes during periods of oxidative stress. This induces a persistent increase in channel open probability, and the resulting constitutive increase in Ca2+ influx amplifies the cross-talk between the mitochondria and the channel. Novel strategies utilising targeted peptide delivery to uncouple mitochondrial ROS and Ca2+ flux via the L-type Ca2+ channel following ischemia-reperfusion have delivered promising results, and have proven capable of restoring appropriate mitochondrial function in myocytes and in vivo.

  7. In vitro Studies on Anti-diabetic and Anti-ulcer Potentials of Jatropha ...

    African Journals Online (AJOL)

    In vitro Studies on Anti-diabetic and Anti-ulcer Potentials of Jatropha gossypifolia (Euphorbiaceae). Hammad Saleem, Irshad Ahmad, M Ashraf, M Shoaib Ali Gill, Muhammad Faisal Nadeem, M Nabeel Shahid, Kashif Barkat ...

  8. S-glutathionylation of glyceraldehyde-3-phosphate dehydrogenase induces formation of C150-C154 intrasubunit disulfide bond in the active site of the enzyme.

    Science.gov (United States)

    Barinova, K V; Serebryakova, M V; Muronetz, V I; Schmalhausen, E V

    2017-12-01

    Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a glycolytic protein involved in numerous non-glycolytic functions. S-glutathionylated GAPDH was revealed in plant and animal tissues. The role of GAPDH S-glutathionylation is not fully understood. Rabbit muscle GAPDH was S-glutathionylated in the presence of H 2 O 2 and reduced glutathione (GSH). The modified protein was assayed by MALDI-MS analysis, differential scanning calorimetry, dynamic light scattering, and ultracentrifugation. Incubation of GAPDH in the presence of H 2 O 2 together with GSH resulted in the complete inactivation of the enzyme. In contrast to irreversible oxidation of GAPDH by H 2 O 2 , this modification could be reversed in the excess of GSH or dithiothreitol. By data of MALDI-MS analysis, the modified protein contained both mixed disulfide between Cys150 and GSH and the intrasubunit disulfide bond between Cys150 and Cys154 (different subunits of tetrameric GAPDH may contain different products). S-glutathionylation results in loosening of the tertiary structure of GAPDH, decreases its affinity to NAD + and thermal stability. The mixed disulfide between Cys150 and GSH is an intermediate product of S-glutathionylation: its subsequent reaction with Cys154 results in the intrasubunit disulfide bond in the active site of GAPDH. The mixed disulfide and the C150-C154 disulfide bond protect GAPDH from irreversible oxidation and can be reduced in the excess of thiols. Conformational changes that were observed in S-glutathionylated GAPDH may affect interactions between GAPDH and other proteins (ligands), suggesting the role of S-glutathionylation in the redox signaling. The manuscript considers one of the possible mechanisms of redox regulation of cell functions. Copyright © 2017 Elsevier B.V. All rights reserved.

  9. In vitro antiplasmodial and antimicrobial potential of Tagetes erecta roots.

    Science.gov (United States)

    Gupta, Pankaj; Vasudeva, Neeru

    2010-11-01

    Among strategies to combat malaria, the search for newer antimalarial compounds is a priority. Traditionally, Tagetes erecta Linn. (Compositae) has been used for the treatment of various diseases and ailments including malaria. Five successive extracts (petroleum ether, chloroform, ethyl acetate, methanol and aqueous) of the roots of T. erecta and a new bithienyl compound, 2-hydroxymethyl-non-3-ynoic acid 2-[2,2']-bithiophenyl-5-ethyl ester from the roots of the plant, were evaluated for in vitro antiplasmodial activity against chloroquine sensitive and resistant strains of Plasmodium falciparum. The extracts were also tested for in vitro antimicrobial activity against seven microbial strains. The antiplasmodial screening was carried out using the schizont maturation inhibition assay. Preliminary antimicrobial screening was carried out using the agar well assay followed by determination of minimum inhibitory concentration (MIC) using two-fold serial dilutions. Among all the extracts tested, the ethyl acetate fraction exhibited significant antiplasmodial efficacy with the 50% inhibitory concentrations (IC(50)) of 0.02 and 0.07 mg/mL against the chloroquine sensitive and resistant strains of Plasmodium falciparum respectively. The new bithienyl compound also showed significant schizonticidal activity against both chloroquine sensitive and resistant strains of Plasmodium falciparum with the IC(50) values of 0.01 and 0.02 mg/mL. Additionally, all extracts exhibited significant antimicrobial activity against three Gram-positive and two Gram-negative bacterial and two fungal strains with MIC values ranging between 12.5-100 µg/mL. The new bithienyl compound was profoundly able to arrest the ring stages of the malarial parasites thereby exerting its antiplasmodial effect. The observations provide support for the ethnobotanical use of the plant.

  10. In vitro anticandidal and antioxidant potential of Mezoneuron benthamianum

    Directory of Open Access Journals (Sweden)

    Scott O Fayemi

    2012-01-01

    Conclusions: The result from this research suggests that M. benthamianum is a potent anti-candida and antioxidant plant and may be a potential anti-candida plant for future prospect in drug development.

  11. 20S proteasome activity is modified via S-glutathionylation based on intracellular redox status of the yeast Saccharomyces cerevisiae: implications for the degradation of oxidized proteins.

    Science.gov (United States)

    Demasi, Marilene; Hand, Adrian; Ohara, Erina; Oliveira, Cristiano L P; Bicev, Renata N; Bertoncini, Clelia A; Netto, Luis E S

    2014-09-01

    Protein S-glutathionylation is a post-translational modification that controls many cellular pathways. Recently, we demonstrated that the α5-subunit of the 20S proteasome is S-glutathionylated in yeast cells grown to the stationary phase in rich medium containing glucose, stimulating 20S core gate opening and increasing the degradation of oxidized proteins. In the present study, we evaluated the correlation between proteasomal S-glutathionylation and the intracellular redox status. The redox status was controlled by growing yeast cells in distinct carbon sources which induced respiratory (glycerol/ethanol) or fermentative (glucose) metabolism. Cells grown under glycerol/ethanol displayed higher reductive power when compared to cells grown under glucose. When purified from cells grown in glucose, 20S proteasome α5-subunit exhibited an intense anti-glutathione labeling. A higher frequency of the open catalytic chamber gate was observed in the S-glutathionylated preparations as demonstrated by transmission electron microscopy. Therefore, cells that had been grown in glucose displayed an increased ability to degrade oxidized proteins. The results of the present study suggest that 20S proteasomal S-glutathionylation is a relevant adaptive response to oxidative stress that is capable to sense the intracellular redox environment, leading to the removal of oxidized proteins via a process that is not dependent upon ubiquitylation and ATP consumption. Copyright © 2014 Elsevier Inc. All rights reserved.

  12. Phytochemicals and in vitro antioxidant potentials of defatted ...

    African Journals Online (AJOL)

    ONOS

    2010-01-18

    Jan 18, 2010 ... Antioxidant potential of Ecklonia cavaon reactive oxygen species scavenging, metal chelating, reducing power and lipid peroxidation inhibition. Food Sci. Technol Int. 12: 27-38. Marcocci l, Maguire JJ, Droy-Lefai MT, Parker L (1994). Antioxidant action of ginkgo biloba extracts EGb761. Biochem. Biosphys.

  13. In vitro assessment of the antimicrobial potential of honey on ...

    African Journals Online (AJOL)

    Background: Honey produced by honeybees (Apis mellifera) is one of the ancient traditional medicines used for treatment and prevention of various illnesses. Objective: To assess the antimicrobial potential of honey on some common bacterial pathogen. Methods: This experimental study was conducted in Jimma University ...

  14. studies on in vitro evaluation of antidiabetic potentials of ...

    African Journals Online (AJOL)

    userpc

    Hence the current work focuses on investigating the anti-diabetic activity of fruit peel of Citrullus lanatus (watermelon) and Punica granatum. (pomegranate) using the above mentioned assays. Our results suggested that methanol extract of Citrullus lanatus and Punica granatum peels showed potential anti-diabetic activity.

  15. In vitro control of Alternaria citri using antifungal potentials of ...

    African Journals Online (AJOL)

    msc06-08

    2012-05-24

    May 24, 2012 ... significantly reduced the fungal biomass of the target fungal pathogen. Generally, 100% culture filtrate of the test ... be used to prevent, mitigate or control plant diseases. (Chandler et al., 2008). Overdose usage of ... bacteria are potential biocontrol agents (Harman 2000;. Harman et al., 2004). Fungal and ...

  16. Phytochemical screening and In vitro antioxidant potentials of ...

    African Journals Online (AJOL)

    Background: Over the last decade, extensive research work has focused on the potential health benefits of antioxidants while many medicinal plant extracts have been evaluated for their antioxidant profile. Medicinal plants selected for this study are widely used in traditional medicine for the treatment of diabetes mellitus in ...

  17. In vitro antidiabetic potential of the fruits of Crataegus pinnatifida

    Science.gov (United States)

    Chowdhury, S.S.; Islam, M.N.; Jung, H.A.; Choi, J.S.

    2014-01-01

    In an attempt to develop alternative medicine for the treatment of diabetes and related complications, the antidiabetic potential of the fruits of Crataegus pinnatifida was evaluated. The antidiabetic potential of the methanol (MeOH) extract as well as different solvent soluble fractions of the fruits of C. pinnatifida was evaluated via α-glucosidase, protein tyrosine phosphatase 1B (PTP1B), rat lens aldose reductase (RLAR), and advanced glycation end products (AGEs) formation inhibitory assays. The MeOH extract showed potent inhibitory activity against α-glucosidase, PTP1B, and AGEs formation with IC50 values of 122.11, 3.66 and 65.83 μg/ml respectively, while it showed moderate inhibitory activity against RLAR with the IC50 value of 160.54 μg/ml. Among different fractions, the ethyl acetate (EtOAc) and the dichloromethane (CH2Cl2) fractions were found as active fractions exhibiting potential α-glucosidase, PTP1B, RLAR inhibitory, and AGEs formation inhibitory activities. Seven compounds including hyperoside, chlorogenic acid, ursolic acid, oleanolic acid, 3-epicorosolic acid, β-sitosterol, β-sitosterol glucoside were isolated from these two fractions. 3-Epicorosolic acid showed both potent α-glucosidase and PTP1B inhibitory activities with IC50 values of 30.18 and 4.08 μg/ml respectively. Moreover, kinetic study revealed that 3-epicorosolic acid showed mixed type inhibition against PTP1B, while it showed uncompetitive inhibition against α-glucosidase. Therefore, these results suggest that the fruits of C. pinnatifida and its constituents have potential antidiabetic activity which might be used as a functional food for the treatment of diabetes and associated complications. PMID:25598795

  18. Attitudes to in vitro meat: A survey of potential consumers in the United States

    OpenAIRE

    Wilks, Matti; Clive J C Phillips

    2017-01-01

    Positivity towards meat consumption remains strong, despite evidence of negative environmental and ethical outcomes. Although awareness of these repercussions is rising, there is still public resistance to removing meat from our diets. One potential method to alleviate these effects is to produce in vitro meat: meat grown in a laboratory that does not carry the same environmental or ethical concerns. However, there is limited research examining public attitudes towards in vitro meat, thus we ...

  19. Glutathione S-Transferase P-Mediated Protein S-Glutathionylation of Resident Endoplasmic Reticulum Proteins Influences Sensitivity to Drug-Induced Unfolded Protein Response.

    Science.gov (United States)

    Ye, Zhi-Wei; Zhang, Jie; Ancrum, Tiffany; Manevich, Yefim; Townsend, Danyelle M; Tew, Kenneth D

    2017-02-20

    S-glutathionylation of cysteine residues, catalyzed by glutathione S-transferase Pi (GSTP), alters structure/function characteristics of certain targeted proteins. Our goal is to characterize how S-glutathionylation of proteins within the endoplasmic reticulum (ER) impact cell sensitivity to ER-stress inducing drugs. We identify GSTP to be an ER-resident protein where it demonstrates both chaperone and catalytic functions. Redox based proteomic analyses identified a cluster of proteins cooperatively involved in the regulation of ER stress (immunoglobulin heavy chain-binding protein [BiP], protein disulfide isomerase [PDI], calnexin, calreticulin, endoplasmin, sarco/endoplasmic reticulum Ca2+-ATPase [SERCA]) that individually co-immunoprecipitated with GSTP (implying protein complex formation) and were subject to reactive oxygen species (ROS) induced S-glutathionylation. S-glutathionylation of each of these six proteins was attenuated in cells (liver, embryo fibroblasts or bone marrow dendritic) from mice lacking GSTP (Gstp1/p2-/-) compared to wild type (Gstp1/p2+/+). Moreover, Gstp1/p2-/- cells were significantly more sensitive to the cytotoxic effects of the ER-stress inducing drugs, thapsigargin (7-fold) and tunicamycin (2-fold). Within the family of GST isozymes, GSTP has been ascribed the broadest range of catalytic and chaperone functions. Now, for the first time, we identify it as an ER resident protein that catalyzes S-glutathionylation of critical ER proteins within this organelle. Of note, this can provide a nexus for linkage of redox based signaling and pathways that regulate the unfolded protein response (UPR). This has novel importance in determining how some drugs kill cancer cells. Contextually, these results provide mechanistic evidence that GSTP can exert redox regulation in the oxidative ER environment and indicate that, within the ER, GSTP influences the cellular consequences of the UPR through S-glutathionylation of a series of key interrelated

  20. Proteomic analysis of S-nitrosylated and S-glutathionylated proteins in wheat seedlings with different dehydration tolerances.

    Science.gov (United States)

    Gietler, Marta; Nykiel, Małgorzata; Orzechowski, Sławomir; Fettke, Joerg; Zagdańska, Barbara

    2016-11-01

    A loss of dehydration tolerance in wheat seedlings on the fifth day following imbibition is associated with a disturbance in cellular redox homeostasis, as documented by a shift of the reduced/oxidized glutathione ratio to a more oxidized state and a significant increase in the ratio of protein thiols to the total thiol group content. Therefore, the identification and characterization of redox-sensitive proteins are important steps toward understanding the molecular mechanisms of the loss of dehydration tolerance. In the present study, proteins that were differentially expressed between fully turgid (control), dehydrated tolerant (four-day-old) and dehydrated sensitive (six-day-old) wheat seedlings were analysed. Protein spots having at least a significant (p < 0.05) two-fold change in protein abundance were selected by Delta2D as differentially expressed, identified by MALDI-TOF and LC-MS/MS, and classified according to their function. The observed changes in the proteomic patterns of the differentially S-nitrosylated and S-glutathionylated proteins were highly specific in dehydration-tolerant and -sensitive wheat seedlings. The metabolic function of these proteins indicates that dehydration tolerance is mainly related to nucleic acids, protein metabolism, and energy metabolism. It has been proven that leaf-specific thionins BTH6 and DB4, chloroplastic 50S ribosomal protein L16, phospholipase A1-II delta, and chloroplastic thioredoxin M2 are both S-nitrosylated and S-glutathionylated upon water deficiency. Our results revealed the existence of interplay between S-nitrosylation and S-glutathionylation, two redox-regulated protein posttranslational modifications that could enhance plant defence mechanisms and/or facilitate the acclimation of plants to unfavourable environmental conditions. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  1. Attitudes to in vitro meat: A survey of potential consumers in the United States.

    Directory of Open Access Journals (Sweden)

    Matti Wilks

    Full Text Available Positivity towards meat consumption remains strong, despite evidence of negative environmental and ethical outcomes. Although awareness of these repercussions is rising, there is still public resistance to removing meat from our diets. One potential method to alleviate these effects is to produce in vitro meat: meat grown in a laboratory that does not carry the same environmental or ethical concerns. However, there is limited research examining public attitudes towards in vitro meat, thus we know little about the capacity for it be accepted by consumers. This study aimed to examine perceptions of in vitro meat and identify potential barriers that might prevent engagement. Through conducting an online survey with US participants, we identified that although most respondents were willing to try in vitro meat, only one third were definitely or probably willing to eat in vitro meat regularly or as a replacement for farmed meat. Men were more receptive to it than women, as were politically liberal respondents compared with conservative ones. Vegetarians and vegans were more likely to perceive benefits compared to farmed meat, but they were less likely to want to try it than meat eaters. The main concerns were an anticipated high price, limited taste and appeal and a concern that the product was unnatural. It is concluded that people in the USA are likely to try in vitro meat, but few believed that it would replace farmed meat in their diet.

  2. Attitudes to in vitro meat: A survey of potential consumers in the United States.

    Science.gov (United States)

    Wilks, Matti; Phillips, Clive J C

    2017-01-01

    Positivity towards meat consumption remains strong, despite evidence of negative environmental and ethical outcomes. Although awareness of these repercussions is rising, there is still public resistance to removing meat from our diets. One potential method to alleviate these effects is to produce in vitro meat: meat grown in a laboratory that does not carry the same environmental or ethical concerns. However, there is limited research examining public attitudes towards in vitro meat, thus we know little about the capacity for it be accepted by consumers. This study aimed to examine perceptions of in vitro meat and identify potential barriers that might prevent engagement. Through conducting an online survey with US participants, we identified that although most respondents were willing to try in vitro meat, only one third were definitely or probably willing to eat in vitro meat regularly or as a replacement for farmed meat. Men were more receptive to it than women, as were politically liberal respondents compared with conservative ones. Vegetarians and vegans were more likely to perceive benefits compared to farmed meat, but they were less likely to want to try it than meat eaters. The main concerns were an anticipated high price, limited taste and appeal and a concern that the product was unnatural. It is concluded that people in the USA are likely to try in vitro meat, but few believed that it would replace farmed meat in their diet.

  3. The odontogenic potential of STRO-1 sorted rat dental pulp stem cells in vitro.

    NARCIS (Netherlands)

    Yang, X.; Dolder, J. van den; Walboomers, X.F.; Zhang, W.; Bian, Z.; Fan, M.; Jansen, J.A.

    2007-01-01

    The presence of heterogeneous cell populations in dental pulp may count for the considerable variation in the outcome of in vitro and in vivo experiments. Here, we intended to determine whether a minor cell sub-population of high proliferation and odontogenic potential existed among a larger

  4. Regulation of cell physiology and pathology by protein S-glutathionylation: lessons learned from the cardiovascular system.

    Science.gov (United States)

    Pimentel, David; Haeussler, Dagmar Johanna; Matsui, Reiko; Burgoyne, Joseph Robert; Cohen, Richard Alan; Bachschmid, Markus Michael

    2012-03-15

    Reactive oxygen and nitrogen species contributing to homeostatic regulation and the pathogenesis of various cardiovascular diseases, including atherosclerosis, hypertension, endothelial dysfunction, and cardiac hypertrophy, is well established. The ability of oxidant species to mediate such effects is in part dependent on their ability to induce specific modifications on particular amino acids, which alter protein function leading to changes in cell signaling and function. The thiol containing amino acids, methionine and cysteine, are the only oxidized amino acids that undergo reduction by cellular enzymes and are, therefore, prime candidates in regulating physiological signaling. Various reports illustrate the significance of reversible oxidative modifications on cysteine thiols and their importance in modulating cardiovascular function and physiology. The use of mass spectrometry, novel labeling techniques, and live cell imaging illustrate the emerging importance of reversible thiol modifications in cellular redox signaling and have advanced our analytical abilities. Distinguishing redox signaling from oxidative stress remains unclear. S-nitrosylation as a precursor of S-glutathionylation is controversial and needs further clarification. Subcellular distribution of glutathione (GSH) may play an important role in local regulation, and targeted tools need to be developed. Furthermore, cellular redundancies of thiol metabolism complicate analysis and interpretation. The development of novel pharmacological analogs that specifically target subcellular compartments of GSH to promote or prevent local protein S-glutathionylation as well as the establishment of conditional gene ablation and transgenic animal models are needed.

  5. The Role of S-Nitrosylation and S-Glutathionylation of Protein Disulphide Isomerase in Protein Misfolding and Neurodegeneration

    Directory of Open Access Journals (Sweden)

    M. Halloran

    2013-01-01

    Full Text Available Neurodegenerative diseases involve the progressive loss of neurons, and a pathological hallmark is the presence of abnormal inclusions containing misfolded proteins. Although the precise molecular mechanisms triggering neurodegeneration remain unclear, endoplasmic reticulum (ER stress, elevated oxidative and nitrosative stress, and protein misfolding are important features in pathogenesis. Protein disulphide isomerase (PDI is the prototype of a family of molecular chaperones and foldases upregulated during ER stress that are increasingly implicated in neurodegenerative diseases. PDI catalyzes the rearrangement and formation of disulphide bonds, thus facilitating protein folding, and in neurodegeneration may act to ameliorate the burden of protein misfolding. However, an aberrant posttranslational modification of PDI, S-nitrosylation, inhibits its protective function in these conditions. S-nitrosylation is a redox-mediated modification that regulates protein function by covalent addition of nitric oxide- (NO- containing groups to cysteine residues. Here, we discuss the evidence for abnormal S-nitrosylation of PDI (SNO-PDI in neurodegeneration and how this may be linked to another aberrant modification of PDI, S-glutathionylation. Understanding the role of aberrant S-nitrosylation/S-glutathionylation of PDI in the pathogenesis of neurodegenerative diseases may provide insights into novel therapeutic interventions in the future.

  6. In vitro fertilizing potential of urethral and epididymal spermatozoa collected from domestic cats (Felis catus).

    Science.gov (United States)

    Prochowska, S; Niżański, W

    2017-03-28

    The aim of this study was to provide a comparative analysis of in vitro fertilizing potential of frozen-thawed urethral and epididymal feline spermatozoa. Both types of semen were collected from 7 cats and cryopreserved in liquid nitrogen. To perform in vitro fertilization, both urethral and epididymal samples from the same individual were thawed and spermatozoa were co-incubated with in vitro matured cat oocytes. Obtained embryos were cultured in vitro for 7 days in a commercial medium. Cleavage rate, morula rate and blastocyst rate were calculated. Experiment was run in 10 replicates. The examined parameters showed no significant differences between urethral and epididymal spermatozoa (p>0.05). Cleavage rate and embryo's development were highly variable between replicates, even for the different sperm samples collected from one individual. There was no significant correlation between fertilizing capacity of two types of spermatozoa collected from the same male. In this study we confirmed that cryopreserved urethral spermatozoa have equally good fertilizing potential as epididymal ones, and both can be successfully used for in vitro fertilization in cats with the use of commercial medium.

  7. Expert consensus on an in vitro approach to assess pulmonary fibrogenic potential of aerosolized nanomaterials.

    Science.gov (United States)

    Clippinger, Amy J; Ahluwalia, Arti; Allen, David; Bonner, James C; Casey, Warren; Castranova, Vincent; David, Raymond M; Halappanavar, Sabina; Hotchkiss, Jon A; Jarabek, Annie M; Maier, Monika; Polk, William; Rothen-Rutishauser, Barbara; Sayes, Christie M; Sayre, Phil; Sharma, Monita; Stone, Vicki

    2016-07-01

    The increasing use of multi-walled carbon nanotubes (MWCNTs) in consumer products and their potential to induce adverse lung effects following inhalation has lead to much interest in better understanding the hazard associated with these nanomaterials (NMs). While the current regulatory requirement for substances of concern, such as MWCNTs, in many jurisdictions is a 90-day rodent inhalation test, the monetary, ethical, and scientific concerns associated with this test led an international expert group to convene in Washington, DC, USA, to discuss alternative approaches to evaluate the inhalation toxicity of MWCNTs. Pulmonary fibrosis was identified as a key adverse outcome linked to MWCNT exposure, and recommendations were made on the design of an in vitro assay that is predictive of the fibrotic potential of MWCNTs. While fibrosis takes weeks or months to develop in vivo, an in vitro test system may more rapidly predict fibrogenic potential by monitoring pro-fibrotic mediators (e.g., cytokines and growth factors). Therefore, the workshop discussions focused on the necessary specifications related to the development and evaluation of such an in vitro system. Recommendations were made for designing a system using lung-relevant cells co-cultured at the air-liquid interface to assess the pro-fibrogenic potential of aerosolized MWCNTs, while considering human-relevant dosimetry and NM life cycle transformations. The workshop discussions provided the fundamental design components of an air-liquid interface in vitro test system that will be subsequently expanded to the development of an alternative testing strategy to predict pulmonary toxicity and to generate data that will enable effective risk assessment of NMs.

  8. S-Glutathionylation of troponin I (fast) increases contractile apparatus Ca2+ sensitivity in fast-twitch muscle fibres of rats and humans

    Science.gov (United States)

    Mollica, J P; Dutka, T L; Merry, T L; Lamboley, C R; McConell, G K; McKenna, M J; Murphy, R M; Lamb, G D

    2012-01-01

    Oxidation can decrease or increase the Ca2+ sensitivity of the contractile apparatus in rodent fast-twitch (type II) skeletal muscle fibres, but the reactions and molecular targets involved are unknown. This study examined whether increased Ca2+ sensitivity is due to S-glutathionylation of particular cysteine residues. Skinned muscle fibres were directly activated in heavily buffered Ca2+ solutions to assess contractile apparatus Ca2+ sensitivity. Rat type II fibres were subjected to S-glutathionylation by successive treatments with 2,2′-dithiodipyridine (DTDP) and glutathione (GSH), and displayed a maximal increase in pCa50 (−log10µCa2+½ at half-maximal force) of ∼0.24 pCa units, with little or no effect on maximum force or Hill coefficient. Partial similar effect was produced by exposure to oxidized gluthathione (GSSG, 10 mm) for 10 min at pH 7.1, and near-maximal effect by GSSG treatment at pH 8.5. None of these treatments significantly altered Ca2+ sensitivity in rat type I fibres. Western blotting showed that both the DTDP–GSH and GSSG–pH 8.5 treatments caused marked S-glutathionylation of the fast troponin I isoform (TnIf) present in type II fibres, but not of troponin C (TnC) or myosin light chain 2. Both the increased Ca2+ sensitivity and glutathionylation of TnIf were blocked by N-ethylmaleimide (NEM). S-Nitrosoglutathione (GSNO) also increased Ca2+ sensitivity, but only in conditions where it caused S-glutathionylation of TnIf. In human type II fibres from vastus lateralis muscle, DTDP–GSH treatment also caused similar increased Ca2+ sensitivity and S-glutathionylation of TnIf. When the slow isoform of TnI in type I fibres of rat was partially substituted (∼30%) with TnIf, DTDP–GSH treatment caused a significant increase in Ca2+ sensitivity (∼0.08 pCa units). TnIf in type II fibres from toad and chicken muscle lack Cys133 present in mammalian TnIf, and such fibres showed no change in Ca2+ sensitivity with DTDP–GSH nor any S-glutathionylation

  9. Evaluation of feeds from tropical origin for in vitro methane production potential and rumen fermentation in vitro

    Directory of Open Access Journals (Sweden)

    Kaushik Pal

    2015-09-01

    Full Text Available Enteric methane arising due to fermentation of feeds in the rumen contributes substantially to the greenhouse gas emissions. Thus, like evaluation of chemical composition and nutritive values of feeds, methane production potential of each feed should be determined. This experiment was conducted to evaluate several feeds for methane production potential and rumen fermentation using in vitro gas production technique so that low methane producing feeds could be utilized to feed ruminants. Protein- and energy-rich concentrates (n=11, cereal and grass forages (n=11, and different straws and shrubs (n=12, which are commonly fed to ruminants in India, were collected from a number of locations. Gas production kinetics, methane production, degradability and rumen fermentation greatly varied (p<0.01 among feeds depending upon the chemical composition. Methane production (mL/g of degraded organic matter was lower (p<0.01 for concentrate than forages, and straws and shrubs. Among shrubs and straws, methane production was lower (p<0.01 for shrubs than straws. Methane production was correlated (p<0.05 with concentrations of crude protein (CP, ether extract and non-fibrous carbohydrate (NFC negatively, and with neutral detergent (NDF and acid detergent fiber (ADF positively. Potential gas production was negatively correlated (p=0.04 with ADF, but positively (p<0.01 with NFC content. Rate of gas production and ammonia concentration were influenced by CP content positively (p<0.05, but by NDF and ADF negatively (p<0.05. Total volatile fatty acid concentration and organic matter degradability were correlated (p<0.05 positively with CP and NFC content, but negatively with NDF and ADF content. The results suggest that incorporation of concentrates and shrubs replacing straws and forages in the diets of ruminants may decrease methane production.

  10. Evaluation of feeds from tropical origin for in vitro methane production potential and rumen fermentation in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Pal, K.; Patra, A. K.; Sahoo, K.

    2015-07-01

    Enteric methane arising due to fermentation of feeds in the rumen contributes substantially to the greenhouse gas emissions. Thus, like evaluation of chemical composition and nutritive values of feeds, methane production potential of each feed should be determined. This experiment was conducted to evaluate several feeds for methane production potential and rumen fermentation using in vitro gas production technique so that low methane producing feeds could be utilized to feed ruminants. Protein- and energy-rich concentrates (n=11), cereal and grass forages (n=11), and different straws and shrubs (n=12), which are commonly fed to ruminants in India, were collected from a number of locations. Gas production kinetics, methane production, degradability and rumen fermentation greatly varied (p<0.01) among feeds depending upon the chemical composition. Methane production (mL/g of degraded organic matter) was lower (p<0.01) for concentrate than forages, and straws and shrubs. Among shrubs and straws, methane production was lower (p<0.01) for shrubs than straws. Methane production was correlated (p<0.05) with concentrations of crude protein (CP), ether extract and non-fibrous carbohydrate (NFC) negatively, and with neutral detergent (NDF) and acid detergent fiber (ADF) positively. Potential gas production was negatively correlated (p=0.04) with ADF, but positively (p<0.01) with NFC content. Rate of gas production and ammonia concentration were influenced by CP content positively (p<0.05), but by NDF and ADF negatively (p<0.05). Total volatile fatty acid concentration and organic matter degradability were correlated (p<0.05) positively with CP and NFC content, but negatively with NDF and ADF content. The results suggest that incorporation of concentrates and shrubs replacing straws and forages in the diets of ruminants may decrease. (Author)

  11. Sodium hyaluronate enhances colorectal tumour cell metastatic potential in vitro and in vivo.

    LENUS (Irish Health Repository)

    Tan, B

    2012-02-03

    BACKGROUND: Sodium hyaluronate has been used intraperitoneally to prevent postoperative adhesions. However, the effect of sodium hyaluronate on tumour growth and metastasis in vitro and in vivo is still unknown. METHODS: Human colorectal tumour cell lines SW480, SW620 and SW707 were treated with sodium hyaluronate (10-500 microg\\/ml) and carboxymethylcellulose (0.125-1 per cent), and tumour cell proliferation and motility were determined in vitro. For the in vivo experiments male BD IX rats were randomized to a sodium hyaluronate group (n = 11; intraperitoneal administration of 0.5 x 10(6) DHD\\/K12 tumour cells and 5 ml 0.4 per cent sodium hyaluronate) or a phosphate-buffered saline group (n = 11; 0.5 x 10(6) DHD\\/K12 tumour cells and 5 ml phosphate-buffered saline intraperitoneally). Four weeks later the intraperitoneal tumour load was visualized directly. RESULTS: In vitro sodium hyaluronate increased tumour cell proliferation and motility significantly. Sodium hyaluronate-induced tumour cell motility appeared to be CD44 receptor dependent, whereas sodium hyaluronate-induced tumour cell proliferation was CD44 receptor independent. In vivo there was a significantly higher total tumour nodule count in the peritoneal cavity of the sodium hyaluronate-treated group compared with the control (P = 0.016). CONCLUSION: Sodium hyaluronate enhances tumour metastatic potential in vitro and in vivo, which suggests that use of sodium hyaluronate to prevent adhesions in colorectal cancer surgery may also potentiate intraperitoneal tumour growth. Presented to the Patey Prize Session of the Surgical Research Society and the annual scientific meeting of the Association of Surgeons of Great Britain and Ireland, Brighton, UK, 4-7 May 1999

  12. A Novel Approach of Synthesizing and Evaluating the Anticancer Potential of Silver Oxide Nanoparticles in vitro.

    Science.gov (United States)

    Banerjee, Kaushik; Das, Satyajit; Choudhury, Pritha; Ghosh, Sarbari; Baral, Rathindranath; Choudhuri, Soumitra Kumar

    2017-01-01

    Development of novel strategies to kill cancer by sparing normal cells is of utmost importance. Apart from their known antimicrobial activity, only limited information has been recorded regarding the antitumor potential of biocompatible silver oxide nanoparticles (AgONPs). There is a need to evaluate the anticancer potential of biocompatible AgONPs in vitro. A new approach of utilizing the leaf extract of Excoecaria agallocha was used to synthesize AgONPs. This was then characterized by ultraviolet-visible spectrophotometry, nanoparticle-tracking analysis, and ζ-potential analysis. Cytotoxicity and apoptotic potential were evaluated with an MTT assay and an annexin V-binding assay against the murine melanoma (B16F10), murine colon cancer (CT26), murine lung adenocarcinoma (3LL), and murine Ehrlich ascites carcinoma (EAC) cell lines. Cellular localization of AgONPs was evaluated on fluorescence microscopy. UV peaks at 270 and 330 nm indicated the formation of nanoparticles (NPs) and the NP-tracking analyzer revealed them to have a size of 228 nm. AgONPs exerted initial cytotoxicity, specifically against all the experimental malignant cells by sparing the normal cell lines. Moreover, AgONPs exert apoptosis equally on all the malignant cells in vitro and ex vivo. This cytotoxicity possibly occurs via the nuclear translocation of AgONPs as analyzed in B16F10 cells. AgONPs utilizing natural sources would be a new medicinal approach against a broad spectrum of malignancy. © 2017 S. Karger AG, Basel.

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

    Directory of Open Access Journals (Sweden)

    Intekhab Islam

    2016-01-01

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

  14. In vitro evaluation of Bifidobacterium strains of human origin for potential use in probiotic functional foods.

    Science.gov (United States)

    Souza, T C; Silva, A M; Drews, J R P; Gomes, D A; Vinderola, C G; Nicoli, J R

    2013-06-01

    The present study investigated some in vitro properties for probiotic use of four strains of bifidobacteria isolated from faeces of healthy children (Bifidobacterium longum 51A, Bifidobacterium breve 1101A, Bifidobacterium pseudolongum 1191A and Bifidobacterium bifidum 1622A). In vitro tests were carried out to compare growth rate, aerotolerance, antagonistic activity against pathogens, antimicrobial susceptibility profile and cell wall hydrophobicity. Mean doubling time of B. longum 51A was shorter compared to the other strains. All strains were aerotolerant up to 72 h of exposure to oxygen. In vitro antagonism showed that B. longum 51A and B. pseudolongum 1191A were able to produce inhibitory diffusible compounds against all pathogenic bacteria tested, but not against Candida albicans. B. longum 51A was sensitive to all the antimicrobials tested, except neomycin. The hydrophobic property of the cell wall was highest for B. bifidum 1622A. Based on these parameters, B. longum 51A showed the best potential for probiotic use among the tested strains, presenting the greatest sensitivity to antimicrobials, the best growth rate and the highest capacity to produce antagonistic substances against various pathogenic microorganisms.

  15. Evaluation of antihyperglycemia and antihypertension potential of native Peruvian fruits using in vitro models.

    Science.gov (United States)

    Pinto, Marcia Da Silva; Ranilla, Lena Galvez; Apostolidis, Emmanouil; Lajolo, Franco Maria; Genovese, Maria Inés; Shetty, Kalidas

    2009-04-01

    Local food diversity and traditional crops are essential for cost-effective management of the global epidemic of type 2 diabetes and associated complications of hypertension. Water and 12% ethanol extracts of native Peruvian fruits such as Lucuma (Pouteria lucuma), Pacae (Inga feuille), Papayita arequipeña (Carica pubescens), Capuli (Prunus capuli), Aguaymanto (Physalis peruviana), and Algarrobo (Prosopis pallida) were evaluated for total phenolics, antioxidant activity based on 2, 2-diphenyl-1-picrylhydrazyl radical scavenging assay, and functionality such as in vitro inhibition of alpha-amylase, alpha-glucosidase, and angiotensin I-converting enzyme (ACE) relevant for potential management of hyperglycemia and hypertension linked to type 2 diabetes. The total phenolic content ranged from 3.2 (Aguaymanto) to 11.4 (Lucuma fruit) mg/g of sample dry weight. A significant positive correlation was found between total phenolic content and antioxidant activity for the ethanolic extracts. No phenolic compound was detected in Lucuma (fruit and powder) and Pacae. Aqueous extracts from Lucuma and Algarrobo had the highest alpha-glucosidase inhibitory activities. Papayita arequipeña and Algarrobo had significant ACE inhibitory activities reflecting antihypertensive potential. These in vitro results point to the excellent potential of Peruvian fruits for food-based strategies for complementing effective antidiabetes and antihypertension solutions based on further animal and clinical studies.

  16. Bioelectric Field Enhancement: The Influence on Membrane Potential and Cell Migration In Vitro.

    Science.gov (United States)

    Purnell, Marcy C; Skrinjar, Terence J

    2016-12-01

    Objective: The extracellular matrix consists of critical components that affect fibroblast polarization and migration. The existence of both intrinsic and extrinsic electrical signals that play essential roles in the development, physiology, regeneration, and pathology of cells was discovered over a century ago. In this study, we study how the Bioelectric Field Enhancement (BEFE) device and its generated electromagnetic field (EMF) by continuous direct current (DC) significantly affect the membrane potential and cell migration of fibroblasts in vitro. Approach: This is an experimental analysis of membrane potential and cell migration of murine fibroblasts when grown in treated media that has been reconstituted with an aqueous solution that has been exposed to an EMF, which is generated by this device versus fibroblasts grown in identically prepared control media that has not been exposed to the EMF. Results: The growth of fibroblasts in the treated media shows a strong percent change in polarization of the plasma membrane and significant increase in cell migration compared to control groups. Innovation: These experiments show the potential for an adjunct wound care therapy using a continuous DC EMF application through a medium of water. Conclusion: Growth media that was reconstituted with an aqueous solution that had been exposed to this DC derived EMF shows significant changes in cell polarity and cell migration of fibroblasts in vitro. The BEFE device has shown enhanced chronic wound healing in anecdotal reports from patients globally for decades when used as a footbath/bath and could lead to a novel EMF application in wound healing.

  17. Potential neoplastic evolution of Vero cells: in vivo and in vitro characterization.

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    Andreani, N A; Renzi, S; Piovani, G; Ajmone Marsan, P; Bomba, L; Villa, R; Ferrari, M; Dotti, S

    2017-10-01

    Vero cell lines are extensively employed in viral vaccine manufacturing. Similarly to all established cells, mutations can occur during Vero cells in vitro amplification which can result in adverse features compromising their biological safety. To evaluate the potential neoplastic evolution of these cells, in vitro transformation test, gene expression analysis and karyotyping were compared among low- (127 and 139 passages) and high-passage (passage 194) cell lines, as well as transformed colonies (TCs). In vivo tumorigenicity was also tested to confirm preliminary in vitro data obtained for low passage lines and TCs. Moreover, Vero cells cultivated in foetal bovine serum-free medium and derived from TCs were analysed to investigate the influence of cultivation methods on tumorigenic evolution. Low-passage Vero developed TCs in soft agar, without showing any tumorigenic evolution when inoculated in the animal model. Karyotyping showed a hypo-diploid modal chromosome number and rearrangements with no difference among Vero cell line passages and TCs. These abnormalities were reported also in serum-free cultivated Vero. Gene expression revealed that high-passage Vero cells had several under-expressed and a few over-expressed genes compared to low-passage ones. Gene ontology revealed no significant enrichment of pathways related to oncogenic risk. These findings suggest that in vitro high passage, and not culture conditions, induces Vero transformation correlated to karyotype and gene expression alterations. These data, together with previous investigations reporting tumour induction in high-passage Vero cells, suggest the use of low-passage Vero cells or cell lines other than Vero to increase the safety of vaccine manufacturing.

  18. Cancer ameliorating potential of Phyllanthus amarus: In vivo and in vitro studies against Aflatoxin B1 toxicity

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    Md. Sultan Ahmad

    2015-10-01

    Conclusion: Ameliorating potential of P. amarus was dose and duration dependant. These extracts significantly reduced the mutagenicity and genotoxicity that were produced due to AFB1 treatment both in vitro and in vivo.

  19. Carbonylation and glutathionylation of proteins in the blue mussel Mytilus edulis detected by proteomic analysis and Western blotting: Actin as a target for oxidative stress.

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    McDonagh, Brian; Tyther, Raymond; Sheehan, David

    2005-07-01

    Protein expression profiles (PEPs) were generated by two-dimensional electrophoresis (2-D SDS-PAGE) for gill and digestive glands of Mytilus edulis sampled from a polluted and reference site in Cork Harbour, Ireland. Similar patterns and expression levels were found for both sites in silver stained gels. However, Western blotting for carbonylated proteins demonstrated higher levels of specific carbonylation of proteins in tissues from animals in the polluted site. Animals from the reference site were acclimated in holding tanks, exposed to 1 mM H2O2 for 24 h, dissected and analysed by 2-D SDS-PAGE. Again, generally similar PEPs were found in control and exposed animals for gill and digestive gland but carbonylation was more pronounced in polluted and exposed animals. Western blotting of extracts after one-dimensional electrophoresis with antibodies to glutathione and actin revealed that gill proteins are glutathionylated more strongly than digestive gland and that this process is more pronounced in polluted animals than in controls. We conclude that carbonylation and glutathionylation can occur in gill and digestive gland in response to oxidative stress in M. edulis. Actin is a major target for both glutathionylation and carbonylation under oxidative stress conditions.

  20. Potencial de multiplicação in vitro de cultivares de morangueiro In vitro multiplication potential of strawberry cultivars

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    Rafael Ucker Brahm

    2004-12-01

    Full Text Available Este trabalho foi realizado com o objetivo de avaliar o potencial de multiplicação in vitro de dez cultivares de morangueiro: Aromas, Bürkley, Camarosa, Campinas, Dover, Milsei-Tudla, Oso Grande, Santa Clara, Sweet Charlie e Vila Nova. Utilizou-se protocolo similar ao dos laboratórios comerciais. A desinfestação dos estolões foi realizada em soluções à base de álcool e hipoclorito de sódio; a cultura dos meristemas em meio semi-sólido MS com 1 mg L-1 BAP, 0,01 mg L-1 ANA e 0,1 mg L-1 AG3; e a multiplicação em meio MS com 1 mg L-1 BAP, à 25 ± 4ºC, 20 µE m-2 s-1 e fotoperíodo de 16 horas. Partiu-se de 10 meristemas de cada cultivar, avaliando-se a taxa de multiplicação e os níveis de contaminação, vitrificação e oxidação durante as fases de estabelecimento (30 dias e de multiplicação (quatro subcultivos. O número estimado de plântulas obtidas por meristema foi: 559 de 'Aromas'; 569 de 'Bürkley'; 516 de 'Camarosa'; 517 de 'Campinas'; 3.907 de 'Dover'; 1.841 de 'Milsei-Tudla'; 943 de 'Oso Grande'; 350 de 'Santa Clara'; 298 de 'Sweet Charlie', e 1.132 de 'Vila Nova'. A quantificação dessa variabilidade genética é importante para o planejamento da produção de matrizes de cada cultivar nos laboratórios de micropropagação.The objective of this research work was to evaluate the in vitro multiplication potential of ten strawberry cultivars : 'Aromas', 'Bürkley', 'Camarosa', 'Campinas', 'Dover', 'Milsei-Tudla', 'Oso Grande', 'Santa Clara', 'Sweet Charlie', and 'Vila Nova'. The procedures used for this purpose were similar to those found in the protocol observed by commercial micropropagation laboratories. The disinfestation of the scions was made by dipping them in an alcohol and sodium hypochlorite solution, the meristem culturing in a semisolid medium containing 1 mg of BAP, 0.01 mg of NAA, and 0.1 mg of G3A per liter and the scions multiplication in an MS medium containing 1 mg of BAP at 25° ± 4° C, 20 µ

  1. In vitro thrombolytic potential of root extracts of four medicinal plants available in Bangladesh

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    Fahad Hussain

    2014-01-01

    Full Text Available Context: Thrombus formation inside the blood vessels obstructs blood flow through the circulatory system leading hypertension, stroke to the heart, anoxia, and so on. Thrombolytic drugs are widely used for the management of cerebral venous sinus thrombosis patients, but they have certain limitations. Medicinal plants and their components possessing antithrombotic activity have been reported before. However, plants that could be used for thrombolysis has not been reported so far. Aims: This study′s aim was to evaluate the thrombolytic potential of selected plants′ root extracts. Settings and Design: Plants were collected, dried, powdered and extracted by methanol and then fractionated by n-hexane for getting the sample root extracts. Venous blood samples were drawn from 10 healthy volunteers for the purposes of investigation. Subjects and Methods: An in vitro thrombolytic model was used to check the clot lysis potential of four n-hexane soluble roots extracts viz., Acacia nilotica, Justicia adhatoda, Azadirachta indica, and Lagerstroemia speciosa along with streptokinase as a positive control and saline water as a negative control. Statistical Analysis Used: Dunnett t-test analysis was performed using SPSS is a statistical analysis program developed by IBM Corporation, USA. on Windows. Results: Using an in vitro thrombolytic model, A. nilotica, L. speciosa, A. indica, and J. adhatoda at 5 mg extract/ml NaCl solution concentration showed 15.1%, 15.49%, 21.26%, and 19.63% clot lysis activity respectively. The reference streptokinase showed 47.21%, and 24.73% clot lysis for 30,000 IU and 15,000 IU concentrations, respectively whereas 0.9% normal saline showed 5.35% clot lysis. Conclusions: The selected extracts of the plant roots possess marked thrombolytic properties that could lyse blood clots in vitro; however, in vivo clot dissolving properties and active components responsible for clot lysis are yet to be discovered.

  2. Quality control and in vitro antioxidant potential of Coriandrum sativum Linn.

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    Mhaveer Singh

    2015-01-01

    Full Text Available Background: Coriandrum sativum Linn., commonly known as coriander, is a well-known spice and drug in India. It has various health-related benefits and used in various Unani formulations. In this present study, quality assessment of coriander fruits was carried out by studying anatomical characters, physicochemical tests, and chemoprofiling using high performance thin layer chromatography (HPTLC and gas chromatography-mass spectroscopy (GC-MS along with in vitro antioxidant potential. Materials and Methods: Standardization was carried out as per the pharmacopeial guidelines. Estimation of heavy metals, pesticides, and aflatoxins was carried out to ascertain the presence of any contaminant in the sample. Chemoprofiling was achieved by thin layer chromatography (TLC by optimizing the mobile phase for different extracts. The most of the pharmacological activities of coriander are based on volatile oil constituents. Hence, GC-MS profiling was also carried out using hexane-soluble fraction of hydro-alcoholic extract. The total phenolic contents and in vitro antioxidant efficacy were determined using previously established methods. Results: The quality control and anatomical studies were very valuable for the identification whereas good antioxidant potential was observed when compared to ascorbic acid. The drug was found free of contaminant when analyzed for pesticides and aflatoxins whereas heavy metals were found under reported limits. Conclusion: The work embodied in this present research can be utilized for the identification and the quality control of the coriander fruit.

  3. Ten years of iPSC: clinical potential and advances in vitro hematopoietic differentiation.

    Science.gov (United States)

    Paes, Bárbara Cristina Martins Fernandes; Moço, Pablo Diego; Pereira, Cristiano Gonçalves; Porto, Geciane Silveira; de Sousa Russo, Elisa Maria; Reis, Luiza Cunha Junqueira; Covas, Dimas Tadeu; Picanço-Castro, Virginia

    2017-06-01

    Ten years have passed since the first publication announcing the generation of induced pluripotent stem cells (iPSCs). Issues related to ethics, immune rejection, and cell availability seemed to be solved following this breakthrough. The development of iPSC technology allows advances in in vitro cell differentiation for cell therapy purpose and other clinical applications. This review provides a perspective on the iPSC potential for cell therapies, particularly for hematological applications. We discuss the advances in in vitro hematopoietic differentiation, the possibilities to employ iPSC in hematology studies, and their potential clinical application in hematologic diseases. The generation of red blood cells and functional T cells and the genome editing technology applied to mutation correction are also covered. We highlight some of the requirements and obstacles to be overcome before translating these cells from research to the clinic, for instance, iPSC variability, genotoxicity, the differentiation process, and engraftment. Also, we evaluate the patent landscape and compile the clinical trials in the field of pluripotent stem cells. Currently, we know much more about iPSC than in 2006, but there are still challenges that must be solved. A greater understanding of molecular mechanisms underlying the generation of hematopoietic stem cells is necessary to produce suitable and transplantable hematopoietic stem progenitor cells from iPSC.

  4. Sulfonamide chalcones: Synthesis and in vitro exploration for therapeutic potential against Brugia malayi.

    Science.gov (United States)

    Bahekar, Sandeep P; Hande, Sneha V; Agrawal, Nikita R; Chandak, Hemant S; Bhoj, Priyanka S; Goswami, Kalyan; Reddy, M V R

    2016-11-29

    Keeping in mind the immense biological potential of chalcones and sulfonamide scaffolds, a library of sulfonamide chalcones has been synthesized and evaluated for in vitro antifilarial assay against human lymphatic filarial parasite Brugia malayi. Experimental evidence showcased for the first time the potential of some sulfonamide chalcones as effective and safe antifilarial lead molecules against human lymphatic filarial parasite B. malayi. Sulfonamide chalcones 4d, 4p, 4q, 4t and 4aa displayed the significantly wide therapeutic window. Particularly chalcones with halogen substitution in aromatic ring proved to be potent antifilarial agents against Brugia malayi. Sulphonamide chalcones with lipophilic methyl moiety (4q and 4aa) at para position of terminal phenyl rings of compounds were found to have remarkable antifilarial activities with therapeutic efficacy. Observed preliminary evidence of apoptosis by effective chalcone derivatives envisaged its fair possibility to inhibit folate pathway with consequent defect in DNA synthesis. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  5. A novel electrical potential sensing method for in vitro stent fracture monitoring and detection.

    Science.gov (United States)

    Park, Chan-Hee; Tijing, Leonard D; Yun, Yeoheung; Kim, Cheol Sang

    2011-01-01

    This article describes a preliminary investigation and prototype fabrication of a novel potential sensing method to continuously monitor vascular stent fractures. A potential measurement system consisting of Wheatstone bridge circuit and signal conditioning circuit was designed for the cardiovascular stent durability and fatigue test. Each end of a bare and polyurethane-covered Nitinol vascular stent was electrically connected to the potential measurement system and then immersed either in simulated body fluid (SBF) media or distilled water at 36.4 ± 1 °C. When the stent experienced fracture (i.e., a cut), its electrical potential decreased with an increase in electrical resistance. This method successfully measured fractures in the stent regardless of location. Furthermore, the number of cycles at the onset of stent fracture was accurately detected and continuously monitored using this technique. Thus, the present fracture detection method, which to our knowledge is the first ever report to use electrical potential measurement for stent durability test, gives a fast, real-time, accurate and efficient detection of fractures in stent during in vitro fatigue and durability test.

  6. S-nitrosylation and S-glutathionylation of Cys134 on troponin I have opposing competitive actions on Ca(2+) sensitivity in rat fast-twitch muscle fibers.

    Science.gov (United States)

    Dutka, T L; Mollica, J P; Lamboley, C R; Weerakkody, V C; Greening, D W; Posterino, G S; Murphy, R M; Lamb, G D

    2017-03-01

    Nitric oxide is generated in skeletal muscle with activity and decreases Ca(2+) sensitivity of the contractile apparatus, putatively by S-nitrosylation of an unidentified protein. We investigated the mechanistic basis of this effect and its relationship to the oxidation-induced increase in Ca(2+) sensitivity in mammalian fast-twitch (FT) fibers mediated by S-glutathionylation of Cys134 on fast troponin I (TnIf). Force-[Ca(2+)] characteristics of the contractile apparatus in mechanically skinned fibers were assessed by direct activation with heavily Ca(2+)-buffered solutions. Treatment with S-nitrosylating agents, S-nitrosoglutathione (GSNO) or S-nitroso-N-acetyl-penicillamine (SNAP), decreased pCa50 ( = -log10 [Ca(2+)] at half-maximal activation) by ~-0.07 pCa units in rat and human FT fibers without affecting maximum force, but had no effect on rat and human slow-twitch fibers or toad or chicken FT fibers, which all lack Cys134. The Ca(2+) sensitivity decrease was 1) fully reversed with dithiothreitol or reduced glutathione, 2) at least partially reversed with ascorbate, indicative of involvement of S-nitrosylation, and 3) irreversibly blocked by low concentration of the alkylating agent, N-ethylmaleimide (NEM). The biotin-switch assay showed that both GSNO and SNAP treatments caused S-nitrosylation of TnIfS-glutathionylation pretreatment blocked the effects of S-nitrosylation on Ca(2+) sensitivity, and vice-versa. S-nitrosylation pretreatment prevented NEM from irreversibly blocking S-glutathionylation of TnIf and its effects on Ca(2+) sensitivity, and likewise S-glutathionylation pretreatment prevented NEM block of S-nitrosylation. Following substitution of TnIf into rat slow-twitch fibers, S-nitrosylation treatment caused decreased Ca(2+) sensitivity. These findings demonstrate that S-nitrosylation and S-glutathionylation exert opposing effects on Ca(2+) sensitivity in mammalian FT muscle fibers, mediated by competitive actions on Cys134 of TnIf. Copyright

  7. Oxidative inhibition of the vascular Na+-K+ pump via NADPH oxidase-dependent β1-subunit glutathionylation: implications for angiotensin II-induced vascular dysfunction.

    Science.gov (United States)

    Liu, Chia-Chi; Karimi Galougahi, Keyvan; Weisbrod, Robert M; Hansen, Thomas; Ravaie, Ramtin; Nunez, Andrea; Liu, Yi B; Fry, Natasha; Garcia, Alvaro; Hamilton, Elisha J; Sweadner, Kathleen J; Cohen, Richard A; Figtree, Gemma A

    2013-12-01

    Glutathionylation of the Na(+)-K(+) pump's β1-subunit is a key molecular mechanism of physiological and pathophysiological pump inhibition in cardiac myocytes. Its contribution to Na(+)-K(+) pump regulation in other tissues is unknown, and cannot be assumed given the dependence on specific β-subunit isoform expression and receptor-coupled pathways. As Na(+)-K(+) pump activity is an important determinant of vascular tone through effects on [Ca(2+)]i, we have examined the role of oxidative regulation of the Na(+)-K(+) pump in mediating angiotensin II (Ang II)-induced increases in vascular reactivity. β1-subunit glutathione adducts were present at baseline and increased by exposure to Ang II in rabbit aortic rings, primary rabbit aortic vascular smooth muscle cells (VSMCs), and human arterial segments. In VSMCs, Ang II-induced glutathionylation was associated with marked reduction in Na(+)-K(+)ATPase activity, an effect that was abolished by the NADPH oxidase inhibitory peptide, tat-gp91ds. In aortic segments, Ang II-induced glutathionylation was associated with decreased K(+)-induced vasorelaxation, a validated index of pump activity. Ang II-induced oxidative inhibition of Na(+)-K(+) ATPase and decrease in K(+)-induced relaxation were reversed by preincubation of VSMCs and rings with recombinant FXYD3 protein that is known to facilitate deglutathionylation of β1-subunit. Knock-out of FXYD1 dramatically decreased K(+)-induced relaxation in a mouse model. Attenuation of Ang II signaling in vivo by captopril (8 mg/kg/day for 7 days) decreased superoxide-sensitive DHE levels in the media of rabbit aorta, decreased β1-subunit glutathionylation, and enhanced K(+)-induced vasorelaxation. Ang II inhibits the Na(+)-K(+) pump in VSMCs via NADPH oxidase-dependent glutathionylation of the pump's β1-subunit, and this newly identified signaling pathway may contribute to altered vascular tone. FXYD proteins reduce oxidative inhibition of the Na(+)-K(+) pump and may have an

  8. Potential inhibition of demineralization in vitro by fluoride-releasing sealants.

    Science.gov (United States)

    Salar, David V; García-Godoy, Franklin; Flaitz, Catherine M; Hicks, M John

    2007-04-01

    The incorporation of fluoride into sealants has been viewed as a viable way to prevent pit-and-fissure caries by potential inhibition of demineralization through the release of fluoride to enamel. The authors conducted a study to examine the effect of a recently introduced fluoride-releasing sealant (ProSeal, Reliance Orthodontic Products, Itasca, Ill.) on enamel demineralization in an in vitro artificial caries system. The authors randomly assigned 45 extracted human third molars to three treatment groups receiving either conventional sealant without fluoride (Group 1), fluoride-releasing sealant (Group 2) or glass ionomer sealant with high fluoride release (Group 3). They placed cavity preparations on the buccal surfaces of the molars and filled them with the assigned material. They placed acid-resistant varnish on the specimens' enamel surfaces to within 1 millimeter of the sealant, leaving a 1-mm rim of sound enamel available for in vitro enamel caries formation. They thermocycled the teeth (500 cycles) in artificial saliva. They subjected the teeth to an in vitro artificial caries challenge for six weeks to produce caries-like lesions in enamel adjacent to the sealant materials. The authors took longitudinal sections from each tooth, immersed them in water and examined them via polarized light microscopy to determine wall lesion frequencies. The mean (+/- standard deviation) lesion depths were 232 +/- 17 micrometers for Group 1, 144 +/- 21 mum for Group 2 and 128 +/- 15 mum for Group 3. The wall lesion frequency was 12 percent for Group 1 and 7 percent for both Groups 2 and 3. There was a significant difference (P sealant substantially reduces the amount of enamel demineralization adjacent to the material. ProSeal provided increased demineralization inhibition compared with a conventional sealant containing no fluoride, but less than that shown by a glass ionomer sealant. ProSeal's physical properties and cariostatic effects may allow for applications beyond

  9. Selection of potential probiotic lactic acid bacteria from fermented olives by in vitro tests.

    Science.gov (United States)

    Argyri, Anthoula A; Zoumpopoulou, Georgia; Karatzas, Kimon-Andreas G; Tsakalidou, Effie; Nychas, George-John E; Panagou, Efstathios Z; Tassou, Chrysoula C

    2013-04-01

    The present study aims to evaluate the probiotic potential of lactic acid bacteria (LAB) isolated from naturally fermented olives and select candidates to be used as probiotic starters for the improvement of the traditional fermentation process and the production of newly added value functional foods. Seventy one (71) lactic acid bacterial strains (17 Leuconostoc mesenteroides, 1 Ln. pseudomesenteroides, 13 Lactobacillus plantarum, 37 Lb. pentosus, 1 Lb. paraplantarum, and 2 Lb. paracasei subsp. paracasei) isolated from table olives were screened for their probiotic potential. Lb. rhamnosus GG and Lb. casei Shirota were used as reference strains. The in vitro tests included survival in simulated gastrointestinal tract conditions, antimicrobial activity (against Listeria monocytogenes, Salmonella Enteritidis, Escherichia coli O157:H7), Caco-2 surface adhesion, resistance to 9 antibiotics and haemolytic activity. Three (3) Lb. pentosus, 4 Lb. plantarum and 2 Lb. paracasei subsp. paracasei strains demonstrated the highest final population (>8 log cfu/ml) after 3 h of exposure at low pH. The majority of the tested strains were resistant to bile salts even after 4 h of exposure, while 5 Lb. plantarum and 7 Lb. pentosus strains exhibited partial bile salt hydrolase activity. None of the strains inhibited the growth of the pathogens tested. Variable efficiency to adhere to Caco-2 cells was observed. This was the same regarding strains' susceptibility towards different antibiotics. None of the strains exhibited β-haemolytic activity. As a whole, 4 strains of Lb. pentosus, 3 strains of Lb. plantarum and 2 strains of Lb. paracasei subsp. paracasei were found to possess desirable in vitro probiotic properties similar to or even better than the reference probiotic strains Lb. casei Shirota and Lb. rhamnosus GG. These strains are good candidates for further investigation both with in vivo studies to elucidate their potential health benefits and in olive fermentation processes

  10. Pterostilbene as a potential novel telomerase inhibitor: molecular docking studies and its in vitro evaluation.

    Science.gov (United States)

    Tippani, Radhika; Prakhya, Laxmi Jaya Shankar; Porika, Mahendar; Sirisha, Kalam; Abbagani, Sadanandam; Thammidala, Christopher

    2014-01-01

    Pterostilbene is a naturally occurring dimethyl ether analog of resveratrol identified in several plant species. Telomerase is important in tumor initiation and cellular immortalization. Given the striking correlations between telomerase activity and proliferation capacity in tumor cells, telomerase had been considered as a potentially important molecular target in cancer therapeutics. Molecular docking studies were performed on pterostilbene with the crystal structure of telomerase (3DU6). Pterostilbene was evaluated for its in vitro cytotoxicity in breast (MCF7) and lung cancer (NCI H-460) cell lines, antimitotic activity in green grams and telomerase activity. Curcumin was used as a standard. Docking results indicated good interaction between pterostilbene and the active site of telomerase and the docked energy of pterostilbene was -7.10 kcal/mol. Pterostilbene showed strong inhibitory effect on in vitro telomerase activity and cell growth in both the cell lines tested in a dose dependent manner. Cancer cells treated with 80 µM pterostilbene exhibited significant telomerase inhibition, after 72 hours (MCF-7 and NCI H-460; 81.52% and 74.69% reduction, respectively, compared to control). The IC50 of pterostilbene for anti-proliferative activity in MCF7 and NCI H-460 cell lines were found to be 30.0 and 47.2 µM, respectively. The best antimitotic activity was obtained with 80 μM of pterostilbene (100% reduction in water imbibition). All the above results were comparable to that of curcumin. The drug-related properties of pterostilbene were calculated using Molinspiration, Osiris Property Explorer and ACD/Chemsketch softwares. Pterostilbene obeyed Lipinski's Rule of Five indicating its therapeutic potential in humans. It was found that the telomerase inhibitory activity exhibited by pterostilbene was dependent of the cell viability and has the potential to be a new drug candidate against breast and lung cancers.

  11. Hepatocytic Differentiation Potential of Human Fetal Liver Mesenchymal Stem Cells: In Vitro and In Vivo Evaluation

    Directory of Open Access Journals (Sweden)

    Hoda El-Kehdy

    2016-01-01

    Full Text Available In line with the search of effective stem cell population that would progress liver cell therapy and because the rate and differentiation potential of mesenchymal stem cells (MSC decreases with age, the current study investigates the hepatogenic differentiation potential of human fetal liver MSCs (FL-MSCs. After isolation from 11-12 gestational weeks’ human fetal livers, FL-MSCs were shown to express characteristic markers such as CD73, CD90, and CD146 and to display adipocytic and osteoblastic differentiation potential. Thereafter, we explored their hepatocytic differentiation potential using the hepatogenic protocol applied for adult human liver mesenchymal cells. FL-MSCs differentiated in this way displayed significant features of hepatocyte-like cells as demonstrated in vitro by the upregulated expression of specific hepatocytic markers and the induction of metabolic functions including CYP3A4 activity, indocyanine green uptake/release, and glucose 6-phosphatase activity. Following transplantation, naive and differentiated FL-MSC were engrafted into the hepatic parenchyma of newborn immunodeficient mice and differentiated in situ. Hence, FL-MSCs appeared to be interesting candidates to investigate the liver development at the mesenchymal compartment level. Standardization of their isolation, expansion, and differentiation may also support their use for liver cell-based therapy development.

  12. In Vitro Differentiation Potential of Human Placenta Derived Cells into Skin Cells

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    Ruhma Mahmood

    2015-01-01

    Full Text Available Skin autografting is the most viable and aesthetic technique for treatment of extensive burns; however, this practice has potential limitations. Harvesting cells from neonatal sources (such as placental tissue is a simple, inexpensive, and noninvasive procedure. In the current study authors sought to evaluate in vitro potential of human placenta derived stem cells to develop into skin-like cells. After extensive washing, amniotic membrane and umbilical cord tissue were separated to harvest amniotic epithelial cells (AECs and umbilical cord mesenchymal stem cells (UC-MSCs, respectively. Both types of cells were characterized for the expression of embryonic lineage markers and their growth characteristics were determined. AECs and UC-MSCs were induced to differentiate into keratinocytes-like and dermal fibroblasts-like cells, respectively. After induction, morphological changes were detected by microscopy. The differentiation potential was further assessed using immunostaining and RT-PCR analyses. AECs were positive for cytokeratins and E-Cadherin while UC-MSCs were positive for fibroblast specific makers. AECs differentiated into keratinocytes-like cells showed positive expression of keratinocyte specific cytokeratins, involucrin, and loricrin. UC-MSCs differentiated into dermal fibroblast-like cells indicated expression of collagen type 3, desmin, FGF-7, fibroblast activation protein alpha, procollagen-1, and vimentin. In conclusion, placenta is a potential source of cells to develop into skin-like cells.

  13. Hippocampal neuron firing and local field potentials in the in vitro 4-aminopyridine epilepsy model.

    Science.gov (United States)

    Gonzalez-Sulser, Alfredo; Wang, Jing; Queenan, Bridget N; Avoli, Massimo; Vicini, Stefano; Dzakpasu, Rhonda

    2012-11-01

    Excessive synchronous neuronal activity is a defining feature of epileptic activity. We previously characterized the properties of distinct glutamatergic and GABAergic transmission-dependent synchronous epileptiform discharges in mouse hippocampal slices using the 4-aminopyridine model of epilepsy. In the present study, we sought to identify the specific hippocampal neuronal populations that initiate and underlie these local field potentials (LFPs). A perforated multielectrode array was used to simultaneously record multiunit action potential firing and LFPs during spontaneous epileptiform activity. LFPs had distinct components based on the initiation site, extent of propagation, and pharmacological sensitivity. Individual units, located in different hippocampal subregions, fired action potentials during these LFPs. A specific neuron subgroup generated sustained action potential firing throughout the various components of the LFPs. The activity of this subgroup preceded the LFPs observed in the presence of antagonists of ionotropic glutamatergic synaptic transmission. In the absence of ionotropic glutamatergic and GABAergic transmission, LFPs disappeared, but units with shorter spike duration and high basal firing rates were still active. These spontaneously active units had an increased level of activity during LFPs and consistently preceded all LFPs recorded before blockade of synaptic transmission. Our findings reveal that neuronal subpopulations with interneuron properties are likely responsible for initiating synchronous activity in an in vitro model of epileptiform discharges.

  14. In Vitro Wound Healing Potential and Identification of Bioactive Compounds from Moringa oleifera Lam

    Directory of Open Access Journals (Sweden)

    Abubakar Amali Muhammad

    2013-01-01

    Full Text Available Moringa oleifera Lam. (M. oleifera from the monogeneric family Moringaceae is found in tropical and subtropical countries. The present study was aimed at exploring the in vitro wound healing potential of M. oleifera and identification of active compounds that may be responsible for its wound healing action. The study included cell viability, proliferation, and wound scratch test assays. Different solvent crude extracts were screened, and the most active crude extract was further subjected to differential bioguided fractionation. Fractions were also screened and most active aqueous fraction was finally obtained for further investigation. HPLC and LC-MS/MS analysis were used for identification and confirmation of bioactive compounds. The results of our study demonstrated that aqueous fraction of M. oleifera significantly enhanced proliferation and viability as well as migration of human dermal fibroblast (HDF cells compared to the untreated control and other fractions. The HPLC and LC-MS/MS studies revealed kaempferol and quercetin compounds in the crude methanolic extract and a major bioactive compound Vicenin-2 was identified in the bioactive aqueous fraction which was confirmed with standard Vicenin-2 using HPLC and UV spectroscopic methods. These findings suggest that bioactive fraction of M. oleifera containing Vicenin-2 compound may enhance faster wound healing in vitro.

  15. Evaluation of in vitro and in vivo anti-arthritic potential of Berberis calliobotrys

    Directory of Open Access Journals (Sweden)

    Alamgeer

    2015-12-01

    Full Text Available The present study was commenced to evaluate the anti-arthritic effect of 70% methanol extract and n-butanol and aqueous fractions of Berberis calliobotrys using both in vitro and in vivo arthritis models. Extract and fractions were investigated in vitro for inhibition of protein (bovine serum and egg albumin denaturation and human red blood cell membrane stabilization. In vivo anti-arthritic activity of extract and fractions at 50, 100 and 200 mg/kg was assessed using turpentine oil and formaldehyde-induced arthritis, while, 200 mg/kg dose was evaluated against complete Freund’s adjuvant-induced arthritis. B. calliobotrys produced significant (p<0.001 dose dependent inhibition of protein denaturation and human red blood cell membrane stabilization. In turpentine oil, formaldehyde and complete Freund’s adjuvant-induced arthritis models, B. calliobotrys significantly (p<0.001 reduced joint and paw swelling. B. calliobotrys markedly improved body weight, hematology profile, radiological and histopathological parameters in complete Freund’s adjuvant model. It could be concluded that B. calliobotrys holds anti-arthritic potential, supporting its traditional use in treatment of rheumatoid arthritis.

  16. Development of next-generation peptide binders using in vitro display technologies and their potential applications

    Directory of Open Access Journals (Sweden)

    Akira eWada

    2013-08-01

    Full Text Available During the last decade, a variety of monoclonal antibodies have been developed and used as molecular targeting drugs in medical therapies. Although antibody drugs tend to have intense pharmacological activities and negligible side effects, several issues in their development and prescription remain to be resolved. Synthetic peptides with affinities and specificities for a desired target have received significant attention as alternatives to antibodies. In vitro display technologies are powerful methods for the selection of such peptides from combinatorial peptide libraries. Various types of peptide binders are being selected with such technologies for use in a wide range of fields from bioscience to medicine. This mini review article focuses on the current state of in vitro display selection of synthetic peptide binders and compares the selected peptides with natural peptides/proteins to provide a better understanding of the target affinities and inhibitory activities derived from their amino acid sequences and structural frameworks. The potential of synthetic peptide binders as alternatives to antibody drugs in therapeutic applications is also reviewed.

  17. Free radical scavenging potential of in vitro raised and greenhouse acclimatized plants of Artemisia amygdalina.

    Science.gov (United States)

    Rasool, R; Ganai, B A; Akbar, S; Kamili, A N

    2013-07-01

    Artemisia amygdalina Decne. (Asteraceae) is a critically endangered and endemic herb of Kashmir Himalayan sub-alpine region and Pakistan. Scientific research throughout the world has evidence to support the tremendous medicinal utility of the genus Artemisia. The natural resources of medicinal plants are being reduced day by day. This study provides the alternative way for medicinal resource utilization and conservation of A. amygdalina. In vitro-raised plants and greenhouse acclimatized plants were obtained by culturing wild explants on Murashige and Skoog's medium. Plant extracts were obtained and subjected to different antioxidant assays: DPPH assay, riboflavin photo-oxidation assay, deoxy ribose assay, ferric thiocyanate assay, thiobarbituric acid assay, post mitochondrial supernatant assay and DNA damage on agarose gel. In vitro grown plants, as well as those acclimatized in the greenhouse reveals antioxidant activity against hydroxyl, superoxide, and lipid peroxyl radicals. This preliminary study revealed the free radical scavenging potential of tissue culture-raised plant extracts of A. amydalina. Copyright © 2013 China Pharmaceutical University. Published by Elsevier B.V. All rights reserved.

  18. Potential applications of nonthermal plasmas against biofilm-associated micro-organisms in vitro.

    Science.gov (United States)

    Puligundla, P; Mok, C

    2017-05-01

    Biofilms as complex microbial communities attached to surfaces pose several challenges in different sectors, ranging from food and healthcare to desalination and power generation. The biofilm mode of growth allows microorganisms to survive in hostile environments and biofilm cells exhibit distinct physiology and behaviour in comparison with their planktonic counterparts. They are ubiquitous, resilient and difficult to eradicate due to their resistant phenotype. Several chemical-based cleaning and disinfection regimens are conventionally used against biofilm-dwelling micro-organisms in vitro. Although such approaches are generally considered to be effective, they may contribute to the dissemination of antimicrobial resistance and environmental pollution. Consequently, advanced green technologies for biofilm control are constantly emerging. Disinfection using nonthermal plasmas (NTPs) is one of the novel strategies having a great potential for control of biofilms of a broad spectrum of micro-organisms. This review discusses several aspects related to the inactivation of biofilm-associated bacteria and fungi by different types of NTPs under in vitro conditions. A brief introduction summarizes prevailing methods in biofilm inactivation, followed by introduction to gas discharge plasmas, active plasma species and their inactivating mechanism. Subsequently, significance and aspects of NTP inactivation of biofilm-associated bacteria, especially those of medical importance, including opportunistic pathogens, oral pathogenic bacteria, foodborne pathogens and implant bacteria, are discussed. The remainder of the review discusses majorly about the synergistic effect of NTPs and their activity against biofilm-associated fungi, especially Candida species. © 2017 The Society for Applied Microbiology.

  19. In vitro wound healing potential and identification of bioactive compounds from Moringa oleifera Lam.

    Science.gov (United States)

    Muhammad, Abubakar Amali; Pauzi, Nur Aimi Syarina; Arulselvan, Palanisamy; Abas, Faridah; Fakurazi, Sharida

    2013-01-01

    Moringa oleifera Lam. (M. oleifera) from the monogeneric family Moringaceae is found in tropical and subtropical countries. The present study was aimed at exploring the in vitro wound healing potential of M. oleifera and identification of active compounds that may be responsible for its wound healing action. The study included cell viability, proliferation, and wound scratch test assays. Different solvent crude extracts were screened, and the most active crude extract was further subjected to differential bioguided fractionation. Fractions were also screened and most active aqueous fraction was finally obtained for further investigation. HPLC and LC-MS/MS analysis were used for identification and confirmation of bioactive compounds. The results of our study demonstrated that aqueous fraction of M. oleifera significantly enhanced proliferation and viability as well as migration of human dermal fibroblast (HDF) cells compared to the untreated control and other fractions. The HPLC and LC-MS/MS studies revealed kaempferol and quercetin compounds in the crude methanolic extract and a major bioactive compound Vicenin-2 was identified in the bioactive aqueous fraction which was confirmed with standard Vicenin-2 using HPLC and UV spectroscopic methods. These findings suggest that bioactive fraction of M. oleifera containing Vicenin-2 compound may enhance faster wound healing in vitro.

  20. In Vitro and In Vivo Infectious Potential of Coxiella burnetii: A Study on Belgian Livestock Isolates

    Science.gov (United States)

    Mori, Marcella; Boarbi, Samira; Michel, Patrick; Bakinahe, Raïssa; Rits, Katleen; Wattiau, Pierre; Fretin, David

    2013-01-01

    Q-fever is a zoonosis caused by the gram-negative obligate intracellular pathogen Coxiella burnetii. Since its discovery, and particularly following the recent outbreaks in the Netherlands, C. burnetii appeared as a clear public health concern. In the present study, the infectious potential displayed by goat and cattle isolates of C. burnetii was compared to a reference strain (Nine Mile) using both in vitro (human HeLa and bovine macrophage cells) and in vivo (BALB/c mice) models. The isolates had distant genomic profiles with one - the goat isolate - being identical to the predominant strain circulating in the Netherlands during the 2007–2010 outbreaks. Infective doses were established with ethidium monoazide-PCR for the first time here applied to C. burnetii. This method allowed for the preparation of reproducible and characterized inocula thanks to its capacity to discriminate between live and dead cells. Globally, the proliferative capacity of the Nine Mile strain in cell lines and mice was higher compared to the newly isolated field strains. In vitro, the bovine C. burnetii isolate multiplied faster in a bovine macrophage cell line, an observation tentatively explained by the preferential specificity of this strain for allogeneic host cells. In the BALB/c mouse model, however, the goat and bovine isolates multiplied at about the same rate indicating no peculiar hypervirulent behavior in this animal model. PMID:23840751

  1. In vitro and in vivo germ line potential of stem cells derived from newborn mouse skin.

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    Paul W Dyce

    Full Text Available We previously reported that fetal porcine skin-derived stem cells were capable of differentiation into oocyte-like cells (OLCs. Here we report that newborn mice skin-derived stem cells are also capable of differentiating into early OLCs. Using stem cells from mice that are transgenic for Oct4 germline distal enhancer-GFP, germ cells resulting from their differentiation are expected to be GFP(+. After differentiation, some GFP(+ OLCs reached 40-45 µM and expressed oocyte markers. Flow cytometric analysis revealed that ∼ 0.3% of the freshly isolated skin cells were GFP(+. The GFP-positive cells increased to ∼ 7% after differentiation, suggesting that the GFP(+ cells could be of in vivo origin, but are more likely induced upon being cultured in vitro. To study the in vivo germ cell potential of skin-derived cells, they were aggregated with newborn ovarian cells, and transplanted under the kidney capsule of ovariectomized mice. GFP(+ oocytes were identified within a subpopulation of follicles in the resulting growth. Our finding that early oocytes can be differentiated from mice skin-derived cells in defined medium may offer a new in vitro model to study germ cell formation and oogenesis.

  2. Equol an isoflavonoid: potential for improved prostate health, in vitro and in vivo evidence

    Science.gov (United States)

    2011-01-01

    Background To determine: in vitro binding affinity of equol for 5alpha-dihydrotestosterone (5alpha-DHT), in vitro effects of equol treatment in human prostate cancer (LNCap) cells, and in vivo effects of equol on rat prostate weight and circulating levels of sex steroid hormones. Methods First, in vitro equol binding affinity for 5alpha-DHT was determined using 14C5alpha-DHT combined with cold 5alpha-DHT (3.0 nM in all samples). These steroids were incubated with increasing concentrations of equol (0-2,000 nM) and analyzed by Sephadex LH-20 column chromatography. 14C5alpha-DHT peak/profiles were determined by scintillation counting of column fractions. Using the 14C5alpha-DHT peak (0 nM equol) as a reference standard, a binding curve was generated by quantifying shifts in the 14C5alpha-DHT peaks as equol concentrations increased. Second, equol's in vitro effects on LNCap cells were determined by culturing cells (48 hours) in the presence of increasing concentrations of dimethyl sulfoxide (DMSO) (vehicle-control), 5alpha-DHT, equol or 5alpha-DHT+equol. Following culture, prostate specific antigen (PSA) levels were quantified via ELISA. Finally, the in vivo effects of equol were tested in sixteen male Long-Evans rats fed a low isoflavone diet. From 190-215 days, animals received 0.1cc s.c. injections of either DMSO-control vehicle (n = 8) or 1.0 mg/kg (body weight) of equol (in DMSO) (n = 8). At 215 days, body and prostate weights were recorded, trunk blood was collected and serum assayed for luteinizing hormone (LH), 5alpha-DHT, testosterone and 17beta-estradiol levels. Results Maximum and half maximal equol binding to 5alpha-DHT occurred at approximately 100 nM and 4.8 nM respectively. LNCap cells cultured in the presence of 5alpha-DHT significantly increased PSA levels. However, in the presence of 5alpha-DHT+equol, equol blocked the significant increases in PSA levels from LNCap cells. In vivo equol treatment significantly decreased rat prostate weights and serum

  3. iEquol an isoflavonoid: potential for improved prostate health, in vitro and in vivo evidence

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    Hamaker Amy N

    2011-01-01

    Full Text Available Abstract Background To determine: in vitro binding affinity of equol for 5alpha-dihydrotestosterone (5alpha-DHT, in vitro effects of equol treatment in human prostate cancer (LNCap cells, and in vivo effects of equol on rat prostate weight and circulating levels of sex steroid hormones. Methods First, in vitro equol binding affinity for 5alpha-DHT was determined using 14C5alpha-DHT combined with cold 5alpha-DHT (3.0 nM in all samples. These steroids were incubated with increasing concentrations of equol (0-2,000 nM and analyzed by Sephadex LH-20 column chromatography. 14C5alpha-DHT peak/profiles were determined by scintillation counting of column fractions. Using the 14C5alpha-DHT peak (0 nM equol as a reference standard, a binding curve was generated by quantifying shifts in the 14C5alpha-DHT peaks as equol concentrations increased. Second, equol's in vitro effects on LNCap cells were determined by culturing cells (48 hours in the presence of increasing concentrations of dimethyl sulfoxide (DMSO (vehicle-control, 5alpha-DHT, equol or 5alpha-DHT+equol. Following culture, prostate specific antigen (PSA levels were quantified via ELISA. Finally, the in vivo effects of equol were tested in sixteen male Long-Evans rats fed a low isoflavone diet. From 190-215 days, animals received 0.1cc s.c. injections of either DMSO-control vehicle (n = 8 or 1.0 mg/kg (body weight of equol (in DMSO (n = 8. At 215 days, body and prostate weights were recorded, trunk blood was collected and serum assayed for luteinizing hormone (LH, 5alpha-DHT, testosterone and 17beta-estradiol levels. Results Maximum and half maximal equol binding to 5alpha-DHT occurred at approximately 100 nM and 4.8 nM respectively. LNCap cells cultured in the presence of 5alpha-DHT significantly increased PSA levels. However, in the presence of 5alpha-DHT+equol, equol blocked the significant increases in PSA levels from LNCap cells. In vivo equol treatment significantly decreased rat prostate

  4. Quantitation of protein S-glutathionylation by liquid chromatography-tandem mass spectrometry: correction for contaminating glutathione and glutathione disulfide.

    Science.gov (United States)

    Bukowski, Michael R; Bucklin, Christopher; Picklo, Matthew J

    2015-01-15

    Protein S-glutathionylation is a posttranslational modification that links oxidative stimuli to reversible changes in cellular function. Protein-glutathione mixed disulfide (PSSG) is commonly quantified by reduction of the disulfide and detection of the resultant glutathione species. This methodology is susceptible to contamination by free unreacted cellular glutathione (GSH) species, which are present in 1000-fold greater concentration. A liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based method was developed for quantification of glutathione and glutathione disulfide (GSSG), which was used for the determination of PSSG in biological samples. Analysis of rat liver samples demonstrated that GSH and GSSG coprecipitated with proteins similar to the range for PSSG in the sample. The use of [(13)C2,(5)N]GSH and [(13)C4,(5)N2]GSSG validated these results and demonstrated that the release of GSH from PSSG did not occur during sample preparation and analysis. These data demonstrate that GSH and GSSG contamination must be accounted for when determining PSSG content in cellular/tissue preparations. A protocol for rinsing samples to remove the adventitious glutathione species is demonstrated. The fragmentation patterns for glutathione were determined by high-resolution mass spectrometry, and candidate ions for detection of PSSG on protein and protein fragments were identified. Published by Elsevier Inc.

  5. Identification of glutathionyl-3-hydroxykynurenine glucoside as a novel fluorophore associated with aging of the human lens.

    Science.gov (United States)

    Garner, B; Vazquez, S; Griffith, R; Lindner, R A; Carver, J A; Truscott, R J

    1999-07-23

    A novel fluorophore was isolated from human lenses using high performance liquid chromatography (HPLC). The new fluorophore was well separated from 3-hydroxykynurenine glucoside (3-OHKG) and its deaminated isoform, 4-(2-amino-3-hydroxyphenyl)-4-oxobutanoic acid O-glucoside, which are known UV filter compounds. The new compound exhibited UV absorbance maxima at 260 and 365 nm, was fluorescent (Ex(360 nm)/Em(500 nm)), and increased in concentration with age. Further analysis of the purified compound by microbore HPLC with in-line electrospray ionization mass spectrometry revealed a molecular mass of 676 Da. This mass corresponds to that of an adduct of GSH with a deaminated form of 3-OHKG. This adduct was synthesized using 3-OHKG and GSH as starting materials. The synthetic glutathionyl-3-hydroxykynurenine glucoside (GSH-3-OHKG) adduct had the same HPLC elution time, thin-layer chromatography R(F) value, UV absorbance maxima, fluorescence characteristics, and mass spectrum as the lens-derived fluorophore. Furthermore, the (1)H and (13)C NMR spectra of the synthetic adduct were entirely consistent with the proposed structure of GSH-3-OHKG. These data indicate that GSH-3-OHKG is present as a novel fluorophore in aged human lenses. The GSH-3-OHKG adduct was found to be less reactive with beta-glucosidase compared with 3-OHKG, and this could be due to a folded conformation of the adduct that was suggested by molecular modeling.

  6. Regulation of protein function by S-nitrosation and S-glutathionylation: processes and targets in cardiovascular pathophysiology.

    Science.gov (United States)

    Belcastro, Eugenia; Gaucher, Caroline; Corti, Alessandro; Leroy, Pierre; Lartaud, Isabelle; Pompella, Alfonso

    2017-11-27

    Decades of chemical, biochemical and pathophysiological research have established the relevance of post-translational protein modifications induced by processes related to oxidative stress, with critical reflections on cellular signal transduction pathways. A great deal of the so-called 'redox regulation' of cell function is in fact mediated through reactions promoted by reactive oxygen and nitrogen species on more or less specific aminoacid residues in proteins, at various levels within the cell machinery. Modifications involving cysteine residues have received most attention, due to the critical roles they play in determining the structure/function correlates in proteins. The peculiar reactivity of these residues results in two major classes of modifications, with incorporation of NO moieties (S-nitrosation, leading to formation of protein S-nitrosothiols) or binding of low molecular weight thiols (S-thionylation, i.e. in particular S-glutathionylation, S-cysteinylglycinylation and S-cysteinylation). A wide array of proteins have been thus analyzed in detail as far as their susceptibility to either modification or both, and the resulting functional changes have been described in a number of experimental settings. The present review aims to provide an update of available knowledge in the field, with a special focus on the respective (sometimes competing and antagonistic) roles played by protein S-nitrosations and S-thionylations in biochemical and cellular processes specifically pertaining to pathogenesis of cardiovascular diseases.

  7. Raman spectrum: A potential biomarker for embryo assessment during in vitro fertilization.

    Science.gov (United States)

    Ding, Jiayi; Xu, Tian; Tan, Xiaofang; Jin, Hua; Shao, Jun; Li, Haibo

    2017-05-01

    The aim of the study was to investigate whether Raman spectrum is consistent with the morphological scoring of the embryo of day 3 during in vitro fertilization (IVF). The spent culture media of embryo of day 3 from 10 patients were collected and analyzed. The samples were analyzed using Raman spectroscopy and graded according to the standard embryo scoring system simultaneously. Data showed that the Raman spectra obtained from the droplet of media were useful, as they can act as the characteristic signature for protein and amino acids. The Raman biospectroscopy-based metabonomics profiling of spent media was consistent with the result of conventional morphological evaluation. In conclusion, this technology offers great potential for the development of tools allowing rapid non-invasive assessment of the quality of embryo of day 3 during IVF.

  8. Phenolic compounds, antioxidant potential and antiproliferative potential of 10 common edible flowers from China assessed using a simulated in vitro digestion-dialysis process combined with cellular assays.

    Science.gov (United States)

    Huang, Weisu; Mao, Shuqin; Zhang, Liuquan; Lu, Baiyi; Zheng, Lufei; Zhou, Fei; Zhao, Yajing; Li, Maiquan

    2017-11-01

    Phenolic compounds could be sensitive to digestive conditions, thus a simulated in vitro digestion-dialysis process and cellular assays was used to determine phenolic compounds and antioxidant and antiproliferative potentials of 10 common edible flowers from China and their functional components. Gallic acid, ferulic acid, and rutin were widely present in these flowers, which demonstrated various antioxidant capacities (DPPH, ABTS, FRAP and CAA values) and antiproliferative potentials measured by the MTT method. Rosa rugosa, Paeonia suffruticosa and Osmanthus fragrans exhibited the best antioxidant and antiproliferative potentials against HepG2, A549 and SGC-7901 cell lines, except that Osmanthus fragrans was not the best against SGC-7901 cells. The in vitro digestion-dialysis process decreased the antioxidant potential by 33.95-90.72% and the antiproliferative potential by 13.22-87.15%. Following the in vitro digestion-dialysis process, phenolics were probably responsible for antioxidant (R2 = 0.794-0.924, P < 0.01) and antiproliferative (R2 = 0.408-0.623, P < 0.05) potential. Moreover, gallic acid may be responsible for the antioxidant potential of seven flowers rich in edible flowers. The antioxidant and antiproliferative potential of 10 edible flowers revealed a clear decrease after digestion and dialysis along with the reduction of phenolics. Nevertheless, they still had considerable antioxidant and antiproliferative potential, which merited further investigation in in vivo studies. © 2017 Society of Chemical Industry. © 2017 Society of Chemical Industry.

  9. Bioavailability and potential carcinogenicity of polycyclic aromatic hydrocarbons from wood combustion particulate matter in vitro.

    Science.gov (United States)

    Gauggel-Lewandowski, Susanne; Heussner, Alexandra H; Steinberg, Pablo; Pieterse, Bart; van der Burg, Bart; Dietrich, Daniel R

    2013-11-25

    Due to increasing energy demand and limited fossil fuels, renewable energy sources have gained in importance. Particulate matter (PM) in general, but also PM from the combustion of wood is known to exert adverse health effects in human. These are often related to specific toxic compounds adsorbed to the PM surface, such as polycyclic aromatic hydrocarbons (PAH), of which some are known human carcinogens. This study focused on the bioavailability of PAHs and on the tumor initiation potential of wood combustion PM, using the PAH CALUX® reporter gene assay and the BALB/c 3T3 cell transformation assay, respectively. For this, both cell assays were exposed to PM and their respective organic extracts from varying degrees of combustion. The PAH CALUX® experiments demonstrated a concentration-response relationship matching the PAHs detected in the samples. Contrary to expectations, PM samples from complete (CC) and incomplete combustion (IC) provided for a stronger and weaker response, respectively, suggesting that PAH were more readily bioavailable in PM from CC. These findings were corroborated via PAH spiking experiments indicating that IC PM contains organic components that strongly adsorb PAH thereby reducing their bioavailability. The results obtained with organic extracts in the cell transformation assay presented the highest potential for carcinogenicity in samples with high PAH contents, albeit PM from CC also demonstrated a carcinogenic potential. In conclusion, the in vitro assays employed emphasize that CC produces PM with low PAH content however with a general higher bioavailability and thus with a nearly similar carcinogenic potential than IC PM. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  10. Eucalyptus Essential Oil as a Natural Food Preservative: In Vivo and In Vitro Antiyeast Potential

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    Amit Kumar Tyagi

    2014-01-01

    Full Text Available In this study, the application of eucalyptus essential oil/vapour as beverages preservative is reported. The chemical composition of eucalyptus oil was determined by gas chromatography-mass spectrometry (GC-MS and solid phase microextraction GC-MS (SPME/GC-MS analyses. GC-MS revealed that the major constituents were 1,8-cineole (80.5%, limonene (6.5%, α-pinene (5%, and γ-terpinene (2.9% while SPME/GC-MS showed a relative reduction of 1,8-cineole (63.9% and an increase of limonene (13.8%, α-pinene (8.87%, and γ-terpinene (3.98%. Antimicrobial potential of essential oil was initially determined in vitro against 8 different food spoilage yeasts by disc diffusion, disc volatilization, and microdilution method. The activity of eucalyptus vapours was significantly higher than the eucalyptus oil. Minimum inhibitory concentration (MIC and minimum fungicidal concentration (MFC varied from 0.56 to 4.50 mg/mL and from 1.13 to 9 mg/mL, respectively. Subsequently, the combined efficacy of essential oil and thermal treatment were used to evaluate the preservation of a mixed fruit juice in a time-dependent manner. These results suggest eucalyptus oil as a potent inhibitor of food spoilage yeasts not only in vitro but also in a real food system. Currently, this is the first report that uses eucalyptus essential oil for fruit juice preservation against food spoiling yeast.

  11. In Vitro Properties of Potential Probiotic Indigenous Lactic Acid Bacteria Originating from Traditional Pickles.

    Science.gov (United States)

    Tokatlı, Mehmet; Gülgör, Gökşen; Bağder Elmacı, Simel; Arslankoz İşleyen, Nurdan; Özçelik, Filiz

    2015-01-01

    The suitable properties of potential probiotic lactic acid bacteria (LAB) strains (preselected among 153 strains on the basis of their potential technological properties) isolated from traditional Çubuk pickles were examined in vitro. For this purpose, these strains (21 Lactobacillus plantarum, 11 Pediococcus ethanolidurans, and 7 Lactobacillus brevis) were tested for the ability to survive at pH 2.5, resistance to bile salts, viability in the presence of pepsin-pancreatin, ability to deconjugate bile salts, cholesterol assimilation, and surface hydrophobicity properties. Most of the properties tested could be assumed to be strain-dependent. However, L. plantarum and L. brevis species were found to possess desirable probiotic properties to a greater extent compared to P. ethanolidurans. In contrast to P. ethanolidurans strains, the tested L. plantarum and L. brevis strains exhibited bile salt tolerance, albeit to different extent. All tested strains showed less resistance to intestinal conditions than gastric juice environment. Based on the survival under gastrointestinal conditions, 22 of the 39 strains were selected for further characterization. The eight strains having the highest cholesterol assimilation and surface hydrophobicity ratios could be taken as promising probiotic candidates for further in vivo studies, because of the strongest variations found among the tested strains with regard to these properties.

  12. In vitro selection of bacteria with potential for use as probiotics in marine shrimp culture

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    Felipe do Nascimento Vieira

    2013-08-01

    Full Text Available The objective of this work was to isolate strains of lactic acid bacteria with probiotic potential from the digestive tract of marine shrimp (Litopenaeus vannamei, and to carry out in vitro selection based on multiple characters. The ideotype (ideal proposed strain was defined by the highest averages for the traits maximum growth velocity, final count of viable cells, and inhibition halo against nine freshwater and marine pathogens, and by the lowest averages for the traits duplication time and resistance of strains to NaCl (1.5 and 3%, pH (6, 8, and 9, and biliary salts (5%. Mahalanobis distance (D² was estimated among the evaluated strains, and the best ones were those with the shortest distances to the ideotype. Ten bacterial strains were isolated and biochemically identified as Lactobacillus plantarum (3, L. brevis (3, Weissella confusa (2, Lactococcus lactis (1, and L. delbrueckii (1. Lactobacillus plantarum strains showed a wide spectrum of action and the largest inhibition halos against pathogens, both Gram-positive and negative, high growth rate, and tolerance to all evaluated parameters. In relation to ideotype, L. plantarum showed the lowest Mahalanobis (D² distance, followed by the strains of W. confusa, L. brevis, L. lactis, and L. delbrueckii. Among the analyzed bacterial strains, those of Lactobacillus plantarum have the greatest potential for use as a probiotic for marine shrimp.

  13. In vitro osteoinductive potential of porous monetite for bone tissue engineering

    Directory of Open Access Journals (Sweden)

    Bernadine Idowu

    2014-05-01

    Full Text Available Tissue engineering–based bone grafts are emerging as a viable alternative treatment modality to repair and regenerate tissues damaged as a result of disease or injury. The choice of the biomaterial component is a critical determinant of the success of the graft or scaffold; essentially, it must induce and allow native tissue integration, and most importantly mimic the hierarchical structure of the native bone. Calcium phosphate bioceramics are widely used in orthopaedics and dentistry applications due to their similarity to bone mineral and their ability to induce a favourable biological response. One such material is monetite, which is biocompatible, osteoconductive and has the ability to be resorbed under physiological conditions. The osteoinductive properties of monetite in vivo are known; however, little is known of the direct effect on osteoinduction of human mesenchymal stem cells in vitro. In this study, we evaluated the potential of monetite to induce and sustain human mesenchymal stem cells towards osteogenic differentiation. Human mesenchymal stem cells were seeded on the monetite scaffold in the absence of differentiating factors for up to 28 days. The gene expression profile of bone-specific markers in cells on monetite scaffold was compared to the control material hydroxyapatite. At day 14, we observed a marked increase in alkaline phosphatase, osteocalcin and osteonectin expressions. This study provides evidence of a suitable material that has potential properties to be used as a tissue engineering scaffold.

  14. In vitro osteoinductive potential of porous monetite for bone tissue engineering

    Science.gov (United States)

    Idowu, Bernadine; Cama, Giuseppe; Deb, Sanjukta

    2014-01-01

    Tissue engineering–based bone grafts are emerging as a viable alternative treatment modality to repair and regenerate tissues damaged as a result of disease or injury. The choice of the biomaterial component is a critical determinant of the success of the graft or scaffold; essentially, it must induce and allow native tissue integration, and most importantly mimic the hierarchical structure of the native bone. Calcium phosphate bioceramics are widely used in orthopaedics and dentistry applications due to their similarity to bone mineral and their ability to induce a favourable biological response. One such material is monetite, which is biocompatible, osteoconductive and has the ability to be resorbed under physiological conditions. The osteoinductive properties of monetite in vivo are known; however, little is known of the direct effect on osteoinduction of human mesenchymal stem cells in vitro. In this study, we evaluated the potential of monetite to induce and sustain human mesenchymal stem cells towards osteogenic differentiation. Human mesenchymal stem cells were seeded on the monetite scaffold in the absence of differentiating factors for up to 28 days. The gene expression profile of bone-specific markers in cells on monetite scaffold was compared to the control material hydroxyapatite. At day 14, we observed a marked increase in alkaline phosphatase, osteocalcin and osteonectin expressions. This study provides evidence of a suitable material that has potential properties to be used as a tissue engineering scaffold. PMID:24904727

  15. In vitro photoprotective effects of Marcetia taxifolia ethanolic extract and its potential for sunscreen formulations

    Directory of Open Access Journals (Sweden)

    Sônia C.C. Costa

    Full Text Available AbstractThe species Marcetia taxifolia (A. St.-Hil. DC., Melastomataceae, which is endemic of the rupestrian fields of northeastern Brazil, contains a significant amount of flavonoids. In this work, the potential of the ethanolic extract of M. taxifolia as the active principle in a sunscreen photoprotection (UV-A and UV-B formulation was investigated. The Liquid Chromatography High Performance-Diode Array Detector quantification (quercetin, total flavonoid content, antioxidant activity through 2.2-diphenyl-1- picrylhydrazil method, photoprotective activity against UV-B and UV-A radiation in vitro (spectrophotometric method and potential for eye irritation using the methodology of the hen egg test-chorioallantoic membrane were performed in the extract. After that, the formulations were prepared using different concentrations of active ethanolic extract (5, 10, 20 and 30% and the evaluation of the sun protection factor was carried out using the same methodology used for the crude extract. The crude extract showed UV-A photoprotection and low eye irritation in the hen egg test-chorioallantoic membrane test. All formulations containing M. taxifolia extract had ≥ 6 sun protection factor. Its shows the possibility to use this extracts as a sunscreen in pharmaceutical preparations.

  16. Equol, a Dietary Daidzein Gut Metabolite Attenuates Microglial Activation and Potentiates Neuroprotection In Vitro.

    Science.gov (United States)

    Subedi, Lalita; Ji, Eunhee; Shin, Dongyun; Jin, Jongsik; Yeo, Joo Hong; Kim, Sun Yeou

    2017-02-27

    Estrogen deficiency has been well characterized in inflammatory disorders including neuroinflammation. Daidzein, a dietary alternative phytoestrogen found in soy (Glycine max) as primary isoflavones, possess anti-inflammatory activity, but the effect of its active metabolite Equol (7-hydroxy-3-(4'-hydroxyphenyl)-chroman) has not been well established. In this study, we investigated the anti-neuroinflammatory and neuroprotective effect of Equol in vitro. To evaluate the potential effects of Equol, three major types of central nervous system (CNS) cells, including microglia (BV-2), astrocytes (C6), and neurons (N2a), were used. Effects of Equol on the expression of inducible nitric oxide synthase (iNOS), cyclooxygenase (COX-2), Mitogen activated protein kinase (MAPK) signaling proteins, and apoptosis-related proteins were measured by western blot analysis. Equol inhibited the lipopolysaccharide (LPS)-induced TLR4 activation, MAPK activation, NF-kB-mediated transcription of inflammatory mediators, production of nitric oxide (NO), release of prostaglandin E2 (PGE-2), secretion of tumor necrosis factor-α (TNF-α) and interleukin 6 (IL-6), in Lipopolysaccharide (LPS)-activated murine microglia cells. Additionally, Equol protects neurons from neuroinflammatory injury mediated by LPS-activated microglia through downregulation of neuronal apoptosis, increased neurite outgrowth in N2a cell and neurotrophins like nerve growth factor (NGF) production through astrocytes further supporting its neuroprotective potential. These findings provide novel insight into the anti-neuroinflammatory effects of Equol on microglial cells, which may have clinical significance in cases of neurodegeneration.

  17. In Vitro Properties of Potential Probiotic Indigenous Lactic Acid Bacteria Originating from Traditional Pickles

    Directory of Open Access Journals (Sweden)

    Mehmet Tokatlı

    2015-01-01

    Full Text Available The suitable properties of potential probiotic lactic acid bacteria (LAB strains (preselected among 153 strains on the basis of their potential technological properties isolated from traditional Çubuk pickles were examined in vitro. For this purpose, these strains (21 Lactobacillus plantarum, 11 Pediococcus ethanolidurans, and 7 Lactobacillus brevis were tested for the ability to survive at pH 2.5, resistance to bile salts, viability in the presence of pepsin-pancreatin, ability to deconjugate bile salts, cholesterol assimilation, and surface hydrophobicity properties. Most of the properties tested could be assumed to be strain-dependent. However, L. plantarum and L. brevis species were found to possess desirable probiotic properties to a greater extent compared to P. ethanolidurans. In contrast to P. ethanolidurans strains, the tested L. plantarum and L. brevis strains exhibited bile salt tolerance, albeit to different extent. All tested strains showed less resistance to intestinal conditions than gastric juice environment. Based on the survival under gastrointestinal conditions, 22 of the 39 strains were selected for further characterization. The eight strains having the highest cholesterol assimilation and surface hydrophobicity ratios could be taken as promising probiotic candidates for further in vivo studies, because of the strongest variations found among the tested strains with regard to these properties.

  18. The Influence of Glutamate on Axonal Compound Action Potential In Vitro.

    Science.gov (United States)

    Abouelela, Ahmed; Wieraszko, Andrzej

    2016-01-01

    Background  Our previous experiments demonstrated modulation of the amplitude of the axonal compound action potential (CAP) by electrical stimulation. To verify assumption that glutamate released from axons could be involved in this phenomenon, the modification of the axonal CAP induced by glutamate was investigated. Objectives  The major objective of this research is to verify the hypothesis that axonal activity would trigger the release of glutamate, which in turn would interact with specific axonal receptors modifying the amplitude of the action potential. Methods  Segments of the sciatic nerve were exposed to exogenous glutamate in vitro, and CAP was recorded before and after glutamate application. In some experiments, the release of radioactive glutamate analog from the sciatic nerve exposed to exogenous glutamate was also evaluated. Results  The glutamate-induced increase in CAP was blocked by different glutamate receptor antagonists. The effect of glutamate was not observed in Ca-free medium, and was blocked by antagonists of calcium channels. Exogenous glutamate, applied to the segments of sciatic nerve, induced the release of radioactive glutamate analog, demonstrating glutamate-induced glutamate release. Immunohistochemical examination revealed that axolemma contains components necessary for glutamatergic neurotransmission. Conclusion  The proteins of the axonal membrane can under the influence of electrical stimulation or exogenous glutamate change membrane permeability and ionic conductance, leading to a change in the amplitude of CAP. We suggest that increased axonal activity leads to the release of glutamate that results in changes in the amplitude of CAPs.

  19. Potential of two Metschnikowia pulcherrima (yeast) strains for in vitro biodegradation of patulin.

    Science.gov (United States)

    Reddy, K R N; Spadaro, D; Gullino, M L; Garibaldi, A

    2011-01-01

    Patulin contamination of apple and other fruit-based foods and beverages is an important food safety issue, as consumption of these commodities throughout the world is great. Studies are therefore necessary to reduce patulin levels to acceptable limits or undetectable levels to minimize toxicity. This study was undertaken to investigate the efficacy of two Metschnikowia pulcherrima strains (MACH1 and GS9) on biodegradation of patulin under in vitro conditions. These yeast strains were tested for their abilities to degrade patulin in liquid medium amended with 5, 7.5, 10, and 15 μg/ml patulin and a yeast cell concentration of 1 × 10(8) cells per ml at 25°C. Of the two strains tested, MACH1 completely (100%) reduced patulin levels within 48 h, and GS9 within 72 h, at all concentrations of patulin. MACH1 effectively degraded the patulin within 24 h by 83 to 87.4%, and GS9 by 73 to 75.6% at 48 h, regardless of concentration. Patulin was not detected in yeast cell walls. This indicates that yeast cell walls did not absorb patulin, and that they completely degraded the toxin. Patulin had no influence on yeast cell concentration during growth. Therefore, these yeast strains could potentially be used for the reduction of patulin in naturally contaminated fruit juices. To our knowledge, this is the first report regarding the potential of M. pulcherrima strains for patulin biodegradation.

  20. In silico and in vitro prediction of gastrointestinal absorption from potential drug eremantholide C.

    Science.gov (United States)

    Caldeira, Tamires G; Saúde-Guimarães, Dênia A; Dezani, André B; Serra, Cristina Helena Dos Reis; de Souza, Jacqueline

    2017-11-01

    Analysis of the biopharmaceutical properties of eremantholide C, sesquiterpene lactone with proven pharmacological activity and low toxicity, is required to evaluate its potential to become a drug. Preliminary analysis of the physicochemical characteristics of eremantholide C was performed in silico. Equilibrium solubility was evaluated using the shake-flask method, at 37.0 °C, 100 rpm during 72 h in biorelevant media. The permeability was analysed using parallel artificial membrane permeability assay, at 37.0 °C, 50 rpm for 5 h. The donor compartment was composed of an eremantholide C solution in intestinal fluid simulated without enzymes, while the acceptor compartment consisted of phosphate buffer. Physicochemical characteristics predicted in silico indicated that eremantholide C has a low solubility and high permeability. In-vitro data of eremantholide C showed low solubility, with values for the dose/solubility ratio (ml): 9448.82, 10 389.61 e 15 000.00 for buffers acetate (pH 4.5), intestinal fluid simulated without enzymes (pH 6.8) and phosphate (pH 7.4), respectively. Also, it showed high permeability, with effective permeability of 30.4 × 10(-6) cm/s, a higher result compared with propranolol hydrochloride (9.23 × 10(-6) cm/s). The high permeability combined with its solubility, pharmacological activity and low toxicity demonstrate the importance of eremantholide C as a potential drug candidate. © 2017 Royal Pharmaceutical Society.

  1. The erosive potential of some alcopops using bovine enamel: an in vitro study.

    Science.gov (United States)

    Ablal, M A; Kaur, J S; Cooper, L; Jarad, F D; Milosevic, A; Higham, S M; Preston, A J

    2009-11-01

    Alcoholic soft drinks have become increasingly popular and have high concentrations of citric acid and alcohol so might have the potential to cause dental erosion. This study aimed to investigate the erosive potential of alcopops on bovine enamel in vitro. Six bovine upper incisors were prepared and sectioned to give six slabs per tooth, 4mm x 4mm each. Each slab was covered with nail varnish, leaving an exposed window (2mm x 2mm). Samples were immersed in 20ml of each of the test solutions for 20min, 1h, and 24h under gentle agitation (100rpm). Enamel surface loss was determined using Quantitative Laser Fluorescence (QLF), Non-contact Profilometry (NCP) and Transverse Microradiography (TMR). Enamel loss occurred with all test drinks and the positive control (p<0.05), and the depth of lesion correlated with pH and time. No significant difference was observed between 20min and 1h exposure, although both times had significantly (p<0.05) greater erosion when compared with baseline. Within each alcopops group significant erosion had occurred at 24h exposure compared with the baseline and previous times. All the tested alcopops resulted in significant enamel loss at 24h (p<0.001) with direct correlation between degree of enamel loss and both pH and increasing exposure time.

  2. In vitro antibacterial potential of Eugenia jambolana seed extracts against multidrug-resistant human bacterial pathogens.

    Science.gov (United States)

    Bag, Anwesa; Bhattacharyya, Subir Kumar; Pal, Nishith Kumar; Chattopadhyay, Rabi Ranjan

    2012-06-20

    The present study was carried out to evaluate the possible in vitro antibacterial potential of extracts of Eugenia jambolana seeds against multidrug-resistant human bacterial pathogens. Agar well diffusion and microbroth dilution assay methods were used for antibacterial susceptibility testing. Kill-kinetics study was done to know the rate and extent of bacterial killing. Phytochemical analysis and TLC-bioautography were performed by colour tests to characterize the putative compounds responsible for this antibacterial activity. Cytotoxic potential was evaluated on human erythrocytes by haemolytic assay method and acute oral toxicity study was done in mice. The plant extracts demonstrated varying degrees of strain specific antibacterial activity against all the test isolates. Further, ethyl acetate fraction obtained from fractionation of most active ethanol extract showed maximum antibacterial effect against all the test isolates. Phytochemical analysis and TLC-bioautography of ethyl acetate fraction revealed that phenolics were the major active phytoconstituents. Ethyl acetate fraction also demonstrated no haemolytic activity on human erythrocytes and no gross behavioural changes as well as toxic symptoms were observed in mice at recommended dosage level. The results provide justification for the use of E. jambolana in folk medicine to treat various infectious diseases and may contribute to the development of novel antimicrobial agents for the treatment of infections caused by these drug-resistant bacterial pathogens. Copyright © 2012 Elsevier GmbH. All rights reserved.

  3. Antidiabetes and antihypertension potential of commonly consumed carbohydrate sweeteners using in vitro models.

    Science.gov (United States)

    Ranilla, Lena Galvez; Kwon, Young-In; Genovese, Maria Ines; Lajolo, Franco Maria; Shetty, Kalidas

    2008-06-01

    Commonly consumed carbohydrate sweeteners derived from sugar cane, palm, and corn (syrups) were investigated to determine their potential to inhibit key enzymes relevant to Type 2 diabetes and hypertension based on the total phenolic content and antioxidant activity using in vitro models. Among sugar cane derivatives, brown sugars showed higher antidiabetes potential than white sugars; nevertheless, no angiotensin I-converting enzyme (ACE) inhibition was detected in both sugar classes. Brown sugar from Peru and Mauritius (dark muscovado) had the highest total phenolic content and 1,1-diphenyl-2-picrylhydrazyl radical scavenging activity, which correlated with a moderate inhibition of yeast alpha-glucosidase without showing a significant effect on porcine pancreatic alpha-amylase activity. In addition, chlorogenic acid quantified by high-performance liquid chromatography was detected in these sugars (128 +/- 6 and 144 +/- 2 microg/g of sample weight, respectively). Date sugar exhibited high alpha-glucosidase, alpha-amylase, and ACE inhibitory activities that correlated with high total phenolic content and antioxidant activity. Neither phenolic compounds or antioxidant activity was detected in corn syrups, indicating that nonphenolic factors may be involved in their significant ability to inhibit alpha-glucosidase, alpha-amylase, and ACE. This study provides a strong biochemical rationale for further in vivo studies and useful information to make better dietary sweetener choices for Type 2 diabetes and hypertension management.

  4. Potential of Ruminant Feed with Appearance of In Vitro Gas Production

    Directory of Open Access Journals (Sweden)

    Firsoni

    2017-10-01

    Full Text Available Indonesia has many kinds of feedstuff with different qualities that can be used as ruminants feed. One way to evaluate it is analyzing the performance of the feed gas production value in vitro. Feed ingredients tested in this study were the flour of coffee hull, peanut hull, field grass, turn leaves, rice straw and fermented rice straw. Samples weighed 200 ± 5 mg, put into a 100 ml syringe glass, added 30 ml buffalo rumen liquor with bicarbonate buffer medium, then incubated in the water bath at 39 ° C for 48 hours. Neway software and random block design with 4 blocks are used to calculate the value of fitted gas and to analyze the variance. The variables measured were gas production 2, 4, 6, 8, 10, 12, 24, 48, 72 and 96 hours, organic material degradable, gas production potential (a+b, gas production rate (k. The highest gas production 24, 48 and 72 hours from the field grass treatment (C was 36.33, 51.12 and 56.29 ml/200 mg DM but 96 hours of rice straw ie 59.60 ml/200 mg DM, while the lowest (24, 48, 72 and 96 hours of coffee skin (6.08, 7.77, 7.61, and 7.68 ml/200 mg DM respectively. The highest gas production potential of rice straw is 69.13 ml/200 mg DM and the lowest of coffee skin is 7.72 ml/200 mg BK. The highest percentage of gas production after 24 hours was obtained Turi leaves (D: 91.46% and the lowest rice straw (E: 41.22%. Rice straw can be suggested to be field grass substitution by processed again to reduce its crude fiber content, while the coffee and peanut hulls need further study, due to low gas production potential of 7.72 and 11.45 ml / 200 mg DM.

  5. Potential of D-Octaarginine-Linked Polymers as an in Vitro Transfection Tool for Biomolecules.

    Science.gov (United States)

    Mohri, Kohta; Morimoto, Naoki; Maruyama, Megumi; Nakamoto, Norimasa; Hayashi, Emi; Nagata, Kengo; Miyata, Kohei; Ochiai, Kyohei; Hiwatari, Ken-ichiro; Tsubaki, Kazufumi; Tobita, Etsuo; Ishimaru, Yuki; Maeda, Sadaaki; Sakuma, Shinji

    2015-08-19

    We have been investigating the potential use of cell-penetrating peptide-linked polymers as a novel penetration enhancer. Since previous in vivo studies demonstrated that poly(N-vinylacetamide-co-acrylic acid) bearing D-octaarginine, a typical cell-penetrating peptide, enhanced membrane permeation of biomolecules, its potential as an in vitro transfection tool was evaluated in this study. A plasmid DNA encoding green fluorescent protein (pGFP-C1), β-galactosidase, and bovine serum albumin (BSA) were used as model biomolecules. Anionic pGFP-C1 interacted electrostatically with cationic d-octaarginine-linked polymers. When the ratio of mass concentration of polymers to that of pGFP-C1 reached 2.5, complexes whose size and zeta potential were approximately 200 nm and 15 mV, respectively, were obtained. GFP expression was observed in cells incubated with complexes prepared under conditions in which the polymer/pDNA concentration ratio exceeded 2.5. The expression level elevated with an increase in the concentration ratio, but physicochemical properties of the complexes remained unchanged. Results suggested that free polymers contributed to pGFP-C1 internalization. Another cell study demonstrated that β-galactosidase premixed with polymers was taken up into cells in its active tetrameric form. Similar electrostatic interaction-driven complex formation was observed for BSA charged negatively in neutral solution. However, it appeared that the internalization processes of BSA differed from those of pGFP-C1. A mass concentration-dependent increase in internalized BSA was observed, irrespective of the polymer/protein concentration ratio. Due to frail interactions, polymers that were released from the complexes and subsequently immobilized on cell membranes might also contribute to membrane permeation of BSA.

  6. In Vitro Study on the Antioxidant Potentials of the Leaves and Fruits of Nauclea latifolia

    Science.gov (United States)

    Ayeleso, Ademola O.; Oguntibeju, Oluwafemi O.; Brooks, Nicole L.

    2014-01-01

    This study was carried out to investigate the in vitro antioxidant potentials of the leaves and fruits of Nauclea latifolia, a straggling shrub or small tree, native to tropical Africa and Asia. Hot water extracts of the leaves and fruits of Nauclea latifolia were assessed for their total polyphenolic, flavanol, and flavonol contents as well as 1-diphenyl-2-picrylhydrazyl (DPPH) scavenging ability, ferric reducing antioxidant power (FRAP), Trolox equivalence antioxidant capacity (TEAC), and oxygen radical absorbance capacity (ORAC) assays. The aqueous extract of the leaves was found to contain higher level of total polyphenols (11.63 ± 0.023 mg GAE/g), flavanol (1.45 ± 0.10 mg CE/g), and flavonol (2.22 ± 0.37 mg QE/g) than the extract of the fruits with values of 1.75 ± 0.02 mg GAE/g (total polyphenol), 0.15 ± 0.01 mg CE/g (flavanol), and 1.00 ± 0.13 mg QE/g (flavonol). Similarly, the aqueous extract of the leaves also exhibited higher DPPH (IC50 20.64 mg/mL), FRAP (86.10 ± 3.46 μmol AAE/g), TEAC (94.83 ± 3.57 μmol TE/g), and ORAC (196.55 ± 0.073 μmol TE/g) than the extract of the fruits with DPPH (IC50 120.33 mg/mL), FRAP (12.23 ± 0.40 μmol AAE/g), TEAC (12.48 ± 0.21 μmol TE/g), and ORAC (58.88 ± 0.073 μmol TE/g). The present study showed that Nauclea latifolia has strong antioxidant potentials with the leaves demonstrating higher in vitro antioxidant activities than the fruits. PMID:25013856

  7. Comparison of the binding potential of various diisocyanates on DNA in vitro.

    Science.gov (United States)

    Peel, M; Marczynski, B; Baur, X

    1997-12-26

    Inhalation of diisocyanate vapors is associated with immediate-type hypersensitivity reactions and direct toxic responses. The genotoxic effects of diisocyanates have not been clarified. The aim of this study was to examine the changes in DNA following in vitro exposure to three most commonly used diisocyanates (toluene diisocyanate, TDI; methylenediphenyl-4,4'-diisocyanate, MDI; and hexamethylene diisocyanate, HDI) and to compare their binding potential using melting behavior of DNA and electrophoresis studies in DNA. Following incubation of DNA with MDI (pure and mix) and HDI we found no differences in the melting behavior compared to the control calf thymus DNA. However, DNA treated with TDI showed differences in the shape of the native DNA curves due to changes in hyperchromicity and exhibited 14% more DNA reconstitution after renaturation. The small changes in the melting behavior of native DNA do not suggest the formation of DNA intrastrand cross-links but rather conformational changes of single- and double-stranded DNA. These conformational changes were further explored by agarose electrophoresis of native and denatured calf thymus DNA. Control and all diisocyanate-exposed DNA showed no differences in the size of native DNA fragments. Conversely, electrophoresis of TDI mix-incubated DNA, following denaturation, showed a distinct reduction in the double-stranded DNA fragment size compared to the control, MDI-denatured (pure and mix), and HDI-denatured DNA. These findings may help to better understand the mechanisms of the genotoxic effect of TDI.

  8. Synaptic potentiation facilitates memory-like attractor dynamics in cultured in vitro hippocampal networks.

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    Mark Niedringhaus

    Full Text Available Collective rhythmic dynamics from neurons is vital for cognitive functions such as memory formation but how neurons self-organize to produce such activity is not well understood. Attractor-based computational models have been successfully implemented as a theoretical framework for memory storage in networks of neurons. Additionally, activity-dependent modification of synaptic transmission is thought to be the physiological basis of learning and memory. The goal of this study is to demonstrate that using a pharmacological treatment that has been shown to increase synaptic strength within in vitro networks of hippocampal neurons follows the dynamical postulates theorized by attractor models. We use a grid of extracellular electrodes to study changes in network activity after this perturbation and show that there is a persistent increase in overall spiking and bursting activity after treatment. This increase in activity appears to recruit more "errant" spikes into bursts. Phase plots indicate a conserved activity pattern suggesting that a synaptic potentiation perturbation to the attractor leaves it unchanged. Lastly, we construct a computational model to demonstrate that these synaptic perturbations can account for the dynamical changes seen within the network.

  9. Effect of gibberellic acid on germination potential in vitro seed Carica quercifolia (St. Hil.. Hieron. (Caricaceae

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    T. Gerber

    2014-03-01

    Full Text Available The species Carica quercifolia (St. Hil.. Hieron. (Caricaceae, native to the region west of Santa Catarina has ornamental potential and fruitful, however, its seeds have low germination rates. Thus, the objective was to evaluate the effect of gibberellic acid (GA on seed germination of C. quercifolia in vitro. Initially, the seeds were subjected to an aseptic treatment with sodium hypochlorite and 70% alcohol, then subjected to five different treatments with gibberellic acid (control, 50, 100, 150 and 200 mg.L-1 and subsequently inoculatedinoculated on MS medium. Each treatment had 12 replications and six sub-repetitions. After inoculation, seeds were kept in a germination chamber with a photoperiod of 16 hours (50 mol photons m-² s-¹ and temperature of 26 ± 2 ° C. The onset of germination was observed at day 35 of culture only in treatments with 100 and 200 mg L-1 GA. After 42 days of cultivation, there was low germination rate in both control seeds and seeds treated with GA, and there were no statistically significant differences. The results suggest that the concentrations of GA were used possibly lower, unable to induce the germination C. quercifolia.

  10. Folic acid modified gelatine coated quantum dots as potential reagents for in vitro cancer diagnostics

    LENUS (Irish Health Repository)

    Gerard, Valerie A

    2011-11-10

    Abstract Background Gelatine coating was previously shown to effectively reduce the cytotoxicity of CdTe Quantum Dots (QDs) which was a first step towards utilising them for biomedical applications. To be useful they also need to be target-specific which can be achieved by conjugating them with Folic Acid (FA). Results The modification of QDs with FA via an original "one-pot" synthetic route was proved successful by a range of characterisation techniques including UV-visible absorption spectroscopy, Photoluminescence (PL) emission spectroscopy, fluorescence life-time measurements, Transmission Electron Microscopy (TEM) and Dynamic Light Scattering (DLS). The resulting nanocomposites were tested in Caco-2 cell cultures which over-express FA receptors. The presence of FA on the surface of QDs significantly improved the uptake by targeted cells. Conclusions The modification with folic acid enabled to achieve a significant cellular uptake and cytotoxicity towards a selected cancer cell lines (Caco-2) of gelatine-coated TGA-CdTe quantum dots, which demonstrated good potential for in vitro cancer diagnostics.

  11. Summary of: an in vitro investigation of the erosive potential of smoothies.

    Science.gov (United States)

    Brand, H S

    2013-02-01

    Recent health promotion campaigns have encouraged the public to consume at least five portions of fruit and vegetables per day. Many see consuming fruit smoothies as a way of achieving this. To ascertain the potential or otherwise for fruit smoothies to bring about dental erosion. Laboratory study. This was an in vitro investigation in which five varieties of shop bought fruit smoothies, including a 'thickie' were investigated, with respect to their initial pH, titratable acidity and effect upon exposure to the surface microhardness and profile of extracted human teeth. In addition their performance was compared to negative (Volvic™ water) and positive (orange juice) control drinks as well as a homemade smoothie, based upon the recipe of one of the commercially bought drinks, from which ingredient omissions were made. The majority of the drinks investigated had a baseline pH below the critical pH of enamel (5.5) and required comparable volumes of 0.1M NaOH to raise their pH to neutrality as the positive control. Only two drinks (Volvic™ still mineral water, the negative control, and the yoghurt, vanilla bean and honey 'thickie') displayed a higher pH, though to neutralise the thickie, a lesser quantity of alkali addition was required. The immersion of the tooth samples in the drinks brought about reductions in their surface hardness (expressed as a percentage change of median hardness) but these were only significant (p benefits of their consumption, their consumption should be confined to mealtimes.

  12. Antioxidant properties of potentially probiotic bacteria: in vitro and in vivo activities.

    Science.gov (United States)

    Amaretti, Alberto; di Nunzio, Mattia; Pompei, Anna; Raimondi, Stefano; Rossi, Maddalena; Bordoni, Alessandra

    2013-01-01

    Thirty-four strains of lactic acid bacteria (seven Bifidobacterium, 11 Lactobacillus, six Lactococcus, and 10 Streptococcus thermophilus) were assayed in vitro for antioxidant activity against ascorbic and linolenic acid oxidation (TAA(AA) and TAA(LA)), trolox-equivalent antioxidant capacity (TEAC), intracellular glutathione (TGSH), and superoxide dismutase (SOD). Wide dispersion of each of TAA(AA), TAA(LA), TEAC, TGSH, and SOD occurred within bacterial groups, indicating that antioxidative properties are strain specific. The strains Bifidobacterium animalis subsp. lactis DSMZ 23032, Lactobacillus acidophilus DSMZ 23033, and Lactobacillus brevis DSMZ 23034 exhibited among the highest TAA(AA), TAA(LA), TEAC, and TGSH values within the lactobacilli and bifidobacteria. These strains were used to prepare a potentially antioxidative probiotic formulation, which was administered to rats at the dose of 10(7), 10(8), and 10(9) cfu/day for 18 days. The probiotic strains colonized the colon of the rats during the trial and promoted intestinal saccharolytic metabolism. The analysis of plasma antioxidant activity, reactive oxygen molecules level, and glutathione concentration, revealed that, when administered at doses of at least 10(8) cfu/day, the antioxidant mixture effectively reduced doxorubicin-induced oxidative stress. Probiotic strains which are capable to limit excessive amounts of reactive radicals in vivo may contribute to prevent and control several diseases associated with oxidative stress.

  13. In vitro corrosion properties and cytocompatibility of Fe-Ga alloys as potential biodegradable metallic materials.

    Science.gov (United States)

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

    2017-02-01

    The in vitro biodegradable properties and cytocompatibility of Fe-Ga alloys including Fe81Ga19, (Fe81Ga19)98B2 and (Fe81Ga19)99.5(TaC)0.5, and pure Fe were investigated for biomedical applications. The microstructure of the alloys was characterized using X-ray diffraction spectroscopy and optical microscopy. The results showed that A2 and D03 phases were detected for the three types of Fe-Ga alloys, and additional Fe2B and TaC phases were found in the (Fe81Ga19)98B2 and (Fe81Ga19)99.5(TaC)0.5 alloys, respectively. The corrosion rates of the Fe-Ga alloys were higher than that of pure Fe, as demonstrated by both potentiodynamic polarization measurements and immersion tests in simulated body fluid. The alloying element Ga lowered the corrosion potential of the Fe matrix and made it more susceptible to corrosion. Severe pitting corrosion developed on the surface of the Fe81Ga19 alloy after the addition of ternary B or TaC due to the multi-phase microstructures. The MC3T3-E1 cells exhibited good adhesion and proliferation behavior on the surfaces of the Fe-Ga alloys after culture for 4h and 24h. Copyright © 2016 Elsevier B.V. All rights reserved.

  14. In vitro probiotic potential of Lactobacillus spp. isolated from fermented milks

    Directory of Open Access Journals (Sweden)

    A.F. Cunha

    2013-12-01

    Full Text Available The potential of in vitro probiotic Lactobacillus spp. was evaluated in fermented milks marketed in Belo Horizonte, MG, Brazil. Of the samples analyzed, 86.7% had at least 10(6 CFU/mL of Lactobacillus spp., complying with the Brazilian quality standards for fermented milks. Furthermore, 56.7% had minimum count ranging from 10(8 to 10(9 CFU/mL, which is in accordance with legal parameters. The remaining 43.3% would not be able to satisfactorily guarantee benefits to consumers. The amount of Lactobacillus spp. varied between batches of products, which may indicate failures in monitoring during manufacture, transport or storage. All strains of Lactobacillus spp. showed some inhibitory activity against the indicator microorganisms, being more pronounced against pathogenic microorganisms than against non-pathogenic (P<0.05. Samples of Lactobacillus spp. showed different profiles of antimicrobial susceptibility, with an occurrence of cases of multidrug resistance. All strains tested showed sensitivity to bile salts (0.3% and resistance to gastric pH (2.0. Lactobacillus spp. of commercial fermented milks should be present in higher amounts in some brands, be resistant to bile salts and have no multiple resistance to antimicrobials.

  15. In vitro proliferation and differentiation of hepatic oval cells and their potential capacity for intrahepatic transplantation

    Energy Technology Data Exchange (ETDEWEB)

    Li, Z.; Chen, J. [Liaocheng People' s Hospital, Department of Hepatobiliary Surgery, Liaocheng, Shandong, China, Department of Hepatobiliary Surgery, Liaocheng People’s Hospital, Liaocheng, Shandong (China); Li, L.; Ran, J.H.; Liu, J. [The First People' s Hospital of Kunming, Kunming, Yunnan, China, The First People’s Hospital of Kunming, Kunming, Yunnan (China); Gao, T.X.; Guo, B.Y. [Dongchangfu Hospital of Women and Child Health Care, Liaocheng, Shandong (China); Li, X.H.; Liu, Z.H.; Liu, G.J.; Gao, Y.C.; Zhang, X.L. [Liaocheng People' s Hospital, Department of Hepatobiliary Surgery, Liaocheng, Shandong, China, Department of Hepatobiliary Surgery, Liaocheng People’s Hospital, Liaocheng, Shandong (China)

    2013-07-30

    Hepatic oval cells (HOCs) are recognized as facultative liver progenitor cells that play a role in liver regeneration after acute liver injury. Here, we investigated the in vitro proliferation and differentiation characteristics of HOCs in order to explore their potential capacity for intrahepatic transplantation. Clusters or scattered HOCs were detected in the portal area and interlobular bile duct in the liver of rats subjected to the modified 2-acetylaminofluorene and partial hepatectomy method. Isolated HOCs were positive for c-kit and CD90 staining (99.8% and 88.8%, respectively), and negative for CD34 staining (3.6%) as shown by immunostaining and flow cytometric analysis. In addition, HOCs could be differentiated into hepatocytes and bile duct epithelial cells after leukemia inhibitory factor deprivation. A two-cuff technique was used for orthotopic liver transplantation, and HOCs were subsequently transplanted into recipients. Biochemical indicators of liver function were assessed 4 weeks after transplantation. HOC transplantation significantly prolonged the median survival time and improved the liver function of rats receiving HOCs compared to controls (P=0.003, Student t-test). Administration of HOCs to rats also receiving liver transplantation significantly reduced acute allograft rejection compared to control liver transplant rats 3 weeks following transplantation (rejection activity index score: control=6.3±0.9; HOC=3.5±1.5; P=0.005). These results indicate that HOCs may be useful in therapeutic liver regeneration after orthotopic liver transplantation.

  16. Folic acid modified gelatine coated quantum dots as potential reagents for in vitro cancer diagnostics.

    Science.gov (United States)

    Gérard, Valérie A; Maguire, Ciaran M; Bazou, Despina; Gun'ko, Yurii K

    2011-11-10

    Gelatine coating was previously shown to effectively reduce the cytotoxicity of CdTe Quantum Dots (QDs) which was a first step towards utilising them for biomedical applications. To be useful they also need to be target-specific which can be achieved by conjugating them with Folic Acid (FA). The modification of QDs with FA via an original "one-pot" synthetic route was proved successful by a range of characterisation techniques including UV-visible absorption spectroscopy, Photoluminescence (PL) emission spectroscopy, fluorescence life-time measurements, Transmission Electron Microscopy (TEM) and Dynamic Light Scattering (DLS). The resulting nanocomposites were tested in Caco-2 cell cultures which over-express FA receptors. The presence of FA on the surface of QDs significantly improved the uptake by targeted cells. The modification with folic acid enabled to achieve a significant cellular uptake and cytotoxicity towards a selected cancer cell lines (Caco-2) of gelatine-coated TGA-CdTe quantum dots, which demonstrated good potential for in vitro cancer diagnostics.

  17. Folic acid modified gelatine coated quantum dots as potential reagents for in vitro cancer diagnostics

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    Gérard Valérie A

    2011-11-01

    Full Text Available Abstract Background Gelatine coating was previously shown to effectively reduce the cytotoxicity of CdTe Quantum Dots (QDs which was a first step towards utilising them for biomedical applications. To be useful they also need to be target-specific which can be achieved by conjugating them with Folic Acid (FA. Results The modification of QDs with FA via an original "one-pot" synthetic route was proved successful by a range of characterisation techniques including UV-visible absorption spectroscopy, Photoluminescence (PL emission spectroscopy, fluorescence life-time measurements, Transmission Electron Microscopy (TEM and Dynamic Light Scattering (DLS. The resulting nanocomposites were tested in Caco-2 cell cultures which over-express FA receptors. The presence of FA on the surface of QDs significantly improved the uptake by targeted cells. Conclusions The modification with folic acid enabled to achieve a significant cellular uptake and cytotoxicity towards a selected cancer cell lines (Caco-2 of gelatine-coated TGA-CdTe quantum dots, which demonstrated good potential for in vitro cancer diagnostics.

  18. In vitro evaluation of the erosive potential of viscosity-modified soft acidic drinks on enamel.

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    Aykut-Yetkiner, Arzu; Wiegand, Annette; Ronay, Valerie; Attin, Rengin; Becker, Klaus; Attin, Thomas

    2014-04-01

    The objective of this in vitro study was to investigate the effect of viscosity-modified soft acidic drinks on enamel erosion. A total of 108 bovine enamel samples (∅ = 3 mm) were embedded in acrylic resin and allocated into six groups (n = 18). Soft acidic drinks (orange juice, Coca-Cola, Sprite) were used both in their regular forms and at a kinetic viscositiy of 5 mm(2)/s, which was adjusted by adding hydroxypropyl cellulose. All solutions were pumped over the enamel surface from a reservoir with a drop rate of 3 ml/min. Each specimen was eroded for 10 min at 20 °C. Erosion of enamel surfaces was measured using profilometry. Data were analyzed using independent t tests and one-way ANOVAs (p viscosity-modified drinks (Coca-Cola, 4.90 ± 0.34 μm; Sprite, 4.46 ± 0.39 μm; orange juice, 1.10 ± 0.22 μm). For both regular and viscosity-modified forms, Coca-Cola and Sprite caused higher enamel loss than orange juice. Increasing the viscosity of acidic soft drinks to 5 mm(2)/s reduced enamel erosion by 12.6-18.7 %. The erosive potential of soft acidic drinks is not only dependent on various chemical properties but also on the viscosity of the acidic solution and can be reduced by viscosity modification.

  19. Synthesis of protocatechuic acid grafted chitosan copolymer: structure characterization and in vitro neuroprotective potential.

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    Xu, Chao; Guan, Shui; Wang, Bo; Wang, Shuping; Wang, Yuxin; Sun, Changkai; Ma, Xuehu; Liu, Tianqing

    2017-12-05

    Excessive free radicals can cause oxidative damage to human tissues, which results in a variety of diseases. Therefore, the development of antioxidant materials is one of the great projects in biomedical field. In this work, antioxidant protocatechuic acid (PCA) monomers were grafted onto chitosan (CS) backbones to develop a PCA grafted chitosan (PCA-g-CS) antioxidant copolymer via the method of free radical-induced grafting reaction. The formation of covalent bonds between PCA and CS were confirmed by FTIR, 1H NMR, XRD and UV-Vis. The antioxidant activity of PCA-g-CS was analyzed by 2, 2-diphenyl-1-picrylhydrazyl (DPPH) and hydroxyl radical scavenging assays. In addition, the cytotoxicity of PCA-g-CS on neuron-like rat phaeochromocytoma (PC12) cells was evaluated by using MTT assay. The neuroprotective effects against hydrogen peroxide (H2O2) and L-glutamic acid (GLU) induced apoptosis in PC12 cells were also investigated. Our results demonstrated that the PCA-g-CS antioxidant copolymer had the ability to scavenge DPPH and hydroxyl radical in vitro. Furthermore, the PCA-g-CS was biocompatible and had neuroprotective effects against free radical-induced apoptosis in PC12 cells. This PCA-g-CS copolymer is firstly synthesized for neuroprotection and the results suggest the PCA-g-CS may be a potential antioxidant material in the treatment of oxidative damage related diseases. Copyright © 2017. Published by Elsevier B.V.

  20. Surface-Modified Liposomal Formulation of Amphotericin B: In vitro Evaluation of Potential Against Visceral Leishmaniasis.

    Science.gov (United States)

    Patere, Shilpa N; Pathak, Pankaj O; Kumar Shukla, Anil; Singh, Rajesh Kumar; Kumar Dubey, Vikash; Mehta, Miten J; Patil, Anand G; Gota, Vikram; Nagarsenker, Mangal S

    2017-04-01

    Surface modification of liposomes with targeting ligands is known to improve the efficacy with reduced untoward effects in treating infective diseases like visceral leishmaniasis (VL). In the present study, modified ligand (ML), designed by modifying polysaccharide with a long chain lipid was incorporated in liposomes with the objective to target amphotericin B (Amp B) to reticuloendothelial system and macrophages. Conventional liposomes (CL) and surface modified liposomes (SML) were characterized for size, shape, and entrapment efficiency (E.E.). Amp B SML with 3% w/w of ML retained the vesicular nature with particle size of ∼205 nm, E.E. of ∼95% and good stability. SML showed increased cellular uptake in RAW 264.7 cells which could be attributed to receptor-mediated endocytosis. Compared to Amp B solution, Amp B liposomes exhibited tenfold increased safety in vitro in RAW 264.7 and J774A.1 cell lines. Pharmacokinetics and biodistribution studies revealed high t 1/2, area under the curve (AUC)0-24, reduced clearance and prolonged retention in liver and spleen with Amp B SML compared to other formulations. In promastigote and amastigote models, Amp B SML showed enhanced performance with low 50% inhibitory concentration (IC50) compared to Amp B solution and Amp B CL. Thus, due to the targeting ability of ML, SML has the potential to achieve enhanced efficacy in treating VL.

  1. In vitro toxicity of reuterin, a potential food biopreservative.

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    Fernández-Cruz, María L; Martín-Cabrejas, Izaskun; Pérez-Del Palacio, José; Gaya, Pilar; Díaz-Navarro, Caridad; Navas, José M; Medina, Margarita; Arqués, Juan L

    2016-10-01

    Reuterin has a high potential as a food preservative due to both its chemical characteristics and its antimicrobial activity against food-borne pathogens and spoilage bacteria. However, there is a lack of information about its toxicity and its capacity to interfere with the metabolism of drugs by inhibiting cytochrome P450 (CYP) activity. The results of this study indicated that reuterin exhibited a moderate cytotoxicity in the human hepatoma cell line HepG2 according to assays measuring three different endpoints in the same set of cells. Reuterin was much less toxic than acrolein and only four times more toxic than diacetyl, a generally recognized as safe flavoring compound. In vitro experiments utilizing human liver microsomes showed that reuterin presents low possibility of displaying in vivo drug interactions by inhibition of CYP3A4, CYP2D6, and CYP2C9. Therefore, reuterin can be considered a promising food biopreservative, although additional toxicology research is needed before permission for use can be granted. Copyright © 2016 Elsevier Ltd. All rights reserved.

  2. Hepatotoxic potential of asarones: In vitro evaluation of hepatotoxicity and quantitative determination in herbal products

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    Dhavalkumar Narendrabha Patel

    2015-02-01

    Full Text Available α and β asarones are natural constituents of some aromatic plants, especially species of the genus Acorus. In addition to beneficial properties of asarones, genotoxicity and carcinogenicity are also reported. Due to potential toxic effects of β-asarone, a limit of exposure from herbal products of approximately 2 μg/kg body weight/day has been set temporarily until a full benefit/risk assessment has been carried out by the European Medicines Agency. Therefore, it is important to monitor levels of β-asarone in herbal products. In this study, we developed a simple, rapid and validated GC-MS method for quantitative determination of asarones and applied it in 20 pediatric herbal products after detecting high concentrations of β-asarone in a product suspected to be implicated in hepatotoxicity in a 3 month old infant. Furthermore, targeted toxicological effects were further investigated in human hepatocytes (THLE-2 cells by employing various in vitro assays, with the goal of elucidating possible mechanisms for the observed toxicity. Results showed that some of the products contained as much as 4 to 25 times greater amounts of β-asarone than the recommended levels. In 4 of 10 samples found to contain asarones, the presence of asarones could not be linked to the labeled ingredients, possibly due to poor quality control. Cell-based investigations in THLE2 cells confirmed the cytotoxicity of -asarone (IC50 = 40.0 ± 2.0 µg/mL which was associated with significant lipid peroxidation and glutathione depletion. This observed cytotoxicity effect is likely due to induction of oxidative stress by asarones. Overall, the results of this study ascertained the usability of this GC-MS method for the quantitative determination of asarones from herbal products, and shed light on the importance of controlling the concentration of potentially toxic asarones in herbal products to safeguard consumer safety. Further investigations of the toxicity of asarones are

  3. Equol, a Dietary Daidzein Gut Metabolite Attenuates Microglial Activation and Potentiates Neuroprotection In Vitro

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    Lalita Subedi

    2017-02-01

    Full Text Available Estrogen deficiency has been well characterized in inflammatory disorders including neuroinflammation. Daidzein, a dietary alternative phytoestrogen found in soy (Glycine max as primary isoflavones, possess anti‐inflammatory activity, but the effect of its active metabolite Equol (7‐hydroxy‐3‐(4′‐hydroxyphenyl‐chroman has not been well established. In this study, we investigated the anti‐neuroinflammatory and neuroprotective effect of Equol in vitro. To evaluate the potential effects of Equol, three major types of central nervous system (CNS cells, including microglia (BV‐2, astrocytes (C6, and neurons (N2a, were used. Effects of Equol on the expression of inducible nitric oxide synthase (iNOS, cyclooxygenase (COX‐2, Mitogen activated protein kinase (MAPK signaling proteins, and apoptosis‐related proteins were measured by western blot analysis. Equol inhibited the lipopolysaccharide (LPS‐induced TLR4 activation, MAPK activation, NF‐kB‐mediated transcription of inflammatory mediators, production of nitric oxide (NO, release of prostaglandin E2 (PGE‐2, secretion of tumor necrosis factor‐α (TNF‐α and interleukin 6 (IL‐6, in Lipopolysaccharide (LPS‐activated murine microglia cells. Additionally, Equol protects neurons from neuroinflammatory injury mediated by LPS‐activated microglia through downregulation of neuronal apoptosis, increased neurite outgrowth in N2a cell and neurotrophins like nerve growth factor (NGF production through astrocytes further supporting its neuroprotective potential. These findings provide novel insight into the anti‐neuroinflammatory effects of Equol on microglial cells, which may have clinical significance in cases of neurodegeneration.

  4. MCH and apomorphine in combination enhance action potential firing of nucleus accumbens shell neurons in vitro

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    F Woodward Hopf

    2013-04-01

    Full Text Available The MCH and dopamine receptor systems have been shown to modulate a number of behaviors related to reward processing, addiction, and neuropsychiatric conditions such as schizophrenia and depression. In addition, MCH and dopamine receptors can interact in a positive manner, for example in the expression of cocaine self-administration. A recent report (Chung et al., 2011a showed that the DA1/DA2 dopamine receptor activator apomorphine suppresses pre-pulse inhibition, a preclinical model for some aspects of schizophrenia. Importantly, MCH can enhance the effects of lower doses of apomorphine, suggesting that co-modulation of dopamine and MCH receptors might alleviate some symptoms of schizophrenia with a lower dose of dopamine receptor modulator and thus fewer potential side effects. Here, we investigated whether MCH and apomorphine could enhance action potential firing in vitro in the nucleus accumbens shell (NAshell, a region which has previously been shown to mediate some behavioral effects of MCH. Using whole-cell patch-clamp electrophysiology, we found that MCH, which has no effect on firing on its own, was able to increase NAshell firing when combined with a subthreshold dose of apomorphine. Further, this MCH/apomorphine increase in firing was prevented by an antagonist of either a DA1 or a DA2 receptor, suggesting that apomorphine acts through both receptor types to enhance NAshell firing. The MCH/apomorphine-mediated firing increase was also prevented by an MCH receptor antagonist or a PKA inhibitor. Taken together, our results suggest that MCH can interact with lower doses of apomorphine to enhance NAshell firing, and thus that MCH and apomorphine might interact in vivo within the NAshell to suppress pre-pulse inhibition.

  5. Antioxidant potential of hydro-methanolic extract of seed of Caesalpinia bonduc: An in vitro study

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    Kishalay Jana

    2011-01-01

    Full Text Available It is well known that the over production of reactive oxygen species is harmful for living organisms and it damages major cellular constituents such as DNA, protein, and lipid. At present, searching of new plant sources having free radical scavenging activity is an important field of research in phytomedicine as natural products are safe and relatively low cost. In this respect, attention has been focused to evaluate the antioxidant potential of hydro-methanolic extract of seed of Caesalpinia bonduc (Caesalpenacae using different in vitro models. To evaluate the antioxidant activity, extract was examined on 2, 2-diphenyl-1-picrylhydrazyl radical scavenging effect, scavenging of hydrogen peroxide, hydroxyl radical scavenging potential, and anti-lipid peroxidation activity by biochemical methods. Total phenol and flavonoids contents in the said extract were measured biochemically as per standard methods. Results were compared with butylated hydroxyl toluene and α-tocopherol. Results indicated that hydro-methanolic extract has strong scavenging activity on 2, 2-diphenyl-1-picrylhydrazyl radical with IC 50 value 157.4 μg/ml, hydroxyl radical with IC 50 value 61.9 μg/ml and hydrogen peroxide with IC 50 value 64.32 μg/ml. Hydro-methanolic extract also showed notable inhibition in lipid peroxidation having IC 50 value 58.87 μg/ml. Phytochemical study focused that the extract is rich in phenolic compounds (24.66 mg gallic acid equivalent/g dried extract and flavonoids (136.65 mg quercetin equivalent/g dried extract. Findings of the experiment indicated that the hydro-methanolic extract of seed of Caesalpinia bonduc is a source of natural antioxidants.

  6. Nutritional content and in vitro antioxidant potential of Garcinia atroviridis (Asam gelugor) leaves and fruits.

    Science.gov (United States)

    Nursakinah, I; Zulkhairi, H A; Norhafizah, M; Hasnah, B; Zamree, Md S; Farrah, Shafeera I; Razif, D; Hamzah, Fansuri H

    2012-12-01

    The objective of this study was to determine antioxidant potential of Garcinia atroviridis leaves and fruits extracts in vitro. Antioxidant activity was assessed using 1,1-diphenyl-2-picrylhydrazyl (DPPH) and ferric reducing antioxidant power (FRAP) assays. Total phenolic content (TPC) of the extracts was estimated as gallic acid equivalent by Folin-Ciocalteau method. Proximate analysis was determined based on the Association of Official Analytical Chemists (AOAC) procedures. Garcinia atroviridis leaves extracted at 100 degrees C/15 min demonstrated the highest TPC value (21.21 +/- 0.28 mg GAE/mg) and was significantly different (p < 0.05) from that of leaves extracted at 60 degrees C/6 h and 40 degrees C/12 h. On the other hand, the fruit extracted at 60 degrees C/6 h showed the highest TPC value (16.23 +/- 0.18 mg GAE/mg) (p < 0.05) compared to the fruit extracted at 40 degrees C/12 h and 100 degrees C/15 h respectively. The antioxidant activities of both samples were positively correlated with the TPC values based on DPPH-radical-scavenging activity and ferric reducing power estimation. Garcinia atroviridis leaf extract contained significantly higher proteins, carbohydrate and ash contents (2.16% +/- 0.08; 15.98% +/- 0.12 and 0.72% +/- 0.07 respectively) than its fruit extract (0.46% +/- 0.08, 8.64% +/- 0.06 and 0.15% +/- 0.06) respectively). The energy content was also found to be higher in the leaf (73.64% +/- 2.15) compared to the fruit (38.38% +/- 1.72) (p < 0.05). The findings indicate that G. atroviridis leaves and fruits have potential for use as a source of natural antioxidants and nutrients for therapeutic purposes against free radical mediated health conditions.

  7. Probiotic potential of Lactobacilli with antagonistic activity against pathogenic strains: An in vitro validation for the production of inhibitory substances

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    Chidre Prabhurajeshwar

    2017-10-01

    Conclusion: Based on the drawn results, T2, T4 and T16 Lactobacillus isolates were recognised as ideal, potential in vitro antimicrobial probiotic isolates against pathogens and studies are needed further in-vivo assessment and human health benefits in their real-life situations.

  8. Combining in vitro protein detection and in vivo antibody detection identifies potential vaccine targets against Staphylococcus aureus during osteomyelitis.

    Science.gov (United States)

    den Reijer, P Martijn; Sandker, Marjan; Snijders, Susan V; Tavakol, Mehri; Hendrickx, Antoni P A; van Wamel, Willem J B

    2017-02-01

    Currently, little is known about the in vivo human immune response against Staphylococcus aureus during a biofilm-associated infection, such as osteomyelitis, and how this relates to protein production in biofilms in vitro. Therefore, we characterized IgG responses in 10 patients with chronic osteomyelitis against 50 proteins of S. aureus, analyzed the presence of these proteins in biofilms of the infecting isolates on polystyrene (PS) and human bone in vitro, and explored the relation between in vivo and in vitro data. IgG levels against 15 different proteins were significantly increased in patients compared to healthy controls. Using a novel competitive Luminex-based assay, eight of these proteins [alpha toxin, Staphylococcus aureus formyl peptide receptor-like 1 inhibitor (FlipR), glucosaminidase, iron-responsive surface determinants A and H, the putative ABC transporter SACOL0688, staphylococcal complement inhibitor (SCIN), and serine-aspartate repeat-containing protein E (SdrE)] were also detected in a majority of the infecting isolates during biofilm formation in vitro. However, 4 other proteins were detected in only a minority of isolates in vitro while, vice versa, 7 proteins were detected in multiple isolates in vitro but not associated with significantly increased IgG levels in patients. Detection of proteins was largely confirmed using a transcriptomic approach. Our data provide further insights into potential therapeutic targets, such as for vaccination, to reduce S. aureus virulence and biofilm formation. At the same time, our data suggest that either in vitro or immunological in vivo data alone should be interpreted cautiously and that combined studies are necessary to identify potential targets.

  9. Therapeutic potentials of naringin on polymethylmethacrylate induced osteoclastogenesis and osteolysis, in vitro and in vivo assessments

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    Li N

    2013-12-01

    Full Text Available Nianhu Li,1,2,* Zhanwang Xu,2,* Paul H Wooley,1,3 Jianxin Zhang,2 Shang-You Yang1,3 1Department of Surgery, Orthopedics, University of Kansas School of Medicine, Wichita, KS, USA; 2Department of Orthopedics, Affiliated Hospital to Shandong University of Traditional Chinese Medicine, Jinan, People's Republic of China; 3Orthopaedic Research Institute, Via Christi Wichita Hospitals, Wichita, KS, USA *The first two authors contributed equally to this work Abstract: Wear debris associated periprosthetic osteolysis represents a major pathological process associated with the aseptic loosening of joint prostheses. Naringin is a major flavonoid identified in grapefruit. Studies have shown that naringin possesses many pharmacological properties including effects on bone metabolism. The current study evaluated the influence of naringin on wear debris induced osteoclastic bone resorption both in vitro and in vivo. The osteoclast precursor cell line RAW 264.7 was cultured and stimulated with polymethylmethacrylate (PMMA particles followed by treatment with naringin at several doses. Tartrate resistant acid phosphatase (TRAP, calcium release, and gene expression profiles of TRAP, cathepsin K, and receptor activator of nuclear factor-kappa B were sequentially evaluated. PMMA challenged murine air pouch and the load bearing tibia titanium pin-implantation mouse models were used to evaluate the effects of naringin in controlling PMMA induced bone resorption. Histological analyses and biomechanical pullout tests were performed following the animal experimentation. The in vitro data clearly demonstrated the inhibitory effects of naringin in PMMA induced osteoclastogenesis. The naringin dose of 10 µg/mL exhibited the most significant influence on the suppression of TRAP activities. Naringin treatment also markedly decreased calcium release in the stimulated cell culture medium. The short-term air pouch mouse study revealed that local injection of naringin

  10. Probing Regenerative Potential of Moringa oleifera Aqueous Extracts Using In vitro Cellular Assays.

    Science.gov (United States)

    Fernandes, Evangeline E; Pulwale, Anubha V; Patil, Gauri A; Moghe, Alpana S

    2016-01-01

    Molecules stimulating regeneration and proliferation of cells are of significance in combating ailments caused due to tissue injury, inflammation, and degenerative disorders. Moringa oleifera is one of the most valued food plants having the profile of important nutrients and impressive range of medicinal uses. To evaluate the potential of M. oleifera aqueous leaf and flower extracts to promote the proliferation of cells and explore their effect on cancer cell lines for assessment of safety. Aqueous leaf and flower extracts of M. oleifera were investigated for effect on rat-derived primary fibroblast, mesenchymal stem cells (MSCs), and cancer cell lines using cell proliferation assay. They were also tested and compared for wound healing, angiogenesis, and hepatoprotective effect using in vitro assays. Statistically significant increase in the proliferation of primary rat fibroblast, MSCs, and angiogenesis was observed after treatment with aqueous flower extract. The aqueous leaf extract determined a comparatively moderate increment in the proliferation of MSCs and angiogenesis. It however showed prominent cytotoxicity to cancer cell lines and a significant hepatoprotective effect. A very clear difference in response of the two extracts to different types of cells was detected in this study. The aqueous flower extract exhibited a higher potential to stimulate cell proliferation while not exerting the same effect on cancer cell lines. The leaf extract on the other hand, had a prominent antitumor and hepatoptotective effects. Moringa oleifera flower extract showed significant ability to promote proliferation of rat fibroblast and mesenchymal stem cells. The extract also had prominent angiogenic and hepatoprotective effects.The extract did not influence proliferation of cancer cell lines indicating its safety for human consumption and use in pharmaceuticals.The Moringa oleifera leaf extract showed relatively less potential to stimulate cells but had prominent cytotoxic

  11. Probing Regenerative Potential of Moringa oleifera Aqueous Extracts Using In vitro Cellular Assays

    Science.gov (United States)

    Fernandes, Evangeline E.; Pulwale, Anubha V.; Patil, Gauri A.; Moghe, Alpana S.

    2016-01-01

    Background: Molecules stimulating regeneration and proliferation of cells are of significance in combating ailments caused due to tissue injury, inflammation, and degenerative disorders. Moringa oleifera is one of the most valued food plants having the profile of important nutrients and impressive range of medicinal uses. Objective: To evaluate the potential of M. oleifera aqueous leaf and flower extracts to promote the proliferation of cells and explore their effect on cancer cell lines for assessment of safety. Materials and Methods: Aqueous leaf and flower extracts of M. oleifera were investigated for effect on rat-derived primary fibroblast, mesenchymal stem cells (MSCs), and cancer cell lines using cell proliferation assay. They were also tested and compared for wound healing, angiogenesis, and hepatoprotective effect using in vitro assays. Results: Statistically significant increase in the proliferation of primary rat fibroblast, MSCs, and angiogenesis was observed after treatment with aqueous flower extract. The aqueous leaf extract determined a comparatively moderate increment in the proliferation of MSCs and angiogenesis. It however showed prominent cytotoxicity to cancer cell lines and a significant hepatoprotective effect. Conclusion: A very clear difference in response of the two extracts to different types of cells was detected in this study. The aqueous flower extract exhibited a higher potential to stimulate cell proliferation while not exerting the same effect on cancer cell lines. The leaf extract on the other hand, had a prominent antitumor and hepatoptotective effects. SUMMARY Moringa oleifera flower extract showed significant ability to promote proliferation of rat fibroblast and mesenchymal stem cells. The extract also had prominent angiogenic and hepatoprotective effects.The extract did not influence proliferation of cancer cell lines indicating its safety for human consumption and use in pharmaceuticals.The Moringa oleifera leaf extract

  12. Honey - a potential agent against Porphyromonas gingivalis: an in vitro study.

    Science.gov (United States)

    Eick, Sigrun; Schäfer, Gesine; Kwieciński, Jakub; Atrott, Julia; Henle, Thomas; Pfister, Wolfgang

    2014-03-25

    Honey has been discussed as a therapeutic option in wound healing since ancient time. It might be also an alternative to the commonly used antimicrobials in periodontitis treatment. The in-vitro study was aimed to determine the antimicrobial efficacy against Porphyromonas gingivalis as a major periodontopathogen. One Manuka and one domestic beekeeper honey have been selected for the study. As a screening, MICs of the honeys against 20 P. gingivalis strains were determined. Contents of methylglyoxal and hydrogen peroxide as the potential antimicrobial compounds were determined. These components (up to 100 mg/l), propolis (up to 200 mg/l) as well as the two honeys (up to 10% w/v) were tested against four P. gingivalis strains in planktonic growth and in a single-species biofilm. 2% of Manuka honey inhibited the growth of 50% of the planktonic P. gingivalis, the respective MIC50 of the German beekeeper honey was 5%. Manuka honey contained 1.87 mg/kg hydrogen peroxide and the domestic honey 3.74 mg/kg. The amount of methylglyoxal was found to be 2 mg/kg in the domestic honey and 982 mg/kg in the Manuka honey. MICs for hydrogen peroxide were 10 mg/l - 100 mg/l, for methylglyoxal 5 - 20 mg/l, and for propolis 20 mg/l - 200 mg/l. 10% of both types of honey inhibited the formation of P. gingivalis biofilms and reduced the numbers of viable bacteria within 42 h-old biofilms. Neither a total prevention of biofilm formation nor a complete eradication of a 42 h-old biofilm by any of the tested compounds and the honeys were found. Honey acts antibacterial against P. gingivalis. The observed pronounced effects of Manuka honey against planktonic bacteria but not within biofilm can be attributed to methylglyoxal as the characteristic antimicrobial component.

  13. In Vitro Study on the Antioxidant Potentials of the Leaves and Fruits of Nauclea latifolia

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    Ademola O. Ayeleso

    2014-01-01

    Nauclea latifolia, a straggling shrub or small tree, native to tropical Africa and Asia. Hot water extracts of the leaves and fruits of Nauclea latifolia were assessed for their total polyphenolic, flavanol, and flavonol contents as well as 1-diphenyl-2-picrylhydrazyl (DPPH scavenging ability, ferric reducing antioxidant power (FRAP, Trolox equivalence antioxidant capacity (TEAC, and oxygen radical absorbance capacity (ORAC assays. The aqueous extract of the leaves was found to contain higher level of total polyphenols (11.63±0.023 mg GAE/g, flavanol (1.45±0.10 mg CE/g, and flavonol (2.22±0.37 mg QE/g than the extract of the fruits with values of 1.75±0.02 mg GAE/g (total polyphenol, 0.15±0.01 mg CE/g (flavanol, and 1.00±0.13 mg QE/g (flavonol. Similarly, the aqueous extract of the leaves also exhibited higher DPPH (IC50 20.64 mg/mL, FRAP (86.10±3.46 μmol AAE/g, TEAC (94.83±3.57 μmol TE/g, and ORAC (196.55±0.073 μmol TE/g than the extract of the fruits with DPPH (IC50 120.33 mg/mL, FRAP (12.23±0.40 μmol AAE/g, TEAC (12.48±0.21 μmol TE/g, and ORAC (58.88±0.073 μmol TE/g. The present study showed that Nauclea latifolia has strong antioxidant potentials with the leaves demonstrating higher in vitro antioxidant activities than the fruits.

  14. 3,4,5-Trichloroaniline Nephrotoxicity in Vitro: Potential Role of Free Radicals and Renal Biotransformation

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    Christopher Racine

    2014-11-01

    Full Text Available Chloroanilines are widely used in the manufacture of drugs, pesticides and industrial intermediates. Among the trichloroanilines, 3,4,5-trichloroaniline (TCA is the most potent nephrotoxicant in vivo. The purpose of this study was to examine the nephrotoxic potential of TCA in vitro and to determine if renal biotransformation and/or free radicals contributed to TCA cytotoxicity using isolated renal cortical cells (IRCC from male Fischer 344 rats as the animal model. IRCC (~4 million cells/mL; 3 mL were incubated with TCA (0, 0.1, 0.25, 0.5 or 1.0 mM for 60–120 min. In some experiments, IRCC were pretreated with an antioxidant or a cytochrome P450 (CYP, flavin monooxygenase (FMO, cyclooxygenase or peroxidase inhibitor prior to incubation with dimethyl sulfoxide (control or TCA (0.5 mM for 120 min. At 60 min, TCA did not induce cytotoxicity, but induced cytotoxicity as early as 90 min with 0.5 mM or higher TCA and at 120 min with 0.1 mM or higher TCA, as evidenced by increased lactate dehydrogenase (LDH release. Pretreatment with the CYP inhibitor piperonyl butoxide, the cyclooxygenase inhibitor indomethacin or the peroxidase inhibitor mercaptosuccinate attenuated TCA cytotoxicity, while pretreatment with FMO inhibitors or the CYP inhibitor metyrapone had no effect on TCA nephrotoxicity. Pretreatment with an antioxidant (α-tocopherol, glutathione, ascorbate or N-acetyl-l-cysteine also reduced or completely blocked TCA cytotoxicity. These results indicate that TCA is directly nephrotoxic to IRCC in a time and concentration dependent manner. Bioactivation of TCA to toxic metabolites by CYP, cyclooxygenase and/or peroxidase contributes to the mechanism of TCA nephrotoxicity. Lastly, free radicals play a role in TCA cytotoxicity, although the exact nature of the origin of these radicals remains to be determined.

  15. Titanium–35niobium alloy as a potential material for biomedical implants: In vitro study

    Energy Technology Data Exchange (ETDEWEB)

    Perez de Andrade, Dennia; Marotta Reis de Vasconcellos, Luana; Chaves Silva Carvalho, Isabel; Ferraz de Brito Penna Forte, Lilibeth; Souza Santos, Evelyn Luzia de [Department of Bioscience and Oral Diagnosis, Institute of Science and Technology, UNESP — Univ Estadual Paulista, State University of São Paulo (UNESP), Av. Engenheiro Francisco José Longo, 777, São José dos Campos 12245-000, SP (Brazil); Falchete do Prado, Renata, E-mail: renatafalchete@hotmail.com [Department of Bioscience and Oral Diagnosis, Institute of Science and Technology, UNESP — Univ Estadual Paulista, State University of São Paulo (UNESP), Av. Engenheiro Francisco José Longo, 777, São José dos Campos 12245-000, SP (Brazil); Santos, Dalcy Roberto dos; Alves Cairo, Carlos Alberto [Division of Materials, Air and Space Institute, CTA, Praça Mal. do Ar Eduardo Gomes, 14, São José dos Campos 12904-000, SP (Brazil); Rodarte Carvalho, Yasmin [Department of Bioscience and Oral Diagnosis, Institute of Science and Technology, UNESP — Univ Estadual Paulista, State University of São Paulo (UNESP), Av. Engenheiro Francisco José Longo, 777, São José dos Campos 12245-000, SP (Brazil)

    2015-11-01

    Research on new titanium alloys and different surface topographies aims to improve osseointegration. The objective of this study is to analyze the behavior of osteogenic cells cultivated on porous and dense samples of titanium–niobium alloys, and to compare them with the behavior of such type of cells on commercial pure titanium. Samples prepared using powder metallurgy were characterized using scanning electron microscopy (SEM), energy dispersive spectroscopy (EDS), X-ray diffraction (XRD), and metallographic and profilometer analyses. Osteogenic cells from newborn rat calvaria were plated over different groups: dense or porous samples composed of Ti or Ti–35niobium (Nb). Cell adhesion, cell proliferation, MTT assay, cell morphology, protein total content, alkaline phosphatase activity, and mineralization nodules were assessed. Results from XRD and EDS analysis confirmed the presence of Ti and Nb in the test alloy. Metallographic analysis revealed interconnected pores, with pore size ranging from 138 to 150 μm. The profilometer analysis detected the greatest rugosity within the dense alloy samples. In vitro tests revealed similar biocompatibility between Ti–35Nb and Ti; furthermore, it was possible to verify that the association of porous surface topography and the Ti–35Nb alloy positively influenced mineralized matrix formation. We propose that the Ti–35Nb alloy with porous topography constitutes a biocompatible material with great potential for use in biomedical implants. - Highlights: • Powder metallurgy is effective in producing porous biomaterials. • Ti–35Nb alloy improved mineralized matrix formation. • Porous surface favored a multidirectional pattern of cell spreading. • Porous surface Ti–35Nb alloy appears to be more favorable to bone formation than existing alloys.

  16. Evaluation of remineralizing potential of commercially available child formula dentifrices: An in vitro study

    Directory of Open Access Journals (Sweden)

    Apurva Jagdish Gujarathi

    2015-01-01

    Full Text Available Aim: The aim of this in vitro study is to evaluate the remineralizing potential of commercially available low fluoride child formula dentifrice on primary teeth. Materials and Methods: Total 30 primary teeth were placed in demineralizing solution for 96 hours to produce artificial carious lesions of approximately 100 μm depth, and then cut longitudinally into 30 sections of 100-150 μm thickness and randomly assigned to three groups. Sections were treated with dentifrices containing Colgate ® (anti tooth decay 500 ppm NaF, Cheerio gel ® 458 ppm MFP and Vicco ® non-fluoridated dentifrice. Lesions were evaluated using polarized light microscopy. Results: Colgate ® (anti tooth decay 500 ppm NaF sections exhibited a statistically significant decrease in lesion depth (P < 0.05, paired t-test, whereas those in Cheerio gel ® 458 ppm MFP showed a decrease in lesion depth but was not statistically significant. Vicco ® non-fluoridated dentifrice showed increase in lesion depth. Statistics: A paired t-test is used to evaluate pre- and post-treatment lesion depth measurements, and Newman-Keuls multiple post hoc procedures was carried out to compare pair-wise difference of pre- and post-treatment lesion depth. Conclusion: The Colgate ® (anti tooth decay 500 ppm NaF dentifrice and Cheerio gel ® 458 ppm MFP demonstrated remineralization of carious lesions by virtue of decrease in lesion depth, whereas Vicco ® non-fluoridated dentifrice showed increase in lesion depth.

  17. Establishment of a Predictive In Vitro Assay for Assessment of the Hepatotoxic Potential of Oligonucleotide Drugs

    National Research Council Canada - National Science Library

    Sewing, Sabine; Boess, Franziska; Moisan, Annie; Bertinetti-Lapatki, Cristina; Minz, Tanja; Hedtjaern, Maj; Tessier, Yann; Schuler, Franz; Singer, Thomas; Roth, Adrian B

    2016-01-01

    .... The mechanisms of SSO induced liver toxicity are poorly understood, and up to now no preclinical in vitro model has been established that allows prediction of the hepatotoxicity risk of a given SSO...

  18. A mechanistic insight into curcumin modulation of the IL-1β secretion and NLRP3 S-glutathionylation induced by needle-like cationic cellulose nanocrystals in myeloid cells.

    Science.gov (United States)

    Guglielmo, Andrew; Sabra, Adham; Elbery, Mostafa; Cerveira, Milena M; Ghenov, Fernanda; Sunasee, Rajesh; Ckless, Karina

    2017-08-25

    Recently we have demonstrated that needle-like cationic cellulose nanocrystals (CNC-AEMA2) evoke immunological responses through NLRP3 inflammasome/IL-1β inflammatory pathway. In this study we demonstrated that curcumin, a naturally occurring polyphenolic compound isolated from Curcuma longa (Zingiberaceae), was able to suppress, at least in part, this immunological response, as observed by diminished IL-1β secretion in CNC-AEMA2-stimulated macrophages primed with LPS. Curcumin is a well-known antioxidant and anti-inflammatory natural compound and in addition to acting as "scavenger" of reactive oxygen species (ROS), it can also upregulates antioxidant enzymes. However, the mechanisms by which this natural compound exerts its protective activity is still under investigation. We hypothesize that curcumin may also affect S-glutathionylation of key proteins involved in the NLRP3 inflammasome/IL-1β pathway, and therefore impact their protein-protein interactions. The goal of this study was to investigate the effects of curcumin on the S-glutathionylation of NLRP3 induced by CNC-AEMA2 in LPS-primed mouse macrophages (J774A.1), as well as interactions among proteins of the NLRP3 inflammasome complex. Our main finding indicates that the addition of curcumin concomitantly with LPS caused the greatest decrease in NLRP3 S-glutathionylation and a respective increase in caspase-1 S-glutathionylation, which appears to favor protein-protein interactions in the NLRP3 complex. Taking together, our results suggest that, at least in part, the anti-inflammatory activity of curcumin is associated with changes in S-glutathionylation of key NLRP3 inflammasome components, and perhaps resulting in sustained complex assembly and suppression of IL-1β secretion. Copyright © 2017 Elsevier B.V. All rights reserved.

  19. The potential use of maturation in vitro of human oocytes in low responder patients.

    Science.gov (United States)

    Requena, A; Neuspiller, F; Cobo, A C; Aragonés, M; Remohí, J; Simón, C; Pellicer, A

    2000-05-01

    To assess whether maturation in vitro of human oocytes (MIVHO) could be an alternative treatment in low responders to ovarian stimulation for in vitro fertilization (IVF). Prospective case-control study. Spontaneously ovulatory women who volunteered were included in our program of MIVHO at the Instituto Valenciano de Infertilidad. Rates of oocyte retrieval, in vitro maturation, fertilization, and development up to the blastocyst stage were studied. A significantly increased rate of oocyte retrieval was found when the pickup was performed before follicular selection. No differences were found when MIVHO was used in a low responder patient with an ovarian content of early antral follicles > 5 as compared to normal responders. MIVHO could be a successful choice in low responder patients with an acceptable number of early antral follicles. Oocyte retrieval should be performed before follicular selection in order to obtain more oocytes.

  20. Honey – a potential agent against Porphyromonas gingivalis: an in vitro study

    Science.gov (United States)

    2014-01-01

    Background Honey has been discussed as a therapeutic option in wound healing since ancient time. It might be also an alternative to the commonly used antimicrobials in periodontitis treatment. The in-vitro study was aimed to determine the antimicrobial efficacy against Porphyromonas gingivalis as a major periodontopathogen. Methods One Manuka and one domestic beekeeper honey have been selected for the study. As a screening, MICs of the honeys against 20 P. gingivalis strains were determined. Contents of methylglyoxal and hydrogen peroxide as the potential antimicrobial compounds were determined. These components (up to 100 mg/l), propolis (up to 200 mg/l) as well as the two honeys (up to 10% w/v) were tested against four P. gingivalis strains in planktonic growth and in a single-species biofilm. Results 2% of Manuka honey inhibited the growth of 50% of the planktonic P. gingivalis, the respective MIC50 of the German beekeeper honey was 5%. Manuka honey contained 1.87 mg/kg hydrogen peroxide and the domestic honey 3.74 mg/kg. The amount of methylglyoxal was found to be 2 mg/kg in the domestic honey and 982 mg/kg in the Manuka honey. MICs for hydrogen peroxide were 10 mg/l - 100 mg/l, for methylglyoxal 5 – 20 mg/l, and for propolis 20 mg/l – 200 mg/l. 10% of both types of honey inhibited the formation of P. gingivalis biofilms and reduced the numbers of viable bacteria within 42 h-old biofilms. Neither a total prevention of biofilm formation nor a complete eradication of a 42 h-old biofilm by any of the tested compounds and the honeys were found. Conclusions Honey acts antibacterial against P. gingivalis. The observed pronounced effects of Manuka honey against planktonic bacteria but not within biofilm can be attributed to methylglyoxal as the characteristic antimicrobial component. PMID:24666777

  1. CXCL10/IP10 is a novel potential in vitro marker of TB infection

    Directory of Open Access Journals (Sweden)

    Ilaria Sauzullo

    2009-06-01

    Full Text Available Introduction IFN-γ is a pivotal cytokine in the immune response to Myc. tuberculosis, infact this is the key cytokine produced in response to antigens specific following tuberculosis exposure causing either active or latent tuberculosis (TB and this observation forms the basis of interferon gamma release assay (IGRA, but there are alternative or additional cytokines and chemokines that could be used to improve detection of Myc. tuberculosis infection.The aim of this study was to evaluate the diagnostic utility of chemokine CXCL10/IP-10 as biomarker of active TB and to compare the results with classical QuantiFERON-Gold assay . Methods CXCL10/IP-10 and IFN-γ responses to stimulation with ESAT-6 and CFP-10 were evaluated in 21 patients with active tuberculosis and in 6 healthy unexposed subjects with no history of TB or TB contact were used as controls healthy controls. QuantiFERON-TB Gold (QFT-G, Cellestis was used for the measurement of IFN-γ levels; CXCL10/IP-10 was detected by ELISA (R&D Systems . Results Of the 21 TB patients included, 11 had a QFT-G positive and 10 had negative QFT-G results.All QFT-G positive patients had increased levels of CXCL10/IP-10 (median, pg/ml in both ESAT-6 and CFP-10 stimulated samples patients compared to healthy controls (1807 and 1111 vs 251 and 188 of controls, respectively (p<0.001 for both. The patients with active TB and QFT-G negative exhibited higher concentrations of CXCL10/IP-10 following antigen stimulation (837 pg/ml for ESAT-6;1674 pg/ml for CFP-10 (p<0.001. Conclusion Our study showed that in all patients with active TB, the CXCL10/IP-10 is expressed in higher amounts than IFN-γ following Myc. tuberculosis antigen-specific stimulation, and CXCL10/IP-10 appeared to be even more sensitive than QuantiFERON TB-Gold in TB patients with negative IFN-γ response. The measurement of chemokine CXCL10/IP-10, although not specific for tuberculosis, may have potential as an alternative or additional marker

  2. Self-assembled amphotericin B-loaded polyglutamic acid nanoparticles: preparation, characterization and in vitro potential against Candida albicans

    OpenAIRE

    Zia Q; Khan AA; Swaleha Z; Owais M

    2015-01-01

    Qamar Zia,1 Aijaz Ahmed Khan,2 Zubair Swaleha,3 Mohammad Owais1 1Interdisciplinary Biotechnology Unit, 2Department of Anatomy, 3Women’s College, Aligarh Muslim University, Aligarh, India Abstract: In the present study, we developed a self-assembled biodegradable polyglutamic acid (PGA)-based formulation of amphotericin B (AmB) and evaluated its in vitro antifungal potential against Candida albicans. The AmB-loaded PGA nanoparticles were prepared in-house and had a mean size dimens...

  3. Antioxidant potential of a unique LAB culture isolated from Harbin dry sausage: In vitro and in a sausage model.

    Science.gov (United States)

    Chen, Qian; Kong, Baohua; Sun, Qinxiu; Dong, Fujia; Liu, Qian

    2015-12-01

    The lactic acid bacteria Pediococcus pentosaceus, Lactobacillus curvatus, Lactobacillus brevis, and Lactobacillus fermentum isolated from Harbin dry sausage were evaluated for their potential antioxidant activity. The in vitro results showed that P. pentosaceus had the strongest H2O2 resistance, radical scavenging activity, reducing power, and inhibition of lipid peroxidation (Pacid-reactive substance and carbonyl formation, while it also reduced the sulfhydryl loss in sausages (Pantioxidant starter culture in fermented meat products. Copyright © 2015. Published by Elsevier Ltd.

  4. Hydroxycinnamate conjugates as potential monolignol replacements: In vitro lignification and cell wall studies with rosmarinic acid

    Science.gov (United States)

    The plasticity of lignin biosynthesis should permit the inclusion of new compatible phenolic monomers such as rosmarinic acid (RA) and analogous catechol derivatives to create cell wall lignins that are less recalcitrant to biomass processing. In vitro lignin polymerization experiments revealed that...

  5. The potential of a niacinamide dominated cosmeceutical formulation on fibroblast activity and wound healing in vitro.

    Science.gov (United States)

    Wessels, Quenton; Pretorius, Etheresia; Smith, Celeste M; Nel, Hugo

    2014-04-01

    Knowledge on the intrinsic mechanisms involved in wound healing provides opportunity for various therapeutic strategies. The manipulation of dermal fibroblast proliferation and differentiation might prove to beneficially augment wound healing. This study evaluated the combined effects of niacinamide, L-carnosine, hesperidin and Biofactor HSP(®) on fibroblast activity. The effects on fibroblast collagen production, cellular proliferation, migration and terminal differentiation were assessed. In addition, the authors determined the effects on in vitro wound healing. The optimal concentrations of actives were determined in vitro. Testing parameters included microscopic morphological cell analysis, cell viability and proliferation determination, calorimetric collagen detection and in vitro wound healing dynamics. Results show that 0·31 mg/ml niacinamide, 0·10 mg/ml L-carnosine, 0·05 mg/ml hesperidin and 5·18 µg/ml Biofactor HSP® proved optimal in vitro. The results show that fibroblast collagen synthesis was increased alongside with cellular migration and proliferation. © 2012 The Authors. International Wound Journal © 2012 Medicalhelplines.com Inc and John Wiley & Sons Ltd.

  6. In vitro and in vivo evaluation of antidiabetic potential of extracts of ...

    African Journals Online (AJOL)

    Recipes were extracted in water according to traditional usage and screened in vitro to assess glucose uptake in C2C12 muscle cells and glucose production by the H4IIE liver cells (through inhibition of glucose-6-phosphatase, the rate limiting enzyme) and in vivo through the oral glucose tolerance test in normal mice (2 ...

  7. Potential of white garlic powder ( Allium sativum L.) to modify in vitro ...

    African Journals Online (AJOL)

    The current study aimed to evaluate the effect of increasing doses of garlic powder (GaP) on in vitro fermentation characteristics. Two successive 24-hour incubations were run, and gas production was measured at the end of each incubation period. Liquid samplings for each dose were reserved to determine ammonia ...

  8. Can Ocimum basilicum L. and Ocimum tenuiflorum L. in vitro culture be a potential source of secondary metabolites?

    Science.gov (United States)

    Bhuvaneshwari, Karuppiah; Gokulanathan, Ananda; Jayanthi, Malayandi; Govindasamy, Vaithiyanathan; Milella, Luigi; Lee, Sungyoung; Yang, Deok Chun; Girija, Shanmugam

    2016-03-01

    In this study Ocimum basilicum L. (OB) and Ocimum tenuiflorum L. (OT) in vitro culture standardisation for increasing eugenol distribution, in comparison to their respective field grown parts was carried out. Eugenol was quantified using an optimised HPLC method and its relation with the total phenolic content (TPC) was measured. In vitro grown leaves and somatic embryos, of both OB and OT were found to contain similar quantities of eugenol (85μg/g approximately), higher than OB and OT field-grown leaves (30.2μg/g and 25.1μg/g respectively). It was also determined that in vitro grown leaves were richer in TPC than the field-grown intact organs. Results demonstrated the prominence of in vitro cultures for eugenol extraction. This study underlines that important food flavouring metabolites (e.g. vanillin, vanillic acids) might be produced, via the eugenol pathway, in Ocimum species that may be a good potential source of eugenol. Copyright © 2015. Published by Elsevier Ltd.

  9. In vitro evaluation of whole faba bean and its seed coat as a potential source of functional food components.

    Science.gov (United States)

    Çalışkantürk Karataş, Selen; Günay, Demet; Sayar, Sedat

    2017-09-01

    In vitro studies were conducted to evaluate the particular nutritional benefits of whole faba bean seed (WFB) and fava bean seed coat (FBSC). Total dietary fiber contents of WFB and FBSC were 27.5% and 82.3%, respectively. FBSC were contained much higher total phenolic substances, condensed tannins, and total antioxidant activity than WFB. Bile acid (BA)-binding capacities of in vitro digested samples and nutritionally important products produced by in vitro fermentation of digestion residues were also studied. The BA-binding capacities of WFB and FBSC were 1.94 and 37.50μmol/100mg, respectively. Total BA bound by FBSC was even higher than the positive standard cholestyramine. Lignin and other constituents of the Klason residue were found to influence BA-binding properties. Moreover, the extent of the in vitro fermentation process showed that, fermentability of FBSC residue was significantly lower than that of WFB residue. Overall, faba bean, especially its seed coat, has great potential as a functional food. Copyright © 2017 Elsevier Ltd. All rights reserved.

  10. The Measurement and Research of Surface Potentials of Human Tooth in vitro

    National Research Council Canada - National Science Library

    Zhang, J

    2001-01-01

    .... To confirm the existence of the surface potentials in extracted tooth and research the development of the potentials, the surface potentials between mid-spots of enamel crown's buccal side and tooth...

  11. Criteria that optimize the potential of murine embryonic stem cells for in vitro and in vivo developmental studies.

    Science.gov (United States)

    Brown, D G; Willington, M A; Findlay, I; Muggleton-Harris, A L

    1992-01-01

    Cultured mouse embryonic stem (ES) cells are used for both in vitro and in vivo studies. The uncommitted pluripotent cells provide a model system with which to study cellular differentiation and development; they can also be used as vectors to carry specific mutations into the mouse genome by homologous recombination. To ensure successful integration into the germ line, competent totipotent diploid ES cell lines are selected using a cell injection bioassay that is both time consuming and technically demanding. The prolonged in vitro culture of rapidly dividing ES cells can lead to accumulated changes and chromosomal abnormalities that will compromise the biological function and abrogate germ line transmission of chimeric mice carrying novel genetic mutations. Such in vitro conditions will vary between individual laboratories; for example, differences in the serums used for maintenance. Using a number of different criteria we attempt in this paper to define the parameters that we found to be key factors for optimization of the biological potential of established ES cell lines. The successful integration into the germ line is dependant on acquiring or deriving a competent totipotent mouse ES diploid cell line. In this paper parameters and criteria are defined which we found to be key factors for the optimization of the biological potential of established ES cell lines.

  12. In vitro and in vivo immunomodulatory potential of Pedicularis longiflora and Allium carolinianum in alloxan-induced diabetes in rats.

    Science.gov (United States)

    Yatoo, Mohd Iqbal; Dimri, Umesh; Gopalakrishnan, Arumugam; Saxena, Archana; Wani, Sarfaraz Ahmad; Dhama, Kuldeep

    2017-10-27

    Pedicularis longiflora Rudolph (Orobanchaceae) and Allium carolinianum Linn (Alliaceae) are two important medicinal plants found in trans-Himalayan Changthang. The immunomodulatory potential of these plants has not been explored. In the present study, we evaluated the in vitro and in vivo immunomodulatory potential of P. longiflora and A. carolinianum in alloxan-induced diabetes in Wistar rats. The ethanol extracts of the aerial parts of P. longiflora and whole plant parts of A. carolinianum were used for studying the in vitro immunomodulatory activity using lymphocyte stimulation and cytokine release assays. For the in vivo study, 5 groups of 6 rats per group, including alloxan-induced diabetic and plant extract-treated rats, were evaluated for cell-mediated immune (CMI) and humoral immune (HMI) responses in a 42-day experimental trial using doses of 500mg/kg b.wt. for P. longiflora and 250mg/kgbwt. for A. carolinianum. For P. longiflora, the median effective dose was found to be 500mg/kg. The in vitro lymphocyte stimulation index for P. longiflora was significantly higher (1.73±0.04, pimmunomodulatory activities than A. carolinianum, especially in alloxan-induced diabetic rats. However, further research is needed to identify the different molecular mechanisms involved in mediating this immunomodulatory response. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  13. Development of assay platforms for in vitro screening of Treg modulating potential of pharmacological compounds

    DEFF Research Database (Denmark)

    Pedersen, Anders Elm; Holmstrøm, Kim; Jørgensen, Flemming

    2015-01-01

    sorting (FACS) sorted CD4 + CD25(high)CD127(dim/-)CD45RA+ naïve Treg cells followed by in vitro expansion. We report on the use of these cells in a short-term assay based on Treg mediated inhibition of the early effector T cell activation markers CD69 and CD154. Additionally, we investigate the use...

  14. Proximate composition, phytochemical analysis, and in vitro antioxidant potentials of extracts of Annona muricata (Soursop)

    OpenAIRE

    Agu, Kingsley C.; Okolie, Paulinus N.

    2017-01-01

    Abstract Numerous bioactive compounds and phytochemicals have been reported to be present Annona muricata (Soursop). Some of these chemical compounds have been linked to the ethnomedicinal properties of the plant and its antioxidant properties. The aim of this study was to assess the proximate composition, phytochemical constituents and in vitro antioxidant properties of A. muricata using standard biochemical procedures. The defatted Annona muricata crude methanolic extracts of the different ...

  15. Exploring the metal phytoremediation potential of three Populus alba L. clones using an in vitro screening.

    Science.gov (United States)

    Di Lonardo, Sara; Capuana, Maurizio; Arnetoli, Miluscia; Gabbrielli, Roberto; Gonnelli, Cristina

    2011-01-01

    This work was planned for providing a useful screening tool for the selection of Populus alba clones suitable for phytoremediation techniques. To this aim, we investigated variation in arsenic, cadmium, copper, and zinc tolerance, accumulation and translocation in three poplar clones through an in vitro screening. Poplars have been widely proposed for phytoremediation, as they are adaptable to grow on contaminated areas and able to accumulate metals. The investigation of possible differences among poplar clones in metal tolerance and accumulation deserves to be deeply studied and exploited for the selection of the more suitable tool for phytoremediation purposes. In vitro multiplied microshoots of a commercial and two autochthonous P. alba clones were subcultured on hormone-free WPM medium for 1 month and then transferred for 2 weeks onto media containing different concentrations of the metals investigated. At the end of the treatments, plantlets were sampled, weighed, and mineralised by wet ashing. Metal concentrations were determined by ICP-OES. For the metal concentration used in the experiments, our clones of P. alba showed variation in metal tolerance, metal accumulation and content. The fast-growing commercial clone, even if rarely showing the highest plant metal concentration, displayed the highest metal content, suggesting biomass production as the key factor in evaluating the phytoextraction capacity of P. alba clones for the metals studied. Data demonstrated that in vitro screening of cuttings represents a valuable way of assessing the ability of different poplar clones to take up, tolerate and survive metal stress.

  16. The Potential of an in Vitro Digestion Method for Predicting Glycemic Response of Foods and Meals

    Directory of Open Access Journals (Sweden)

    Konstantina Argyri

    2016-04-01

    Full Text Available Increased interest in glycemic response derives from its linkage with chronic diseases, including obesity and type 2 diabetes. Our objective was to develop an in vitro method that predicts glycemic response. We proposed a simulated gastrointestinal digestion protocol that uses the concentration of dialyzable glucose (glucose in the soluble low molecular weight fraction of digests as an index for the prediction of glycemic response. For protocol evaluation, dialyzable glucose from 30 foods or meals digested in vitro were compared with published values for their glycemic index (GI (nine foods, glycemic load (GL (16 foods and glycemic response (14 meals. The correlations were significant when comparing dialyzable glucose with GL (Spearman’s rho = 0.953, p < 0.001, GI (Spearman’s rho = 0.800, p = 0.010 and glycemic response (Spearman’s rho = 0.736, p = 0.003. These results demonstrate that despite limitations associated with in vitro approaches, the proposed protocol may be a useful tool for predicting glycemic response of foods or meals.

  17. Stress-Induced Protein S-Glutathionylation and S-Trypanothionylation in African Trypanosomes-A Quantitative Redox Proteome and Thiol Analysis.

    Science.gov (United States)

    Ulrich, Kathrin; Finkenzeller, Caroline; Merker, Sabine; Rojas, Federico; Matthews, Keith; Ruppert, Thomas; Krauth-Siegel, R Luise

    2017-09-20

    Trypanosomatids have a unique trypanothione-based thiol redox metabolism. The parasite-specific dithiol is synthesized from glutathione and spermidine, with glutathionylspermidine as intermediate catalyzed by trypanothione synthetase. In this study, we address the oxidative stress response of African trypanosomes with special focus on putative protein S-thiolation. Challenging bloodstream Trypanosoma brucei with diamide, H 2 O 2 or hypochlorite results in distinct levels of reversible overall protein S-thiolation. Quantitative proteome analyses reveal 84 proteins oxidized in diamide-stressed parasites. Fourteen of them, including several essential thiol redox proteins and chaperones, are also enriched when glutathione/glutaredoxin serves as a reducing system indicating S-thiolation. In parasites exposed to H 2 O 2 , other sets of proteins are modified. Only three proteins are S-thiolated under all stress conditions studied in accordance with a highly specific response. H 2 O 2 causes primarily the formation of free disulfides. In contrast, in diamide-treated cells, glutathione, glutathionylspermidine, and trypanothione are almost completely protein bound. Remarkably, the total level of trypanothione is decreased, whereas those of glutathione and glutathionylspermidine are increased, indicating partial hydrolysis of protein-bound trypanothione. Depletion of trypanothione synthetase exclusively induces protein S-glutathionylation. Total mass analyses of a recombinant peroxidase treated with T(SH) 2 and either diamide or hydrogen peroxide verify protein S-trypanothionylation as stable modification. Our data reveal for the first time that trypanosomes employ protein S-thiolation when exposed to exogenous and endogenous oxidative stresses and trypanothione, despite its dithiol character, forms protein-mixed disulfides. The stress-specific responses shown here emphasize protein S-trypanothionylation and S-glutathionylation as reversible protection mechanism in these

  18. Field and action potential recordings in heart slices: correlation with established in vitro and in vivo models

    Science.gov (United States)

    Himmel, Herbert M; Bussek, Alexandra; Hoffmann, Michael; Beckmann, Rolf; Lohmann, Horst; Schmidt, Matthias; Wettwer, Erich

    2012-01-01

    BACKGROUND AND PURPOSE Action potential (AP) recordings in ex vivo heart preparations constitute an important component of the preclinical cardiac safety assessment according to the ICH S7B guideline. Most AP measurement models are sensitive, predictive and informative but suffer from a low throughput. Here, effects of selected anti-arrhythmics (flecainide, quinidine, atenolol, sotalol, dofetilide, nifedipine, verapamil) on field/action potentials (FP/AP) of guinea pig and rabbit ventricular slices are presented and compared with data from established in vitro and in vivo models. EXPERIMENTAL APPROACH Data from measurements of membrane currents (hERG, INa), AP/FP (guinea pig and rabbit ventricular slices), AP (rabbit Purkinje fibre), haemodynamic/ECG parameters (conscious, telemetered dog) were collected, compared and correlated to complementary published data (focused literature search). KEY RESULTS The selected anti-arrhythmics, flecainide, quinidine, atenolol, sotalol, dofetilide, nifedipine and verapamil, influenced the shape of AP/FP of guinea pig and rabbit ventricular slices in a manner similar to that observed for rabbit PF. The findings obtained from slice preparations are in line with measurements of membrane currents in vitro, papillary muscle AP in vitro and haemodynamic/ECG parameters from conscious dogs in vivo, and were also corroborated by published data. CONCLUSION AND IMPLICATIONS FP and AP recordings from heart slices correlated well with established in vitro and in vivo models in terms of pharmacology and predictability. Heart slice preparations yield similar results as papillary muscle but offer enhanced throughput for mechanistic investigations and may substantially reduce the use of laboratory animals. PMID:22074238

  19. Hypoxia increases mouse satellite cell clone proliferation maintaining both in vitro and in vivo heterogeneity and myogenic potential.

    Science.gov (United States)

    Urbani, Luca; Piccoli, Martina; Franzin, Chiara; Pozzobon, Michela; De Coppi, Paolo

    2012-01-01

    Satellite cells (SCs) are essential for postnatal muscle growth and regeneration, however, their expansion potential in vitro is limited. Recently, hypoxia has been used to enhance proliferative abilities in vitro of various primary cultures. Here, by isolating SCs from single mouse hindlimb skeletal myofibers, we were able to distinguish two subpopulations of clonally cultured SCs (Low Proliferative Clones--LPC--and High Proliferative Clones--HPC), which, as shown in rat skeletal muscle, were present at a fixed proportion. In addition, culturing LPC and HPC at a low level of oxygen we observed a two fold increased proliferation both for LPC and HPC. LPC showed higher myogenic regulatory factor (MRF) expression than HPC, particularly under the hypoxic condition. Notably, a different myogenic potential between LPC and HPC was retained in vivo: green fluorescent protein (GFP)+LPC transplantation in cardiotoxin-injured Tibialis Anterior led to a higher number of new GFP+muscle fibers per transplanted cell than GFP+HPC. Interestingly, the in vivo myogenic potential of a single cell from an LPC is similar if cultured both in normoxia and hypoxia. Therefore, starting from a single satellite cell, hypoxia allows a larger expansion of LPC than normal O(2) conditions, obtaining a consistent amount of cells for transplantation, but maintaining their myogenic regeneration potential.

  20. Combination effects of epidermal growth factor and glial cell line-derived neurotrophic factor on the in vitro developmental potential of porcine oocytes

    DEFF Research Database (Denmark)

    Valleh, Mehdi Vafaye; Rasmussen, Mikkel Aabech; Hyttel, Poul

    2016-01-01

    The developmental potential of in vitro matured porcine oocytes is still lower than that of oocytes matured and fertilized in vivo. Major problems that account for the lower efficiency of in vitro production include the improper nuclear and cytoplasmic maturation of oocytes. With the aim of impro...

  1. Developmental potential of vitrified holstein cattle embryos fertilized in vitro with sex-sorted sperm.

    Science.gov (United States)

    Xu, J; Guo, Z; Su, L; Nedambale, T L; Zhang, J; Schenk, J; Moreno, J F; Dinnyés, A; Ji, W; Tian, X C; Yang, X; Du, F

    2006-07-01

    In vitro fertilization (IVF) is a feasible way to utilize sex-sorted sperm to produce offspring of a predetermined sex in the livestock industry. The objective of the present study was to examine the effects of various factors on bovine IVF and to systematically improve the efficiency of IVF production using sex-sorted sperm. Both bulls and sorting contributed to the variability among differential development rates of embryos fertilized by sexed sperm. Increased sorting pressures (275.8 to 344.75 kPa) did not have a significant effect on the in vitro fertility of the sorted sperm; neither did an extended period of 9 to 14 h from semen collection to sorting. As few as 600 sorted sperm were used to fertilize an oocyte, resulting in blastocyst development of 33.2%. Postwarming of vitrified sexed IVF embryos resulted in high morphological survival (96.3%) and hatching (84.4%) rates, similar to those fertilized by nonsexed sperm (93.1 and 80.6%, respectively). A 40.9% pregnancy rate was established following the transfer of 3,627 vitrified, sexed embryos into synchronized recipients. This was not different from the rates with nonsexed IVF (41.9%, n = 481), or in vivo-produced (53.1%, n = 192) embryos. Of 458 calves born, 442 (96.5%) were female and 99.6% appeared normal. These technologies (sperm sexing-IVF-vitrification-embryo transfer) provide farmers, as well as the livestock industry, with a valuable option for herd expansion and heifer replacement programs. In summary, calves were produced using embryos fertilized by sex-sorted sperm in vitro and cryopreserved by rapid cooling vitrification.

  2. Prediction and in vitro verification of potential CTL epitopes conserved among PRRSV-2 strains

    DEFF Research Database (Denmark)

    Welner, Simon; Nielsen, Morten; Rasmussen, Michael

    2017-01-01

    binding predictions. Predicted binders were prioritized according to genomic conservation and SLA coverage using the PopCover algorithm. From this, 53 peptides were acquired for further analysis. Binding affinity and stability of a subset of 101 peptide-SLA combinations were validated in vitro for 4...... of the 5 SLAs. Eventually, 23% of the predicted peptide-SLA combinations showed to form complexes with a dissociation half-life ≥30 min. Additionally, combining the two prediction methods proved to be more robust across alleles than either method used alone in terms of predicted-to-observed correlations...

  3. Electroresponsive properties and membrane potential trajectories of three types of inspiratory neurons in the newborn mouse brain stem in vitro

    DEFF Research Database (Denmark)

    Rekling, J C; Champagnat, J; Denavit-Saubié, M

    1996-01-01

    with the aim of extending the classification of inspiratory neurons to include analysis of active membrane properties. 2. The slice generated a regular rhythmic motor output recorded as burst of action potentials on a XII nerve root with a peak to peak time of 11.5 +/- 3.4 s and a duration of 483 +/- 54 ms......1. The electrophysiological properties of inspiratory neurons were studied in a rhythmically active thick-slice preparation of the newborn mouse brain stem maintained in vitro. Whole cell patch recordings were performed from 60 inspiratory neurons within the rostral ventrolateral part of the slice...... (means +/- SD, n = 50). Based on the electroresponsive properties and membrane potential trajectories throughout the respiratory cycle, three types of inspiratory neurons could be distinguished. 3. Type-1 neurons were spiking in the interval between the inspiratory potentials (n = 9) or silent...

  4. Chemopreventive potential of β-Sitosterol in experimental colon cancer model - an In vitro and In vivo study

    Directory of Open Access Journals (Sweden)

    Paulraj Gabriel M

    2010-06-01

    Full Text Available Abstract Background Asclepias curassavica Linn. is a traditional medicinal plant used by tribal people in the western ghats, India, to treat piles, gonorrhoea, roundworm infestation and abdominal tumours. We have determined the protective effect of β-sitosterol isolated from A. curassavica in colon cancer, using in vitro and in vivo models. Methods The active molecule was isolated, based upon bioassay guided fractionation, and identified as β-sitosterol on spectral evidence. The ability to induce apoptosis was determined by its in vitro antiradical activity, cytotoxic studies using human colon adenocarcinoma and normal monkey kidney cell lines, and the expression of β-catenin and proliferating cell nuclear antigen (PCNA in human colon cancer cell lines (COLO 320 DM. The chemopreventive potential of β-sitosterol in colon carcinogenesis was assessed by injecting 1,2-dimethylhydrazine (DMH, 20 mg/kg b.w. into male Wistar rats and supplementing this with β-sitosterol throughout the experimental period of 16 weeks at 5, 10, and 20 mg/kg b.w. Results β-sitosterol induced significant dose-dependent growth inhibition of COLO 320 DM cells (IC50 266.2 μM, induced apoptosis by scavenging reactive oxygen species, and suppressed the expression of β-catenin and PCNA antigens in human colon cancer cells. β-sitosterol supplementation reduced the number of aberrant crypt and crypt multiplicity in DMH-initiated rats in a dose-dependent manner with no toxic effects. Conclusion We found doses of 10-20 mg/kg b.w. β-sitosterol to be effective for future in vivo studies. β-sitosterol had chemopreventive potential by virtue of its radical quenching ability in vitro, with minimal toxicity to normal cells. It also attenuated β-catenin and PCNA expression, making it a potential anticancer drug for colon carcinogenesis.

  5. Chemopreventive potential of beta-Sitosterol in experimental colon cancer model--an in vitro and In vivo study.

    Science.gov (United States)

    Baskar, Albert A; Ignacimuthu, Savarimuthu; Paulraj, Gabriel M; Al Numair, Khalid S

    2010-06-04

    Asclepias curassavica Linn. is a traditional medicinal plant used by tribal people in the western ghats, India, to treat piles, gonorrhoea, roundworm infestation and abdominal tumours. We have determined the protective effect of beta-sitosterol isolated from A. curassavica in colon cancer, using in vitro and in vivo models. The active molecule was isolated, based upon bioassay guided fractionation, and identified as beta-sitosterol on spectral evidence. The ability to induce apoptosis was determined by its in vitro antiradical activity, cytotoxic studies using human colon adenocarcinoma and normal monkey kidney cell lines, and the expression of beta-catenin and proliferating cell nuclear antigen (PCNA) in human colon cancer cell lines (COLO 320 DM). The chemopreventive potential of beta-sitosterol in colon carcinogenesis was assessed by injecting 1,2-dimethylhydrazine (DMH, 20 mg/kg b.w.) into male Wistar rats and supplementing this with beta-sitosterol throughout the experimental period of 16 weeks at 5, 10, and 20 mg/kg b.w. beta-sitosterol induced significant dose-dependent growth inhibition of COLO 320 DM cells (IC50 266.2 microM), induced apoptosis by scavenging reactive oxygen species, and suppressed the expression of beta-catenin and PCNA antigens in human colon cancer cells. beta-sitosterol supplementation reduced the number of aberrant crypt and crypt multiplicity in DMH-initiated rats in a dose-dependent manner with no toxic effects. We found doses of 10-20 mg/kg b.w. beta-sitosterol to be effective for future in vivo studies. beta-sitosterol had chemopreventive potential by virtue of its radical quenching ability in vitro, with minimal toxicity to normal cells. It also attenuated beta-catenin and PCNA expression, making it a potential anticancer drug for colon carcinogenesis.

  6. Chemopreventive potential of β-Sitosterol in experimental colon cancer model - an In vitro and In vivo study

    Science.gov (United States)

    2010-01-01

    Background Asclepias curassavica Linn. is a traditional medicinal plant used by tribal people in the western ghats, India, to treat piles, gonorrhoea, roundworm infestation and abdominal tumours. We have determined the protective effect of β-sitosterol isolated from A. curassavica in colon cancer, using in vitro and in vivo models. Methods The active molecule was isolated, based upon bioassay guided fractionation, and identified as β-sitosterol on spectral evidence. The ability to induce apoptosis was determined by its in vitro antiradical activity, cytotoxic studies using human colon adenocarcinoma and normal monkey kidney cell lines, and the expression of β-catenin and proliferating cell nuclear antigen (PCNA) in human colon cancer cell lines (COLO 320 DM). The chemopreventive potential of β-sitosterol in colon carcinogenesis was assessed by injecting 1,2-dimethylhydrazine (DMH, 20 mg/kg b.w.) into male Wistar rats and supplementing this with β-sitosterol throughout the experimental period of 16 weeks at 5, 10, and 20 mg/kg b.w. Results β-sitosterol induced significant dose-dependent growth inhibition of COLO 320 DM cells (IC50 266.2 μM), induced apoptosis by scavenging reactive oxygen species, and suppressed the expression of β-catenin and PCNA antigens in human colon cancer cells. β-sitosterol supplementation reduced the number of aberrant crypt and crypt multiplicity in DMH-initiated rats in a dose-dependent manner with no toxic effects. Conclusion We found doses of 10-20 mg/kg b.w. β-sitosterol to be effective for future in vivo studies. β-sitosterol had chemopreventive potential by virtue of its radical quenching ability in vitro, with minimal toxicity to normal cells. It also attenuated β-catenin and PCNA expression, making it a potential anticancer drug for colon carcinogenesis. PMID:20525330

  7. Articular cartilage-derived cells hold a strong osteogenic differentiation potential in comparison to mesenchymal stem cells in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Salamon, Achim, E-mail: achim.salamon@med.uni-rostock.de [Department of Cell Biology, Rostock University Medical Center, Schillingallee 69, D-18057 Rostock (Germany); Jonitz-Heincke, Anika, E-mail: anika.jonitz@med.uni-rostock.de [Biomechanics and Implant Technology Research Laboratory, Department of Orthopedics, Rostock University Medical Center, Doberaner Straße 142, D-18057 Rostock (Germany); Adam, Stefanie, E-mail: stefanie.adam@med.uni-rostock.de [Department of Cell Biology, Rostock University Medical Center, Schillingallee 69, D-18057 Rostock (Germany); Rychly, Joachim, E-mail: joachim.rychly@med.uni-rostock.de [Department of Cell Biology, Rostock University Medical Center, Schillingallee 69, D-18057 Rostock (Germany); Müller-Hilke, Brigitte, E-mail: brigitte.mueller-hilke@med.uni-rostock.de [Institute of Immunology, Rostock University Medical Center, Schillingallee 68, D-18057 Rostock (Germany); Bader, Rainer, E-mail: rainer.bader@med.uni-rostock.de [Biomechanics and Implant Technology Research Laboratory, Department of Orthopedics, Rostock University Medical Center, Doberaner Straße 142, D-18057 Rostock (Germany); Lochner, Katrin, E-mail: katrin.lochner@med.uni-rostock.de [Biomechanics and Implant Technology Research Laboratory, Department of Orthopedics, Rostock University Medical Center, Doberaner Straße 142, D-18057 Rostock (Germany); Peters, Kirsten, E-mail: kirsten.peters@med.uni-rostock.de [Department of Cell Biology, Rostock University Medical Center, Schillingallee 69, D-18057 Rostock (Germany)

    2013-11-01

    Cartilaginous matrix-degenerative diseases like osteoarthritis (OA) are characterized by gradual cartilage erosion, and also by increased presence of cells with mesenchymal stem cell (MSC) character within the affected tissues. Moreover, primary chondrocytes long since are known to de-differentiate in vitro and to be chondrogenically re-differentiable. Since both findings appear to conflict with each other, we quantitatively assessed the mesenchymal differentiation potential of OA patient cartilage-derived cells (CDC) towards the osteogenic and adipogenic lineage in vitro and compared it to that of MSC isolated from adipose tissue (adMSC) of healthy donors. We analyzed expression of MSC markers CD29, CD44, CD105, and CD166, and, following osteogenic and adipogenic induction in vitro, quantified their expression of osteogenic and adipogenic differentiation markers. Furthermore, CDC phenotype and proliferation were monitored. We found that CDC exhibit an MSC CD marker expression pattern similar to adMSC and a similar increase in proliferation rate during osteogenic differentiation. In contrast, the marked reduction of proliferation observed during adipogenic differentiation of adMSC was absent in CDC. Quantification of differentiation markers revealed a strong osteogenic differentiation potential for CDC, however almost no capacity for adipogenic differentiation. Since in the pathogenesis of OA, cartilage degeneration coincides with high bone turnover rates, the high osteogenic differentiation potential of OA patient-derived CDC may affect clinical therapeutic regimens aiming at autologous cartilage regeneration in these patients. - Highlights: • We analyze the mesenchymal differentiation capacity of cartilage-derived cells (CDC). • CDC express mesenchymal stem cell (MSC) markers CD29, CD44, CD105, and CD166. • CDC and MSC proliferation is reduced in adipogenesis and increased in osteogenesis. • Adipogenic differentiation is virtually absent in CDC, but

  8. Isolation and in vitro selection of actinomycetes strains as potential probiotics for aquaculture

    Directory of Open Access Journals (Sweden)

    Milagro García Bernal

    2015-02-01

    Full Text Available Aim: This study was designed to describe a series of in vitro tests that may aid the discovery of probiotic strains from actinomycetes. Materials and Methods: Actinomycetes were isolated from marine sediments using four different isolation media, followed by antimicrobial activity and toxicity assessment by the agar diffusion method and the hemolysis of human blood cells, respectively. Extracellular enzymatic production was monitored by the hydrolysis of proteins, lipids and carbohydrates. Tolerance to different pH values and salt concentrations was also determined, followed by hydrophobicity analysis and genetic identification of the most promising strains. Results: Five out of 31 isolated strains showed antimicrobial activity against three Vibrio species. Three non-hemolytic strains (N7, RL8 and V4 among these active isolates yielded positive results in hydrophobicity tests and exhibited good growth at salt concentrations ranging from 0% to 10%, except strain RL8, which required a salt concentration >0.6%. Although these strains did not grow at pH<3, they showed different enzymatic activities. Phylogenetic analysis revealed that strains N7 and V4 have more than 99% identity with several Streptomyces species, whereas the closest matches to strain RL8 are Streptomyces panacagri and Streptomyces flocculus, with 98% and 98.2% similarity, respectively. Conclusion: Three actinomycetes strains showing probiotic-like properties were discovered using several in vitro tests that can be easily implemented in different institutions around the world.

  9. Optimization of conditions for in vitro development of Trichoderma viride-based biofilms as potential inoculants.

    Science.gov (United States)

    Triveni, Sodimalla; Prasanna, Radha; Saxena, Anil Kumar

    2012-09-01

    Biofilms represent mixed communities present in a diverse range of environments; however, their utility as inoculants is less investigated. Our investigation was aimed towards in vitro development of biofilms using fungal mycelia (Trichoderma viride) as matrices and nitrogen-fixing and P-solubilizing bacteria as partners, as a prelude to their use as biofertilizers (biofilmed biofertilizers, BBs) and biocontrol agents for different crops. The most suitable media in terms of population counts, fresh mass and dry biomass for Trichoderma and Bacillus subtilis/Pseudomonas fluorescens was found to be Pikovskaya broth ± 1 % CaCO(3), while for Trichoderma and Azotobacter chroococcum, Jensen's medium was most optimal. The respective media were then used for optimization of the inoculation rate of the partners in terms of sequence of addition of partners, fresh/dry mass of biofilms and population counts of partners for efficient film formation. Microscopic observations revealed significant differences in the progress of growth of biofilms and dual cultures. In the biofilms, the bacteria were observed growing intermingled within the fungal mycelia mat. Further, biofilm formation was compared under static and shaking conditions and the fresh mass of biofilms was higher in the former. Such biofilms are being further characterized under in vitro conditions, before using them as inoculants with crops.

  10. Comparison of the antioxidant potential of antiparkinsonian drugs in different in vitro models

    Directory of Open Access Journals (Sweden)

    Carine Coneglian de Farias

    2014-12-01

    Full Text Available Parkinson's disease (PD is characterized by progressive degeneration of dopaminergic neurons in the substantia nigra pars compacta. Furthermore, oxidative stress plays a role in PD, causing or contributing to the neurodegenerative process. Currently PD has only symptomatic treatment and still nothing can be done to stop the degenerative process of the disease. This study aimed to comparatively evaluate the antioxidant capacity of pramipexole, selegeline and amantadine in different in vitrostudies and to offer possible explanations on the molecular antioxidant mechanisms of these drugs. In vitro, the antioxidant capacity of the drugs was assessed by the ability of antiparkinsonian drugs to decrease or scavenge ROS in the neutrophil respiratory burst, ability of antiparkinsonian drugs to donate hydrogen and stabilize the free radical 2,2-diphenyl-1-picryl-hydrazyl (DPPH•, to scavenge 2,2'-azino-di-(3-ethylbenzthiazoline-6-sulphonic acid (ABTS+ and evaluation of the ferric reducing antioxidant power (FRAP. This study demonstrated that both pramipexole and selegiline, but not amantadine, have antioxidant effects in vitro by scavenging superoxide anion on the respiratory burst, donating electron in the ABTS+ assay and presenting ferric reduction antioxidant power. This chemical structure-related antioxidant capacity suggests a possible neuroprotective mechanism of these drugs beyond their already recognized mechanism of action.

  11. Antioxidant potential of bitter cumin (Centratherum anthelminticum (L. Kuntze seeds in in vitro models

    Directory of Open Access Journals (Sweden)

    Naidu Kamatham A

    2011-05-01

    Full Text Available Abstract Background Bitter cumin (Centratherum anthelminticum (L. Kuntze, is a medicinally important plant. Earlier, we have reported phenolic compounds, antioxidant, and anti-hyperglycemic, antimicrobial activity of bitter cumin. In this study we have further characterized the antioxidative activity of bitter cumin extracts in various in vitro models. Methods Bitter cumin seeds were extracted with a combination of acetone, methanol and water. The antioxidant activity of bitter cumin extracts were characterized in various in vitro model systems such as DPPH radical, ABTS radical scavenging, reducing power, oxidation of liposomes and oxidative damage to DNA. Results The phenolic extracts of bitter cumin at microgram concentration showed significant scavenging of DPPH and ABTS radicals, reduced phosphomolybdenum (Mo(VI to Mo(V, ferricyanide Fe(III to Fe(II, inhibited liposomes oxidation and hydroxyl radical induced damage to prokaryotic genomic DNA. The results showed a direct correlation between phenolic acid content and antioxidant activity. Conclusion Bitter cumin is a good source of natural antioxidants.

  12. An evaluation of the sonoporation potential of low-boiling point phase-change ultrasound contrast agents in vitro.

    Science.gov (United States)

    Fix, Samantha M; Novell, Anthony; Yun, Yeoheung; Dayton, Paul A; Arena, Christopher B

    2017-01-01

    Phase-change ultrasound contrast agents (PCCAs) offer a solution to the inherent limitations associated with using microbubbles for sonoporation; they are characterized by prolonged circulation lifetimes, and their nanometer-scale sizes may allow for passive accumulation in solid tumors. As a first step towards the goal of extravascular cell permeabilization, we aim to characterize the sonoporation potential of a low-boiling point formulation of PCCAs in vitro. Parameters to induce acoustic droplet vaporization and subsequent microbubble cavitation were optimized in vitro using high-speed optical microscopy. Sonoporation of pancreatic cancer cells in suspension was then characterized at a range of pressures (125-600 kPa) and pulse lengths (5-50 cycles) using propidium iodide as an indicator molecule. We achieved sonoporation efficiencies ranging from 8 ± 1% to 36 ± 4% (percent of viable cells), as evidenced by flow cytometry. Increasing sonoporation efficiency trended with increasing pulse length and peak negative pressure. We conclude that PCCAs can be used to induce the sonoporation of cells in vitro, and our results warrant further investigation into the use of PCCAs as extravascular sonoporation agents in vivo.

  13. The potentiality of two-dimensional preantral follicle culture as an in vitro model in predicting premature ovarian failure.

    Science.gov (United States)

    Zhang, Ting; Chen, Ying; Yang, Yang; Wang, Zhonghui; Pan, Qi; Xu, Sichong; Sun, Zuyue

    2017-09-05

    The purpose of this study is to identify the potential of a two-dimensional preantral follicle culture as an in vitro model of predicting premature ovarian failure. The two-dimensional preantral follicles culture method was established by cultivating preantral follicles collected from ICR F1 hybrids (aged 12-14days) for 12days. The preantral follicles were incubated with 0.54mg/ml cyclophosphamide, 0.5mg/ml busulfan, 0.12mg/ml cisplatin, 3.12mg/ml 4-vinylcyclohexene diepoxide, 5mg/ml D (+) galactose, and 0.5mg/ml hydrocortisone for 24h at culture days 2, 6 and 11. The diameter of follicles, the cumulus-oocyte complex number and the maturity of oocytes were recorded as the parameters to detect follicular maturation induced by the culture agents. The results indicated that, except for busulfan, D (+) galactose, and hydrocortisone, such test articles could significantly decrease follicular growth (pculture could be utilized as a potential in vitro system to mimic the POF model. It may also be employed in screening potential ovarian toxic agents, reducing laboratory animal use and promoting animal welfare. Copyright © 2017 Elsevier GmbH. All rights reserved.

  14. Evaluation of Nystatin Containing Chitosan Hydrogels as Potential Dual Action Bio-Active Restorative Materials: in Vitro Approach

    Directory of Open Access Journals (Sweden)

    V. Tamara Perchyonok

    2014-11-01

    Full Text Available Healing is a specific biological process related to the general phenomenon of growth and tissue regeneration and is a process generally affected by several systemic conditions or as detrimental side-effects of chemotherapy- and radiotherapy-induced inflammation of the oral mucosa. The objectives of this study is to evaluate the novel chitosan based functional drug delivery systems, which can be successfully incorporated into “dual action bioactive restorative materials”, capable of inducing in vitro improved wound healing prototype and containing an antibiotic, such as nystatin, krill oil as an antioxidant and hydroxyapatite as a molecular bone scaffold, which is naturally present in bone and is reported to be successfully used in promoting bone integration when implanted as well as promoting healing. The hydrogels were prepared using a protocol as previously reported by us. The physico-chemical features, including surface morphology (SEM, release behaviors, stability of the therapeutic agent-antioxidant-chitosan, were measured and compared to the earlier reported chitosan-antioxidant containing hydrogels. Structural investigations of the reactive surface of the hydrogel are reported. Release of nystatin was investigated for all newly prepared hydrogels. Bio-adhesive studies were performed in order to assess the suitability of these designer materials. Free radical defense capacity of the biomaterials was evaluated using established in vitro model. The bio-adhesive capacity of the materials in the in vitro system was tested and quantified. It was found that the favorable synergistic effect of free radical built-in defense mechanism of the new functional materials increased sustainable bio-adhesion and therefore acted as a functional multi-dimensional restorative material with potential application in wound healing in vitro.

  15. Human Spinal Bone Dust as a Potential Local Autograft: In vitro Potent Anabolic Effect on Human Osteoblasts.

    Science.gov (United States)

    Gao, Ryan; Street, Matthew; Tay, Mei Lin; Callon, Karen E; Naot, Dorit; Lock, Alistair; Munro, Jacob T; Cornish, Jillian; Ferguson, John; Musson, David

    2017-07-18

    In Vitro Study. To evaluate the effect that factors released from human posterior spinal bone dust have on primary human osteoblast growth and maturation. Bone dust, created during spinal fusion surgeries has the potential to be used as an autologous bone graft by providing a source of viable autologous osteoblasts and mesenchymal stem cells with osteogenic potential. To date, no information is available on whether bone dust also provides a source of anabolic factors with the potential to enhance osteoblast proliferation and maturation, which would enhance its therapeutic potential. Bone dust was collected from consenting patients undergoing elective posterior spinal fusion surgeries, and primary human osteoblasts were cultured from patients undergoing elective hip or knee arthroplasty. Growth factors and cytokines released by bone dust were quantified using enzyme-linked immunosorbent assay (ELISA). Primary human osteoblast proliferation and gene expression in response to bone dust were assessed using H-thymidine incorporation and real-time polymerase chain reaction (qPCR), respectively. Human bone dust released anabolic cytokines (IL-1β and IL-6) and growth factors (TGF-β, VEGF, FGF-Basic and PDGF-BB) in increasing concentrations over a 7-day period. In vitro, the anabolic factors released by bone dust increased osteoblast proliferation by 7-fold, compared with osteoblasts cultured alone. In addition, the factors released from bone dust up-regulated a number of osteoblastic genes integral to osteoblast differentiation, maturation and angiogenesis. This study is the first to demonstrate that human posterior spinal bone dust released anabolic factors that potently enhance osteoblast proliferation and the expression of genes that favor bone healing and bone union. Given that bone dust is anabolic and its harvest is fast, simple, and safe to perform, spinal surgeons should be encouraged to 'recycle' bone dust and harness the regenerative potential of this free

  16. Probiotic potential of Lactobacilli with antagonistic activity against pathogenic strains: An in vitro validation for the production of inhibitory substances.

    Science.gov (United States)

    Prabhurajeshwar, Chidre; Chandrakanth, Revanasiddappa Kelmani

    2017-10-01

    Probiotics, live cells with different beneficiary characteristics, have been extensively studied and explored commercially in many different products in the world. Their benefits to human and animal health have proven in hundreds of scientific studies. Based on rich bibliographic material, Curd is the potential source of probiotic Lactobacilli. The aim of the present study was to observe Lactobacilli with probiotic potential activities from different curd samples for isolation, identification and characterization of Lactobacillus species. Among the samples, thirty lactic acid bacterial strains were isolated, sixteen (16/30) best Lactobacillus isolates were selected by preliminary screening as potential probiotic for acid and bile tolerance, further confirmed using 16s rRNA identification. All the selected Lactobacillus isolates were then characterized in vitro for their probiotic characteristics and antimicrobial activities against pathogens and aggregation studies. The results indicated that selected potential probiotic isolates (T2, T4 and T16) were screened and confirmed as Lactobacillus. The isolates produced positive tolerance to excited pH, NaCl and bile salts, also revealed noticeable antimicrobial activities against pathogens. All the Lactobacillus isolates were susceptible to clinical antibiotics used. Besides, T2 isolate was constituted to retain stronger auto and co-aggregation and cell surface hydrophobicity capacity. Based on the drawn results, T2, T4 and T16 Lactobacillus isolates were recognised as ideal, potential in vitro antimicrobial probiotic isolates against pathogens and studies are needed further in-vivo assessment and human health benefits in their real-life situations. Copyright © 2017 Chang Gung University. Published by Elsevier B.V. All rights reserved.

  17. Identification of potential biomarkers in donor cows for in vitro embryo production by granulosa cell transcriptomics

    DEFF Research Database (Denmark)

    Mazzoni, Gianluca; Salleh, Suraya M; Freude, Kristine

    2017-01-01

    The Ovum Pick Up-In vitro Production (OPU-IVP) of embryos is an advanced reproductive technology used in cattle production but the complex biological mechanisms behind IVP outcomes are not fully understood. In this study we sequenced RNA of granulosa cells collected from Holstein cows at oocyte...... aspiration prior to IVP, to identify candidate genes and biological mechanisms for favourable IVP-related traits in donor cows where IVP was performed separately for each animal. We identified 56 genes significantly associated with IVP scores (BL rate, kinetic and morphology). Among these, BEX2, HEY2, RGN......, TNFAIP6 and TXNDC11 were negatively associated while Mx1 and STC1 were positively associated with all IVP scores. Functional analysis highlighted a wide range of biological mechanisms including apoptosis, cell development and proliferation and four key upstream regulators (COX2, IL1, PRL, TRIM24...

  18. In vitro assessment of TAT - Ciliary Neurotrophic Factor therapeutic potential for peripheral nerve regeneration.

    Science.gov (United States)

    Barbon, Silvia; Stocco, Elena; Negro, Alessandro; Dalzoppo, Daniele; Borgio, Luca; Rajendran, Senthilkumar; Grandi, Francesca; Porzionato, Andrea; Macchi, Veronica; De Caro, Raffaele; Parnigotto, Pier Paolo; Grandi, Claudio

    2016-10-15

    In regenerative neurobiology, Ciliary Neurotrophic Factor (CNTF) is raising high interest as a multifunctional neurocytokine, playing a key role in the regeneration of injured peripheral nerves. Despite its promising trophic and regulatory activity, its clinical application is limited by the onset of severe side effects, due to the lack of efficient intracellular trafficking after administration. In this study, recombinant CNTF linked to the transactivator transduction domain (TAT) was investigated in vitro and found to be an optimized fusion protein which preserves neurotrophic activity, besides enhancing cellular uptake for therapeutic advantage. Moreover, a compelling protein delivery method was defined, in the future perspective of improving nerve regeneration strategies. Following determination of TAT-CNTF molecular weight and concentration, its specific effect on neural SH-SY5Y and PC12 cultures was assessed. Cell proliferation assay demonstrated that the fusion protein triggers PC12 cell growth within 6h of stimulation. At the same time, the activation of signal transduction pathway and enhancement of cellular trafficking were found to be accomplished in both neural cell lines after specific treatment with TAT-CNTF. Finally, the recombinant growth factor was successfully loaded on oxidized polyvinyl alcohol (PVA) scaffolds, and more efficiently released when polymer oxidation rate increased. Taken together, our results highlight that the TAT domain addiction to the protein sequence preserves CNTF specific neurotrophic activity in vitro, besides improving cellular uptake. Moreover, oxidized PVA could represent an ideal biomaterial for the development of nerve conduits loaded with the fusion protein to be delivered to the site of nerve injury. Copyright © 2016 Elsevier Inc. All rights reserved.

  19. In-vitro anti-microbial and brine-shrimp lethality potential of the ...

    African Journals Online (AJOL)

    The brine-shrimp lethality assay analyzed using the Finney probit method showed that the crude ethanolic extracts of the leaves and stem displayed LD50 values at 192 ppm and 182 ppm respectively. These findings indicate the potential of the plant as panacea for infectious diseases and also reveal a novel potential in the ...

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

    Directory of Open Access Journals (Sweden)

    Masako Fujioka-Kobayashi

    2016-11-01

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

  1. Hydrogen gas treatment prolongs replicative lifespan of bone marrow multipotential stromal cells in vitro while preserving differentiation and paracrine potentials

    Energy Technology Data Exchange (ETDEWEB)

    Kawasaki, Haruhisa [Department of Pathology, The Ohio State University, Columbus, OH 43210 (United States); Davis Heart and Lung Research Institute, The Ohio State University, Columbus, OH 43210 (United States); Guan, Jianjun [Department of Materials Science and Engineering, The Ohio State University, Columbus, OH 43210 (United States); Tamama, Kenichi, E-mail: kenichi.tamama@osumc.edu [Department of Pathology, The Ohio State University, Columbus, OH 43210 (United States); Davis Heart and Lung Research Institute, The Ohio State University, Columbus, OH 43210 (United States); Center for Stem Cell and Regenerative Medicine, The Ohio State University, Columbus, OH 43210 (United States)

    2010-07-02

    Cell therapy with bone marrow multipotential stromal cells/mesenchymal stem cells (MSCs) represents a promising approach in the field of regenerative medicine. Low frequency of MSCs in adult bone marrow necessitates ex vivo expansion of MSCs after harvest; however, such a manipulation causes cellular senescence with loss of differentiation, proliferative, and therapeutic potentials of MSCs. Hydrogen molecules have been shown to exert organ protective effects through selective reduction of hydroxyl radicals. As oxidative stress is one of the key insults promoting cell senescence in vivo as well as in vitro, we hypothesized that hydrogen molecules prevent senescent process during MSC expansion. Addition of 3% hydrogen gas enhanced preservation of colony forming early progenitor cells within MSC preparation and prolonged the in vitro replicative lifespan of MSCs without losing differentiation potentials and paracrine capabilities. Interestingly, 3% hydrogen gas treatment did not decrease hydroxyl radical, protein carbonyl, and 8-hydroxydeoxyguanosine, suggesting that scavenging hydroxyl radical might not be responsible for these effects of hydrogen gas in this study.

  2. Prediction of the developmental potential of hamster embryos in vitro by precise timing of the third cell cycle.

    Science.gov (United States)

    Gonzales, D S; Pinheiro, J C; Bavister, B D

    1995-09-01

    Time-lapse videomicrography was used to determine the timing of early developmental events in hamster embryos in vitro. The time intervals from pronuclear envelope breakdown to the completion of the first cleavage (Dt2), second cleavage (Dt4 = 2-4 cells), third cleavage (Dt8 = 4-8 cells), blastocyst formation, and zona escape were precisely measured to determine whether the variable 'time' (t) can be used to predict the developmental potential of preimplantation embryos. The range of the developmental time interval (Dt) from the second to the third cleavage divisions (Dt8) provided the best indicator for predicting the probabilities of blastocyst formation and zona escape (P = 0.015 and 0.041, respectively). Dt8 was subdivided into consecutive time cutoff points of < or = 750, < or = 800, < or = 850 and < or = 900 min. Of the embryos that took < or = 750 min to complete the third cleavage division, 92% developed into blastocysts and 69% escaped from their zonae pellucidae. When the completion of Dt8 extended to < or = 900 min, the percentages decreased to 75% and 49% for blastocyst formation and zona escape, respectively. This study identifies a specific developmental time interval and a model whereby time can be used as a noninvasive parameter to predict embryo developmental potential in vitro.

  3. In vitro effects of waterpipe smoke condensate on endothelial cell function: a potential risk factor for vascular disease.

    Science.gov (United States)

    Rammah, Mayyasa; Dandachi, Farah; Salman, Rola; Shihadeh, Alan; El-Sabban, Marwan

    2013-05-23

    Despite its increasing popularity, little is known about the health effects of waterpipe smoking (WPS), particularly on the cardiovascular system. To investigate the role of WPS as a risk factor for vascular disease, we evaluated its effect on endothelial cell function, which is an early event in vascular disease pathogenesis. We assessed the changes in cell viability, ROS generation, inflammatory and vasodilatory markers and in vitro angiogenesis of human aortic endothelial cells in response to waterpipe smoke condensate exposure. Mainstream waterpipe smoke condensate (WSC) was generated using a standard laboratory machine protocol. Compared to control, WSC induced cell cycle arrest, apoptosis, and oxidative stress in human primary endothelial cells. In addition, we assayed for impaired endothelium-dependent vasodilation and induced inflammation by studying the effect of WPS on the content and activity of AMPK, eNOS proteins and NF-κB p65 ser536 phosphorylation, respectively. WSC inhibited AMPK/eNOS phosphorylation and induced phosphorylation of p65. Moreover, we evaluated endothelial cells repair mechanism related properties that include migration/invasion and in vitro tube formation upon treatment with WSC. WSC reduced the motility and inhibited angiogenic potential of HAEC cells. WPS induced endothelial cell dysfunction as evident by exerting oxidative stress, inflammation, and impaired endothelial vasodilatory function and repair mechanisms. All together these data provide evidence for the potential contribution of WPS to endothelial dysfunction and thus to vascular disease. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  4. Potentiation of 5-fluorouracil encapsulated in zeolites as drug delivery systems for in vitro models of colorectal carcinoma.

    Science.gov (United States)

    Vilaça, Natália; Amorim, Ricardo; Machado, Ana F; Parpot, Pier; Pereira, Manuel F R; Sardo, Mariana; Rocha, João; Fonseca, António M; Neves, Isabel C; Baltazar, Fátima

    2013-12-01

    The studies of potentiation of 5-fluorouracil (5-FU), a traditional drug used in the treatment of several cancers, including colorectal (CRC), were carried out with zeolites Faujasite in the sodium form, with different particle sizes (NaY, 700nm and nanoNaY, 150nm) and Linde type L in the potassium form (LTL) with a particle size of 80nm. 5-FU was loaded into zeolites by liquid-phase adsorption. Characterization by spectroscopic techniques (FTIR, (1)H NMR and (13)C and (27)Al solid-state MAS NMR), chemical analysis, thermal analysis (TGA), nitrogen adsorption isotherms and scanning electron microscopy (SEM), demonstrated the successful loading of 5-FU into the zeolite hosts. In vitro drug release studies (PBS buffer pH 7.4, 37°C) revealed the release of 80-90% of 5-FU in the first 10min. To ascertain the drug release kinetics, the release profiles were fitted to zero-order, first-order, Higuchi, Hixson-Crowell, Korsmeyer-Peppas and Weibull kinetic models. The in vitro dissolution from the drug delivery systems (DDS) was explained by the Weibull model. The DDS efficacy was evaluated using two human colorectal carcinoma cell lines, HCT-15 and RKO. Unloaded zeolites presented no toxicity to both cancer cells, while all DDS allowed an important potentiation of the 5-FU effect on the cell viability. Immunofluorescence studies provided evidence for zeolite-cell internalization. Copyright © 2013 Elsevier B.V. All rights reserved.

  5. Evaluation of the spoilage potential of bacteria isolated from chilled chicken in vitro and in situ.

    Science.gov (United States)

    Wang, Guang-Yu; Wang, Hu-Hu; Han, Yi-Wei; Xing, Tong; Ye, Ke-Ping; Xu, Xing-Lian; Zhou, Guang-Hong

    2017-05-01

    Microorganisms play an important role in the spoilage of chilled chicken. In this study, a total of 53 isolates, belonging to 7 species of 3 genera, were isolated using a selective medium based on the capacity to spoil chicken juice. Four isolates, namely Aeromonas salmonicida 35, Pseudomonas fluorescens H5, Pseudomonas fragi H8 and Serratia liquefaciens 17, were further characterized to assess their proteolytic activities in vitro using meat protein extracts and to evaluate their spoilage potential in situ. The in vitro studies showed that A. salmonicida 35 displayed the strongest proteolytic activity against both sarcoplasmic and myofibrillar proteins. However, the major spoilage isolate in situ was P. fragi H8, which exhibited a fast growth rate, slime formation and increased pH and total volatile basic nitrogen (TVBN) on chicken breast fillets. The relative amounts of volatile organic compounds (VOCs) originating from the microorganisms, including alcohols, aldehydes, ketones and several sulfur compounds, increased during storage. In sum, this study demonstrated the characteristics of 4 potential spoilage bacteria on chilled yellow-feather chicken and provides a simple and convenient method to assess spoilage bacteria during quality management. Copyright © 2016 Elsevier Ltd. All rights reserved.

  6. Inhibition of Autophagy Potentiates Atorvastatin-Induced Apoptotic Cell Death in Human Bladder Cancer Cells in Vitro

    Directory of Open Access Journals (Sweden)

    Minyong Kang

    2014-05-01

    Full Text Available Statins are cholesterol reduction agents that exhibit anti-cancer activity in several human cancers. Because autophagy is a crucial survival mechanism for cancer cells under stress conditions, cooperative inhibition of autophagy acts synergistically with other anti-cancer drugs. Thus, this study investigates whether combined treatment of atorvastatin and autophagy inhibitors results in enhancing the cytotoxic effects of atorvastatin, upon human bladder cancer cells, T24 and J82, in vitro. To measure cell viability, we performed the EZ-Cytox cell viability assay. We examined apoptosis by flow cytometry using annexin-V/propidium iodide (PI and western blot using procaspase-3 and poly (ADP-ribose polymerase (PARP antibodies. To examine autophagy activation, we evaluated the co-localization of LC3 and LysoTracker by immunocytochemistry, as well as the expression of LC3 and p62/sequestosome-1 (SQSTM1 by western blot. In addition, we assessed the survival and proliferation of T24 and J82 cells by a clonogenic assay. We found that atorvastatin reduced the cell viability of T24 and J82 cells via apoptotic cell death and induced autophagy activation, shown by the co-localization of LC3 and LysoTracker. Moreover, pharmacologic inhibition of autophagy significantly enhanced atorvastatin-induced apoptosis in T24 and J82 cells. In sum, inhibition of autophagy potentiates atorvastatin-induced apoptotic cell death in human bladder cancer cells in vitro, providing a potential therapeutic approach to treat bladder cancer.

  7. Erosive potential of soft drinks on human enamel: An in vitro study

    Directory of Open Access Journals (Sweden)

    Yin-Lin Wang

    2014-11-01

    Conclusion: All tested soft drinks were found to be erosive. Soft drinks with high calcium contents have significantly lower erosive potential. Low pH value and high citrate content may cause more surface enamel loss. As the erosive time increased, the titratable acidity to pH 7 may be a predictor of the erosive potential for acidic soft drinks. The erosive potential of the soft drinks may be predicted based on the types of acid content, pH value, titratable acidity, and ion concentration.

  8. Calcium-dependent plateau potentials in rostral ambiguus neurons in the newborn mouse brain stem in vitro

    DEFF Research Database (Denmark)

    Rekling, J C; Feldman, J L

    1997-01-01

    Calcium-dependent plateau potentials in rostral ambiguus neurons in the newborn mouse brain stem in vitro. J. Neurophysiol. 78: 2483-2492, 1997. The nucleus ambiguus contains vagal and glossopharyngeal motoneurons and preganglionic neurons involved in respiration, swallowing, vocalization......, and control of heart beat. Here we show that the rostral compact formation's ambiguus neurons, which control the esophageal phase of swallowing, display calcium-dependent plateau potentials in response to tetanic orthodromic stimulation or current injection. Whole cell recordings were made from visualized...... neurons in the rostral nucleus ambiguus using a slice preparation from the newborn mouse. Biocytin-labeling revealed dendritic trees with pronounced rostrocaudal orientations confined to the nucleus ambiguus, a morphological profile matching that of vagal motoneurons projecting to the esophagus. Single...

  9. In vivo and In vitro Identification of Endocannabinoid Signaling in Periodontal Tissues and Their Potential Role in Local Pathophysiology.

    Science.gov (United States)

    Konermann, Anna; Jäger, Andreas; Held, Stefanie A E; Brossart, P; Schmöle, Anne

    2017-03-14

    The endocannabinoid system (ECS) with its binding receptors CB1 and CB2 impacts multiple pathophysiologies not only limited to neuronal psychoactivity. CB1 is assigned to cerebral neuron action, whereas CB2 is mainly expressed in different non-neuronal tissues and associated with immunosuppressive effects. Based on these tissue-selective CB receptor roles, it was the aim of this study to analyze potential expression in periodontal tissues under physiological conditions and inflammatory states. In vivo, CB receptor expression was investigated on human periodontal biopsies with or without bacterial inflammation and on rat maxillae with or without sterile inflammation. In vitro analyses were performed on human periodontal ligament (PDL) cells at rest or under mechanical strain via qRT-PCR, Western blot, and immunocytochemistry. P periodontal tissues, both adjusted by different entities of periodontal inflammation and by mechanical stress. This indicates potential ECS function as regulatory tool in controlling of periodontal pathophysiology.

  10. The Measurement and Research of Surface Potentials of Human Tooth in vitro

    Science.gov (United States)

    2001-10-25

    dental hard tissue, no mater enamel , dentin or cementum, is formed mostly by the mineral, hydroxyapatite . It is soaked in the electrolyte surroundings...property made by Klein and Amberson, 1932, had suggested that the dental enamel was an electrostatic ion screen and it had permselectivity[1]. Then...developed the surface potentials of tooth. A preliminary study of the dental electrochemical property in vivo had found that the potential of enamel lesion

  11. Evaluating the anti-inflammatory potential of Tectaria cicutaria L. rhizome extract in vitro as well as in vivo.

    Science.gov (United States)

    Choudhari, Amit S; Raina, Prerna; Deshpande, Manasi M; Wali, Ashok G; Zanwar, Anand; Bodhankar, Subhash L; Kaul-Ghanekar, Ruchika

    2013-10-28

    The rhizome of Tectaria cicutaria has been used in the folklore system of Indian traditional medicine (Ayurveda) for the treatment of various disorders such as rheumatic pain, chest complaints, burns, sprain, poisonous bites, tonsilitis, toothache, gum complaints, cuts and wounds. The present work has for the first time tried to elucidate the anti-inflammatory potential of aqueous extract of Tectaria cicutaria rhizome (TCRaq) in vitro as well as in vivo. Anti-inflammatory potential of TCRaq was analyzed in vivo in carrageenan induced rat paw edema model. Serum antioxidant status in TCRaq-treated as well as untreated control rodents was measured by oxygen radical absorbance capacity (ORAC) assay. In vitro experiments for analyzing the anti-inflammatory potential of TCRaq were performed on murine macrophage cell line, RAW 264.7. Analysis of nitric oxide release in RAW 264.7 cells was done by Griess reaction. RT-PCR and western blotting experiment was performed to analyze the expression of iNOS. Expression of COX-2 and NFκB proteins was evaluated by western blotting. TCRaq significantly reduced the paw volume in Sprague-Dawley rats at a dose of 200mg/kg body weight, which was comparable with the standard diclofenac treatment. The rats treated with TCRaq showed a significant increase in the serum antioxidant levels compared to the untreated control animals. TCRaq was able to reduce the nitric oxide (NO) levels in RAW 264.7 cells that had been stimulated with lipopolysaccharide (LPS). This was accompanied by a corresponding decrease in iNOS expression at mRNA and protein level. Interestingly, TCRaq was found to decrease the expression of COX-2 as well as the nuclear translocation of NFκB in RAW 264.7 cells. Our study signifies the anti-inflammatory potential of Tectaria cicutaria and scientifically validates its traditional use in inflammatory conditions. © 2013 Elsevier Ireland Ltd. All rights reserved.

  12. In vitro inhibitory potential of selected Malaysian plants against key enzymes involved in hyperglycemia and hypertension.

    Science.gov (United States)

    Loh, S P; Hadira, O

    2011-04-01

    This study was conducted to determine the inhibitory potential of selected Malaysian plants against key enzymes related to type 2 diabetes and hypertension. The samples investigated were pucuk ubi (Manihot esculenta), pucuk betik (Carica papaya), ulam raja (Cosmos caudatus), pegaga (Centella asiatica) and kacang botol (Psophocarpus tetragonolobus). The inhibitory potential of hexane and dichloromethane extracts against the enzymes were determined by using alpha-amylase, alpha-glucosidase and angiotensin I-converting enzyme (ACE) inhibition assay. In alpha-amylase inhibition assay, the inhibitory potential was highest in pucuk ubi for both hexane (59.22%) and dichloromethane extract (54.15%). Hexane extract of pucuk ubi (95.01%) and dichloromethane extract of kacang botol (38.94%) showed the highest inhibitory potential against alpha-glucosidase, while in ACE inhibition assay, the inhibitory potential was highest in hexane extract of pegaga (48.45%) and dichloromethane extract of pucuk betik (59.77%). This study suggests a nutraceutical potential of some of these plants for hyperglycemia and hypertension prevention associated with type 2 diabetes.

  13. In vitro characterization of the immunotoxic potential of several perfluorinated compounds (PFCs)

    Energy Technology Data Exchange (ETDEWEB)

    Corsini, Emanuela, E-mail: emanuela.corsini@unimi.it [Laboratory of Toxicology, Department of Pharmacological Sciences, Università degli Studi di Milano, Via Balzaretti 9, 20133 Milano (Italy); Sangiovanni, Enrico [Laboratory of Pharmacognosy, Department of Pharmacological Sciences, Università degli Studi di Milano, Via Balzaretti 9, 20133 Milano (Italy); Avogadro, Anna; Galbiati, Valentina; Viviani, Barbara; Marinovich, Marina; Galli, Corrado L. [Laboratory of Toxicology, Department of Pharmacological Sciences, Università degli Studi di Milano, Via Balzaretti 9, 20133 Milano (Italy); Dell' Agli, Mario [Laboratory of Pharmacognosy, Department of Pharmacological Sciences, Università degli Studi di Milano, Via Balzaretti 9, 20133 Milano (Italy); Germolec, Dori R. [National Toxicology Program, National Institute of Environmental Health Sciences, NIH, RTP, NC (United States)

    2012-01-15

    We have previously shown that PFOA and PFOS directly suppress cytokine secretion in immune cells, with different mechanisms of action. In particular, we have demonstrated a role for PPAR-α in PFOA-induced immunotoxicity, and that PFOS has an inhibitory effect on LPS-induced I-κB degradation. These studies investigate the immunomodulatory effects of four other PFCs, namely PFBS, PFOSA, PFDA, and fluorotelomer using in vitro assays. The release of the pro-inflammatory cytokines IL-6 and TNF-α was evaluated in lipolysaccharide (LPS)-stimulated human peripheral blood leukocytes (hPBL) and in the human promyelocytic cell line THP-1, while the release of IL-10 and IFN-γ was evaluated in phytohemagglutinin (PHA)-stimulated hPBL. All PFCs suppressed LPS-induced TNF-α production in hPBL and THP-1 cells, while IL-6 production was suppressed by PFOSA, PFOS, PFDA and fluorotelomer. PFBS, PFOSA, PFOS, PFDA and fluorotelomer inhibited PHA-induced IL-10 release, while IFN-γ secretion was affected by PFOSA, PFOS, PFDA and fluorotelomer. Leukocytes obtained from female donors appear to be more sensitive to the in vitro immunotoxic effects of PFCs when their responses are compared to the results obtained using leukocytes from male donors. Mechanistic investigations demonstrated that inhibition of TNF-α release in THP-1 cells occurred at the transcriptional level. All PFCs, including PFOA and PFOS, decreased LPS-induced NF-κB activation. With the exception of PFOA, none of the PFCs tested was able to activate PPARα driven transcription in transiently transfected THP-1 cells, excluding a role for PPARα in the immunomodulation observed. PFBS and PFDA prevented LPS-induced I-κB degradation. Overall, these studies suggest that PFCs affect NF-κB activation, which directly suppresses cytokine secretion by immune cells. Our results indicate that PFOA is the least active of the PFCs examined followed by PFBS, PFDA, PFOS, PFOSA and fluorotelomer. -- Research Highlights: ► PFCs

  14. In vitro assessment of pharmaceutical potential of ethosomes entrapped with terbinafine hydrochloride.

    Science.gov (United States)

    Iizhar, Syed Ahmed; Syed, Ismail Ahmed; Satar, Rukhsana; Ansari, Shakeel Ahmed

    2016-05-01

    The present study investigates the entrapment of terbinafine hydrochloride (TH) in ethosomal vesicles via unsonicated and sonication method. Carbopol 934P was incorporated in the best formulation, F6, obtained by sonication method. The formulated ethosomal gel obtained as such i.e. F6(∗) was exploited to achieve a zero order release profile of TH. The composition includes phospholipid, ethanol and propylene glycol. Drug entrapment efficiency (DEE), in-vitro and ex-vivo drug diffusion studies, FT-IR and stability studies of the prepared ethosomes were investigated. The size and shape of F6 ethosomes vesicles were characterized by SEM. In-vitro drug release studies were performed using sigma dialysis membrane in phosphate buffer, pH 7.4 for 12 h while drug content was determined by HPLC. DEE was ranked from 55.33 ± 1.32% to 69.11 ± 2.11%. Highest DEE was seen with F6 ethosomal formulation with a vesicle size of 248 ± 1.02 nm. FT-IR studies confirmed that there was no chemical interaction between drug and excipients used in the formulation. Ex-vivo result suggested that drug diffusion observed after 12 h from F6(∗) and marketed cream (MR) formulations was 74.01 ± 0.62% and 61.45 ± 0.86%, respectively. The results of similarity factor (f 2 values) for MR and F6(∗) ethosomal gel were 85.14 and 42.63, respectively. It revealed that F6(∗) showed dissimilar dissolution profiles. Transdermal flux value for F6(∗) and MR was found to be 144.61 ± 1.28 μg/cm(2)/h and 121.6 ± 1.16 μg/cm(2)/h, respectively. This study disclosed that F6(∗) resides at targeted site for a relatively longer period of time thereby signifying the improved patient compliance.

  15. In vitro inhibitory effect on digestive enzymes and antioxidant potential of commonly consumed fruits.

    Science.gov (United States)

    Podsędek, Anna; Majewska, Iwona; Redzynia, Małgorzata; Sosnowska, Dorota; Koziołkiewicz, Maria

    2014-05-21

    Dietary inhibitors of fats and carbohydrates degrading enzymes can reduce obesity and type 2 diabetes. In this study, we screened crude extracts from 30 commonly consumed fruits to test their in vitro inhibitory effect against key enzymes relevant for obesity (pancreatic lipase) and type 2 diabetes (α-glucosidase and α-amylase), total phenolic content (Folin-Ciocalteu method), and antioxidant capacity (ABTS and FRAP). The IC50 values of the fruits tested varied from 39.91 to >400 mg/mL, from 1.04 to >80 mg/mL, and from 0.72 to 135.07 mg/mL against α-glucosidase, α-amylase, and pancreatic lipase, respectively. Antioxidant capacity ranged from 0.66 to 124.66 μmol of TE/g of fruit and strongly correlated with phenolic content, while the enzyme inhibition was poorly correlated with total phenolic and antioxidant capacity. Among fruits tested, blue honeysuckle and red gooseberry exhibited the highest inhibitory activity with respect to the carbohydrate degrading enzymes, while lingonberry had the strongest anti-lipase activity.

  16. Hydroxycinnamate Conjugates as Potential Monolignol Replacements: In vitro Lignification and Cell Wall Studies with Rosmarinic Acid

    Energy Technology Data Exchange (ETDEWEB)

    Yuki, Tobimatsu; Sasikumar, Elumalai; Grabber, John H.; Davidson, Christy L.; Xuejun, Pan; John, Ralph

    2012-04-01

    The plasticity of lignin biosynthesis should permit the inclusion of new compatible phenolic monomers, such as rosmarinic acid (RA) and analogous catechol derivatives, into cell-wall lignins that are consequently less recalcitrant to biomass processing. In vitro lignin polymerization experiments revealed that RA readily underwent peroxidase-catalyzed copolymerization with monolignols and lignin oligomers to form polymers with new benzodioxane inter-unit linkages. Incorporation of RA permitted extensive depolymerization of synthetic lignins by mild alkaline hydrolysis, presumably by cleavage of ester intra-unit linkages within RA. Copolymerization of RA with monolignols into maize cell walls by in situ peroxidases significantly enhanced alkaline lignin extractability and promoted subsequent cell wall saccharification by fungal enzymes. Incorporating RA also improved cell wall saccharification by fungal enzymes and by rumen microflora even without alkaline pretreatments, possibly by modulating lignin hydrophobicity and/or limiting cell wall cross-linking. Consequently, we anticipate that bioengineering approaches for partial monolignol substitution with RA and analogous plant hydroxycinnamates would permit more efficient utilization of plant fiber for biofuels or livestock production.

  17. Processing black mulberry into jam: effects on antioxidant potential and in vitro bioaccessibility.

    Science.gov (United States)

    Tomas, Merve; Toydemir, Gamze; Boyacioglu, Dilek; Hall, Robert D; Beekwilder, Jules; Capanoglu, Esra

    2017-08-01

    Black mulberries (Morus nigra) were processed into jam on an industrialised scale, including the major steps of: selection of frozen black mulberries, adding glucose-fructose syrup and water, cooking, adding citric acid and apple pectin, removing seeds, and pasteurisation. Qualitative and quantitative determinations of antioxidants in black mulberry samples were performed using spectrophotometric methods, as well as HPLC- and LC-QTOF-MS-based measurements. These analyses included the determination of total polyphenolic content, % polymeric colour, total and individual anthocyanin contents, antioxidant capacity, and in vitro bioaccessibility in processing samples. Jam processing led to a significant reduction in total phenolics (88%), total flavonoids (89%), anthocyanins (97%), and antioxidant capacity (88-93%) (P anthocyanin contents, determined using HPLC analysis, also showed a significant decrease (∼99% loss). In contrast, % recovery of bioaccessible total phenolics, anthocyanins, and antioxidant capacity (ABTS assay) increased after jam processing (16%, 12%, and 37%, respectively). Fruit processing resulted in losses of polyphenols, anthocyanins, and antioxidant capacity of black mulberry jam. Optimisation of food processing could help to protect the phenolic compounds in fruits which might be helpful for the food industry to minimise the antioxidant loss and improve the final product quality. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

  18. Identification of potential biomarkers in donor cows for in vitro embryo production by granulosa cell transcriptomics

    Science.gov (United States)

    Mazzoni, Gianluca; Salleh, Suraya M.; Freude, Kristine; Pedersen, Hanne S.; Stroebech, Lotte; Callesen, Henrik; Hyttel, Poul; Kadarmideen, Haja N.

    2017-01-01

    The Ovum Pick Up-In vitro Production (OPU-IVP) of embryos is an advanced reproductive technology used in cattle production but the complex biological mechanisms behind IVP outcomes are not fully understood. In this study we sequenced RNA of granulosa cells collected from Holstein cows at oocyte aspiration prior to IVP, to identify candidate genes and biological mechanisms for favourable IVP-related traits in donor cows where IVP was performed separately for each animal. We identified 56 genes significantly associated with IVP scores (BL rate, kinetic and morphology). Among these, BEX2, HEY2, RGN, TNFAIP6 and TXNDC11 were negatively associated while Mx1 and STC1 were positively associated with all IVP scores. Functional analysis highlighted a wide range of biological mechanisms including apoptosis, cell development and proliferation and four key upstream regulators (COX2, IL1, PRL, TRIM24) involved in these mechanisms. We found a range of evidence that good IVP outcome is positively correlated with early follicular atresia. Furthermore we showed that high genetic index bulls can be used in breeding without reducing the IVP performances. These findings can contribute to the development of biomarkers from follicular fluid content and to improving Genomic Selection (GS) methods that utilize functional information in cattle breeding, allowing a widespread large scale application of GS-IVP. PMID:28403200

  19. In Vitro Testing of Potential Entamoeba histolytica Pyruvate Phosphate Dikinase Inhibitors.

    Science.gov (United States)

    Saidin, Syazwan; Othman, Nurulhasanah; Noordin, Rahmah

    2017-10-01

    Adverse effects and resistance to metronidazole have motivated the search for new antiamoebic agents against Entamoeba histolytica. Control of amoeba growth may be achieved by inhibiting the function of the glycolytic enzyme and pyruvate phosphate dikinase (PPDK). In this study, we screened 10 compounds using an in vitro PPDK enzyme assay. These compounds were selected from a virtual screening of compounds in the National Cancer Institute database. The antiamoebic activity of the selected compounds was also evaluated by determining minimal inhibitory concentrations (MICs) and IC50 values using the nitro-blue tetrazolium reduction assay. Seven of the 10 compounds showed inhibitory activities against the adenosine triphosphate (ATP)/inorganic phosphate binding site of the ATP-grasp domain. Two compounds, NSC349156 (pancratistatin) and NSC228137 (7-ethoxy-4-[4-methylphenyl] sulfonyl-3-oxido-2, 1, 3-benzoxadiazol-3-ium), exhibited inhibitory effects on the growth of E. histolytica trophozoites with MIC values of 25 and 50 μM, and IC50 values of 14 and 20.7 μM, respectively.

  20. SCREENING APPROACHES FOR METHANE MITIGATING POTENTIAL OF TANNIN-CONTAINING PLANTS UNDER IN VITRO RUMEN ENVIRONMENT

    Directory of Open Access Journals (Sweden)

    A. Jayanegara

    2014-10-01

    Full Text Available The aim of the present study was to conduct univariate, bivariate and multivariate (principalcomponent analysis, PCA approaches in the screening of tannin-containing plants from variouscollection sites for their CH4 mitigating properties. Plant samples were obtained from various collectionsites in different countries, i.e. Indonesia (n = 27 species, Mongolia (n = 14, Switzerland (n = 16 andGermany (n = 3. The plants were incubated in vitro with buffered-rumen fluid at 39oC for 24 h. Totalgas production was recorded as an indicator of feed quality and emission of CH4 was measured. Resultsshowed that, based on bivariate screening, generally, plants possessed low CH4 production had lowquality or low total gas production except Rhus typhina, i.e. 43 ml/200 mg DM. The loading plot of PCAshowed that all phenolic fractions were in the opposite direction with CH4 and total gas production.Plants clustered together in reverse direction to that of CH4 were Bergenia crassifolia root and leaf,Swietenia mahagoni, Clidemia hirta, Peltiphyllum peltatum, Acacia villosa and R. typhina. It wasconluded that, for tannin-containing plants, screenings based on univariate, bivariate and multivariateapproaches in relation to ruminal CH4 emission led to similar results.

  1. Synthesis and in vitro evaluation of dioxopyrrolopyrroles as potential low-affinity fluorescent Ca2+ indicators

    Directory of Open Access Journals (Sweden)

    Nesibe Avcıbaşi

    2004-01-01

    1,4-dihydropyrrolo[3,4-c]pyrrole-1,4-dione (DPP3 have been synthesized and evaluated for their Ca2+ binding properties via fluorimetric titrations. The in vitro dissociation constant Kd measured at 21 ∘C in 100 mM KCl buffered solution, pH 7.05, for the Ca2+ –DPP1 complex is 10 μM; for Ca2+ –DPP2 and Ca2+ –DPP3 a Kd value of 20 μM is found. All three indicators form 1 : 1 complexes with Ca2+. The fluorescence quantum yields of the uncomplexed forms of DPP1, DPP2 and DPP3 are 1.2×10−2, 3.4×10−2 and 3.6×10−2, respectively. After binding to Ca2+ these values increase to 4.8×10−2, 5.0×10−2 and 5.1×10−2, respectively.

  2. Full-term potential of goat in vitro produced embryos after different cryopreservation methods.

    Science.gov (United States)

    Ferreira-Silva, José Carlos; Moura, Marcelo Tigre; Silva, Túlio Diego; Oliveira, Luis Rennan Sampaio; Chiamenti, Adauto; Figueirêdo Freitas, Vicente José; Oliveira, Marcos Antonio Lemos

    2017-04-01

    Cryopreservation of preimplantation embryos represents a major challenge due to their shape and relatively large cells. Embryo source and cryopreservation method are key factors to cryotolerance efficiency and few reports have investigated more promising protocols for goat embryos. The study was aimed to compare different cryopreservation methods for goat in vitro produced (IVP) embryos. Goat blastocysts were subjected to conventional freezing (CF), Dimethyl sulfoxide vitrification (DMSO-V) and Dimethylformamide vitrification (DMF-V). Cryopreserved blastocysts were assessed for re-expansion, cell viability and in vivo development rates. Blastocyst re-expansion after cryopreservation was similar between groups, but cell viability was lower for DMF-V (32%) than CF (68%) and DMSO-V (60%). Pregnancy and delivery rates were similar for CF (60% and 50%) and DMSO-V (50% and 45%) and higher then DMF-V (20% and 15%), respectively. Finally, kidding rates were also indistinguishable for CF (40%) and DMSO-V (35%), but higher then DMF-V (12.5%). In conclusion, conventional freezing and vitrification using DMSO have similar efficiencies for cryopreservation of goat IVP embryos and cryoprotectant for vitrification affects its outcome. Copyright © 2017 Elsevier Inc. All rights reserved.

  3. Influence of mitochondrial membrane potential of spermatozoa on in vitro fertilisation outcome.

    Science.gov (United States)

    Marchetti, P; Ballot, C; Jouy, N; Thomas, P; Marchetti, C

    2012-04-01

    To determine whether the outcome of in vitro fertilisation (IVF) is influenced by the percentage of spermatozoa with functional mitochondria, a total of 91 random couples undergoing IVF were included. Mitochondrial function was determined by flow cytometry and expressed as percentage of spermatozoa. Conventional sperm parameters were studied by light microscopy. Reproductive outcome parameters were fertilisation rate, embryo quality and clinical pregnancy. It was found that the fertilisation rate was correlated with the percentage of spermatozoa (r = 0.24, P = 0.01) as well as with the percentage of highly motile spermatozoa. However, we did not find any relationship between the percentage of spermatozoa and embryo quality. Nevertheless, no patient who exhibited less than 64% of spermatozoa achieved pregnancy. It is concluded that determination of Δψ(m) provides accurate information to guide physicians to identify male patients for whom IVF will be unlikely to result in pregnancy. Therefore, we suggest that the percentage of spermatozoa may contribute to identify the most appropriate treatment for an individual patient. © 2011 Blackwell Verlag GmbH.

  4. Green tea catechins reduced the glycaemic potential of bread: an in vitro digestibility study.

    Science.gov (United States)

    Goh, Royston; Gao, Jing; Ananingsih, Victoria K; Ranawana, Viren; Henry, Christiani Jeyakumar; Zhou, Weibiao

    2015-08-01

    Green tea catechins are potent inhibitors of enzymes for carbohydrate digestion. However, the potential of developing low glycaemic index bakery food using green tea extract has not been investigated. Results of this study showed that addition of green tea extract (GTE) at 0.45%, 1%, and 2% concentration levels significantly reduced the glycaemic potential of baked and steamed bread. The average retention levels of catechins in the baked and steamed bread were 75.3-89.5% and 81.4-99.3%, respectively. Bread fortified with 2% GTE showed a significantly lower level of glucose release during the first 90 min of pancreatic digestion as well as a lower content of rapidly digested starch (RDS) content. A significantly negative correlation was found between the catechin retention level and the RDS content of bread. The potential of transforming bread into a low GI food using GTE fortification was proven to be promising. Copyright © 2015 Elsevier Ltd. All rights reserved.

  5. Potential of a lytic bacteriophage to disrupt Acinetobacter baumannii biofilms in vitro.

    Science.gov (United States)

    Liu, Yannan; Mi, Zhiqiang; Niu, Wenkai; An, Xiaoping; Yuan, Xin; Liu, Huiying; Wang, Yong; Feng, Yuzhong; Huang, Yong; Zhang, Xianglilan; Zhang, Zhiyi; Fan, Hang; Peng, Fan; Li, Puyuan; Tong, Yigang; Bai, Changqing

    2016-10-01

    The ability of Acinetobacter baumannii to form biofilms and develop antibiotic resistance makes it difficult to control infections caused by this bacterium. In this study, we explored the potential of a lytic bacteriophage to disrupt A. baumannii biofilms. The potential of the lytic bacteriophage to disrupt A. baumannii biofilms was assessed by performing electron microscopy, live/dead bacterial staining, crystal violet staining and by determining adenosine triphosphate release. The bacteriophage inhibited the formation of and disrupted preformed A. baumannii biofilms. Results of disinfection assay showed that the lytic bacteriophage lysed A. baumannii cells suspended in blood or grown on metal surfaces. These results suggest the potential of the lytic bacteriophage to disrupt A. baumannii biofilms.

  6. In vitro determination of the antimicrobial potential of homemade preparations based on medicinal plants used to treat infectious diseases

    Directory of Open Access Journals (Sweden)

    GISELE MEDEIROS BASTOS

    2011-06-01

    Full Text Available The majority of the population in developing countries uses plants or plant preparations in their basic health care. Many plant species used nowadays in folk medicine have been proved to have antimicrobial properties. However, several factors, such as incorrect preparation of the plants, can interfere with the effectiveness of the treatment. The purpose of this study was to assess the use of homemade preparations of medicinal plants in the treatment of infectious diseases, by in vitro determination of their antimicrobial potential. Based on recipes elicited by questionnaires that were previously applied to a participant population, the samples were prepared in a similar manner and analyzed by the agar diffusion method. Members of 41 families, whose children attend a center of education that serves several needy communities in the city of Fortaleza (Ceará, Brazil, were interviewed; of these, 97.6% said they had used herbal therapy as a means to treat infectious diseases. In replies to a total of 39 questionnaires, 97 different homemade preparations of medicinal plants were cited. Out of 45 samples subjected to an in vitro assessment of antimicrobial activity, 25 (55.6% had some inhibitory effect on the growth of at least one of the microorganisms used. Most of the plants with known antimicrobial properties and cited by respondents showed variations in their in vitro activity, according to the manner in which they were prepared. Keywords: Medicinal plants. Products with Antimicrobial Action. Traditional Medicine. Homemade preparation. RESUMO Determinação in vitro do potencial antimicrobiano de preparações caseiras de plantas medicinais utilizadas para o tratamento de doenças infecciosas Grande parte da população de países em desenvolvimento utiliza plantas ou preparações vegetais nos cuidados básicos à saúde. Muitas das espécies vegetais utilizadas na medicina popular apresentam propriedades antimicrobianas comprovadas; no entanto

  7. Isatin-benzoazine molecular hybrids as potential antiproliferative agents: synthesis and in vitro pharmacological profiling

    Directory of Open Access Journals (Sweden)

    Abdel-Aziz HA

    2017-08-01

    Full Text Available Hatem A Abdel-Aziz,1 Wagdy M Eldehna,2 Adam B Keeton,3 Gary A Piazza,3 Adnan A Kadi,4 Mohamed W Attwa,4 Ali S Abdelhameed,4 Mohamed I Attia4,5 1Department of Applied Organic Chemistry, National Research Centre, Giza, 2Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Kafrelsheikh University, Kafrelsheikh, Egypt; 3Department of Oncologic Sciences and Pharmacology, Drug Discovery Research Center, Mitchell Cancer Institute, University of South Alabama, Mobile, AL, USA; 4Department of Pharmaceutical Chemistry, College of Pharmacy, King Saud University, Riyadh, Saudi Arabia; 5Medicinal and Pharmaceutical Chemistry Department, Pharmaceutical and Drug Industries Research Division, National Research Centre, Giza, Egypt Abstract: In continuation of our endeavor with respect to the development of potent and effective isatin-based anticancer agents, we adopted the molecular hybridization approach to design and synthesize four different sets of isatin-quinazoline (6a–f and 7a–e/phthalazine (8a–f/quinoxaline (9a–f hybrids. The antiproliferative activity of the target hybrids was assessed towards HT-29 (colon, ZR-75 (breast and A-549 (lung human cancer cell lines. Hybrids 8b–d emerged as the most active antiproliferative congener in this study. Compound 8c induced apoptosis via increasing caspase 3/7 activity by about 5-fold in the A-549 human cancer cell line. In addition, it exhibited an increase in the G1 phase and a decrease in the S and G2/M phases in the cell cycle effect assay. Furthermore, it displayed an inhibitory concentration 50% value of 9.5 µM against multidrug-resistant NCI-H69AR lung cancer cell line. The hybrid 8c was also subjected to in vitro metabolic investigations through its incubation with rat liver microsomes and analysis of the resulting metabolites with the aid of liquid chromatography-mass spectrometry. Keywords: isatins, hybridization approach, antiproliferative, apoptosis

  8. An In vitro Study of Bio-Control and Plant Growth Promotion Potential of Salicaceae Endophytes.

    Science.gov (United States)

    Kandel, Shyam L; Firrincieli, Andrea; Joubert, Pierre M; Okubara, Patricia A; Leston, Natalie D; McGeorge, Kendra M; Mugnozza, Giuseppe S; Harfouche, Antoine; Kim, Soo-Hyung; Doty, Sharon L

    2017-01-01

    Microbial communities in the endosphere of Salicaceae plants, poplar (Populus trichocarpa) and willow (Salix sitchensis), have been demonstrated to be important for plant growth promotion, protection from biotic and abiotic stresses, and degradation of toxic compounds. Our study aimed to investigate bio-control activities of Salicaceae endophytes against various soil borne plant pathogens including Rhizoctonia solani AG-8, Fusarium culmorum, Gaeumannomyces graminis var. tritici, and Pythium ultimum. Additionally, different plant growth promoting traits such as biological nitrogen fixation (BNF), indole-3-acetic acid (IAA) biosynthesis, phosphate solubilization, and siderophore production were assessed in all bio-control positive strains. Burkholderia, Rahnella, Pseudomonas, and Curtobacterium were major endophyte genera that showed bio-control activities in the in-vitro assays. The bio-control activities of Burkholderia strains were stronger across all tested plant pathogens as compared to other stains. Genomes of sequenced Burkholderia strains WP40 and WP42 were surveyed to identify the putative genes involved in the bio-control activities. The ocf and hcnABC gene clusters responsible for biosynthesis of the anti-fungal metabolites, occidiofungin and hydrogen cyanide, are present in the genomes of WP40 and WP42. Nearly all endophyte strains showing the bio-control activities produced IAA, solubilized tricalcium phosphate, and synthesized siderophores in the culture medium. Moreover, some strains reduced acetylene into ethylene in the acetylene reduction assay, a common assay used for BNF. Salicaceae endophytes could be useful for bio-control of various plant pathogens, and plant growth promotion possibly through the mechanisms of BNF, IAA production, and nutrient acquisition.

  9. Proinflammatory effects of bacterial lipoprotein on human neutrophil activation status, function and cytotoxic potential in vitro.

    LENUS (Irish Health Repository)

    Power, C

    2012-02-03

    Bacterial lipoprotein (BLP) is the most abundant protein in gram-negative bacterial cell walls, heavily outweighing lipopolysaccharide (LPS). Herein we present findings demonstrating the potent in vitro effects of BLP on neutrophil (PMN) activation status, function, and capacity to transmigrate an endothelial monolayer. PMNs are the principal effectors of the initial host response to injury or infection and constitute a significant threat to invading bacterial pathogens. The systemic inflammatory response syndrome (SIRS) is characterised by significant host tissue injury mediated, in part, by uncontrolled regulation of PMN cytotoxic activity. We found that BLP-activated human PMN as evidenced by increased CD11b\\/CD18 (Mac-1) expression. Up-regulation of PMN Mac-1 in response to BLP occurred independently of membrane-bound CD14 (mCD14). A similar up-regulation of intercellular adhesion molecule-1 (ICAM-1) on endothelial cells was observed whilst E-Selectin expression was unaffected. PMN transmigration across a human umbilical vein endothelial cell (HUVEC) monolayer was markedly increased after treating either PMN\\'s or HUVEC independently with BLP. This increased transmigration did not occur as a result of any direct effect of BLP on HUVEC monolayer permeability, assessed objectively using the passage of FITC-labeled Dextran-70. BLP primed PMN for enhanced respiratory burst and superoxide anion production in response to PMA, but did not influence phagocytosis of opsonized Escherichia coli. BLP far exceeds LPS as a gram-negative bacterial wall component, these findings therefore implicate BLP as an additional putative mediator of SIRS arising from gram-negative infection.

  10. In-vitro fermentation characteristics and methane reduction potential of mustard cake (Brassica juncea L.

    Directory of Open Access Journals (Sweden)

    S. M. Durge

    2016-10-01

    Full Text Available Aim: To assess the effect of mustard cake (Brassica juncea L. levels in concentrate mixtures and in composite feed mixtures (CFMs on in-vitro fermentation characteristics and methane production. Materials and Methods: Five concentrate mixtures were prepared with containing 30% oil cake, where linseed cake was replaced by mustard cake at the rate of 0%, 7.5%, 15.0%, 22.5%, and 30% in concentrate mixture. Mustard cake contained glucosinolate 72.58 μmol/g oil free dry matter (DM and contents in diet were 0, 5.4, 10.9, 16.3, and 21.8 μmol/g of concentrate mixture, respectively. Concentrate mixture containing 15.0% mustard cake was found to produced minimum methane which was then used for the preparation of CFM containing 0%, 25%, 50%, and 75% levels with gram straw. Result: Increased levels of mustard cake in concentrate mixtures had a linear decrease (p<0.05 in the total gas production, and the 15% inclusion showed lowest methane concentration (quadratic, p<0.01. The degradability of DM and organic matter (OM of concentrate mixtures did not change, however, pH and NH3-N concentrations of the fermentation medium showed linear (p<0.05 reductions with increased mustard cake levels. Increased levels of 15% mustard cake containing concentrate mixture in CFMs exhibited a trend (p=0.052 of increased gas production, whereas methane concentration in total gas, methane produced and degradability of DM and OM were also displayed a linear increase (p<0.05. However, the pH, NH3-N, and total volatile fatty acid levels decreased linearly (p<0.05 with increased levels of concentrate in CFMs. Conclusion: Reduction in methane production was evidenced with the inclusion of mustard cake in concentrate mixture at 15% level, and the CFMs with 25% concentrate, which contained 15% mustard cake, exhibited an improved fermentation and reduced methane production.

  11. In vitro migratory potential of rat quiescent hepatic stellate cells and its augmentation by cell activation.

    Science.gov (United States)

    Ikeda, K; Wakahara, T; Wang, Y Q; Kadoya, H; Kawada, N; Kaneda, K

    1999-06-01

    In liver injury, hepatic stellate cells are considered to depart from the sinusoidal wall and accumulate in the necrotic lesion through migration and proliferation. In this study, we investigated the migratory capacity of quiescent stellate cells in vitro and analyzed the relationship with proliferative response. Freshly isolated stellate cells that were seeded in the upper chamber of Cell Culture Insert (Becton Dickenson, Franklin Lakes, NJ) started to migrate to the lower chamber at 1 day and increased in migration index to 19% at 2 days. Cells in the lower chamber were stretched in shape with many lipid droplets and showed quiescent properties, i.e., negative expression of alpha-smooth muscle actin (alpha-SMA) or platelet-derived growth factor receptor-beta (PDGFR-beta). Migratory capacity in quiescent cells was also shown in the Matrigel-coated insert. Matrix metalloproteinase-2 (MMP-2) messenger RNA expression was low just after isolation, but was enhanced as migration became prominent. Migrating cells further showed higher proliferative activity than resting ones. The presence of PDGF/BB and Kupffer cells accelerated stellate cell migration by the chemotactic mechanism and concurrently augmented proliferation, whereas that of dexamethasone and interferon-gamma (IFN-gamma) attenuated migration as a result of general suppression effects. Compared with quiescent ones, alpha-SMA and PDGFR-beta-positive activated stellate cells obtained by 14-day culture exhibited more rapid and prominent migration, being regulated by mediators in a similar manner as described previously. These data indicate that quiescent stellate cells undergo migration, which is linked to proliferation and enhanced by PDGF/BB and Kupffer cells, suggesting the involvement of this function in the initial phase of development of postnecrotic fibrosis.

  12. Osteogenic potential of dualblocks cultured with human periodontal ligament stem cells: in vitro and synchrotron microtomography study.

    Science.gov (United States)

    Manescu, A; Giuliani, A; Mohammadi, S; Tromba, G; Mazzoni, S; Diomede, F; Zini, N; Piattelli, A; Trubiani, O

    2016-02-01

    In the present study, the early stages of in vitro bone formation in collagenated porcine scaffolds cultured with human periodontal ligament cells were investigated. The comparison between the osteogenic potential of this structure in basal and differentiating culture media was explored to predict the mechanism of its biological behavior as graft in human defect. Results were validated by synchrotron radiation X-Ray phase contrast computed microtomography (micro-CT). As the periodontal disease plays a key role in systemic and oral diseases, it is crucial to find advanced therapeutic clinical interventions to repair periodontal defects. This has been recently explored using cells and tissues developed in vitro that should ideally be immunologically, functionally, structurally and mechanically identical to the native tissue. In vitro cultures of human periodontal ligament cells, easily obtained by scraping of alveolar crestal and horizontal fibers of the periodontal ligament, were seeded on to collagenated porcine blocks constituted by natural cancellous and cortical bone. 3D images were obtained by synchrotron radiation micro-CT and processed with a phase-retrieval algorithm based on the transport of intensity equation. Starting from the second week of culture, newly formed mineralized bone was detected in all the scaffolds, both in basal and differentiating media. Bone mineralization was proved to occur preferentially in the trabecular portion and in differentiating media. The chosen method, supported by phase contrast micro-CT analysis, successfully and quantitatively monitored the early stages of bone formation and the rate of the bioscaffold resorption in basal and differentiating culture media. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  13. Enhanced Metastatic Potential in a 3D Tissue Scaffold toward a Comprehensive in Vitro Model for Breast Cancer Metastasis.

    Science.gov (United States)

    Balachander, Gowri Manohari; Balaji, Sai A; Rangarajan, Annapoorni; Chatterjee, Kaushik

    2015-12-23

    Metastasis is clinically the most challenging and lethal aspect of breast cancer. While animal-based xenograft models are expensive and time-consuming, conventional two-dimensional (2D) cell culture systems fail to mimic in vivo signaling. In this study we have developed a three-dimensional (3D) scaffold system that better mimics the topography and mechanical properties of the breast tumor, thus recreating the tumor microenvironment in vitro to study breast cancer metastasis. Porous poly(ε-caprolactone) (PCL) scaffolds of modulus 7.0 ± 0.5 kPa, comparable to that of breast tumor tissue were fabricated, on which MDA-MB-231 cells proliferated forming tumoroids. A comparative gene expression analysis revealed that cells growing in the scaffolds expressed increased levels of genes implicated in the three major events of metastasis, viz., initiation, progression, and the site-specific colonization compared to cells grown in conventional 2D tissue culture polystyrene (TCPS) dishes. The cells cultured in scaffolds showed increased invasiveness and sphere formation efficiency in vitro and increased lung metastasis in vivo. A global gene expression analysis revealed a significant increase in the expression of genes involved in cell-cell and cell-matrix interactions and tissue remodeling, cancer inflammation, and the PI3K/Akt, Wnt, NF-kappaB, and HIF1 signaling pathways-all of which are implicated in metastasis. Thus, culturing breast cancer cells in 3D scaffolds that mimic the in vivo tumor-like microenvironment enhances their metastatic potential. This system could serve as a comprehensive in vitro model to investigate the manifold mechanisms of breast cancer metastasis.

  14. In vitro and In vivo Studies on Stilbene Analogs as Potential Treatment Agents for Colon Cancer

    Science.gov (United States)

    Based upon the potential of resveratrol as a cancer chemopreventive agent, 27 stilbenes analogs were synthesized and tested against colon cancer cell line HT-29. Among these compounds, amino derivative (Z)-4-(3,5-dimethoxystyryl) aniline (4), (Z)-methyl 4-(3,5-dimethoxystyryl) benzoate (6) and (Z)-1...

  15. Summary of: an in vitro investigation of the erosive potential of smoothies: commentary

    NARCIS (Netherlands)

    Brand, H.S.

    2013-01-01

    Introduction Recent health promotion campaigns have encouraged the public to consume at least five portions of fruit and vegetables per day. Many see consuming fruit smoothies as a way of achieving this. Objective To ascertain the potential or otherwise for fruit smoothies to bring about dental

  16. Antioxidant potential of water hyacinth (Eichornia crassipes): In vitro antioxidant activity and phenolic composition

    DEFF Research Database (Denmark)

    Surendraraj, Alagarsamy; Farvin, Sabeena; Anandan, R.

    2011-01-01

    the various parts of E. crassipes. Out of the 11 phenolic acids analysed, ethanolic extracts contained high amounts gallic, protocatechuic, gentisic and phydroxybenzoic acid, whereas, water extracts contained less amounts of varied number of phenolic acids. Ethanolic extracts of flower, which contained...... aquatic weed, could be a potential natural antioxidant source for food, feed and pharmaceutical applications....

  17. Antioxidant potential of selected supplements in vitro and the problem of its extrapolation for in vivo

    Directory of Open Access Journals (Sweden)

    Julija Ogrin Papić

    2012-04-01

    Full Text Available Introduction: antioxidants, free radicals and oxidative stress have been studied extensively for quite some time but their role in diseases and their prevention has not been clearly determined. Because commercialantioxidants do not need to pass clinical tests in order to be sold over the counter we have decided to test the antioxidant potential of different commercial preparations with the antioxidative properties.Methods: pH, rH and oxidant-reduction potential of different preparations in aqueous solution was measured. Afterwards antioxidant potential using FormPlus® after adding the preparation to human blood as a morecomplex environment with different homeostasis mechanisms was determined.Results: all the results showed expected change compared to the control but the results in aqueous solution did not match the results obtained from the human blood, as was expected.Conclusion: from the experiments it can be concluded that while the preparations did show antioxidant activity, it is very difficult and even wrong to predict the antioxidant potential of an antioxidant preparationadded to human blood, let alone in a living organism, based just on the results obtained in aqueous solution. Further possibilities for research include more extensive studies of antioxidant preparations in more complex environment and last but not least in test organisms or in human trials.

  18. In vitro determination of the anti-aging potential of four southern Africa medicinal plants: Poster

    CSIR Research Space (South Africa)

    Ndlovu, G

    2013-09-01

    Full Text Available studies were explored for their potential use as anti-aging reagents. In this study the anti-collagenase, anti-hyaluronidase and anti-elaste activity of the plants Clerodendrum glabrum (PA, Schotia brachypetala (SB), Psychotria capensis (PC...

  19. Screening of antimicrobial potential of in vitro calli and adult leaf ...

    African Journals Online (AJOL)

    pc

    2013-02-27

    Feb 27, 2013 ... methods could lead to maintain bioactive potential of plants and to formulate antimicrobial drugs of ... not much branched. Leaves are elliptic, oblong and acute at the top. Flowers are in umbel shape, peduncle arises between the petiole (Rastogi and Mehrotra, 1998). The ... w: 220 v; light intensity 36 µ mol.

  20. A novel in vitro whole plant system for analysis of polyphenolics and their antioxidant potential in cultivars of Ocimum basilicum.

    Science.gov (United States)

    Srivastava, Shivani; Cahill, David M; Conlan, Xavier A; Adholeya, Alok

    2014-10-15

    Plants are an important source for medicinal compounds. Chemical screening and selection is critical for identification of compounds of interest. Ocimum basilicum (Basil) is a rich source of polyphenolics and exhibits high diversity, therefore bioprospecting of a suitable cultivar is a necessity. This study reports on the development of a true to type novel "in vitro system" and its comparison with a conventional system for screening and selection of cultivars for high total phenolics, individual polyphenolics, and antioxidant content. We have shown for the first time using online acidic potassium permanganate chemiluminescence that extracts from Ocimum basilicum showed antioxidant potential. The current study identified the cultivar specific composition of polyphenolics and their antioxidant properties. Further, a distinct relationship between plant morphotype and polyphenolic content was also found. Of the 15 cultivars examined, "Holy Green", "Red Rubin", and "Basil Genovese" were identified as high polyphenolic producing cultivars while "Subja" was determined to be a low producer. The "in vitro system" enabled differentiation of the cultivars in their morphology, polyphenolic content, and antioxidant activity and is a cheap and efficient method for bioprospecting studies.

  1. A low molecular weight ES-20 protein released in vivo and in vitro with diagnostic potential in lymph node tuberculosis

    Directory of Open Access Journals (Sweden)

    Shende N

    2008-01-01

    Full Text Available Purpose: To determine role of antigens released in vivo and in vitro in immunodiagnosis of tuberculosis (TB. Methods: In vivo released circulating tuberculosis antigen (CTA was obtained from TB sera by ammonium sulphate precipitation and in vitro released excretory-secretory (ES antigens from Mycobacterium tuberculosis culture filtrate. CTA and ES antigens were fractionated by SDS-PAGE and electro-eluted gel fractions were analysed for antigen by ELISA. Results: Low molecular weight proteins CTA-9 and ES-9 showed high titre of antigen activity. To explore the diagnostic potential of low molecular weight ES antigen, M. tuberculosis ES antigen was further fractionated by gel filtration chromatography followed by purification on anion exchange column using fast protein liquid chromatography and a highly seroreactive ESG-5D (ES-20 antigen was obtained. Competitive inhibition showed that CTA-9 and ES-9 antigens inhibit the binding of ES-20 antigen to its antibody. Seroanalysis showed sensitivity of 83 and 80% for ES-20 antigen and antibody detection, respectively, in pulmonary TB and 90% in lymph node TB. Conclusions: Seroreactivity studies using M. tuberculosis ES-20 antigen showed usefulness in detection of TB; in particular, lymph node TB.

  2. In Vitro and In Vivo Investigation of the Potential of Amorphous Microporous Silica as a Protein Delivery Vehicle

    Directory of Open Access Journals (Sweden)

    Amol Chaudhari

    2013-01-01

    Full Text Available Delivering growth factors (GFs at bone/implant interface needs to be optimized to achieve faster osseointegration. Amorphous microporous silica (AMS has a potential to be used as a carrier and delivery platform for GFs. In this work, adsorption (loading and release (delivery mechanism of a model protein, bovine serum albumin (BSA, from AMS was investigated in vitro as well as in vivo. In general, strong BSA adsorption to AMS was observed. The interaction was stronger at lower pH owing to favorable electrostatic interaction. In vitro evaluation of BSA release revealed a peculiar release profile, involving a burst release followed by a 6 h period without appreciable BSA release and a further slower release later. Experimental data supporting this observation are discussed. Apart from understanding protein/biomaterial (BSA/AMS interaction, determination of in vivo protein release is an essential aspect of the evaluation of a protein delivery system. In this regard micropositron emission tomography (μ-PET was used in an exploratory experiment to determine in vivo BSA release profile from AMS. Results suggest stronger in vivo retention of BSA when adsorbed on AMS. This study highlights the possible use of AMS as a controlled protein delivery platform which may facilitate osseointegration.

  3. Assessment of toxic potential of mycotoxin contaminated bread during in vitro human digestion on human B lymphoid cell line.

    Science.gov (United States)

    Monaci, Linda; Garbetta, Antonella; Angelis, Elisabetta De; Visconti, Angelo; Minervini, Fiorenza

    2015-01-05

    Ingestion of food is considered a major route of exposure to many contaminants including mycotoxins. The amount of mycotoxin resisting to the digestion process and potentially absorbable by the systemic circulation is only a smaller part of that ingested. In vitro digestion models turn useful for evaluating mycotoxins bioaccessibility during the intestinal transit and can be intended as a valuable tool for the assessment of mycotoxin bioavailability in food. In this paper we describe a study aimed at investigating toxicity of in vitro gastro-duodenal digests of mycotoxin contaminated bread collected along the digestion time-course. Toxicity tests were carried out on a sensitive RPMI lymphoid B cell line chosen as the most suitable lineage to assess toxicity retained by gastro-duodenal digests. In parallel, a chemical quantification of T-2 and HT-2 toxins contaminating the bread digests was accomplished during the gastric and duodenal transit. The digestive fluids undergoing chemical and toxicological analysis were collected at the beginning and end of gastric phase, and after completion of the duodenal phase. Results proved that a correlation between HT-2 content and toxicity did exist although a more persistent toxic activity was displayed in the later stage of the duodenal phase. This persistent toxicity might be explained by the co-occurrence of unknown HT-2-related conjugates or metabolites formed during digestion. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  4. Influence of cultivar and concentration of selected phenolic constituents on the in vitro chemiopreventive potential of olive oil extracts.

    Science.gov (United States)

    Fabiani, Roberto; Sepporta, Maria V; Mazza, Teresa; Rosignoli, Patrizia; Fuccelli, Raffaela; De Bartolomeo, Angelo; Crescimanno, Marilena; Taticchi, Agnese; Esposto, Sonia; Servili, Maurizio; Morozzi, Guido

    2011-08-10

    One of the main olive oil phenolic compounds, hydroxytyrosol (3,4-DHPEA), exerts in vitro chemopreventive activities (antiproliferative and pro-apoptotic) on tumor cells through the accumulation of H(2)O(2) in the culture medium. However, the phenol composition of virgin olive oil is complex, and 3,4-DHPEA is present at low concentrations when compared to other secoiridoids. In this study, the in vitro chemopreventive activities of complex virgin olive oil phenolic extracts (VOO-PE, derived from the four Italian cultivars Nocellara del Belice, Coratina, Ogliarola, and Taggiasca) were compared to each other and related to the amount of the single phenolic constituents. A great chemopreventive potential among the different VOO-PE was found following this order: Ogliarola > Coratina > Nocellara > Taggiasca. The antiproliferative and pro-apoptotic activities of VOO-PE were positively correlated to the secoiridoid content and negatively correlated to the concentration of both phenyl alcohols and lignans. All extracts induced H(2)O(2) accumulation in the culture medium, but this phenomenon was not responsible for their pro-apoptotic activity. When tested in a complex mixture, the olive oil phenols exerted a more potent chemopreventive effect compared to the isolated compounds, and this effect could be due either to a synergistic action of components or to any other unidentified extract constituent.

  5. Fasting cycles potentiate the efficacy of gemcitabine treatment in in vitro and in vivo pancreatic cancer models.

    Science.gov (United States)

    D'Aronzo, Martina; Vinciguerra, Manlio; Mazza, Tommaso; Panebianco, Concetta; Saracino, Chiara; Pereira, Stephen P; Graziano, Paolo; Pazienza, Valerio

    2015-07-30

    Pancreatic cancer (PC) is ranked as the fourth leading cause of cancer-related deaths worldwide. Despite recent advances in treatment options, a modest impact on the outcome of the disease is observed so far. Short-term fasting cycles have been shown to potentiate the efficacy of chemotherapy against glioma. The aim of this study was to assess the effect of fasting cycles on the efficacy of gemcitabine, a standard treatment for PC patients, in vitro and in an in vivo pancreatic cancer mouse xenograft model. BxPC-3, MiaPaca-2 and Panc-1 cells were cultured in standard and fasting mimicking culturing condition to evaluate the effects of gemcitabine. Pancreatic cancer xenograft mice were subjected to 24h starvation prior to gemcitabine injection to assess the tumor volume and weight as compared to mice fed ad libitum. Fasted pancreatic cancer cells showed increased levels of equilibrative nucleoside transporter (hENT1), the transporter of gemcitabine across the cell membrane, and decreased ribonucleotide reductase M1 (RRM1) levels as compared to those cultured in standard medium. Gemcitabine was more effective in inducing cell death on fasted cells as compared to controls. Consistently, xenograft pancreatic cancer mice subjected to fasting cycles prior to gemcitabine injection displayed a decrease of more than 40% in tumor growth. Fasting cycles enhance gemcitabine effect in vitro and in the in vivo PC xenograft mouse model. These results suggest that restrictive dietary interventions could enhance the efficacy of existing cancer treatments in pancreatic cancer patients.

  6. Novel Benzothiazole, Benzimidazole and Benzoxazole Derivatives as Potential Antitumor Agents: Synthesis and Preliminary in Vitro Biological Evaluation

    Directory of Open Access Journals (Sweden)

    Li Yang

    2012-01-01

    Full Text Available In a previous hit-to-lead research program targeting anticancer agents, two promising lead compounds, 1a and 1b, were found. However, the poor solubility of 1a and 1b made difficult further in vivo studies. To solve this problem, a lead optimization was conducted through introducing N-methyl-piperazine groups at the 2-position and 6-position. To our delight, the optimized analogue 1d showed comparable antiproliferative activity in vitro with better solubility, compared with 1a. Based on this result, the replacement of the benzothiazole scaffold with benzimidazole and benzoxazole moieties afforded 1f and 1g, whose activities were fundamentally retained. In the preliminary in vitro biological evaluation, the immunofluorescence staining of HCT116 cells indicated that 1d, 1f and 1g led to cytosolic vacuolization which was not induced by 1a at low micromolecular concentrations. These results suggest that these optimized compounds might potentially constitute a novel class of anticancer agents, which merit further studies.

  7. Screening of toxic potential of graphene family nanomaterials using in vitro and alternative in vivo toxicity testing systems.

    Science.gov (United States)

    Chatterjee, Nivedita; Yang, Ji Su; Park, Kwangsik; Oh, Seung Min; Park, Jeonggue; Choi, Jinhee

    2015-01-01

    The widely promising applications of graphene nanomaterials raise considerable concerns regarding their environmental and human health risk assessment. The aim of the current study was to evaluate the toxicity profiling of graphene family nananomaterials (GFNs) in alternative in vitro and in vivo toxicity testing models. The GFNs used in this study are graphene nanoplatelets ([GNPs]-pristine, carboxylate [COOH] and amide [NH2]) and graphene oxides (single layer [SLGO] and few layers [FLGO]). The human bronchial epithelial cells (Beas2B cells) as in vitro system and the nematode Caenorhabditis elegans as in vivo system were used to profile the toxicity response of GFNs. Cytotoxicity assays, colony formation assay for cellular toxicity and reproduction potentiality in C. elegans were used as end points to evaluate the GFNs' toxicity. In general, GNPs exhibited higher toxicity than GOs in Beas2B cells, and among the GNPs the order of toxicity was pristine>NH2>COOH. Although the order of toxicity of the GNPs was maintained in C. elegans reproductive toxicity, but GOs were found to be more toxic in the worms than GNPs. In both systems, SLGO exhibited profoundly greater dose dependency than FLGO. The possible reason of their differential toxicity lay in their distinctive physicochemical characteristics and agglomeration behavior in the exposure media. The present study revealed that the toxicity of GFNs is dependent on the graphene nanomaterial's physical forms, surface functionalizations, number of layers, dose, time of exposure and obviously, on the alternative model systems used for toxicity assessment.

  8. An in vitro analysis of the cariogenic and erosive potential of pediatric liquid analgesics

    OpenAIRE

    Shaam Saeed; Nada Bshara; Juliana Trak; Ghiath Mahmoud

    2015-01-01

    Background: Analgesics such as Ibuprofen and Paracetamol, which are clinically used for the treatment of fever and/or pain, are among the most frequently used pediatric medicines. However, the properties of these preparations determine their cariogenic and erosive potential. Aims: The main objective of this study was to analyze the pH, viscosity and total sugar content in a variety of Syrian pediatric liquid analgesics (PLA). Setting and Design: A total of 16 available liquid analgesics that ...

  9. In vitro potentiation of cephalosporins by alafosfalin against urinary tract bacteria.

    OpenAIRE

    Arisawa, M; Ohshima, J; Ohsawa, E; Maruyama, H B

    1982-01-01

    Potentiating activity of alafosfalin was examined in detail with 8 cephalosporins and mecillinam against 164 urinary bacteria representing 8 genera. Alafosfalin was generally comparable in activity to cefamandole and mecillinam but superior to other cephalosporins tested. When the minimal fractional inhibitory concentration indices were compared, synergism was observed with all of the beta-lactams tested for all species except Streptococcus faecalis. Marked synergism was observed with Escheri...

  10. Antioxidant potential of water hyacinth (Eichornia crassipes): In vitro antioxidant activity and phenolic composition

    DEFF Research Database (Denmark)

    Surendraraj, A.; Farvin, Sabeena; Anandan, R.

    2013-01-01

    and in the antioxidant activities of extracts from the various parts of E. crassipes. Out of the 11 phenolic acids analyzed, ethanolic extracts contained high amounts of gallic, protocatechuic, gentisic, and p-hydroxybenzoic acid, whereas, water extracts contained less amounts of a varied number of phenolic acids...... oil. Our results demonstrate that E. crassipes, an underutilized aquatic weed, could be a potential natural antioxidant source for food, feed, and pharmaceutical applications. © 2013 Copyright Taylor & Francis Group, LLC....

  11. Evaluating the potential genotoxicity of phthalates esters (PAEs) in perfumes using in vitro assays.

    Science.gov (United States)

    Al-Saleh, Iman; Al-Rajudi, Tahreer; Al-Qudaihi, Ghofran; Manogaran, Pulicat

    2017-10-01

    We previously reported high levels of phthalate esters (PAEs) added as solvents or fixatives in 47 brands of perfumes. Diethyl phthalate was the most abundant compound (0.232-23,649 ppm), and 83.3% of the perfumes had levels >1 ppm, the threshold limit cited by a Greenpeace investigation. All samples had dimethyl phthalate levels higher than its threshold limit of 0.1 ppm, and 88, 38, and 7% of the perfumes had benzyl butyl phthalate, di(2-ethylhexyl) phthalate, and dibutyl phthalate levels, respectively, above their threshold limits. The role of PAEs as endocrine disruptors has been well documented, but their effect on genotoxic behavior has received little attention. We used in vitro single-cell gel electrophoresis (comet) and micronucleus (MN) assays with human lymphoblastoid TK6 cells to evaluate the genotoxic potency of 42 of the same perfumes and to determine its association with PAEs. All perfumes induced more DNA damage than a negative control (NEG), ≥ 90% of the samples caused more damage than cells treated with the vehicles possibly used in perfume's preparations such as methanol (ME) and ethanol (ET), and 11.6% of the perfumes caused more DNA damage than a positive control (hydrogen peroxide). Chromosome breakage expressed as MN frequency was higher in cells treated with 71.4, 64.3, 57.1, and 4.8% of the perfumes than in NEG, cells treated with ME or ET, and another positive control (x-rays), respectively. The genotoxic responses in the comet and MN assays were not correlated. The comet assay indicated that the damage in TK6 cells treated with five PAEs at concentrations of 0.05 and 0.2 ppm either individually or as a mixture did not differ significantly from the damage in cells treated with the perfumes. Unlike the comet assay, the sensitivity of the MN assay to PAEs was weak at both low and high concentrations, and MN frequencies were generally low. This study demonstrates for the first time the possible contribution of PAEs in perfumes to DNA

  12. Potential of magnetic resonance-guided focused ultrasound for intracranial hemorrhage: an in vitro feasibility study.

    Science.gov (United States)

    Harnof, Sagi; Hananel, Arik; Zilby, Zion; Kulbatski, Iris; Hadani, Moshe; Kassell, Neal

    2014-01-01

    Intracranial hemorrhage has a mortality rate of up to 40-60% due to the lack of effective treatment. Magnetic resonance-guided focused ultrasound may offer a breakthrough noninvasive technology, by allowing accurate delivery of focused ultrasound, under the guidance of real-time magnetic resonance imaging. The purpose of the current study was to optimize the acoustic parameters of magnetic resonance-guided focused ultrasound for effective clot liquefaction, in order to evaluate the feasibility of magnetic resonance-guided focused ultrasound for thrombolysis. Body (1·1 MHz) and brain (220 kHz) magnetic resonance-guided focused ultrasound systems (InSightec Ltd, Tirat Carmel, Israel) were used to treat tube-like (4 cc), round (10 cc), and bulk (300 cc) porcine blood clots in vitro, using burst sonications of one-second to five-seconds, a duty cycle of 5-50%, and peak acoustic powers between 600 and 1200 W. Liquefied volumes were measured as hyperintense regions on T2-weighted magnetic resonance images for body unit sonications (duration of one-second, duty cycle of 10%, and power of 500-1200 W). Liquefaction efficiency was calculated for brain unit sonications (duration of one-second, duty cycle of 10%, power of 600 W, and burst length between 0·1 ms and 100 ms). Liquified lesion volume increased as power was raised, without a thermal rise. For brain unit sonications, a power setting of 600 W and ultrashort sonications (burst length between 0·1 and 1·0 ms) resulted in liquefaction efficacy above 50%, while longer burst duration yielded lower efficacy. These results demonstrate the feasibility of obtaining reproducible, rapid, efficient, and accurate blood clot lysis using the magnetic resonance-guided focused ultrasound system. Further in vivo studies are needed to validate the feasibility of magnetic resonance-guided focused ultrasound as a treatment modality for intracranial hemorrhage. © 2013 The Authors. International Journal of

  13. In vitro interaction between homocysteine and copper ions: Potential redox implications.

    Science.gov (United States)

    Carrasco-Pozo, Catalina; Alvarez-Lueje, Alejandro; Olea-Azar, Claudio; López-Alarcón, Camilo; Speisky, Hernán

    2006-10-01

    Homocysteine (Hcys) has been implicated in various oxidative stress-related disorders. The presence of a thiol on its structure allows Hcys to exert a double-edge redox action. Depending on whether Cu2+ ions occur concomitantly, Hcys can either promote or prevent free radical generation and its consequences. We have addressed in vitro the interaction between Hcys and Cu2+ ions, in terms of the consequences that such interaction may have on the free radical scavenging properties of Hcys and on the redox state and redox activity of the metal. To this end, we investigated the free radical-scavenging, O2(*-)-generating, and ascorbate-oxidizing properties of the interacting species by assessing the bleaching of ABTS*+ radicals, the reduction of O2(*-)-dependent cytochrome c, and the copper-dependent oxidation of ascorbate, respectively. In addition, electron paramagnetic resonance and Cu(I)-bathocuproine formation were applied to assess the formation of paramagnetic complexes and the metal redox state. Upon a brief incubation, the Hcys/Cu2+ interaction led to a decrease in the free radical-scavenging properties of Hcys, and to a comparable loss of the thiol density. Both effects were partial and were not modified by increasing the incubation time, despite the presence of Cu2+ excess. Depending on the molar Hcys:Cu2+ ratio, the interaction resulted in the formation of mixtures that appear to contain time-stable and ascorbate-reducible Cu(II) complexes (for ratios up to 2:1), and ascorbate- and oxygen-redox-inactive Cu(I) complexes (for ratios up to 4:1). Increasing the interaction ratio beyond 4:1 was associated with the sudden appearance of an O2(*-)-generating activity. The data indicate that depending on the molar ratio of interaction, Hcys and Cu2+ react to form copper complexes that can promote either antioxidant or pro-oxidant actions. We speculate that the redox activity arising from a large molar Hcys excess may partially underlie the association between hyper

  14. Characterization of in vitro phenotypes of Burkholderia pseudomallei and Burkholderia mallei strains potentially associated with persistent infection in mice.

    Science.gov (United States)

    Bernhards, R C; Cote, C K; Amemiya, K; Waag, D M; Klimko, C P; Worsham, P L; Welkos, S L

    2017-03-01

    Burkholderia pseudomallei (Bp) and Burkholderia mallei (Bm), the agents of melioidosis and glanders, respectively, are Tier 1 biothreats. They infect humans and animals, causing disease ranging from acute and fatal to protracted and chronic. Chronic infections are especially challenging to treat, and the identification of in vitro phenotypic markers which signal progression from acute to persistent infection would be extremely valuable. First, a phenotyping strategy was developed employing colony morphotyping, chemical sensitivity testing, macrophage infection, and lipopolysaccharide fingerprint analyses to distinguish Burkholderia strains. Then mouse spleen isolates collected 3-180 days after infection were characterized phenotypically. Isolates from long-term infections often exhibited increased colony morphology differences and altered patterns of antimicrobial sensitivity and macrophage infection. Some of the Bp and Bm persistent infection isolates clearly displayed enhanced virulence in mice. Future studies will evaluate the potential role and significance of these phenotypic markers in signaling the establishment of a chronic infection.

  15. Lantana camara Linn root extract-mediated gold nanoparticles and their in vitro antioxidant and cytotoxic potentials.

    Science.gov (United States)

    Ramkumar, Rajendiran; Balasubramani, Govindasamy; Raja, Ramalingam Karthik; Raja, Manickam; Govindan, Raji; Girija, Easwaradas Kreedapathy; Perumal, Pachiappan

    2017-06-01

    The Lantana camara Linn root extract derived gold nanoparticles (Au NPs) were characterized by Ultraviolet-Visible spectroscopy, X-ray diffraction, fourier transform-infrared, high resolution transmission electron microscopy, selected area electron diffraction pattern and energy dispersive X-ray analyses. In DPPH assay, the inhibitory concentration (IC50) of Au NPs and gallic acid was 24.17 and 5.39 μg/ml, whereas, for cytotoxicity assay, the IC50 of Au NPs was 17.72 and 32.98 μg/ml on MBA-MB-231 and Vero cells, respectively. Thus, the Au NPs possess significant in vitro antioxidant and cytotoxic properties which could be considered as potential alternate for the development of anticancer drug in future.

  16. Potential therapeutic benefit of C1-esterase inhibitor in neuromyelitis optica evaluated in vitro and in an experimental rat model.

    Directory of Open Access Journals (Sweden)

    Lukmanee Tradtrantip

    Full Text Available Neuromyelitis optica (NMO is an autoimmune demyelinating disease of the central nervous system in which binding of anti-aquaporin-4 (AQP4 autoantibodies (NMO-IgG to astrocytes causes complement-dependent cytotoxicity (CDC and inflammation resulting in oligodendrocyte and neuronal injury. There is compelling evidence for a central role of complement in NMO pathogenesis. Here, we evaluated the potential of C1-esterase inhibitor (C1-inh for complement-targeted therapy of NMO. C1-inh is an anti-inflammatory plasma protein with serine protease inhibition activity that has a broad range of biological activities on the contact (kallikrein, coagulation, fibrinolytic and complement systems. C1-inh is approved for therapy of hereditary angioedema (HAE and has been studied in a small safety trial in acute NMO relapses (NCT 01759602. In vitro assays of NMO-IgG-dependent CDC showed C1-inh inhibition of human and rat complement, but with predicted minimal complement inhibition activity at a dose of 2000 units in humans. Inhibition of complement by C1-inh was potentiated by ∼10-fold by polysulfated macromolecules including heparin and dextran sulfate. In rats, intravenous C1-inh at a dose 30-fold greater than that approved to treat HAE inhibited serum complement activity by <5%, even when supplemented with heparin. Also, high-dose C1-inh did not reduce pathology in a rat model of NMO produced by intracerebral injection of NMO-IgG. Therefore, although C1r and C1s are targets of C1-inh, our in vitro data with human serum and in vivo data in rats suggest that the complement inhibition activity of C1-inh in serum is too low to confer clinical benefit in NMO.

  17. Evaluation of indigenous grains from the Peruvian Andean region for antidiabetes and antihypertension potential using in vitro methods.

    Science.gov (United States)

    Ranilla, Lena Galvez; Apostolidis, Emmanouil; Genovese, Maria Ines; Lajolo, Franco Maria; Shetty, Kalidas

    2009-08-01

    The health-relevant functionality of 10 thermally processed Peruvian Andean grains (five cereals, three pseudocereals, and two legumes) was evaluated for potential type 2 diabetes-relevant antihyperglycemia and antihypertension activity using in vitro enzyme assays. Inhibition of enzymes relevant for managing early stages of type 2 diabetes such as hyperglycemia-relevant alpha-glucosidase and alpha-amylase and hypertension-relevant angiotensin I-converting enzyme (ACE) were assayed along with the total phenolic content, phenolic profiles, and antioxidant activity based on the 1,1-diphenyl-2-picrylhydrazyl radical assay. Purple corn (Zea mays L.) (cereal) exhibited high free radical scavenging-linked antioxidant activity (77%) and had the highest total phenolic content (8 +/- 1 mg of gallic acid equivalents/g of sample weight) and alpha-glucosidase inhibitory activity (51% at 5 mg of sample weight). The major phenolic compound in this cereal was protocatechuic acid (287 +/- 15 microg/g of sample weight). Pseudocereals such as Quinoa (Chenopodium quinoa Willd) and Kañiwa (Chenopodium pallidicaule Aellen) were rich in quercetin derivatives (1,131 +/- 56 and 943 +/- 35 microg [expressed as quercetin aglycone]/g of sample weight, respectively) and had the highest antioxidant activity (86% and 75%, respectively). Andean legumes (Lupinus mutabilis cultivars SLP-1 and H-6) inhibited significantly the hypertension-relevant ACE (52% at 5 mg of sample weight). No alpha-amylase inhibitory activity was found in any of the evaluated Andean grains. This in vitro study indicates the potential of combination of Andean whole grain cereals, pseudocereals, and legumes to develop effective dietary strategies for managing type 2 diabetes and associated hypertension and provides the rationale for animal and clinical studies.

  18. Evaluation of the Erosive Potential of Selected Isotonic Drinks: In Vitro Studies.

    Science.gov (United States)

    Ostrowska, Aneta; Szymański, Witold; Kołodziejczyk, Łukasz; Bołtacz-Rzepkowska, Elżbieta

    2016-01-01

    Isotonic drinks are an important component of the diet of athletes. Sports drinks cause the body to maintain proper hydration and supplement minerals which are lost in sweat during excessive exercising. Aside from the benefits of isotonic drinks, it is important to be aware of the harmful effects of citric acid within the products, which could cause enamel erosion. The aim of the study was to evaluate the erosive potential of sports drinks using confocal scanning laser microscopy (CLSM). The studies measured the change of surface roughness of the dental enamel after etching using Isostar, Powerade and Gatorade drinks, and Fortuna orange juice. Measurements were repeated after 1, 2 and 3 h of exposure to the selected liquid. The evaluation of calcium compound contents was carried out using the complexonometric method. The surface roughness measurements of dental enamel showed that the lowest values of the parameters Ra and Rz were obtained for Isostar and orange juice. The research of the calcium content in the selected beverages showed the highest value in Isostar (320.0 mg/L) and the lowest in Powerade (40.0 mg/L) and Gatorade (21.0 mg/L). Our study confirms that Isostar is the safest sports drink, among the analyzed beverages, for athletes, because it causes the least erosive changes in dental enamel. It is recommended to supplement beverages to reduce their potential for erosion using calcium compounds.

  19. Influence of Tunisian Ficus carica fruit variability in phenolic profiles and in vitro radical scavenging potential

    Directory of Open Access Journals (Sweden)

    Emna Faleh

    2012-11-01

    Full Text Available Ficus carica L., Moraceae, is one of the first plants that were cultivated by humans, being the fruit an important crop worldwide for dry and fresh consumption. In this work, phenolics and antioxidant potential of dried fruits of seventeen Tunisian F. carica varieties, from green, red and black phenotypes, were assessed for the first time. HPLC-DAD analysis was performed. All samples presented a similar qualitative profile. The phenolics content ranged between 29.18 and 55.56 mg/kg (in black and red phenotypes, respectively and quercetin-3-O-rutinoside was always the major compound. The antioxidant potential against DPPH•, superoxide and nitric oxide radicals of three varieties representing each phenotype was checked. All samples exhibited activity against the first two radicals in a concentration-dependent way, "Bayoudi" variety being the most effective one (IC25 values of 10.32 and 2.89 µg/ mL, respectively. Nevertheless, only "Hammouri" variety presented some capacity to scavenge nitric oxide radical. Our results reveal nice perspectives for these typical fruits, as they present an interesting phenolic composition and good antiradical activity and may encourage their consumption for health protection.

  20. Potential of a cyclone prototype spacer to improve in vitro dry powder delivery.

    Science.gov (United States)

    Parisini, Irene; Cheng, Sean J; Symons, Digby D; Murnane, Darragh

    2014-05-01

    Low inspiratory force in patients with lung disease is associated with poor deagglomeration and high throat deposition when using dry powder inhalers (DPIs). The potential of two reverse flow cyclone prototypes as spacers for commercial carrier-based DPIs was investigated. Cyclohaler®, Accuhaler® and Easyhaler® were tested with and without the spacers between 30 and 60 Lmin−1. Deposition of particles in the next generation impactor and within the devices was determined by high performance liquid chromatography. Reduced induction port deposition of the emitted particles from the cyclones was observed due to the high retention of the drug within the spacers (e.g. salbutamol sulphate (SS): 67.89 ± 6.51% at 30 Lmin−1 in Cheng 1). Fine particle fractions of aerosol as emitted from the cyclones were substantially higher than the DPIs alone. Moreover, the aerodynamic diameters of particles emitted from the cyclones were halved compared to the DPIs alone (e.g. SS from the Cyclohaler® at 4 kPa: 1.08 ± 0.05 μm vs. 3.00 ± 0.12 μm, with and without Cheng 2, respectively) and unaltered with increased flow rates. This work has shown the potential of employing a cyclone spacer for commercial carrier-based DPIs to improve inhaled drug delivery.

  1. Potential anticancer activity of myricetin in human T24 bladder cancer cells both in vitro and in vivo.

    Science.gov (United States)

    Sun, Fang; Zheng, Xiang Yi; Ye, Jia; Wu, Ting Ting; Wang, Jian li; Chen, Weilin

    2012-01-01

    Myricetin, a naturally occurring phytochemical, has potent anticancer-promoting activity and contributes to the chemopreventive potential of several foods. In this preliminary study, we evaluate the chemopreventive potential of myricetin against bladder cancer and its mechanism of action. The results of a MTT assay showed that myricetin was able to inhibit the viability and proliferation of T24 cells in a dose- and time-dependent manner. It also promoted cell cycle arrest at G2/M in a dose-dependent manner and induced apoptosis detected by flow cytometry and DNA fragmentation analysis. Treatment with myricetin led to G2/M cell cycle arrest in T24 cells by downregulation of Cyclin B1 and cyclin-dependent kinase cdc2. Myricetin-induced apoptosis correlates with the modulation of Bcl-2 family proteins and activation of the caspase-3. Myricetin also inhibited the phosphorylation of Akt, whereas the phosphorylation of p38 MAPK was enhanced. Myricetin had a significantly reduced T24 cell migration that was accompanied by a decreasing MMP-9 expression in vitro. Furthermore, myricetin treatment significantly inhibited the tumor growth on T24 bladder cancer xenografts model. These findings suggest that myricetin has potential anticancer activity and could be an important chemoprevention agent for bladder cancer.

  2. Spectroscopic analysis of embryo culture media for predicting reproductive potential in patients undergoing in vitro fertilization.

    Science.gov (United States)

    Baştu, Ercan; Parlatan, Uğur; Başar, Günay; Yumru, Harika; Bavili, Nima; Sağ, Fatih; Bulgurcuoğlu, Sibel; Buyru, Faruk

    2017-09-01

    To predict the reproductive potential of embryos via Raman spectroscopy evaluation of the spent culture media as well as with a conventional morphologic evaluation. Women of reproductive age (n=31) who were treated for unexplained infertility and scheduled for single embryo transfer were invited to participate in this prospective study. After the embryos were removed from the culture, the spent culture media were stored at -80 °C after snap-freezing in liquid nitrogen. Fifteen patients were clinically pregnant, and 16 patients were clinically non-pregnant. Clinical pregnancy was predicted using Raman spectroscopy in 93% (14/15) of clinically pregnant patients, and in 62.5% (10 out of 16) of clinically non-pregnant patients. The sensitivity of the Raman spectroscopic analysis was 93% and the specificity was 62.5%. Metabolomic evaluation of spent embryo culture media is an emerging technique with promising objective results. However, there is clearly room for improvement.

  3. A new in vitro method for testing the interproximal cleaning potential of toothbrushing.

    Science.gov (United States)

    Bruun, C; Ekstrand, K R; Andreasen, K B

    1998-01-01

    A new laboratory method was developed and used to evaluate the approximal penetration of different toothbrushing techniques. Before brushing, the approximal surfaces on teeth numbers 33 to 38 were covered by a colored coating. After brushing from both the buccal and lingual sides, the teeth were removed and the remaining coatings on the approximal surfaces were photographed and magnified, and their areas were determined by computerization. Two conventional toothbrushing techniques and two toothbrushing techniques specially designed to enhance interproximal access were used. It was found that considerable areas on the approximal surfaces were left untouched by the toothbrush bristles, regardless of the toothbrushing technique employed. The findings suggest that improvements in toothbrush design will be a more important contribution to the attainment of effective interproximal brushing than the development of new brushing techniques. This test method shares many similarities with in vivo conditions and seems also well suited for evaluating interproximal cleaning potential of new toothbrushes.

  4. In vitro antioxidant, antimicrobial, cytotoxic potential of gold and silver nanoparticles prepared using Embelia ribes.

    Science.gov (United States)

    Dhayalan, Manikandan; Denison, Michael Immanuel Jesse; L, Anitha Jegadeeshwari; Krishnan, Kathiravan; N, Nagendra Gandhi

    2017-02-01

    In recent years, the green synthesis of gold (GNPs) and silver (SNPs) nanoparticles has gained great interest among chemists and researchers. The present study reports an eco-friendly, cost-effective, rapid and easy method for the synthesis of gold and silver nanoparticles using the seed extract of Embelia ribes (SEEr) as capping and reducing agent. The synthesised GNPs and SNPs were characterised using the following techniques: UV-vis spectroscopy, DLS, HR-TEM, FT-IR and XRD. The free radical scavenging potential of GNPs and SNPs was measured by DPPH assay and Phosphomolybdenum assay. Further, the antimicrobial activity against two micro-organisms were tested using disc diffusion method and cytotoxicity of GNPs and SNPs was determined against MCF-7 cell lines at different concentrations by MTT assay. Both the GNPs and SNPs prepared from E. ribes comparatively showed promising results thereby proving their clinical importance.

  5. Extracts from New Zealand Undaria pinnatifida Containing Fucoxanthin as Potential Functional Biomaterials against Cancer in Vitro

    Directory of Open Access Journals (Sweden)

    Sheng Kelvin Wang

    2014-03-01

    Full Text Available This study tested extracts from New Zealand seaweed Undaria pinnatifida containing fucoxanthin, in parallel with pure fucoxanthin, in nine human cancer cell lines, for anticancer activity. Growth inhibition effects of extracts from Undaria pinnatifida were found in all types of cancer cell lines in dose- and time- dependent manners. Cytotoxicity of fucoxanthin in three human non-cancer cell lines was also tested. Compared with pure fucoxanthin, our extracts containing low level of fucoxanthin were found to be more effective in inhibiting the growth of lung carcinoma, colon adenocarcinoma and neuroblastoma. Our results suggest that fucoxanthin is a functional biomaterial that may be used as a chemopreventive phytochemical or in combination chemotherapy. Furthermore, we show for the first time that some unknown compounds with potential selective anti-cancer effects may exist in extracts of New Zealand Undaria pinnatifida, and New Zealand Undaria pinnatifida could be used as a source for either functional biomaterial extraction or production of functional food.

  6. Intrinsic differentiation potential of adolescent human tendon tissue: an in-vitro cell differentiation study

    Directory of Open Access Journals (Sweden)

    Weinans Harrie

    2007-02-01

    Full Text Available Abstract Background Tendinosis lesions show an increase of glycosaminoglycan amount, calcifications, and lipid accumulation. Therefore, altered cellular differentiation might play a role in the etiology of tendinosis. This study investigates whether adolescent human tendon tissue contains a population of cells with intrinsic differentiation potential. Methods Cells derived from adolescent non-degenerative hamstring tendons were characterized by immunohistochemistry and FACS-analysis. Cells were cultured for 21 days in osteogenic, adipogenic, and chondrogenic medium and phenotypical evaluation was carried out by immunohistochemical and qPCR analysis. The results were compared with the results of similar experiments on adult bone marrow-derived stromal cells (BMSCs. Results Tendon-derived cells stained D7-FIB (fibroblast-marker positive, but α-SMA (marker for smooth muscle cells and pericytes negative. Tendon-derived cells were 99% negative for CD34 (endothelial cell marker, and 73% positive for CD105 (mesenchymal progenitor-cell marker. In adipogenic medium, intracellular lipid vacuoles were visible and tendon-derived fibroblasts showed upregulation of adipogenic markers FABP4 (fatty-acid binding protein 4 and PPARG (peroxisome proliferative activated receptor γ. In chondrogenic medium, some cells stained positive for collagen 2 and tendon-derived fibroblasts showed upregulation of collagen 2 and collagen 10. In osteogenic medium Von Kossa staining showed calcium deposition although osteogenic markers remained unaltered. Tendon-derived cells and BMCSs behaved largely comparable, although some distinct differences were present between the two cell populations. Conclusion This study suggests that our population of explanted human tendon cells has an intrinsic differentiation potential. These results support the hypothesis that there might be a role for altered tendon-cell differentiation in the pathophysiology of tendinosis.

  7. In vitro comparative analysis of monocrotophos degrading potential of Aspergillus flavus, Fusarium pallidoroseum and Macrophomina sp.

    Science.gov (United States)

    Jain, Rachna; Garg, Veena; Yadav, Deepak

    2014-06-01

    Fungal degradation is emerging as a new powerful tool for the removal of potent neurotoxin pesticide, monocrotophos. Therefore, the present study is aimed at comparative characterization of monocrotophos degrading ability of three different fungal strains. Fungal strains were isolated from local agricultural soil by enrichment culture method, screened by gradient culture and identified as Aspergillus flavus, Fusarium pallidoroseum and Macrophomina sp. Growth kinetics revealed a direct positive influence of monocrotophos on the viability of fungal isolates. Fungal degradation was studied in phosphorus free liquid culture medium supplemented with 150 mg L(-1) concentration of monocrotophos for a period of 15 days under optimized culture conditions. Degradation of MCP followed first order kinetics with kdeg of 0.007, 0.002 and 0.005 day(-1) and half life (t1/2) of 4.21, 12.64 and 6.32 days for A. flavus, F. pallidoroseum and Macrophomina sp. respectively. To the best of our knowledge, it is the first report signifying the potential of monocrotophos degradation by Fusarium and Macrophomina sp. The results were further confirmed by HPTLC and FTIR which indicates disappearance of monocrotophos by hydrolytic cleavage of vinyl phosphate bond. Degradation of monocrotophos by fungal isolates was accompanied by the release of extracellular alkaline phosphatases, inorganic phosphates and ammonia. The overall comparative analysis followed the order of A. flavus > Macrophomina sp. > F. pallidoroseum. Therefore, it could be concluded from the study that these three different fungal strains could be effectively used as a potential candidate for the removal of monocrotophos from contaminated sites.

  8. An in vitro analysis of the cariogenic and erosive potential of pediatric liquid analgesics.

    Science.gov (United States)

    Saeed, Shaam; Bshara, Nada; Trak, Juliana; Mahmoud, Ghiath

    2015-01-01

    Analgesics such as Ibuprofen and Paracetamol, which are clinically used for the treatment of fever and/or pain, are among the most frequently used pediatric medicines. However, the properties of these preparations determine their cariogenic and erosive potential. The main objective of this study was to analyze the pH, viscosity and total sugar content in a variety of Syrian pediatric liquid analgesics (PLA). A total of 16 available liquid analgesics that belong to the Paracetamol and Ibuprofen group were analysed. The endogenous pH was measured using a digital pH meter, the viscosity was measured using a digital rotational viscometer and the total sugar content was performed according to Fehling method. Data were presented by means of descriptive statistics (mean, standard deviation, minimum and maximum values). The mean endogenous pH of PLA was 4.63 ± 0.57 ranging between 3.93 and 5.68, and almost all of analgesics (93.8%) had pH values ≤5.5. The mean viscosity of PLA was 243.56 ± 186.6 cP and varied between 20.5 cP and 640.5 cP. Sugars were detected in 11 (68.75%) analgesics, and varied considerably among sugar-containing analgesics from 5.38 to 69.4 (g/100 mL) with a mean concentration of 24.97 ± 23.24 g/100 mL. PLA are potentially cariogenic and erosive because of low pH, high viscosity and high total sugar content. This may increase our concerns about the dental health of children who take liquid analgesics frequently or when long-term treatment is indicated.

  9. An in vitro analysis of the cariogenic and erosive potential of pediatric liquid analgesics

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    Shaam Saeed

    2015-01-01

    Full Text Available Background: Analgesics such as Ibuprofen and Paracetamol, which are clinically used for the treatment of fever and/or pain, are among the most frequently used pediatric medicines. However, the properties of these preparations determine their cariogenic and erosive potential. Aims: The main objective of this study was to analyze the pH, viscosity and total sugar content in a variety of Syrian pediatric liquid analgesics (PLA. Setting and Design: A total of 16 available liquid analgesics that belong to the Paracetamol and Ibuprofen group were analysed. Materials and Methods: The endogenous pH was measured using a digital pH meter, the viscosity was measured using a digital rotational viscometer and the total sugar content was performed according to Fehling method. Statistical Analysis: Data were presented by means of descriptive statistics (mean, standard deviation, minimum and maximum values. Results: The mean endogenous pH of PLA was 4.63 ± 0.57 ranging between 3.93 and 5.68, and almost all of analgesics (93.8% had pH values ≤5.5. The mean viscosity of PLA was 243.56 ± 186.6 cP and varied between 20.5 cP and 640.5 cP. Sugars were detected in 11 (68.75% analgesics, and varied considerably among sugar-containing analgesics from 5.38 to 69.4 (g/100 mL with a mean concentration of 24.97 ± 23.24 g/100 mL. Conclusion: PLA are potentially cariogenic and erosive because of low pH, high viscosity and high total sugar content. This may increase our concerns about the dental health of children who take liquid analgesics frequently or when long-term treatment is indicated.

  10. Proliferation and differentiation potential of mouse adult hepatic progenitor cells cultured in vitro.

    Science.gov (United States)

    Song, Lujun; Wang, Hongshan; Gao, Xiaodong; Shen, Kuntang; Niu, Weixin; Qin, Xinyu

    2010-02-01

    This study aimed to isolate the stem cells or progenitors, if exist, from normal adult mouse liver and investigate their potential of proliferation and differentiation. Hepatocytes were isolated by modified two-step liver perfusion method and centrifugation, and then cultured in modified serumcontaining DMEM for observation more than 60 days. Immunofluorescence technique was applied to check the hepatocytes and to examine the formation of colonies with albumin, alpha-fetoprotein (AFP) and cytokeratin 19 (CK19). Results showed that some hepatocytes that were strongly positive for hepatocyte specific markers albumin on Day 1 in culture, could be activated at Days 2-3, followed by rapid proliferation and formation of colonies. The colonies could expand continually for more than 60 days. On Day 5, all the cells in the colony expressed hepatic stem cell (HSC) markers AFP. With the time of culture, some cells in colonies lost ability to divide at Days 13-15, and differentiated into cells which had a large cytoplasm and some two nuclei, similar to the appearance of mature hepatocytes morphologically. These differentiated cells demonstrated strong expression of albumin. Around Day 30, some big cells appeared in colonies and expressed bile duct cell marker CK19. Therefore, this subpopulation of mouse hepatocytes could acquire some characteristics of immature hepatocytes and showed the profile of hepatic progenitor cells with a high proliferating ability and bi-potential of differentiation. They were isolated from normal adult mouse, hence, named adult hepatic progenitor cells (AHPCs). Mouse AHPCs may be used as an HSC model for hepatocytes transplantation and hepatopathy study.

  11. In Vitro Protective Potentials of Annona muricata Leaf Extracts Against Sodium Arsenite-induced Toxicity.

    Science.gov (United States)

    George, Vazhappilly Cijo; Kumar, Devanga Ragupathi Naveen; Suresh, Palamadai Krishnan; Kumar, Rangasamy Ashok

    2015-01-01

    Sodium arsenite (NaAsO2) is a metalloid which is present widely in the environment and its chronic exposure can contribute to the induction of oxidative stress, resulting in disturbances in various metabolic functions including liver cell death. Hence, there is a need to develop drugs from natural sources, which can reduce arsenic toxicity. While there have been reports regarding the antioxidant and protective potentials of Annona muricataleaf extracts, our study is the first ofits kind to extend these findings by specifically evaluating its ability to render protection against sodium arsenite (NaAsO2) induced toxicity (10 μM) in WRL-68 (human hepatic cells) and human erythrocytes by employing XTT and haemolysis inhibition assays respectively. The methanolic extract exhibited higher activity than the aqueous extract in both assays. The results showed a dose-dependent decrease in arsenic toxicity in both WRL-68 cells and erythrocytes, suggesting the protective nature of Annona muricatato mitigate arsenic toxicity. Hence the bioactive extracts can further be scrutinized for the identification and characterization of their principal contributors.

  12. In Vitro Enzyme Inhibition Potentials and Antioxidant Activity of Synthetic Flavone Derivatives

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    Mohammad Shoaib

    2015-01-01

    Full Text Available Free radicals are produced by an important chemical process known as oxidation that in turn initiates chain reactions to damage the cells and originate oxidative stress. Flavones have got special position in research field of natural and synthetic organic chemistry due to their biological capabilities as antioxidant. The antioxidants are known to possess extensive biological effects that include antiviral, antibacterial, anti-inflammatory, antithrombotic, and vasodilatory activities. The simple flavone (F1 and substituted flavone derivatives (F2–F5 have been synthesized from o-hydroxyacetophenone and benzaldehyde derivatives in good yield. The structures have been established by different spectroscopic techniques like 1H NMR, 13C NMR, IR, and elemental analysis. Antioxidant profile of these compounds was established using DPPH and H2O2 free radical scavenging assay. The findings showed that halogenated flavones showed more enzyme inhibitions and antioxidant activities than simple flavones and are potential candidates for the treatment of wide range of diseases.

  13. In Vitro Evaluation of Spider Silk Meshes as a Potential Biomaterial for Bladder Reconstruction.

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    Anne Steins

    Full Text Available Reconstruction of the bladder by means of both natural and synthetic materials remains a challenge due to severe adverse effects such as mechanical failure. Here we investigate the application of spider major ampullate gland-derived dragline silk from the Nephila edulis spider, a natural biomaterial with outstanding mechanical properties and a slow degradation rate, as a potential scaffold for bladder reconstruction by studying the cellular response of primary bladder cells to this biomaterial. We demonstrate that spider silk without any additional biological coating supports adhesion and growth of primary human urothelial cells (HUCs, which are multipotent bladder cells able to differentiate into the various epithelial layers of the bladder. HUCs cultured on spider silk did not show significant changes in the expression of various epithelial-to-mesenchymal transition and fibrosis associated genes, and demonstrated only slight reduction in the expression of adhesion and cellular differentiation genes. Furthermore, flow cytometric analysis showed that most of the silk-exposed HUCs maintain an undifferentiated immunophenotype. These results demonstrate that spider silk from the Nephila edulis spider supports adhesion, survival and growth of HUCs without significantly altering their cellular properties making this type of material a suitable candidate for being tested in pre-clinical models for bladder reconstruction.

  14. In Vitro Evaluation of Spider Silk Meshes as a Potential Biomaterial for Bladder Reconstruction

    Science.gov (United States)

    Steins, Anne; Dik, Pieter; Müller, Wally H.; Vervoort, Stephin J.; Reimers, Kerstin; Kuhbier, Jörn W.; Vogt, Peter M.; van Apeldoorn, Aart A.; Coffer, Paul J.; Schepers, Koen

    2015-01-01

    Reconstruction of the bladder by means of both natural and synthetic materials remains a challenge due to severe adverse effects such as mechanical failure. Here we investigate the application of spider major ampullate gland-derived dragline silk from the Nephila edulis spider, a natural biomaterial with outstanding mechanical properties and a slow degradation rate, as a potential scaffold for bladder reconstruction by studying the cellular response of primary bladder cells to this biomaterial. We demonstrate that spider silk without any additional biological coating supports adhesion and growth of primary human urothelial cells (HUCs), which are multipotent bladder cells able to differentiate into the various epithelial layers of the bladder. HUCs cultured on spider silk did not show significant changes in the expression of various epithelial-to-mesenchymal transition and fibrosis associated genes, and demonstrated only slight reduction in the expression of adhesion and cellular differentiation genes. Furthermore, flow cytometric analysis showed that most of the silk-exposed HUCs maintain an undifferentiated immunophenotype. These results demonstrate that spider silk from the Nephila edulis spider supports adhesion, survival and growth of HUCs without significantly altering their cellular properties making this type of material a suitable candidate for being tested in pre-clinical models for bladder reconstruction. PMID:26689371

  15. Evaluation of in vitro Probiotic Potential of Pediococcus pentosaceus OZF Isolated from Human Breast Milk.

    Science.gov (United States)

    Osmanagaoglu, Ozlem; Kiran, Fadime; Ataoglu, Haluk

    2010-10-01

    This study was conducted to evaluate the probiotic properties of Pediococcus pentosaceus OZF isolated from human breast milk. The results obtained so far suggest that the strain is resistant to low pH, bile salt, pepsin and pancreatin, so it could survive while passing through the upper part of the gastrointestinal tract and reveal its potential probiotic action on host organism. The strain was non-pathogenic (γ-hemolytic), produced anti-Listerial bacteriocin, exhibited a strong autoaggregating phenotype (85.71%) and demonstrated 6.26 and 12.99% coaggregation with Salmonella enterica serotype Typhimurium SL 1344 and Escherichia coli LMG 3083 (ETEC), respectively. The degree of adhesion of Ped. pentosaceus OZF to the human Caco-2 cell line was investigated and when compared to the adhesion of pathogenic strains tested, it was shown to inhibit the growth of human enterotoxigenic E. coli LMG 3083 (ETEC) and of Salm. Typhimurium SL 1344. Ped. pentosaceus OZF seems to adhere to human intestinal cells via mechanisms that involve different combinations of carbohydrate and lipid factors on the bacteria and eukaryotic cell surface. The percentage of adhesion to n-hexadecane was 34% showing that the surface was rather hydrophilic. Higher affinity displayed by Ped. pentosaceus OZF for chloroform demonstrates the basic property of a cell, which may be due to the presence of carboxylic groups on the cell surface.

  16. Novel vitamin B12-producing Enterococcus spp. and preliminary in vitro evaluation of probiotic potentials.

    Science.gov (United States)

    Li, Ping; Gu, Qing; Wang, Yuejiao; Yu, Yue; Yang, Lanlan; Chen, Jieyan V

    2017-08-01

    Vitamin B12 is an essential nutrient required for crucial metabolic processes in humans. Vitamin B12-producing lactic acid bacteria (LAB) have been attracting increased attentions currently because of the generally recognized as safe (GRAS) status. Most of recent studies focused on Lactobacillus, and little is known about B12-producing Enterococcus. In the present study, five Enterococcus strains isolated from infant feces were identified as vitamin B12 producers. Among them, Enterococcus faecium LZ86 had the highest B12 production (499.8 ± 83.7 μg/L), and the B12 compound from LZ86 was identified as the biological active adenosylcobalamin, using reversed phase high-performance liquid (RP-HPLC) chromatogram. We examined basic probiotic and safety properties of E. faecium LZ86 and found that it was able to survive harsh environmental conditions (hot temperature, cold temperature, ethanol and osmotic stresses), tolerate gastric acid (pH 2.0, 3 h) and bile salts (0.3%), and adhere to Caco-2 cells. We also showed that E. faecium LZ86 is devoid of transferable antibiotic resistance and potential virulence factors. Together, here we report a B12-producing E. faecium strain LZ86 firstly, which has desirable probiotic properties and may serve as a good candidate for vitamin B12 fortification in food industry.

  17. In vitro evaluation of wound healing and antimicrobial potential of ozone therapy.

    Science.gov (United States)

    Borges, Gabriel Álvares; Elias, Silvia Taveira; da Silva, Sandra Márcia Mazutti; Magalhães, Pérola Oliveira; Macedo, Sergio Bruzadelli; Ribeiro, Ana Paula Dias; Guerra, Eliete Neves Silva

    2017-03-01

    Although ozone therapy is extensively applied when wound repair and antimicrobial effect are necessary, little is known about cellular mechanisms regarding this process. Thus, this study aimed to evaluate ozone cytotoxicity in fibroblasts (L929) and keratinocytes (HaCaT) cell lines, its effects on cell migration and its antimicrobial activity. Cells were treated with ozonated phosphate-buffered saline (8, 4, 2, 1, 0.5 and 0.25 μg/mL ozone), chlorhexidine 0.2% or buffered-solution, and cell viability was determined through MTT assay. The effect of ozone on cell migration was evaluated through scratch wound healing and transwell migration assays. The minimum inhibitory concentrations for Candida albicans and Staphylococcus aureus were determined. Ozone showed no cytotoxicity for the cell lines, while chlorhexidine markedly reduced cell viability. Although no significant difference between control and ozone-treated cells was observed in the scratch assay, a considerable increase in fibroblasts migration was noticed on cells treated with 8 μg/mL ozonated solution. Ozone alone did not inhibit growth of microorganisms; however, its association with chlorhexidine resulted in antimicrobial activity. This study confirms the wound healing and antimicrobial potential of ozone therapy and presents the need for studies to elucidate the molecular mechanisms through which it exerts such biological effects. Copyright © 2017 European Association for Cranio-Maxillo-Facial Surgery. Published by Elsevier Ltd. All rights reserved.

  18. Mechanisms underlying the wound healing potential of propolis based on its in vitro antioxidant activity.

    Science.gov (United States)

    Cao, Xue-Ping; Chen, Yi-Fan; Zhang, Jiang-Lin; You, Meng-Meng; Wang, Kai; Hu, Fu-Liang

    2017-10-15

    Propolis is a resinous substance collected by honeybees, Apis mellifera, from various plant sources. Having various pharmacological and biological activities, it has been used in folk medicine and complementary therapies since ancient times. To evaluate the effects and underlying mechanism of the protective effects of the ethanol extract of Chinese propolis (EECP) on L929 cells injured by hydrogen peroxide (H2O2). The wound healing activities of EECP in L929 cells with H2O2-induced damage were investigated. The main components of EECP were analyzed by RP-HPLC, and the free radical scavenging capacity and reducing power were also measured. The effects of EECP on the expression of antioxidant-related genes in fibroblast L929 cells were determined using qRT-PCR and western blotting. EECP had significant protective effects against cell death induced by H2O2 and significantly inhibited the decline of collagen mRNA expression caused by H2O2 in L929 cells. EECP induced the expression of antioxidant-related genes, such as HO-1, GCLM, and GCLC, which has great implications for the potential of propolis to alleviate oxidative stress in wound tissues. The protective effects of propolis have great implications for using propolis as a wound healing regent. Copyright © 2017 Elsevier GmbH. All rights reserved.

  19. Antitumor potential of conjugable valinomycins bearing hydroxyl sites: in vitro studies.

    Science.gov (United States)

    Iacobazzi, Rosa M; Annese, Cosimo; Azzariti, Amalia; D'Accolti, Lucia; Franco, Massimo; Fusco, Caterina; La Piana, Gianluigi; Laquintana, Valentino; Denora, Nunzio

    2013-12-12

    Following our pioneering studies on the direct and efficient introduction of derivatizable hydroxyl handles into the valinomycin (VLM, 1) structure, a K(+)-ionophore with potent antitumor activity, the ensuing conjugable analogues (HyVLMs 2, 3, and 4) have herein been compared to the parent macrocycle for their potential antiproliferative effects on a panel of cancer cell lines, namely, human MCF-7, A2780, and HepG2, as well as rat C6 cells. On the basis of IC50 values, we find that hydroxyl analogues 3 and 4 are only moderately less active than 1, while analogue 2 experiences a heavily diminished activity. Cytofluorimetric analyses of MCF-7 cells treated with HyVLMs suggest that the latter depolarize mitochondria, thus retaining the typical VLM behavior. It is likely that C6 cells, for which the exceptionally potent cytotoxicity of VLM has never reported previously, follow the same fate, as evidenced by alteration of mitochondrial morphology upon incubation with each ionophore.

  20. Potential anti-osteoporotic effects of herbal extracts on osteoclasts, osteoblasts and chondrocytes in vitro

    Science.gov (United States)

    2014-01-01

    Background Osteoporosis (OP) is one of the most serious diseases in the modern world, and OP patients frequently suffer from fragility fractures in the hip, spine and wrist, resulting in a limited quality of life. Although bisphosphonates (BPs) are the most effective class of anti-bone-resorptive drugs currently available and the most commonly prescribed for the clinical treatment of OP, they are known to cause serious side effects such as bisphosphonate-related osteonecrosis of the jaw. Novel therapeutic materials that can replace the use of BPs have therefore been developed. Methods We commenced an institutional collaborative project in which candidates of herbal extracts were selected from more than 400 bioactive herbal products for their potential therapeutic effects not only in OP, but also in oral and skeletal diseases. In the present study, we report on 3 Chinese medical herbal extracts from the root barks of Melia azedarach, Corydalis turtschaninovii, and Cynanchum atratum. Results All of these extracts inhibited osteoclast proliferation and induced apoptosis by up-regulation of caspase activity and increase of mitochondrial pro-apoptotic proteins expression. Furthermore, the extracts enhanced differentiation, but did not affect proliferation of both osteoblasts and chondrocytes. The osteo-inducible effect was also observed in cultured primary bone marrow cells. Conclusions Although these extracts have been utilized in traditional Chinese medicine for hundreds of years, there are no reports to our knowledge, on their therapeutic effects in OP. In this study, we elucidate the potency of these herbal extracts as novel candidates for OP therapy. PMID:24438322

  1. MC70 potentiates doxorubicin efficacy in colon and breast cancer in vitro treatment.

    Science.gov (United States)

    Azzariti, Amalia; Quatrale, Anna E; Porcelli, Letizia; Colabufo, Nicola A; Cantore, Mariangela; Cassano, Giuseppe; Gasparre, Giuseppe; Iannelli, Giuseppina; Tommasi, Stefania; Panaro, Maria A; Paradiso, Angelo

    2011-11-16

    A major limitation of cancer treatment is the ability of cancer cells to develop resistance to chemotherapeutic drugs, by the establishment of multidrug resistance. Here, we characterize MC70 as ABC transporters inhibitor and anticancer agent, alone or with chemotherapy. MC70 was analyzed for its interaction with ABCB1, ABCG2 and ABCC1 by specific transport assays. In breast and colon cancer cell lines, cell growth and apoptosis were measured by MTT assay and DNA laddering Elisa kit, respectively. Cell cycle perturbation and cellular targets modulation were analyzed by Flow-cytometry and Western blotting, respectively. MC70 interacted with ABC transporters. In breast cancer cells, MC70 slightly inhibited cell proliferation strongly enhancing doxorubicin effectiveness. By contrast, MC70 was found to inhibit cell growth in colon cancer cells without affecting doxorubicin efficacy and in combination with topoisomerase I inhibitors it could be a promising therapeutic approach. What is more, it was also observed that MC70 induced apoptosis, canceled in favor of necrosis when given in combination with high doses of doxorubicin. MC70 inhibited cell migration probably through its interaction with sigma-1 receptor. Modulations of i) cell cycle, ii) pAkt and the phosphorylation of the three MAPKs were highlighted, while any activity was excluded at transcription level, thus accounting for the phenotypic effects observed. MC70 might be considered as a new potential anticancer agent capable to i) enhance chemotherapy effectiveness and ii) to play a contributory role in the treatment of chemotherapy resistant tumors. Copyright © 2011 Elsevier B.V. All rights reserved.

  2. Short-term in vitro and in vivo analyses for assessing the tumor-promoting potentials of cigarette smoke condensates.

    Science.gov (United States)

    Curtin, Geoffrey M; Hanausek, Margaret; Walaszek, Zbigniew; Mosberg, Arnold T; Slaga, Thomas J

    2004-09-01

    Previous studies found that repeated application of smoke condensate from tobacco-burning reference cigarettes to chemically initiated SENCAR mouse skin promoted the development of tumors in a statistically significant and dose-dependent manner, while condensate from prototype cigarettes that primarily heat tobacco promoted statistically fewer tumors. Based on the recognized correlation between sustained, potentiated epidermal hyperplasia and tumor promotion, we conducted tests to examine the utility of selected short-term analyses for discriminating between condensates exhibiting significantly different promotion activities. In vitro analyses assessing the potential for inducing cytotoxicity (ATP bioluminescence) or free radical production (cytochrome c reduction, salicylate trapping) demonstrated significant reductions when comparing condensate collected from prototype cigarettes to reference condensate. Short-term in vivo analyses conducted within the context of a mouse skin, tumor-promotion protocol (i.e., comparative measures of epidermal thickness, proliferative index, myeloperoxidase activity, leukocyte invasion, mutation of Ha-ras, and formation of modified DNA bases) provided similar results. Reference condensate induced statistically significant and dose-dependent increases (relative to vehicle control) for nearly all indices examined, while prototype condensate possessed a significantly reduced potential for inducing changes that we regarded as consistent with sustained epidermal hyperplasia and/or inflammation. Collectively, these data support the contention that selected short-term analyses associated with sustained hyperplasia and/or inflammation are capable of discriminating between smoke condensates with dissimilar tumor-promotion potentials. Moreover, our results suggest that comparative measures of proliferative index and myeloperoxidase activity, both possessing favorable correlation coefficients relative to tumor formation (i.e., > or = 0

  3. Effect and mechanism of thrombospondin-1 on the angiogenesis potential in human endothelial progenitor cells: an in vitro study.

    Directory of Open Access Journals (Sweden)

    Qing Qin

    Full Text Available Coronary collateral circulation plays a protective role in patients with coronary artery disease (CAD. We investigated whether thrombospondin-1(TSP-1 has an inhibitory effect on angiogenesis potential in endothelial progenitor cells(EPCs and tested whether TSP-1 are altered in plasma of patients who had chronic total occlusion (CTO in at least one coronary artery and with different collateral stages(according to Rentrop grading system.We isolated early and late EPCs from human cord blood and investigated a dose-dependent effect of TSP-1 on their angiogenesis potential by Matrigel angiogenesis assay. We found that TSP-1 (5 µg/ml inhibited early EPCs incorporation into tubules after pretreatment for 1, 6 and 12 hours, respectively (83.3±11.9 versus 50.0±10.1 per field for 1 hour,161.7±12.6 versus 124.0±14.4 for 6 hours, 118.3±12.6 versus 68.0±20.1 for 12 hours, p<0.05. TSP-1 also inhibited late EPCs tubule formation at 1 µg/ml (6653.4±422.0 µm/HPFversus 5552.8±136.0 µm/HPF, p<0.05, and the inhibition was further enhanced at 5 µg/ml (6653.4±422.0 µm/HPF versus 2118.6±915.0 µm/HPF p<0.01. To explore the mechanism involved, a small interfering RNA was used. In vitro, CD47 siRNA significantly attenuated TSP-1's inhibition of angiogenesis on late EPCs and similar results were obtained after functional blocking by anti-CD47 antibody. Then we investigated pathways downstream of CD47 and found TSP-1 regulated VEGF-induced VEGFR2 phosphorylation via CD47. Furthermore, we examined plasma TSP-1 levels in patients with CTO who developed different stages of collaterals and found a paradoxical higher level of TSP-1 in patients with good collaterals compared with bad ones (612.9±554.0 ng/ml versus 224.4±132.4 ng/ml, p<0.05.TSP-1 inhibited angiogenesis potential of early and late EPCs in vitro. This inhibition may be regulated by TSP-1's interaction with CD47, resulting in down regulation of VEGFR2 phosphorylation. In patients with CTO, there

  4. In vitro antioxidant potential and deoxyribonucleic acid protecting activity of CNB-001, a novel pyrazole derivative of curcumin

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    Richard L Jayaraj

    2014-01-01

    Full Text Available Background: Free radicals are underpinned to initiate cascade of toxic events leading to oxidative stress and resultant cell death in many neurodegenerative disorders. Now-a-days antioxidants have become mandatory in the treatment of various diseases apart from the drug′s modes of action. CNB-001, a novel hybrid molecule synthesized by combining curcumin and cyclohexyl bisphenol A is known to possess various biological activities, but the antioxidative property of the compound has not yet been elucidated. Aim: The present study is aimed to analyze various free radicals scavenging by employing in vitro antioxidant assays and to evaluate the deoxyribonucleic acid (DNA protecting the ability of CNB-001 against hydroxyl radicals. Materials and methods: The in vitro antioxidant potential of CNB-001 was evaluated by analyzing its ability to scavenge DPPH, ABTS, nitric oxide, superoxide, hydrogen peroxide, superoxide anion, hydroxyl, hydrogen peroxide radicals and reducing power using spectroscopic method. The DNA protecting activity of CNB-001 was also evaluated on pUC19 plasmid DNA subjected to hydroxyl radicals using standard agarose gel electrophoresis. Results: From the assays, it was observed that CNB-001 scavenged free radicals effectively in a dose dependent manner. CNB-001 scavenged 2,2-diphenyl-1-picrylhydrazyl (IC50 = 44.99 μg/ml, 2,2-azinobis (3-ethylbenzothiazoline-6-sulfonic acid (IC50 = 17.99 μg/ml, nitric oxide (IC50 = 1.36 μg/ml, superoxide radical (IC50 = 77.17 μg/ml, hydrogen peroxide (IC50 = 492.7 μg/ml, superoxide (IC50 = 36.92 μg/ml and hydroxyl (IC50 = 456.5 μg/ml radicals effectively and the reducing power was found to be 11.53 μg/ml. CNB-001 showed considerable protecting activity against plasmid DNA (pUC19 strand scission by ·OH at dose dependent manner. Conclusion: Results from these assays concluded that CNB-001 has a good antioxidant potential by reducing reactive oxygen and reactive nitrogen radicals and it

  5. In vitro assessment of the tooth staining potential of endodontic materials in a bovine tooth model.

    Science.gov (United States)

    Dettwiler, Christian A; Walter, Mirella; Zaugg, Lucia K; Lenherr, Patrik; Weiger, Roland; Krastl, Gabriel

    2016-12-01

    Traumatized teeth requiring endodontic intervention may discolor as a result of the treatment performed. Thus, the aim was to investigate the discoloration potential of different endodontic cements, dressings, and irrigants used in dental traumatology. Cylindrical cavities were prepared in 330 bovine enamel-dentine blocks (10 × 10 × 3.5 mm), leaving 2 mm of enamel and dentine on the labial side. The specimens were randomly assigned to 22 groups (n = 15). The cavities were filled with a range of endodontic materials, sealed with composite and stored in physiological saline. The color of the labial enamel surface was measured with a spectrophotometer at 7 time intervals: before (T0) and after the placement of the test material (T1 = baseline), after 1 week (T2), 1 month (T3), 3 months (T4), 6 months (T5), and 12 months (T6). The color difference values (ΔE) were calculated and further analyzed by anova and the Tukey-Kramer post hoc test (α = 0.05). After 12 months, significant staining was observed among the endodontic cements only in the Portland cement group with additional bismuth oxide (H-MED PC BiOx; ΔE 22.2). Specimens with other commercially available calcium silicate cements containing bismuth oxide were not significantly discolored. All specimens with endodontic dressings and irrigants were color stable except the tested triple antibiotic paste (M-TreVitaMix; ΔE 14.9) and the double antibiotic paste (N-BiMix; ΔE 14.9). Both mixtures did not contain tetracycline derivatives. The presence of bismuth oxide in calcium silicate cements was not shown to be a reliable predictor for tooth discoloration. Antibiotic pastes without tetracycline derivatives do not guarantee the color stability of teeth. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  6. Gonadotropins facilitate potential differentiation of human bone marrow mesenchymal stem cells into Leydig cells in vitro.

    Science.gov (United States)

    Hou, Lin; Dong, Qiang; Wu, Yun-Jian; Sun, Yuan-Xing; Guo, Yan-Yu; Huo, Yue-Hong

    2016-01-01

    Infertility due to low testosterone levels has increased in recent years. This has impacted the social well-being of the patients. This study was undertaken to investigate the potential of gonadotropins in facilitating differentiation of human bone marrow mesenchymal stem cells (BMSCs) into Leydig cells in vitro. BMSCs were isolated, cultured, and their biological characteristics were observed. BMSCs were induced with gonadotropins in vitro and their ability to differentiate into Leydig cells was studied. The level of expression of 3-beta hydroxysteroid dehydrogenase (3β-HSD) and secretion of testosterone were determined using flow cytometry and enzyme-linked immunosorbent assay, respectively, and the results were compared between the experimental and control groups. The cultured BMSCs showed a typical morphology of the fibroblast-like colony. The growth curve of cells formed an S-shape. After inducing the cells for 8-13 days, the cells in the experimental group increased in size and showed typical characteristics of Leydig cells, and the growth occurred in spindle or stellate shapes. Cells from the experimental group highly expressed 3β-HSD, and there was a gradual increase in the number of Leydig cells. The control group did not express 3β-HSD. The level of testosterone in the experimental group was higher than the control group (p group secreted higher levels of testosterone with increased culture time. The expression of Leydig cell-specific markers in the experimental group was significantly higher (p < 0.05). With these findings, BMSCs can be considered a new approach for the treatment of patients with low androgen levels. Copyright © 2015. Published by Elsevier Taiwan.

  7. Proximate composition, phytochemical analysis, and in vitro antioxidant potentials of extracts ofAnnona muricata(Soursop).

    Science.gov (United States)

    Agu, Kingsley C; Okolie, Paulinus N

    2017-09-01

    Numerous bioactive compounds and phytochemicals have been reported to be present Annona muricata (Soursop). Some of these chemical compounds have been linked to the ethnomedicinal properties of the plant and its antioxidant properties. The aim of this study was to assess the proximate composition, phytochemical constituents and in vitro antioxidant properties of A. muricata using standard biochemical procedures. The defatted Annona muricata crude methanolic extracts of the different parts of the plant were used for the estimation of proximate composition and phytochemical screening. The crude methanolic extracts of the different parts of the plant were also fractionated using solvent-solvent partitioning. Petroleum ether, chloroform, ethyl acetate, methanol, and methanol-water (90:10) were the solvents used for the fractionation. The different fractions obtained were then used to perform in vitro antioxidant analyses including, 1, 1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging ability, ferric reducing properties, and hydroxyl radical scavenging ability. The leaf methanolic extract had a higher lipid content, whereas its chloroform fraction demonstrated a better ability to quench DPPH free radical. The root-bark methanol-water, leaf methanol, fruit pulp chloroform, and leaf petroleum ether fractions demonstrated potent ferric reducing properties. The leaf and stem-bark petroleum ether fractions demonstrated better hydroxyl-free radical scavenging abilities. The leaf and fruit pulp of Annona muricata have a very potent antioxidant ability compared to the other parts of the plant. This can be associated with the rich phytochemicals and other phytoconstituents like phenols, flavonoids, alkaloids, and essential lipids, etc. Significant correlations were observed between the antioxidant status and phytochemicals present. These results thus suggest that some of the reported ethnomedicinal properties of this plant could be due to its antioxidant potentials.

  8. In vitro evaluation of potential drug interactions mediated by cytochrome P450 and transporters for luseogliflozin, an SGLT2 inhibitor.

    Science.gov (United States)

    Chino, Yukihiro; Hasegawa, Masatoshi; Fukasawa, Yoshiki; Mano, Yoko; Bando, Kagumi; Miyata, Atsunori; Nakai, Yasuhiro; Endo, Hiromi; Yamaguchi, Jun-Ichi

    2017-04-01

    1. We evaluated potential in vitro drug interactions of luseogliflozin, a sodium-glucose cotransporter 2 (SGLT2) inhibitor, mediated by CYP inhibition, CYP induction and drug transporters using human liver microsomes, primary hepatocytes and recombinant cells-expressing efflux or uptake transporters, respectively. 2. Human CYP inhibition studies indicated that luseogliflozin was a weak inhibitor for CYP2C19 with an IC50 value of 58.3 μM, whereas it was not an inhibitor of the other eight major isoforms that were tested. The exposure of primary hepatocytes to luseogliflozin for 72 hrs weakly induced CYP3A4 at a concentration of 10 μM, whereas it did not induce CYP1A2 or CYP2B6 at concentrations of 0.1-10 μM. 3. An in vitro transport study suggested that luseogliflozin is a substrate for human P-glycoprotein (P-gp), but not for breast cancer resistance protein (BCRP), organic anion transporting polypeptide (OATP) 1B1 and OATP1B3, organic anion transporter (OAT) 1 and OAT3, or organic cation transporter (OCT) 2. Luseogliflozin weakly inhibited OATP1B3 with an IC50 value of 93.1 μM, but those for other transporters are greater than 100 μM. 4. Based on the therapeutic plasma concentration of the drug, clinically relevant drug interactions are unlikely to occur between luseogliflozin and coadministered drugs mediated by CYPs and/or transporters.

  9. C5a enhances dysregulated inflammatory and angiogenic responses to malaria in vitro: potential implications for placental malaria.

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    Andrea Conroy

    Full Text Available Placental malaria (PM is a leading cause of maternal and infant mortality. Although the accumulation of parasitized erythrocytes (PEs and monocytes within the placenta is thought to contribute to the pathophysiology of PM, the molecular mechanisms underlying PM remain unclear. Based on the hypothesis that excessive complement activation may contribute to PM, in particular generation of the potent inflammatory peptide C5a, we investigated the role of C5a in the pathogenesis of PM in vitro and in vivo.Using primary human monocytes, the interaction between C5a and malaria in vitro was assessed. CSA- and CD36-binding PEs induced activation of C5 in the presence of human serum. Plasmodium falciparum GPI (pfGPI enhanced C5a receptor expression (CD88 on monocytes, and the co-incubation of monocytes with C5a and pfGPI resulted in the synergistic induction of cytokines (IL-6, TNF, IL-1beta, and IL-10, chemokines (IL-8, MCP-1, MIP1alpha, MIP1beta and the anti-angiogenic factor sFlt-1 in a time and dose-dependent manner. This dysregulated response was abrogated by C5a receptor blockade. To assess the potential role of C5a in PM, C5a plasma levels were measured in malaria-exposed primigravid women in western Kenya. Compared to pregnant women without malaria, C5a levels were significantly elevated in women with PM.These results suggest that C5a may contribute to the pathogenesis of PM by inducing dysregulated inflammatory and angiogenic responses that impair placental function.

  10. Total antioxidant potential of juices, beverages and hot drinks consumed in Egypt screened by DPPH in vitro assay

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    Ramadan-Hassanien, Mohamed Fawzy

    2008-09-01

    Full Text Available Plant foods contain different classes and types of antioxidants and knowledge of their total antioxidant potential (TAP, which is the cumulative capacity of food components to scavenge free radicals, would be useful for epidemiological purposes.To accomplish this, a variety of fruit juices, hot drinks and beverages commonly consumed in Egypt were analyzed using in vitro DPPH assay. The order of effectiveness of fruit juices in inhibiting free radicals was as follows: red grapes juice > mango juice > guava juice > cocktail juice > pineapple juice > orange juice > cherry juice > apple juice. Among beverages and hot drinks, teas followed by coffees had the greatest TAP. These data confirm grape juice, teas and coffees as good dietary sources of antioxidants.Las plantas comestibles contienen diferentes clases y tipos de antioxidantes y el conocimiento de su potencial antioxidante total (TAP, que es la capacidad acumulativa de los componentes de los alimentos para captar radicales libres, debería ser útil en estudios epidemiológicos. De acuerdo a esto, una variedad de zumos de fruta, bebidas calientes y bebidas consumidas habitualmente en Egipto fueron analizadas usando un ensayo in vitro con DPPH. El orden de efectividad de los zumos de frutas en inhibir los radicales libres fue el siguiente: zumo de uva tinta > zumo de mango > zumo de guayaba > zumo de macedonia de frutas > zumo de piña >zumo de naranja > zumo de cereza > zumo de manzana. Entre las bebidas y bebidas calientes, el té seguido por el café son los que tuvieron mayores TAPs. Estos datos confirman que el zumo de uva, el té y el café son buenas fuentes de antioxidantes.

  11. Pyrazinecarboxamides as Potential Elicitors of Flavonolignan and Flavonoid Production in Silybum marianum and Ononis arvensis Cultures In Vitro

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    Martin Dolezal

    2011-11-01

    Full Text Available The effect of new synthetic pyrazinecarboxamide derivatives as potential elicitors of flavonolignan and flavonoid production in Silybum marianum and Ononis arvensis cultures in vitro was investigated. Both tested elicitors increased the production of flavonolignans in S. marianum callus and suspension cultures and flavonoids in O. arvensis callus and suspension cultures. Compound I, 5-(2-hydroxybenzoyl-pyrazine-2-carboxamide, has shown to be an effective elicitor of flavonolignans and taxifoline production in Silybum marianum culture in vitro. The maximum content of silydianin (0.11% in S. marianum suspension culture was induced by 24 h elicitor application in concentration of 1.159 × 10−3 mol/L. The maximum content of silymarin complex (0.08% in callus culture of S. marianum was induced by 168 h elicitor application of a concentration 1.159 × 10−4 mol/L, which represents contents of silydianin (0.03%, silychristin (0.01% and isosilybin A (0.04% compared with control. All three tested concentrations of compound II, N-(2-bromo-3-methylphenyl-5-tert-butylpyrazin-2-carboxamide increased the flavonoid production in callus culture of O. arvensis in a statistically significant way. The best elicitation effect of all elicitor concentrations had the weakest c3 concentration (8.36 × 10−6 mol/L after 168 h time of duration. The maximum content of flavonoids (about 5,900% in suspension culture of O. arvensis was induced by 48 h application of c3 concentration (8.36 × 10−6 mol/L.

  12. Fasting cycles potentiate the efficacy of gemcitabine treatment in in vitro and in vivo pancreatic cancer models

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    Mazza, Tommaso; Panebianco, Concetta; Saracino, Chiara; Pereira, Stephen P.; Graziano, Paolo; Pazienza, Valerio

    2015-01-01

    Background/aims Pancreatic cancer (PC) is ranked as the fourth leading cause of cancer-related deaths worldwide. Despite recent advances in treatment options, a modest impact on the outcome of the disease is observed so far. Short-term fasting cycles have been shown to potentiate the efficacy of chemotherapy against glioma. The aim of this study was to assess the effect of fasting cycles on the efficacy of gemcitabine, a standard treatment for PC patients, in vitro and in an in vivo pancreatic cancer mouse xenograft model. Materials and Methods BxPC-3, MiaPaca-2 and Panc-1 cells were cultured in standard and fasting mimicking culturing condition to evaluate the effects of gemcitabine. Pancreatic cancer xenograft mice were subjected to 24h starvation prior to gemcitabine injection to assess the tumor volume and weight as compared to mice fed ad libitum. Results Fasted pancreatic cancer cells showed increased levels of equilibrative nucleoside transporter (hENT1), the transporter of gemcitabine across the cell membrane, and decreased ribonucleotide reductase M1 (RRM1) levels as compared to those cultured in standard medium. Gemcitabine was more effective in inducing cell death on fasted cells as compared to controls. Consistently, xenograft pancreatic cancer mice subjected to fasting cycles prior to gemcitabine injection displayed a decrease of more than 40% in tumor growth. Conclusion Fasting cycles enhance gemcitabine effect in vitro and in the in vivo PC xenograft mouse model. These results suggest that restrictive dietary interventions could enhance the efficacy of existing cancer treatments in pancreatic cancer patients. PMID:26176887

  13. Repositioning FDA Drugs as Potential Cruzain Inhibitors from Trypanosoma cruzi: Virtual Screening, In Vitro and In Vivo Studies

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    Isidro Palos

    2017-06-01

    Full Text Available Chagas disease (CD is a neglected disease caused by the parasite Trypanosoma cruzi, which affects underdeveloped countries. The current drugs of choice are nifurtimox and benznidazole, but both have severe adverse effects and less effectivity in chronic infections; therefore, the need to discover new drugs is essential. A computer-guided drug repositioning method was applied to identify potential FDA drugs (approved and withdrawn as cruzain (Cz inhibitors and trypanocidal effects were confirmed by in vitro and in vivo studies. 3180 FDA drugs were virtually screened using a structure-based approach. From a first molecular docking analysis, a set of 33 compounds with the best binding energies were selected. Subsequent consensus affinity binding, ligand amino acid contact clustering analysis, and ranked position were used to choose four known pharmacological compounds to be tested in vitro. Mouse blood samples infected with trypomastigotes from INC-5 and NINOA strains were used to test the trypanocidal effect of four selected compounds. Among these drugs, one fibrate antilipemic (etofyllin clofibrate and three β-lactam antibiotics (piperacillin, cefoperazone, and flucloxacillin showed better trypanocidal effects (LC50 range 15.8–26.1 μg/mL in comparison with benznidazole and nifurtimox (LC50 range 33.1–46.7 μg/mL. A short-term in vivo evaluation of these compounds showed a reduction of parasitemia in infected mice (range 90–60% at 6 h, but this was low compared to benznidazole (50%. This work suggests that four known FDA drugs could be used to design and obtain new trypanocidal agents.

  14. In vitro and in vivo antimalarial potential of oleoresin obtained from Copaifera reticulata Ducke (Fabaceae) in the Brazilian Amazon rainforest.

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    de Souza, Giovana A G; da Silva, Nazaré C; de Souza, Juarez; de Oliveira, Karen R M; da Fonseca, Amanda L; Baratto, Leopoldo C; de Oliveira, Elaine C P; Varotti, Fernando de Pilla; Moraes, Waldiney P

    2017-01-15

    In view of the wide variety of the flora of the Amazon region, many plants have been studied in the search for new antimalarial agents. Copaifera reticulata is a tree distributed throughout the Amazon region which contains an oleoresin rich in sesquiterpenes and diterpenes with β-caryophyllene as the major compound. The oleoresin has demonstrated antiparasitic activity against Leishmania amazonensis. Because of this previously reported activity, this oleoresin would be expected to also have antimalarial activity. In this study we evaluated the in vitro and in vivo antimalarial potential of C. reticulata oleoresin. In vitro assays were done using P. falciparum W2 and 3D7 strains and the human fibroblast cell line 26VA Wi-4. For in vivo analysis, BALB/c mice were infected with approximately 106 erythrocytes parasitized by P. berghei and their parasitemia levels were observed over 7 days of treatment with C. reticulata; hematological and biochemical parameters were analyzed at the end of experiment. The oleoresin of C. reticulata containing the sesquiterpenes β-caryophyllene (41.7%) and β-bisabolene (18.6%) was active against the P. falciparum W2 and 3D7 strains (IC50 = 1.66 and 2.54 µg/ml, respectively) and showed low cytotoxicity against the 26VA Wi-4 cell line (IC50 > 100 µg/ml). The C. reticulata oleoresin reduced the parasitemia levels of infected animals and doses of 200 and 100 mg/kg/day reached a rate of parasitemia elimination resembling that obtained with artemisinin 100 mg/kg/day. In addition, treatment with oleoresin improved the hypoglycemic, hematologic, hepatic and renal parameters of the infected animals. The oleoresin of C. reticulata has antimalarial properties and future investigations are necessary to elucidate its mechanism of action. Copyright © 2016 Elsevier GmbH. All rights reserved.

  15. Evaluation of anticataract potential of Triphala in selenite-induced cataract: In vitro and in vivo studies

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    Suresh Kumar Gupta

    2010-01-01

    Full Text Available Triphala (TP is composed of Emblica officinalis, Terminalia chebula, and Terminalia belerica. The present study was undertaken to evaluate its anticataract potential in vitro and in vivo in a selenite-induced experimental model of cataract. In vitro enucleated rat lenses were maintained in organ culture containing Dulbecco′s Modified Eagles Medium alone or with the addition of 100΅M selenite. These served as the normal and control groups, respectively. In the test group, the medium was supplemented with selenite and different concentrations of TP aqueous extract. The lenses were incubated for 24 h at 37°C. After incubation, the lenses were processed to estimate reduced glutathione (GSH, lipid peroxidation product, and antioxidant enzymes. In vivo selenite cataract was induced in 9-day-old rat pups by subcutaneous injection of sodium selenite (25 μmole/kg body weight. The test groups received 25, 50, and 75 mg/kg of TP intraperitoneally 4 h before the selenite challenge. At the end of the study period, the rats′ eyes were examined by slit-lamp. TP significantly (P < 0.01 restored GSH and decreased malondialdehyde levels. A significant restoration in the activities of antioxidant enzymes such as superoxide dismutase (P < 0.05, catalase (P < 0.05, glutathione peroxidase (P < 0.05, and glutathione-s-transferase (P < 0.005 was observed in the TP-supplemented group compared to controls. In vivo TF 25mg/kg developed only 20% nuclear cataract as compared to 100% in control. TP prevents or retards experimental selenite-induced cataract. This effect may be due to antioxidant activity. Further studies are warranted to explore its role in human cataract.

  16. Self-assembled amphotericin B-loaded polyglutamic acid nanoparticles: preparation, characterization and in vitro potential against Candida albicans.

    Science.gov (United States)

    Zia, Qamar; Khan, Aijaz Ahmed; Swaleha, Zubair; Owais, Mohammad

    2015-01-01

    In the present study, we developed a self-assembled biodegradable polyglutamic acid (PGA)-based formulation of amphotericin B (AmB) and evaluated its in vitro antifungal potential against Candida albicans. The AmB-loaded PGA nanoparticles were prepared in-house and had a mean size dimension of around 98±2 nm with a zeta potential of -35.2±7.3 mV. Spectroscopic studies revealed that the drug predominantly acquires an aggregated form inside the formulation with an aggregation ratio above 2. The PGA-based AmB formulation was shown to be highly stable in phosphate-buffered saline as well as in serum (only 10%-20% of the drug was released after 10 days). The AmB-PGA nanoparticles were less toxic to red blood cells (formulation showed potent antimicrobial activity similar to that of Fungizone against C. albicans. Interestingly, AmB-bearing PGA nanoparticles were found to inhibit biofilm formation to a considerable extent. In summary, AmB-PGA nanoparticles showed highly attenuated toxicity when compared with Fungizone, while retaining equivalent active antifungal properties. This study indicates that the AmB-PGA preparation could be a promising treatment for various fungal infections.

  17. Analysis of the Cariogenic Potential of Various Almond Milk Beverages using a Streptococcus mutans Biofilm Model in vitro.

    Science.gov (United States)

    Lee, Janelle; Townsend, Janice A; Thompson, Tatyana; Garitty, Thomas; De, Arpan; Yu, Qingzhao; Peters, Brian M; Wen, Zezhang T

    2017-12-15

    To evaluate the cariogenic properties of almond milk beverages, 6 almond milks, along with soy and whole bovine milk, were analyzed for their abilities to support Streptococcus mutans biofilm formation and acid production, and their capacity to buffer changes in pH. Biofilm formation by S. mutans was analyzed using an in vitro 96-well plate model and measured by crystal violet staining. Acid production by S. mutans was evaluated by a colorimetric L-lactate assay and pH measurement of bacterial cultures. Buffering capacity was assessed by a pH titration assay. Soy milk supported the most biofilm growth, while the least was observed with unsweetened almond milk (both p almond milk yielded the lowest pH (4.56 ± 0.66), followed by soy milk and bovine milk; the highest pH was with unsweetened almond milk (6.48 ± 0.5). When analyzed by pH titration, the unsweetened almond milk displayed the weakest buffering capacity while bovine milk showed the highest (p almond milk beverages, except those that are sweetened with sucrose, possess limited cariogenic properties, while soy milk exhibits the most cariogenic potential. As milk alternatives become increasingly popular, dentists must counsel their patients that almond milks, especially sucrose-sweetened varieties, have cariogenic potential. For patients who are lactose-intolerant or suffer from milk allergy, almond milks may be a better alternative than soy-based products. © 2017 S. Karger AG, Basel.

  18. VNTR diversity in Yersinia pestis isolates from an animal challenge study reveals the potential for in vitro mutations during laboratory cultivation

    Science.gov (United States)

    Vogler, Amy J.; Nottingham, Roxanne; Busch, Joseph D.; Sahl, Jason W.; Shuey, Megan M.; Foster, Jeffrey T.; Schupp, James M.; Smith, Susan; Rocke, Tonie E.; Klein, Paul; Wagner, David M.

    2016-01-01

    Underlying mutation rates and other evolutionary forces shape the population structure of bacteria in nature. Although easily overlooked, similar forces are at work in the laboratory and may influence observed mutations. Here, we investigated tissue samples and Yersinia pestis isolates from a rodent laboratory challenge with strain CO92 using whole genome sequencing and multi-locus variable-number tandem repeat (VNTR) analysis (MLVA). We identified six VNTR mutations that were found to have occurred in vitro during laboratory cultivation rather than in vivo during the rodent challenge. In contrast, no single nucleotide polymorphism (SNP) mutations were observed, either in vivo or in vitro. These results were consistent with previously published mutation rates and the calculated number of Y. pestis generations that occurred during the in vitro versus the in vivo portions of the experiment. When genotyping disease outbreaks, the potential for in vitro mutations should be considered, particularly when highly variable genetic markers such as VNTRs are used.

  19. Assessment of antimicrobial potential of 10% ginger extract against Streptococcus mutans, Candida albicans, and Enterococcus faecalis: an in vitro study.

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    Giriraju, Anjan; Yunus, G Y

    2013-01-01

    Streptococcus mutans, Candida albicans, and Enterococcus faecalis are the three oral microorganisms most commonly implicated in the causation of oral infections. All these oral microorganisms have shown resistant to routinely used antimicrobials. There is a need for an antimicrobial agent which is effective, safe, and economical. Zingiber officinale, commonly known as ginger is one such plant product which has been used from ancient time. It has been shown to possess promising inhibitory effect on many of the oral microorganisms. On review of dental literature, there was scarcity of studies which had tried to assess antimicrobial potential of ginger extract against S. mutans, E. faecalis, and C. albicans; hence, the present study was designed. To evaluate the in vitro antimicrobial potential of 10% ginger extract against S. mutans, E. faecalis, and C. albicans. Laboratory setting and experimental design. In the first part of the study, 10% ethanolic ginger extract was prepared in the laboratory of Pharmacy College. It was then subjected to microbiological assay to determine its zone of inhibition using Agar disk diffusion test and minimum inhibitory concentration (MIC) using serial broth dilution method against S. mutans, C. albicans, and E. faecalis. 10% ethanolic ginger extract showed: (a) Maximum zone of inhibition of 8 mm, 14 mm, and 11 mm against S. mutans, C. albicans, and E. faecalis respectively. (b) MIC of 1.25%, 2.5%, and 2.5% against S. mutans, C. albicans, and E. faecalis respectively. 10% ethanolic ginger extract was found to possess antimicrobial potential against all the three pathogens used in the study.

  20. The effect of aluminium and sodium impurities on the in vitro toxicity and pro-inflammatory potential of cristobalite

    Science.gov (United States)

    Nattrass, C.; Horwell, Claire J.; Damby, David; Brown, David; Stone, Vicki

    2017-01-01

    BackgroundExposure to crystalline silica (SiO2), in the form of quartz, tridymite or cristobalite, can cause respiratory diseases, such as silicosis. However, the observed toxicity and pathogenicity of crystalline silica is highly variable. This has been attributed to a number of inherent and external factors, including the presence of impurities. In cristobalite-rich dusts, substitutions of aluminium (Al) for silicon (Si) in the cristobalite structure, and impurities occluding the silica surface, have been hypothesised to decrease its toxicity. This hypothesis is tested here through the characterisation and in vitro toxicological study of synthesised cristobalite with incremental amounts of Al and sodium (Na) dopants. MethodsSamples of synthetic cristobalite with incremental amounts of Al and Na impurities, and tridymite, were produced through heating of a silica sol-gel. Samples were characterised for mineralogy, cristobalite purity and abundance, particle size, surface area and surface charge. In vitro assays assessed the ability of the samples to induce cytotoxicity and TNF-α production in J774 macrophages, and haemolysis of red blood cells. ResultsAl-only doped or Al+Na co-doped cristobalite contained between 1 and 4 oxide wt% Al and Na within its structure. Co-doped samples also contained Al- and Na-rich phases, such as albite. Doping reduced cytotoxicity to J774 macrophages and haemolytic capacity compared to non-doped samples. Al-only doping was more effective at decreasing cristobalite reactivity than Al+Na co-doping. The reduction in the reactivity of cristobalite is attributed to both structural impurities and a lower abundance of crystalline silica in doped samples. Neither non-doped nor doped crystalline silica induced production of the pro-inflammatory cytokine TNF-α in J774 macrophages. ConclusionsImpurities can reduce the toxic potential of cristobalite and may help explain the low reactivity of some cristobalite-rich dusts. Whilst further work

  1. Studies of the in vitro anticancer, antimicrobial and antioxidant potentials of selected Yemeni medicinal plants from the island Soqotra

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    Bednarski Patrick J

    2009-03-01

    Full Text Available Abstract Background Recent years have witnessed that there is a revival of interest in drug discovery from medicinal plants for the maintenance of health in all parts of the world. The aim of this work was to investigate 26 plants belonging to 17 families collected from a unique place in Yemen (Soqotra Island for their in vitro anticancer, antimicrobial and antioxidant activities. Methods The 26 plants were extracted with methanol and hot water to yield 52 extracts. Evaluation for in vitro anticancer activity was done against three human cancer cell lines (A-427, 5637 and MCF-7 by using an established microtiter plate assay based on cellular staining with crystal violet. Antimicrobial activity was tested against three Gram-positive bacteria, two Gram-negative bacteria, one yeast species and three multiresistant Staphylococcus strains by using an agar diffusion method and the determination of MIC against three Gram-positive bacteria with the broth micro-dilution assay. Antioxidant activity was investigated by measuring the scavenging activity of the DPPH radical. Moreover, a phytochemical screening of the methanolic extracts was done. Results Notable cancer cell growth inhibition was observed for extracts from Ballochia atro-virgata, Eureiandra balfourii and Hypoestes pubescens, with IC50 values ranging between 0.8 and 8.2 μg/ml. The methanol extracts of Acanthospermum hispidum, Boswellia dioscorides, Boswellia socotrana, Commiphora ornifolia and Euphorbia socotrana also showed noticeable antiproliferative potency with IC50 values Acacia pennivenia, Boswellia dioscorides, Boswellia socotrana, Commiphora ornifolia, Euclea divinorum, Euphorbia socotrana, Leucas samhaensis, Leucas virgata, Rhus thyrsiflora, and Teucrium sokotranum with inhibition zones > 15 mm and MIC values ≤ 250 μg/ml. In addition, the methanolic extracts of Acacia pennivenia, Boswellia dioscorides, Boswellia socotrana and Commiphora ornifolia showed good antioxidant potential

  2. The effect of aluminium and sodium impurities on the in vitro toxicity and pro-inflammatory potential of cristobalite.

    Science.gov (United States)

    Nattrass, C; Horwell, C J; Damby, D E; Brown, D; Stone, V

    2017-11-01

    Exposure to crystalline silica (SiO2), in the form of quartz, tridymite or cristobalite, can cause respiratory diseases, such as silicosis. However, the observed toxicity and pathogenicity of crystalline silica is highly variable. This has been attributed to a number of inherent and external factors, including the presence of impurities. In cristobalite-rich dusts, substitutions of aluminium (Al) for silicon (Si) in the cristobalite structure, and impurities occluding the silica surface, have been hypothesised to decrease its toxicity. This hypothesis is tested here through the characterisation and in vitro toxicological study of synthesised cristobalite with incremental amounts of Al and sodium (Na) dopants. Samples of synthetic cristobalite with incremental amounts of Al and Na impurities, and tridymite, were produced through heating of a silica sol-gel. Samples were characterised for mineralogy, cristobalite purity and abundance, particle size, surface area and surface charge. In vitro assays assessed the ability of the samples to induce cytotoxicity and TNF-α production in J774 macrophages, and haemolysis of red blood cells. Al-only doped or Al+Na co-doped cristobalite contained between 1 and 4 oxide wt% Al and Na within its structure. Co-doped samples also contained Al- and Na-rich phases, such as albite. Doping reduced cytotoxicity to J774 macrophages and haemolytic capacity compared to non-doped samples. Al-only doping was more effective at decreasing cristobalite reactivity than Al+Na co-doping. The reduction in the reactivity of cristobalite is attributed to both structural impurities and a lower abundance of crystalline silica in doped samples. Neither non-doped nor doped crystalline silica induced production of the pro-inflammatory cytokine TNF-α in J774 macrophages. Impurities can reduce the toxic potential of cristobalite and may help explain the low reactivity of some cristobalite-rich dusts. Whilst further work is required to determine if these

  3. Phytochemical Compositions and In vitro Assessments of Antioxidant and Antidiabetic Potentials of Fractions from Ehretia cymosa Thonn.

    Science.gov (United States)

    Ogundajo, Akintayo; Ashafa, Anofi Tom

    2017-10-01

    Ehretia cymosa Thonn. is a popular medicinal plant used in different parts of West Africa for the treatment of various ailments including diabetes mellitus. The current study investigates bioactive constituents and in vitro antioxidant and antidiabetic potentials of fractions from extract of E. cymosa. Phytochemical investigation and antioxidant assays were carried out using standard procedures. Antidiabetic potential was assessed by evaluating the inhibitory effects of the fractions on the activities of α-amylase and α-glucosidase, while bioactive constituent's identification was carried out using gas chromatography-mass spectrometric (GC-MS) analysis. The phytochemistry tests of the fractions revealed the presence of tannins, phenols, flavonoids, steroids, terpene, alkaloid, and cardiac glycosides. Methanol fraction shows higher phenolic (27.44 mg gallic acid/g) and flavonoid (235.31 mg quercetin/g) contents, while ethyl acetate fraction revealed higher proanthocyanidins (28.31 mg catechin/g). Methanol fraction displayed higher (P fractions displayed higher inhibition (P fraction also inhibited α-amylase and α-glucosidase in competitive and noncompetitive modes, respectively. The GC-MS chromatogram of the methanol fraction revealed 24 compounds, which include phytol (1.78%), stearic acid (1.02%), and 2-hexadecyloxirane (34.18%), which are known antidiabetic and antioxidant agents. The results indicate E. cymosa leaves as source of active phytochemicals with therapeutic potentials in the management of diabetes. E. cymosa fractions possess antioxidant and antidiabetic activities. Hence, it is a source of active phytochemicals with therapeutic potentials in the management of diabetesThe high flavonoid, phenolic, and proanthocyanidin contents of fractions from E. cymosa also contribute to its antioxidant and antidiabetic propertiesMethanol fraction of E. cymosa displayed better antidiabetic activities compared to acarbose as revealed by their half maximal

  4. In Vitro characterization of Lactococcus lactis strains Isolated from Iranian Traditional Dairy Products as a Potential Probiotic

    Directory of Open Access Journals (Sweden)

    Fatemeh Nejati

    2015-12-01

    Full Text Available Few studies have been reported regarding probiotic properties of Lactococcus lactis strains although they are extensively used as starter cultures in the production of dairy products. In this study 8 wild isolates of Lactococcus lactis were evaluated in vitro with regard to resistance to simulated gastric and intestinal juices, adherence ability to Caco-2 cells and HT29-MTX-E12 cell lines, anti-microbial activity, hydrophobicity and antibiotic susceptibility. The results revealed that all isolates had better survival after exposure to simulated gastrointestinal tract stresses in comparison to control probiotic Lactobacillus rhamnosus GG. Regarding adherence efficiency, almost all isolates exhibited similar adherence with control. Three isolates showed antibacterial activity against Gram-positive pathogens (Staphylococcus aureus and Listeria monocytogenes through spot-agar method. Almost all isolates (seven out of eight showed similar hydrophobicity to control probiotic. Regarding to antibiotic resistance, all isolates were susceptible to gentamicin, ampicillin, ciprofloxacin, erythromycin, tetracycline, penicillin, kanamycin and nitrofurantoin. Although, further investigations are necessary, it was concluded that strains derived from raw milk and home-made dairy products could be a remarkable reservoir for identification of new potential probiotic strains.

  5. In vitro study on the safety of near infrared laser therapy in its potential application as postmastectomy lymphedema treatment.

    Science.gov (United States)

    Cialdai, Francesca; Landini, Ida; Capaccioli, Sergio; Nobili, Stefania; Mini, Enrico; Lulli, Matteo; Monici, Monica

    2015-10-01

    Clinical studies demonstrated the effectiveness of laser therapy in the management of postmastectomy lymphedema, a discomforting disease that can arise after surgery/radiotherapy and gets progressively worse and chronic. However, safety issues restrict the possibility to treat cancer patients with laser therapy, since the effects of laser radiation on cancer cell behavior are not completely known and the possibility of activating postmastectomy residual cancer cells must be considered. This paper reports the results of an in vitro study aimed to investigate the effect of a class IV, dual-wavelength (808 nm and 905 nm), NIR laser system on the behavior of two human breast adenocarcinoma cell lines (namely, MCF7 and MDA-MB361 cell lines), using human dermal fibroblasts as normal control. Cell viability, proliferation, apoptosis, cell cycle and ability to form colonies were analyzed in order to perform a cell-based safety testing of the laser treatment in view of its potential application in the management of postmastectomy lymphedema. The results showed that, limited to the laser source, treatment conditions and experimental models used, laser radiation did not significantly affect the behavior of human breast adenocarcinoma cells, including their clonogenic efficiency. Although these results do not show any significant laser-induced modification of cancer cell behavior, further studies are needed to assess the possibility of safely applying NIR laser therapy for the management of postmastectomy lymphedema. Copyright © 2015 Elsevier B.V. All rights reserved.

  6. Development of an in vitro tumor spheroid culture model amenable to high-throughput testing of potential anticancer nanotherapeutics.

    Science.gov (United States)

    Solomon, Melani A; Lemera, Jenkins; D'Souza, Gerard G M

    2016-09-01

    Three-dimensional tumor spheroid cultures are a better representative of in vivo solid tumors than monolayer cultures and should be used for testing potential nanotherapeutics in vitro. To develop techniques to test the disposition and efficacy of nanocarrier formulations in spheroids in a cost-effective manner amenable to high-throughput testing. Spheroids were obtained using a modified liquid overlay technique in a 96-well plate. Several nanocarrier formulations were prepared and tested in the spheroid model. The disposition of the formulations in the spheroids was determined by confocal microscopy while the effect of the drug-loaded formulations was assessed in terms of the cell viability, loss of membrane integrity, induction of caspases and inhibition of growth of the spheroids. The surface charge of the formulations influenced the accumulation of the nanocarrier and drug in the spheroid, with the cationic formulation accumulating to the greatest extent. Also, the smallest particle size formulation, micelles, penetrated to the greatest extent in the spheroid. The iRGD tumor-penetrating peptide co-administered with unmodified liposomes exhibited both high accumulation and penetration. The effect studies revealed that the formulations that penetrated or accumulated to the highest extent in the spheroid exhibited better antitumor activity compared to the other formulations. The 96-well plate format spheroid model developed in the study can be used toward the rational selection of nanocarrier therapeutics prior to their testing in in vivo models.

  7. Design, synthesis and in vitro cytotoxicity study of benzodiazepine-mustard conjugates as potential brain anticancer agents

    Directory of Open Access Journals (Sweden)

    Rajesh K. Singh

    2017-01-01

    Full Text Available The combination of two pharmacological entities in a single compound has been utilized as a promising drug design strategy for site-specificity. So two nitrogen mustard agents were synthesized by conjugating mustard with the benzodiazepine nucleus in the hope to obtain central nervous system (CNS antitumor agents. The benzodiazepine part is aimed to serve as a CNS active carrier enabling the alkylating moiety to cross the BBB by altering its physicochemical properties. Structures of all the synthesized compounds were confirmed by IR, NMR (1H & 13C, mass spectral and elemental studies. The benzodiazepine-mustard conjugates are oily at room temperature and quite stable when stored in refrigerator for 2 months. Both compounds when evaluated for NBP alkylating activity against chlorambucil, proved to be active alkylating agents. The compounds were markedly active when subjected to in vitro biological evaluation using an MTT colorimetric assay against four human cancer cell lines (A-549, COLO 205, U-87 MG and IMR-32. The physicochemical ADME studies were also analyzed using Qikprop 2.5 tools of Schodinger software which further indicates that both compounds can be potential candidates for the treatment of brain tumor.

  8. In Vitro Potential of Equine DEFA1 and eCATH1 as Alternative Antimicrobial Drugs in Rhodococcosis Treatment

    Science.gov (United States)

    Jung, Sascha; Bruhn, Oliver; Goux, Didier; Leippe, Matthias; Leclercq, Roland; Laugier, Claire; Grötzinger, Joachim; Cauchard, Julien

    2012-01-01

    Rhodococcus equi, the causal agent of rhodococcosis, is a severe pathogen of foals but also of immunodeficient humans, causing bronchopneumonia. The pathogen is often found together with Klebsiella pneumoniae or Streptococcus zooepidemicus in foals. Of great concern is the fact that some R. equi strains are already resistant to commonly used antibiotics. In the present study, we evaluated the in vitro potential of two equine antimicrobial peptides (AMPs), eCATH1 and DEFA1, as new drugs against R. equi and its associated pathogens. The peptides led to growth inhibition and death of R. equi and S. zooepidemicus at low micromolar concentrations. Moreover, eCATH1 was able to inhibit growth of K. pneumoniae. Both peptides caused rapid disruption of the R. equi membrane, leading to cell lysis. Interestingly, eCATH1 had a synergic effect together with rifampin. Furthermore, eCATH1 was not cytotoxic against mammalian cells at bacteriolytic concentrations and maintained its high killing activity even at physiological salt concentrations. Our data suggest that equine AMPs, especially eCATH1, may be promising candidates for alternative drugs to control R. equi in mono- and coinfections. PMID:22232283

  9. Synthesis and In Vitro Characterization of Fe3+-Doped Layered Double Hydroxide Nanorings as a Potential Imageable Drug Delivery System

    Directory of Open Access Journals (Sweden)

    Lijun Wang

    2017-09-01

    Full Text Available Highly dispersed Fe3+-doped layered double hydroxide (LDH-Fe nanorings were obtained by a simple coprecipitation-acid etching approach. The morphology, structure, magnetic resonance imaging (MRI performance in vitro, drug loading and releasing, Fe3+ leakage, and cytotoxicity of the as-prepared LDH-Fe nanorings were characterized. The LDH-Fe nanorings showed good water dispersity and a well-crystallized structure. The DLS average size of nanoparticles was measured to be 94.5 nm. Moreover, the MRI tests showed a favourable T1-weighted MRI performance of the LDH-Fe nanoring with r1 values of 0.54 and 1.68, and low r2/r1 ratios of 10.1 and 6.3, pre- and after calcination, respectively. The nanoparticles also showed high model drug (ibuprofen loading capacities, low Fe3+ leakage, and negligible cytotoxicity. All these results demonstrate the potential of LDH-Fe nanorings as an imageable drug delivery system.

  10. Novel chenodeoxycholic acid-sodium alginate matrix in the microencapsulation of the potential antidiabetic drug, probucol. An in vitro study.

    Science.gov (United States)

    Mooranian, Armin; Negrulj, Rebecca; Mikov, Momir; Golocorbin-Kon, Svetlana; Arfuso, Frank; Al-Salami, Hani

    2015-01-01

    We previously designed, developed and characterized a novel microencapsulated formulation as a platform for the targeted delivery of Probucol (PB) in an animal model of Type 2 Diabetes. The objective of this study is to optimize this platform by incorporating Chenodeoxycholic acid (CDCA), a bile acid with good permeation-enhancing properties, and examine its effect in vitro. Using sodium alginate (SA), we prepared PB-SA (control) and PB-CDCA-SA (test) microcapsules. CDCA resulted in better structural and surface characteristics, uniform morphology, and stable chemical and thermal profiles, while size and rheological parameters remained unchanged. PB-CDCA-SA microcapsules showed good excipients' compatibilities, as evidenced by differential scanning calorimetry (DSC), Fourier transform infrared spectroscopy, scanning electron microscopy and energy dispersive X-ray spectroscopy studies. CDCA reduced microcapsule swelling at pH 7.8 at both 37 °C and 25 °C and improved PB-release. CDCA improved the characteristics and release properties of PB-microcapsules and may have potential in the targeted oral delivery of PB.

  11. Magnesium stearate, a widely-used food additive, exhibits a lack of in vitro and in vivo genotoxic potential.

    Science.gov (United States)

    Hobbs, Cheryl A; Saigo, Kazuhiko; Koyanagi, Mihoko; Hayashi, Shim-Mo

    2017-01-01

    Magnesium stearate is widely used in the production of dietary supplement and pharmaceutical tablets, capsules and powders as well as many food products, including a variety of confectionery, spices and baking ingredients. Although considered to have a safe toxicity profile, there is no available information regarding its potential to induce genetic toxicity. To aid safety assessment efforts, magnesium sulfate was evaluated in a battery of tests including a bacterial reverse mutation assay, an in vitro chromosome aberration assay, and an in vivo erythrocyte micronucleus assay. Magnesium stearate did not produce a positive response in any of the five bacterial strains tested, in the absence or presence of metabolic activation. Similarly, exposure to magnesium stearate did not lead to chromosomal aberrations in CHL/IU Chinese hamster lung fibroblasts, with or without metabolic activation, or induce micronuclei in the bone marrow of male CD-1 mice. These studies have been used by the Japanese government and the Joint FAO/WHO Expert Committee on Food Additives in their respective safety assessments of magnesium stearate. These data indicate a lack of genotoxic risk posed by magnesium stearate consumed at current estimated dietary exposures. However, health effects of cumulative exposure to magnesium via multiple sources present in food additives may be of concern and warrant further evaluation.

  12. In vitro Antimicrobial Assay of Actinomycetes in Rice AgainstXanthomonas oryzae pv. oryzicola and as Potential Plant Growth Promoter

    Directory of Open Access Journals (Sweden)

    Erneeza Mohd Hata

    2015-12-01

    Full Text Available ABSTRACT The aim of this work was to invitro assay the antimicrobial activity of actinomycetes in rice against Xanthomonas oryzae pv. oryzicola and as potential plant growth promoter. A total of 92 actinomycete strains were isolated from different rice plant components and field locations. Of these, only 21.74% showed antagonistic activity against the Xoc pathogen. Molecular identification via 16s rRNA amplification revealed that 60% of the active antagonistic strains belonged to the genus Streptomyces. Isolates that demonstrated the highest antagonistic activity were also able to produce hydrolytic enzymes and plant growth-promoting hormones. Combination of preliminary screening based on in vitro antagonistic, hydrolytic enzyme and plant growth hormone activity facilitated the best selection of actinomycete candidates as evidenced by strains classification using cluster analysis (Ward's Method. Results from the preliminary screening showed that actinomycetes, especially Streptomycetes, could offer a promising source for both biocontrol and plant growth-promotion agents against BLS disease in rice.

  13. 77 FR 41406 - Evaluation of In Vitro Tests for Identifying Eye Injury Hazard Potential of Chemicals and...

    Science.gov (United States)

    2012-07-13

    ... assess the validation status of in vitro tests and integrated non-animal testing strategies proposed for... vitro alternatives to animals for eye safety assessments is an ICCVAM priority (ICCVAM, 2008). See http... test and integrated non-animal ] testing strategies proposed for identifying eye injury hazard...

  14. Evaluation of in vitro antioxidant potential anti-inflammatory activity and melanogenesis inhibition of Artocarpus hirsutus Lam. extracts

    Directory of Open Access Journals (Sweden)

    Mahadeva Nayak

    2015-08-01

    Full Text Available Artocarpus hirsutus Lam. belongs to Moraceae family and is endemic to Western Ghats and Kerala in India. This species is found to be effective in traditional medicine for the treatment of ulcer diarrhea and pimples. However extensive biological evaluation on each component of this specific species rarely appears in the literature which restricts its applicability as medicinal herb. The leaf bark and wood of Artocarpus hirsutus Lam. were separately extracted with hot ethanol. The wood extract was further fractionated to isolate major active molecule whose structure was determined from its NMR spectra and LCMS analysis. All the extracts of A. hirsutus Lam. were then studied in vitro to evaluate their potential on tyrosinase inhibition free radical scavenging activity by 11-Diphenyl-2-picrylhydrazyl DPPH method and oxygen radical absorbance capacity ORAC. Furthermore their effects on melanogenesis inhibition were also evaluated by using murine melanoma cells. Activity guided fractionation of wood extract yielded a pure molecule that was characterized as oxyresveratrol. It was observed that antioxidant activity was higher in wood extract compared to the leaf and bark extracts. Isolated pure oxyresveratrol exhibited a significant antioxidant potential with ORAC value of 366532570 mol Trolox equivalentg and having an IC50 of 4.3 gmL for DPPH free radical scavenging activity. This molecule was found to be effective for the tyrosinase inhibition with an IC50 of 0.1 gmL and melanogenesis inhibition in cultured melanoma cells by 44.62 at 0.2 gmL. Oxyresveratrol also exhibited significant inhibition of lipopolysaccharide LPS induced tumour necrosis factor alpha TNF-amp945 secretion from J774A1 murine macrophage cell lines. This study provides substantial evidence for the presence of oxyresveratrol in the wood of A. hirsutus Lam. with promising anti-inflammatory antioxidant and skin lightening property.

  15. Characterization of Lactobacillus plantarum as a Potential Probiotic In vitro and Use of a Dairy Product (Yogurt as Food Carrier

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    Jawad Kadhim Isa

    2017-01-01

    Full Text Available Background and Objective: The current study was undertaken to check in vitro different characteristics of Lactobacillus plantarum as potential probiotic. These characteristics include viability of probiotic and pH during cold storage, tolerance to acid and bile, and antibiotic resistance.Material and Methods: Samples of yogurt were stored at 4°C and analyzed in time 0, 1, 3, 7, 10 and 14 days of storage. In these periods, probiotic and starter cultures were enumerated and the pH parameter was analyzed.Results and Conclusion: A gradual decline in pH was noticed throughout the storage. Counting of starter cultures decreased by 0.42 log cycle, and the probiotic's viability decreased by 0.68 log cycle at the end of storage, whereas the probiotic's viability inthe samples subjected to re-pasteurization decreased by 0.30, 0.22 log cycles in the selective and reference media, respectively. The average viable cell counts of Lactobacillus plantarum decreased by 0.76, and 0.28 log cycles after incubation period (3 h at 37ºC in the simulated gastric juice (pH 2.0 and 3.0, respectively. Generally, probiotic can maintain its viability by 76.672% in (1.0% w v -1 bile. Lactobacillus plantarum was resistant to gentamicin, streptomycin, and vancomycin but susceptible to chloramphenicol, and tetracycline. Depended on these characteristics, Lactobacillus plantarum showed probiotic potential.Conflict of interest: The authors declare no conflict of interest.

  16. In Vitro Assessment of the Probiotic Potential of Lactococcus lactis LMG 7930 against Ruminant Mastitis-Causing Pathogens.

    Directory of Open Access Journals (Sweden)

    Federica Armas

    Full Text Available Mastitis in dairy ruminants is considered to be the most expensive disease to farmers worldwide. Recently, the intramammary infusion of lactic acid bacteria has emerged as a potential new alternative to antibiotics for preventing and treating bovine mastitis. In this study we have investigated in vitro the probiotic potential of Lactococcus lactis LMG 7930, a food-grade and nisin-producing strain, against mastitis-causing pathogens. We have characterized its carbohydrate fermentation and antibiotic susceptibility profiles, cell surface properties and antimicrobial activity, as well as its capabilities to adhere to and inhibit the invasion of pathogens into the bovine mammary epithelial cell line BME-UV1d. We found that L. lactis LMG 7930 was sensitive to tested drugs, according to the EFSA Panel on Additives and Products or Substances used in Animal Feed (FEEDAP, and showed an improved carbohydrate fermentation capacity compared to starter strains. Moreover, the strain exhibited antagonistic properties towards many of the pathogens tested. It presented medium surface hydrophobicity, a low basic property and no electron acceptor capability. It showed low auto-aggregation and no co-aggregation abilities towards any of the tested pathogens. The strain was one of the most adhesive to bovine mammary epithelial cells among tested bacteria, but its internalisation was low. The strain did not affect significantly pathogen invasion; however, a trend to decrease internalization of some pathogens tested was observed. In conclusion, our results suggest that this strain might be a promising candidate for the development of new strategies of mastitis control in ruminants. Future investigations are needed to evaluate its safety and efficacy under field conditions.

  17. Dithranol-loaded lipid-core nanocapsules improve the photostability and reduce the in vitro irritation potential of this drug

    Energy Technology Data Exchange (ETDEWEB)

    Savian, Ana L. [Programa de Pós-Graduação em Ciências Farmacêuticas, Universidade Federal de Santa Maria, Av. Roraima, 1000, Santa Maria, RS 97105-900 (Brazil); Rodrigues, Daiane [Curso de Farmácia, Centro de Ciências da Saúde, Universidade Federal de Santa Maria, Av. Roraima, 1000, Santa Maria, RS 97105-900 (Brazil); Weber, Julia; Ribeiro, Roseane F. [Programa de Pós-Graduação em Ciências Farmacêuticas, Universidade Federal de Santa Maria, Av. Roraima, 1000, Santa Maria, RS 97105-900 (Brazil); Motta, Mariana H. [Curso de Farmácia, Centro de Ciências da Saúde, Universidade Federal de Santa Maria, Av. Roraima, 1000, Santa Maria, RS 97105-900 (Brazil); Schaffazick, Scheila R.; Adams, Andréa I.H. [Programa de Pós-Graduação em Ciências Farmacêuticas, Universidade Federal de Santa Maria, Av. Roraima, 1000, Santa Maria, RS 97105-900 (Brazil); Andrade, Diego F. de; Beck, Ruy C.R. [Programa de Pós-Graduação em Ciências Farmacêuticas, Universidade Federal do Rio Grande do Sul, Av. Ipiranga, 2752, Porto Alegre, RS 90610-000 (Brazil); and others

    2015-01-01

    Dithranol is a very effective drug for the topical treatment of psoriasis. However, it has some adverse effects such as irritation and stain in the skin that make its application and patient adherence to treatment difficult. The aims of this work were to prepare and characterize dithranol-loaded nanocapsules as well as to evaluate the photostability and the irritation potential of these nanocarriers. Lipid-core nanocapsules containing dithranol (0.5 mg/mL) were prepared by interfacial deposition of preformed polymer. EDTA (0.05%) or ascorbic acid (0.02%) was used as antioxidants. After preparation, dithranol-loaded lipid-core nanocapsules showed satisfactory characteristics: drug content close to the theoretical concentration, encapsulation efficiency of about 100%, nanometric mean size (230–250 nm), polydispersity index below 0.25, negative zeta potential, and pH values from 4.3 to 5.6. In the photodegradation study against UVA light, we observed a higher stability of the dithranol-loaded lipid-core nanocapsules comparing to the solution containing the free drug (half-life times around 4 and 1 h for the dithranol-loaded lipid-core nanocapsules and free drug solution containing EDTA, respectively; half-life times around 17 and 7 h for the dithranol-loaded lipid-core nanocapsules and free drug solution containing ascorbic acid, respectively). Irritation test by HET-CAM method was conducted to evaluate the safety of the formulations. From the results it was found that the nanoencapsulation of the drug decreased its toxicity compared to the effects observed for the free drug. - Highlights: • Strategy to prepare lipid-core nanocapsules containing dithranol • Evaluation of the nanoencapsulation effect on the photostability and irritation • Evaluation of the in vitro release of dithranol-loaded lipid-core nanocapsules.

  18. Intralaboratory validation of four in vitro assays for the prediction of the skin sensitizing potential of chemicals.

    Science.gov (United States)

    Bauch, Caroline; Kolle, Susanne N; Fabian, Eric; Pachel, Christina; Ramirez, Tzutzuy; Wiench, Benjamin; Wruck, Christoph J; van Ravenzwaay, Bennard; Landsiedel, Robert

    2011-09-01

    Allergic contact dermatitis is induced by repeated skin contact with an allergen. Assessment of the skin sensitizing potential of chemicals, agrochemicals, and especially cosmetic ingredients is currently performed with the use of animals. Animal welfare and EU legislation demand animal-free alternatives reflected in a testing and marketing ban for cosmetic ingredients beginning in 2013. The underlying mechanisms of induction and elicitation of skin sensitization are complex and a chemical needs to comply several properties being skin sensitizing. To account for the multitude of events in the induction of skin sensitization an in vitro test system will consist of a battery of various tests. Currently, we performed intralaboratory validations of four assays addressing three different events during induction of skin sensitization. (1) The Direct Peptide Reactivity Assay (DPRA) according to Gerberick and co-workers (Gerberick et al., 2004) using synthetic peptides and HPLC analysis. (2) Two dendritic cell activation assays based on the dendritic cell like cell lines U-937 and THP-1 and flow cytometric detection of the maturation markers CD54 and/or CD86 (Ashikaga et al., 2006; Python et al., 2007; Sakaguchi et al., 2006). (3) Antioxidant response element (ARE)-dependent gene activity in a HaCaT reporter gene cell line (Emter et al., 2010). We present the results of our intralaboratory validation of these assays with 23 substances of known sensitizing potential. The sensitivity, specificity, and accuracy of the individual tests were obtained by comparison to human epidemiological data as well as to data from animal tests such as the local lymph node assay. Copyright © 2011 Elsevier Ltd. All rights reserved.

  19. In Vitro Assessment of the Probiotic Potential of Lactococcus lactis LMG 7930 against Ruminant Mastitis-Causing Pathogens

    Science.gov (United States)

    Armas, Federica; Camperio, Cristina

    2017-01-01

    Mastitis in dairy ruminants is considered to be the most expensive disease to farmers worldwide. Recently, the intramammary infusion of lactic acid bacteria has emerged as a potential new alternative to antibiotics for preventing and treating bovine mastitis. In this study we have investigated in vitro the probiotic potential of Lactococcus lactis LMG 7930, a food-grade and nisin-producing strain, against mastitis-causing pathogens. We have characterized its carbohydrate fermentation and antibiotic susceptibility profiles, cell surface properties and antimicrobial activity, as well as its capabilities to adhere to and inhibit the invasion of pathogens into the bovine mammary epithelial cell line BME-UV1d. We found that L. lactis LMG 7930 was sensitive to tested drugs, according to the EFSA Panel on Additives and Products or Substances used in Animal Feed (FEEDAP), and showed an improved carbohydrate fermentation capacity compared to starter strains. Moreover, the strain exhibited antagonistic properties towards many of the pathogens tested. It presented medium surface hydrophobicity, a low basic property and no electron acceptor capability. It showed low auto-aggregation and no co-aggregation abilities towards any of the tested pathogens. The strain was one of the most adhesive to bovine mammary epithelial cells among tested bacteria, but its internalisation was low. The strain did not affect significantly pathogen invasion; however, a trend to decrease internalization of some pathogens tested was observed. In conclusion, our results suggest that this strain might be a promising candidate for the development of new strategies of mastitis control in ruminants. Future investigations are needed to evaluate its safety and efficacy under field conditions. PMID:28068371

  20. Osteogenic Potential Differentiation of Human Amnion Mesenchymal Stem Cell with Chitosan-Carbonate Apatite Scaffold (In Vitro Study

    Directory of Open Access Journals (Sweden)

    Michael J.K. Kamadjaja

    2016-09-01

    Full Text Available Background: Tissue engineering based approaches have received much attention. Incorporation of chitosan and carbonate apatite (CA improve its capability. Human mesenchymal stem cells (hMSCs is viable for xenogenic transplantation. The purpose of this study was to fabricate and evaluate the osteogenic potential diferentiation of human amnion mesenchymal stem cell with carbonate apatite–chitosan scaffolds (CA-ChSs for tissue engineering. Method: Human amniotic membrane was procured from using cesarean section. Soncini’s protocol was employed for the isolation procedure. The cells cultured on collagen-coated dishes using Dulbecco's minimal essential medium (DMEM/F12 (1:1. A chitosan powder of medium molecular weight deacetylated chitin, poly(D(glucosamine was used and mixed with CA. Immunocytochemistry and flowcytometry used for phenotypic characterization of hAMSC. Result: Amniotic membrane obtained using cesarean section under aseptic condition did not exhibit any growth of cell cultures which were not contaminated. Immunocytochemistry testing revealed that the target cells expressed strong mesenchymal stem cell marker CD 105. Characterization at passage 10 showed that CD44 was the most significant and abundant surface receptors. The number of viable cells in chitosan-carbonate apatite was 66.59%. Scanning electron microscope (SEM observation revealed that CA-ChSs had three-dimensional structure with many pores and hAMSc could attached and proliferation among the porosity of the scaffold. The formation of calcium in the cell as an indicator of osteoblast cells was detected using Alizarin Red solution. Conclusion: hAMSc harvested from human amniotic membrane seeding in CA-ChSs had the capability for in vitro osteogenesis makes them be the one of the potential options for bone tissue engineering.

  1. Potential hazards to embryo implantation: A human endometrial in vitro model to identify unwanted antigestagenic actions of chemicals

    Energy Technology Data Exchange (ETDEWEB)

    Fischer, L.; Deppert, W.R. [Department of Obstetrics and Gynecology, University Hospital Freiburg (Germany); Pfeifer, D. [Department of Hematology and Oncology, University Hospital Freiburg (Germany); Stanzel, S.; Weimer, M. [Department of Biostatistics, German Cancer Research Center, Heidelberg (Germany); Hanjalic-Beck, A.; Stein, A.; Straßer, M.; Zahradnik, H.P. [Department of Obstetrics and Gynecology, University Hospital Freiburg (Germany); Schaefer, W.R., E-mail: wolfgang.schaefer@uniklinik-freiburg.de [Department of Obstetrics and Gynecology, University Hospital Freiburg (Germany)

    2012-05-01

    Embryo implantation is a crucial step in human reproduction and depends on the timely development of a receptive endometrium. The human endometrium is unique among adult tissues due to its dynamic alterations during each menstrual cycle. It hosts the implantation process which is governed by progesterone, whereas 17β-estradiol regulates the preceding proliferation of the endometrium. The receptors for both steroids are targets for drugs and endocrine disrupting chemicals. Chemicals with unwanted antigestagenic actions are potentially hazardous to embryo implantation since many pharmaceutical antiprogestins adversely affect endometrial receptivity. This risk can be addressed by human tissue-specific in vitro assays. As working basis we compiled data on chemicals interacting with the PR. In our experimental work, we developed a flexible in vitro model based on human endometrial Ishikawa cells. Effects of antiprogestin compounds on pre-selected target genes were characterized by sigmoidal concentration–response curves obtained by RT-qPCR. The estrogen sulfotransferase (SULT1E1) was identified as the most responsive target gene by microarray analysis. The agonistic effect of progesterone on SULT1E1 mRNA was concentration-dependently antagonized by RU486 (mifepristone) and ZK137316 and, with lower potency, by 4-nonylphenol, bisphenol A and apigenin. The negative control methyl acetoacetate showed no effect. The effects of progesterone and RU486 were confirmed on the protein level by Western blotting. We demonstrated proof of principle that our Ishikawa model is suitable to study quantitatively effects of antiprogestin-like chemicals on endometrial target genes in comparison to pharmaceutical reference compounds. This test is useful for hazard identification and may contribute to reduce animal studies. -- Highlights: ► We compare progesterone receptor-mediated endometrial effects of chemicals and drugs. ► 4-Nonylphenol, bisphenol A and apigenin exert weak

  2. Contribution of glutaredoxin-1 to S-glutathionylation of endothelial nitric oxide synthase for mesenteric nitric oxide generation in experimental necrotizing enterocolitis.

    Science.gov (United States)

    Shang, Qingjuan; Bao, Lei; Guo, Hongjie; Hao, Fabao; Luo, Qianfu; Chen, Jiaping; Guo, Chunbao

    2017-10-01

    Endothelial nitric oxide synthase (eNOS) is critical for intestinal microcirculatory perfusion and therefore plays a key role in the development of necrotizing enterocolitis (NEC). eNOS-derived nitric oxide (NO) is inhibited by S-glutathionylation of eNOS (eNOS-SSG), which can be reversed by glutaredoxin-1 (Grx1). Therefore, the objective of this study was to investigate the interplay between Grx1 and eNOS in regulating the following inflammation signal during the development of NEC. Primary mouse intestinal microvascular endothelial cells (MIMECs) and peritoneal macrophages were subjected to lipopolysaccharide treatment, and Grx1-/- mice were subjected to an NEC-inducing regimen of formula feeding in combination with hypoxia and hypothermia. The eNOS-SSG level and its activity were assessed using immunoprecipitated assay and NO production evaluation. NO-mediated Toll-like receptor 4 (TLR4) signaling and inflammation injury were further defined. NEC severity was significantly increased in Grx1-/- mice. Grx1-/- mice with NEC showed significantly decreased NO and increased O2•- production with increases in eNOS-SSG. Furthermore, TLR4 signaling, which is required for the development of NEC, was enhanced in the Grx1-deficient mice. These results suggest that eNOS-SSG within the MIMECs inhibited NO production and enhanced TLR4 activity, which were implicated in the pathogenesis of NEC. Grx1 deficiency increases the severity of NEC in association with eNOS-SSG. Copyright © 2016 Elsevier Inc. All rights reserved.

  3. Fermentation properties and potential prebiotic activity of Bimuno? galacto-oligosaccharide (65 % galacto-oligosaccharide content) on in vitro gut microbiota parameters

    OpenAIRE

    Grimaldi, R.; Swann, Jonathan R.; Vulevic, Jelena; Gibson, Glenn R.; Costabile, Adele

    2016-01-01

    Prebiotic oligosaccharides have the ability to generate important changes in the gut microbiota composition that may confer health benefits to the host. Reducing the impurities in prebiotic mixtures could expand their applications in food industries and improve their selectivity and prebiotic effect on the potential beneficial bacteria such as bifidobacteria and lactobacilli. This study aimed to determine the in vitro potential fermentation properties of a 65 % galacto-oligosaccharide (GOS) c...

  4. In Vitro and Clinical Evaluations of the Drug-Drug Interaction Potential of a Metabotropic Glutamate 2/3 Receptor Agonist Prodrug with Intestinal Peptide Transporter 1.

    Science.gov (United States)

    Pak, Y Anne; Long, Amanda J; Annes, William F; Witcher, Jennifer W; Knadler, Mary Pat; Ayan-Oshodi, Mosun A; Mitchell, Malcolm I; Leese, Phillip; Hillgren, Kathleen M

    2017-02-01

    Despite peptide transporter 1 (PEPT1) being responsible for the bioavailability for a variety of drugs, there has been little study of its potential involvement in drug-drug interactions. Pomaglumetad methionil, a metabotropic glutamate 2/3 receptor agonist prodrug, utilizes PEPT1 to enhance absorption and bioavailability. In vitro studies were conducted to guide the decision to conduct a clinical drug interaction study and to inform the clinical study design. In vitro investigations determined the prodrug (LY2140023 monohydrate) is a substrate of PEPT1 with K m value of approximately 30 µM, whereas the active moiety (LY404039) is not a PEPT1 substrate. In addition, among the eight known PEPT1 substrates evaluated in vitro, valacyclovir was the most potent inhibitor (IC 50 = 0.46 mM) of PEPT1-mediated uptake of the prodrug. Therefore, a clinical drug interaction study was conducted to evaluate the potential interaction between the prodrug and valacyclovir in healthy subjects. No effect of coadministration was observed on the pharmacokinetics of the prodrug, valacyclovir, or either of their active moieties. Although in vitro studies showed potential for the prodrug and valacyclovir interaction via PEPT1, an in vivo study showed no interaction between these two drugs. PEPT1 does not appear to easily saturate because of its high capacity and expression in the intestine. Thus, a clinical interaction at PEPT1 is unlikely even with a compound with high affinity for the transporter. Copyright © 2017 by The Author(s).

  5. Identification and nanoentrapment of polyphenolic phytocomplex from Fraxinus angustifolia: in vitro and in vivo wound healing potential.

    Science.gov (United States)

    Moulaoui, Kenza; Caddeo, Carla; Manca, Maria Letizia; Castangia, Ines; Valenti, Donatella; Escribano, Elvira; Atmani, Djebbar; Fadda, Anna Maria; Manconi, Maria

    2015-01-07

    The aim of the present study was to elucidate the polyphenolic composition of Fraxinus angustifolia leaf and bark extracts, and to evaluate their efficacy in wound healing. Quercetin, catechin, rutin and tannic acid were identified as the main components of the extracts. In order to improve their skin bioavailability, the polyphenolic phytocomplexes were incorporated in different nanovesicles, namely ethosomes and phospholipid vesicles containing Transcutol(®) P (Trc) or ethylene glycol (EG). The latter had never been used before as a component of phospholipid vesicles, and it was found to play a key role in improving extract efficacy in wound healing. Results of cryogenic transmission electron microscopy (cryo-TEM), Photon Correlation Spectroscopy (PCS) and Small-Angle X-ray Scattering (SAXS) showed that ethosomes and EG-PEVs were small, monodispersed, unilamellar vesicles, while Trc-PEVs were larger, less homogeneously dispersed and multilamellar, with a large bilayer thickness. Free extracts did not show relevant ability to protect in vitro human keratinocytes from H2O2 damages, while when entrapped in nanovesicles, they significantly inhibited H2O2 stress damages, probably related to a higher level of cell uptake. On the other hand, in vivo results showed that the highest antioxidant and anti-inflammatory effects were provided by the phytocomplexes in EG-PEVs, which favoured wound healing. Moreover, non-entrapped F. angustifolia extracts showed a marginal effect, comparable to that of free quercetin dispersion (control). In conclusion, our results depict that these extracts may find potential applications in biomedicine. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

  6. Triterpenoids from Ocimum labiatum Activates Latent HIV-1 Expression In Vitro: Potential for Use in Adjuvant Therapy

    Directory of Open Access Journals (Sweden)

    Petrina Kapewangolo

    2017-10-01

    Full Text Available Latent HIV reservoirs in infected individuals prevent current treatment from eradicating infection. Treatment strategies against latency involve adjuvants for viral reactivation which exposes viral particles to antiretroviral drugs. In this study, the effect of novel triterpenoids isolated from Ocimum labiatum on HIV-1 expression was measured through HIV-1 p24 antigen capture in the U1 latency model of HIV-1 infection and in peripheral blood mononuclear cells (PBMCs of infected patients on combination antiretroviral therapy (cART. The mechanism of viral reactivation was determined through the compound’s effect on cytokine production, histone deacetylase (HDAC inhibition, and protein kinase C (PKC activation. Cytotoxicity of the triterpenoids was determined using a tetrazolium dye and flow cytometry. The isolated triterpene isomers, 3-hydroxy-4,6a,6b,11,12,14b-hexamethyl-1,2,3,4,6,6a,6b,7,8,8a,9,10,11,12,12a,14,14a,14b-octadecahydropicene-4,8a-dicarboxylic acid (HHODC, significantly (p < 0.05 induced HIV-1 expression in a dose-dependent manner in U1 cells at non-cytotoxic concentrations. HHODC also induced viral expression in PBMCs of HIV-1 infected patients on cART. In addition, the compound up-regulated the production of interleukin (IL-2, IL-6, tumour necrosis factor (TNF-α, and interferon (IFN-γ but had no effect on HDAC and PKC activity, suggesting cytokine upregulation as being involved in latency activation. The observed in vitro reactivation of HIV-1 introduces the adjuvant potential of HHODC for the first time here.

  7. In vitro and in vivo antioxidant potential of milks, yoghurts, fermented milks and cheeses: a narrative review of evidence.

    Science.gov (United States)

    Fardet, Anthony; Rock, Edmond

    2017-10-02

    The antioxidant potential (AP) is an important nutritional property of foods, as increased oxidative stress is involved in most diet-related chronic diseases. In dairy products, the protein fraction contains antioxidant activity, especially casein. Other antioxidants include: antioxidant enzymes; lactoferrin; conjugated linoleic acid; coenzyme Q10; vitamins C, E, A and D3; equol; uric acid; carotenoids; and mineral activators of antioxidant enzymes. The AP of dairy products has been extensively studied in vitro, with few studies in animals and human subjects. Available in vivo studies greatly differ in their design and objectives. Overall, on a 100 g fresh weight-basis, AP of dairy products is close to that of grain-based foods and vegetable or fruit juices. Among dairy products, cheeses present the highest AP due to their higher protein content. AP of milk increases during digestion by up to 2·5 times because of released antioxidant peptides. AP of casein is linked to specific amino acids, whereas β-lactoglobulin thiol groups play a major role in the AP of whey. Thermal treatments such as ultra-high temperature processing have no clear effect on the AP of milk. Raw fat-rich milks have higher AP than less fat-rich milk, because of lipophilic antioxidants. Probiotic yoghurts and fermented milks have higher AP than conventional yoghurt and milk because proteolysis by probiotics releases antioxidant peptides. Among the probiotics, Lactobacillus casei/acidophilus leads to the highest AP. The data are insufficient for cheese, but fermentation-based changes appear to make a positive impact on AP. In conclusion, AP might participate in the reported dairy product-protective effects against some chronic diseases.

  8. Evaluating the inhibitory potential of Withania somnifera on platelet aggregation and inflammation enzymes: An in vitro and in silico study.

    Science.gov (United States)

    M, Madhusudan; Zameer, Farhan; Naidu, Akhilender; M N, Nagendra Prasad; Dhananjaya, Bhadrapura Lakkappa; Hegdekatte, Raghavendra

    2016-09-01

    Context Withania somnifera (L.) Dunal is traditionally used for treating various ailments, but lacks scientific evaluation. Objective This study evaluates Withania somnifera (WS) for its effect on platelet activity and inflammatory enzymes. Materials and methods Aqueous and ethanolic (1:1) leaf extracts were subjected to in vitro indirect haemolytic activity using Naja naja venom, human platelet aggregation was quantified for lipid peroxidation using arachidonic acid (AA) as agonist and 5-lipoxygenase (5-LOX) levels were determined using standard spectrometric assays. Further, molecular docking was performed by the ligand fit method using molegro software package (Molegro ApS, Aarhus, Denmark). Results The study found that aqueous and ethanol extracts have very negligible effect (15%) with an IC50 value of 13.8 mg/mL on PLA2 from Naja naja venom. Further, extracts of WS also had very little effect (18%) with an IC50 value of 16.6 mg/mL on malondialdehyde (MDA) formation. However, a 65% inhibition of 5-LOX with an IC50 value of 0.92 mg/mL was observed in 1:1 ethanol extracts. The same was evident from SAR model with the active ingredient withaferin A binding predominantly on Phe 77, Tyr 98, Arg 99, Asp 164, Leu 168, Ser 382, Arg 395, Tyr 396 and Tyr 614 with an atomic contact energy value of -128.96 compared to standard phenidone (-103.61). Thus, the current study validates the application of WS for inflammatory diseases. Conclusion This study reveals the inhibitory potential of W. somnifera on inflammatory enzymes and platelet aggregation. Thus, WS can serve as a newer, safer and affordable medicine for inflammatory diseases.

  9. Wound-healing potential of human umbilical cord blood-derived mesenchymal stromal cells in vitro--a pilot study.

    Science.gov (United States)

    You, Hi-Jin; Namgoong, Sik; Han, Seung-Kyu; Jeong, Seong-Ho; Dhong, Eun-Sang; Kim, Woo-Kyung

    2015-11-01

    Our previous studies demonstrated that human bone marrow-derived mesenchymal stromal cells have great potential for wound healing. However, it is difficult to clinically utilize cultured stem cells. Recently, human umbilical cord blood-derived mesenchymal stromal cells (hUCB-MSCs) have been commercialized for cartilage repair as a first cell therapy product that uses allogeneic stem cells. Should hUCB-MSCs have a superior effect on wound healing as compared with fibroblasts, which are the main cell source in current cell therapy products for wound healing, they may possibly replace fibroblasts. The purpose of this in vitro study was to compare the wound-healing activity of hUCB-MSCs with that of fibroblasts. This study was particularly designed to compare the effect of hUCB-MSCs on diabetic wound healing with those of allogeneic and autologous fibroblasts. Healthy (n = 5) and diabetic (n = 5) fibroblasts were used as the representatives of allogeneic and autologous fibroblasts for diabetic patients in the control group. Human UCB-MSCs (n = 5) were used in the experimental group. Cell proliferation, collagen synthesis and growth factor (basic fibroblast growth factor, vascular endothelial growth factor and transforming growth factor-β) production were compared among the three cell groups. Human UCB-MSCs produced significantly higher amounts of vascular endothelial growth factor and basic fibroblast growth factor when compared with both fibroblast groups. Human UCB-MSCs were superior to diabetic fibroblasts but not to healthy fibroblasts in collagen synthesis. There were no significant differences in cell proliferation and transforming growth factor-β production. Human UCB-MSCs may have greater capacity for diabetic wound healing than allogeneic or autologous fibroblasts, especially in angiogenesis. Copyright © 2015 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  10. Role of transient receptor potential ankyrin 1 receptors in rodent models of meningeal nociception - Experiments in vitro.

    Science.gov (United States)

    Denner, A C; Vogler, B; Messlinger, K; De Col, R

    2017-05-01

    The TRP channel ankyrin type 1 (TRPA1) is a nonselective cation channel known to be activated by environmental irritants, cold and endogenous mediators of inflammation. Activation of TRPA1 in trigeminal afferents innervating meningeal structures has recently been suggested to be involved in the generation of headaches. Two in vitro models of meningeal nociception were employed using the hemisected rodent head preparation, (1) recording of single meningeal afferents and (2) release of calcitonin gene-related peptide (CGRP) from the cranial dura mater. The role of TRPA1 was examined using the TRPA1 agonists acrolein and mustard oil (MO). BCTC, an inhibitor of TRP vanilloid type 1 receptor channels (TRPV1), and the TRPA1 inhibitor HC030031 as well as mice with genetically deleted TRPA1 and TRPV1 proteins, were used to differentiate between effects. Acrolein did not cause discharge activity in meningeal Aδ- or C-fibres but increased the electrical activation threshold. Acrolein was also effective in releasing CGRP from the dura of TRPV1(-/-) but not of TRPA1(-/-) mice. MO increased the discharge activity of afferent fibres from rat as well as C57 wild-type and TRPA1(-/-) but not TRPV1(-/-) mice. The effect was higher in C57 compared to TRPA1(-/-) mice. Sole TRPA1 receptor channel activation releases CGRP and increases the activation threshold of meningeal afferents but does not generate propagated activity, and so would be capable of causing local effects like vasodilatation but not pain generation. In contrast, combined TRPA1 and TRPV1 activation may be rather pronociceptive supporting headache generation. Sole activation of TRPA1 receptor channels increases the activation threshold but does not cause propagated action potentials in meningeal afferents. TRPA1 agonists cause CGRP release from rodent dura mater. Peripheral TRPA1 receptors may have a pronociceptive function in trigeminal nociception only in combination with TRPV1. © 2016 European Pain Federation

  11. Infection of human endothelium in vitro by cytomegalovirus causes enhanced expression of purinergic receptors : A potential virus escape mechanism?

    NARCIS (Netherlands)

    Zandberg, Mariet; van Son, Willem J.; Harmsen, Martin C.; Bakker, Winston W.

    2007-01-01

    Background. Human cytomegalovirus (CMV) uses different strategies to escape from human host defense reactions. Previously we have observed that infection of endothelial cells with CMV in vitro leads to enhanced activity of endothelial ectonucleotidases. These ectoenzymes are responsible for

  12. Experimental Assessment of Moringa oleifera Leaf and Fruit for Its Antistress, Antioxidant, and Scavenging Potential Using In Vitro and In Vivo Assays

    Directory of Open Access Journals (Sweden)

    Suaib Luqman

    2012-01-01

    Full Text Available We have investigated effect of Moringa oleifera leaf and fruit extracts on markers of oxidative stress, its toxicity evaluation, and correlation with antioxidant properties using in vitro and in vitro assays. The aqueous extract of leaf was able to increase the GSH and reduce MDA level in a concentration-dependent manner. The ethanolic extract of fruit showed highest phenolic content, strong reducing power and free radical scavenging capacity. The antioxidant capacity of ethanolic extract of both fruit and leaf was higher in the in vitro assay compared to aqueous extract which showed higher potential in vivo. Safety evaluation studies showed no toxicity of the extracts up to a dose of 100 mg/kg body weight. Our results support the potent antioxidant activity of aqueous and ethanolic extract of Moringa oleifera which adds one more positive attribute to its known pharmacological importance.

  13. Experimental Assessment of Moringa oleifera Leaf and Fruit for Its Antistress, Antioxidant, and Scavenging Potential Using In Vitro and In Vivo Assays

    Science.gov (United States)

    Luqman, Suaib; Srivastava, Suchita; Kumar, Ritesh; Maurya, Anil Kumar; Chanda, Debabrata

    2012-01-01

    We have investigated effect of Moringa oleifera leaf and fruit extracts on markers of oxidative stress, its toxicity evaluation, and correlation with antioxidant properties using in vitro and in vitro assays. The aqueous extract of leaf was able to increase the GSH and reduce MDA level in a concentration-dependent manner. The ethanolic extract of fruit showed highest phenolic content, strong reducing power and free radical scavenging capacity. The antioxidant capacity of ethanolic extract of both fruit and leaf was higher in the in vitro assay compared to aqueous extract which showed higher potential in vivo. Safety evaluation studies showed no toxicity of the extracts up to a dose of 100 mg/kg body weight. Our results support the potent antioxidant activity of aqueous and ethanolic extract of Moringa oleifera which adds one more positive attribute to its known pharmacological importance. PMID:22216055

  14. Characterization of the In Vitro Kinetic Interaction of Chlorpyrifos-Oxon with Rat Salivary Cholinesterase: A Potential Biomonitoring Matrix

    Energy Technology Data Exchange (ETDEWEB)

    Kousba, Ahmed A.(BATTELLE (PACIFIC NW LAB)); Poet, Torka S.(BATTELLE (PACIFIC NW LAB)); Timchalk, Charles (Pacific Northwest National Laboratory)

    2003-02-12

    Chlorpyrifos (CPF) is a commonly used organophosphate insecticide (OP). The primary mechanism of action for CPF involves the inhibition of acetylcholinesterase (AChE) by the active metabolite, CPF-oxon, with subsequent accumulation of acetylcholine (ACh) resulting in a wide range of neutotoxicity. CPF-oxon, can likewise inhibit other non-target cholinesterases (ChE) such as butyrylcholinesterase (BuChE), which represents a detoxification mechanism and a potential biomarker of exposure/response. Biological monitoring for OPs has focused on measuring parent chemical or metabolite in blood and urine or blood ChE inhibition. Salivary biomonitoring has recently been explored as a practical method for examination of chemical exposure; however, there are a limited number of studies exploring its use for OPs. To evaluate the use of salivary ChE as a biological monitor for OP exposure, the current study characterized salivary ChE activity in Sprague-Dawley rats through its comparison with brain and plasma ChE using BW284C51 and iso-OMPA as selective inhibitors of AChE and BuChE, respectively. The study also estimated the kinetic constants describing BuChE interaction with CPF-oxon. A modified Ellman assay in conjunction with pharmacodynamic (PD) modeling was used to characterize the in vitro titration of diluted rat salivary ChE enzyme with CPF-oxon. The results indicated that, more than 95% of rat salivary ChE activity was associated with BuChE activity, total BuChE active site concentration was 0.0012 0.00013 nmol/ml saliva, reactivation rate constant (Kr) was 0.068 0.008 h-1 and inhibitory (Ki) rate constant of 8.825 and 9.80 nM-1h-1 determined experimentally and using model optimization respectively. These study results would be helpful for further evaluating the potential utility of salivary ChE as a practical tool for biological monitor of OP exposures.

  15. Avaliação in vitro do potencial antioxidante de frutas e hortaliças In vitro assessment of the antioxidant potential of fruits and vegetables

    Directory of Open Access Journals (Sweden)

    Simone Pieniz

    2009-04-01

    Full Text Available O efeito protetor exercido por frutas e hortaliças tem sido atribuído à presença de compostos antioxidantes. Objetivou-se, neste estudo, avaliar in vitro a capacidade antioxidante de um grupo de frutas e hortaliças, cruas e cozidas, através da diminuição da peroxidação lipídica, induzida por ferro em fígado de ratos. Foram utilizados fígados de ratos homogeneizados, que foram submetidos à oxidação pelo ferro. As frutas e hortaliças foram utilizadas como antioxidantes, a fim de combater o estresse oxidativo induzido pelo ferro. O método utilizado neste trabalho foi a Reação ao Ácido Tiobarbitúrico (TBARS, tendo como marcador para avaliar o estresse oxidativo o Malonaldeído (MDA. De acordo com os resultados obtidos, observou-se que houve uma diminuição significativa do estresse oxidativo no grupo das frutas e das hortaliças cruas e cozidas com ferro, quando o fígado foi submetido à oxidação deste micronutriente. No grupo das frutas e das hortaliças cruas e cozidas sem ferro, ocorreu redução significativa do estresse oxidativo, apenas em determinadas frutas e hortaliças. O consumo de uma dieta rica em frutas e hortaliças contribui com a defesa antioxidante do organismo, inibindo danos oxidativos em macromoléculas in vitro.The protector effect of fruits and vegetables has been attributed to the presence of antioxidant compounds. The objective of this study was to evaluate the in vitro antioxidant activity of a group of raw and cooked fruits and vegetables, through the decrease of lipid peroxidation, induced by iron in rat livers. Homogenized liver of rats that were submitted to iron oxidation were used in this experiment. The fruits and vegetables were used as antioxidants, in order to combat the oxidative stress induced by the iron. The method used in this experiment was the thiobarbituric acid reaction (TBARS, with malondialdehyde (MDA used as a marker to evaluate the oxidative stress. In accordance with the

  16. Morphological transformation induced by silver nanoparticles in Balb/c 3T3 A31-1-1 mouse cell model to evaluate in vitro carcinogenic potential.

    Science.gov (United States)

    Choo, Wun Hak; Moon, Byeonghak; Song, Sulhwa; Oh, Seung Min

    2017-10-07

    Carcinogenesis is a complex process involving in genotoxic and non-genotoxic pathways. Carcinogenic potential of AgNPs has been predicted by genotoxic effects using several in vitro and in vivo models. However, there is no little information on non-genotoxic effects of AgNPs for carcinogenesis. In vitro cell transformation assay (CTA) can provide specific and sensitive evidence to predict the tumorigenic potential of a chemical, which cannot be supplied by genotoxicity testing. Therefore, we carried out CTA in Balb/c 3T3 A31-1-1 cells to evaluate the carcinogenic potential of silver nanoparticle (AgNPs). Colony forming efficiency (CFE) assay, and crystal violet (CV) assay were carried out to find cytotoxicity of AgNPs. Cytokinesis-block micronucleus assay (CBMN) and CTA in Balb/c 3T3 A31-1-1 cells were performed to predict in vitro carcinogenic potential of AgNPs. In CBMN assay, AgNPs (10.6 ug/mL) induced a significant increase of the micronucleus formation indicating that AgNPs had genotoxicity and could be an initiator for carcinogenesis. In CTA assay to assess carcinogenic potential of AgNPs, cells exposed to AgNPs for 72 h significantly induced morphological neoplastic transformation at all treated doses (0.17, 0.66, 2.65, 5.3, and 10.6 ug/mL) and Tf (transformation frequency) showed a significant increase in a dose-dependent manner. These results indicated that short-term exposure (72 h) to AgNPs had in vitro carcinogenetic potency in Balb/c 3T3 A31-1-1 cells.

  17. Polyurethane/hydroxypropyl cellulose electrospun nanofiber mats as potential transdermal drug delivery system: characterization studies and in vitro assays.

    Science.gov (United States)

    Gencturk, A; Kahraman, E; Güngör, S; Özhan, G; Özsoy, Y; Sarac, A S

    2017-05-01

    Donepezil hydrochloride containing polyurethane/hydroxypropyl cellulose (PU/HPC) nanofibers were prepared by the electrospinning for transdermal drug delivery. PU/HPC nanofibers were characterized with SEM, DSC, and Pascal mercury porosimetry. Drug-excipient interaction was studied by ATR-FTIR. In vitro release of PU/HPC nanofiber mat (10:2:1) exhibited Korsmeyer-Peppas release kinetics controlled by the diffusion of drug. In vitro permeation studies across skin resembling synthetic membrane demonstrated the flux of model drug. The in vitro cytotoxicity data obtained via MTT assay indicated that PU/HPC nanofiber mat could be well tolerated by the skin and the components was not irritant for the skin.

  18. Effect of postmortem time interval on in vitro culture potential of goat skin tissues stored at room temperature.

    Science.gov (United States)

    Singh, Mahipal; Ma, Xiaoling; Sharma, Anil

    2012-09-01

    Animal cloning using somatic cell nuclear transfer technology has renewed the interest in postmortem tissue storage, since these tissues can be used to reintroduce the lost genes back into the breeding pool in animal agriculture, preserve the genetic diversity, and revive the endangered species. However, for successful cloning of animals, integrity of nuclear DNA is essential. Cell viability and their potential to in vitro culture ensure nuclear integrity. The aim of this study was to determine the limits of postmortem time interval within which live cells can be recovered from goat skin tissues. To test the postmortem tissue storage limits, we cultured 2-3 mm(2) skin pieces (n = 70) from the ears of three breeds of goats (n = 7) after 0, 2, 4, and 6 days of postmortem storage at 24°C. After 10 days of culture, outgrowth of fibroblast-like cells (>50 cells) around the explants was scored. All the explants irrespective of breed displayed outgrowth of cells on the dish containing fresh tissues (i.e., day 0 of storage). However, the number of explants exhibiting outgrowth reduced with increasing time interval. Only 53.85 % explants displayed outgrowth after 2 days of tissue storage. The number of explants displaying outgrowth was much smaller after 4 (16.67 %) and 6 days (13.3 %) of storage. In general, the number of outgrowing cells per explant, on a given day, also decreased with increasing postmortem storage time interval. To test the differences between cell cultures, we established secondary cultures from one of the goats exhibiting outgrowth of cells after 6 days of tissue storage and compared them to similar cells from fresh tissues. Comparison of both the cell lines revealed similar cell morphology and growth curves and had doubling times of 23.04 and 22.56 h, respectively. These results suggest that live cells can be recovered from goat (and perhaps other animal) tissues stored at room temperature even after 6 days of their death with comparable growth

  19. Antiamylase, Anticholinesterases, Antiglycation, and Glycation Reversing Potential of Bark and Leaf of Ceylon Cinnamon (Cinnamomum zeylanicum Blume In Vitro

    Directory of Open Access Journals (Sweden)

    Sirimal Premakumara Galbada Arachchige

    2017-01-01

    Full Text Available Ethanol (95% and dichloromethane : methanol (DCM : M, 1 : 1 v/v bark extracts (BEs and leaf extracts (LEs of authenticated Ceylon cinnamon (CC were studied for antiamylase, antiglucosidase, anticholinesterases, and antiglycation and glycation reversing potential in bovine serum albumin- (BSA- glucose and BSA-methylglyoxal models in vitro. Further, total proanthocyanidins (TP were quantified. Results showed significant differences (p<0.05 between bark and leaf extracts for the studied biological activities (except antiglucosidase and TP. BEs showed significantly high (p<0.05 activities for antiamylase (IC50: 214±2–215±10 μg/mL, antibutyrylcholinesterase (IC50: 26.62±1.66–36.09±0.83 μg/mL, and glycation reversing in BSA-glucose model (EC50: 94.33±1.81–107.16±3.95 μg/mL compared to LEs. In contrast, glycation reversing in BSA-methylglyoxal (EC50: ethanol: 122.15±6.01 μg/mL and antiglycation in both BSA-glucose (IC50: ethanol: 15.22±0.47 μg/mL and BSA-methylglyoxal models (IC50: DCM : M: 278.29±8.55 μg/mL were significantly high (p<0.05 in leaf. Compared to the reference drugs used some of the biological activities were significantly (p<0.05 high (BEs: BChE inhibition and ethanol leaf: BSA-glucose mediated antiglycation, some were comparable (BEs: BSA-glucose mediated antiglycation, and some were moderate (BEs and LEs: antiamylase, AChE inhibition, and BSA-MGO mediated antiglycation; DCM : M leaf: BSA-glucose mediated antiglycation. TP were significantly high (p<0.05 in BEs compared to LEs (BEs and LEs: 1097.90±73.01–1381.53±45.93 and 309.52±2.81–434.24±14.12 mg cyanidin equivalents/g extract, resp.. In conclusion, both bark and leaf of CC possess antidiabetic properties and thus may be useful in managing diabetes and its complications.

  20. Antiamylase, Anticholinesterases, Antiglycation, and Glycation Reversing Potential of Bark and Leaf of Ceylon Cinnamon (Cinnamomum zeylanicum Blume) In Vitro

    Science.gov (United States)

    Abeysekera, Walimuni Prabhashini Kaushalya Mendis; Ratnasooriya, Wanigasekera Daya

    2017-01-01

    Ethanol (95%) and dichloromethane : methanol (DCM : M, 1 : 1 v/v) bark extracts (BEs) and leaf extracts (LEs) of authenticated Ceylon cinnamon (CC) were studied for antiamylase, antiglucosidase, anticholinesterases, and antiglycation and glycation reversing potential in bovine serum albumin- (BSA-) glucose and BSA-methylglyoxal models in vitro. Further, total proanthocyanidins (TP) were quantified. Results showed significant differences (p < 0.05) between bark and leaf extracts for the studied biological activities (except antiglucosidase) and TP. BEs showed significantly high (p < 0.05) activities for antiamylase (IC50: 214 ± 2–215 ± 10 μg/mL), antibutyrylcholinesterase (IC50: 26.62 ± 1.66–36.09 ± 0.83 μg/mL), and glycation reversing in BSA-glucose model (EC50: 94.33 ± 1.81–107.16 ± 3.95 μg/mL) compared to LEs. In contrast, glycation reversing in BSA-methylglyoxal (EC50: ethanol: 122.15 ± 6.01 μg/mL) and antiglycation in both BSA-glucose (IC50: ethanol: 15.22 ± 0.47 μg/mL) and BSA-methylglyoxal models (IC50: DCM : M: 278.29 ± 8.55 μg/mL) were significantly high (p < 0.05) in leaf. Compared to the reference drugs used some of the biological activities were significantly (p < 0.05) high (BEs: BChE inhibition and ethanol leaf: BSA-glucose mediated antiglycation), some were comparable (BEs: BSA-glucose mediated antiglycation), and some were moderate (BEs and LEs: antiamylase, AChE inhibition, and BSA-MGO mediated antiglycation; DCM : M leaf: BSA-glucose mediated antiglycation). TP were significantly high (p < 0.05) in BEs compared to LEs (BEs and LEs: 1097.90 ± 73.01–1381.53 ± 45.93 and 309.52 ± 2.81–434.24 ± 14.12 mg cyanidin equivalents/g extract, resp.). In conclusion, both bark and leaf of CC possess antidiabetic properties and thus may be useful in managing diabetes and its complications. PMID:28951761

  1. Nutrient content, in vitro ruminal fermentation characteristics and methane reduction potential of tropical tannin-containing leaves.

    Science.gov (United States)

    Bhatta, Raghavendra; Saravanan, Mani; Baruah, Luna; Sampath, Koratekere T

    2012-12-01

    Plant tannins as rumen modifiers are better than chemicals or antibiotic-based modifiers since these compounds are natural products which are environmentally friendly and therefore have a better acceptance with regard to feed safety issues. Tropical plants containing phenols such as tannins were found to suppress or eliminate protozoa from the rumen and reduce methane and ammonia production. The screening of these plants is an important step in the identification of new compounds and feed additives which might contribute to mitigate rumen methanogenesis. The present study was carried out to determine the efficacy of tannins from tropical tree leaves for their methane reduction properties. Activity of tannins, as represented by the increase in gas volume with the addition of polyethylene glycol (PEG)-6000 as a tannin binder (tannin bioassay) was highest in Ficus bengalensis (555%), followed by Azardirachta indica (78.5%). PEG addition did not alter (P > 0.05) methane percentage in Ficus racemosa, Glyricidia maculata, Leucena leucocephala, Morus alba and Semaroba glauca, confirming that tannins in these samples did not affect methanogenesis. The increase (P 0.05) in the protozoa population in Autocarpus integrifolia, Ficus bengalensis, Jatropha curcus, Morus alba and Sesbania grandiflora, demonstrating that methane reduction observed in these samples per se was not due to defaunation effect of the tannin. The increase in total volatile fatty acid concentration in samples with PEG ranged from 0.6% to > 70%. The highest increase (%) in NH(3)-N was recorded in Azardirachta indica (67.4), followed by Ficus mysoriensis (35.7) and Semaroba glauca (32.6) leaves, reflecting strong protein binding properties of tannin. The results of our study established that in vitro methanogenesis was not essentially related to the density of protozoa population. Tropical tree leaves containing tannins such as Autocarpus integrifolia, Jatropha curcus and Sesbania grandiflora have the

  2. Paradoxically, iron overload does not potentiate doxorubicin-induced cardiotoxicity in vitro in cardiomyocytes and in vivo in mice

    Energy Technology Data Exchange (ETDEWEB)

    Guenancia, Charles [INSERM UMR866, University of Burgundy, LPPCM, Faculties of Medicine and Pharmacy, Dijon (France); Cardiology Department, University Hospital, Dijon (France); Li, Na [INSERM UMR866, University of Burgundy, LPPCM, Faculties of Medicine and Pharmacy, Dijon (France); Hachet, Olivier [INSERM UMR866, University of Burgundy, LPPCM, Faculties of Medicine and Pharmacy, Dijon (France); Cardiology Department, University Hospital, Dijon (France); Rigal, Eve [INSERM UMR866, University of Burgundy, LPPCM, Faculties of Medicine and Pharmacy, Dijon (France); Cottin, Yves [INSERM UMR866, University of Burgundy, LPPCM, Faculties of Medicine and Pharmacy, Dijon (France); Cardiology Department, University Hospital, Dijon (France); Dutartre, Patrick; Rochette, Luc [INSERM UMR866, University of Burgundy, LPPCM, Faculties of Medicine and Pharmacy, Dijon (France); Vergely, Catherine, E-mail: cvergely@u-bourgogne.fr [INSERM UMR866, University of Burgundy, LPPCM, Faculties of Medicine and Pharmacy, Dijon (France)

    2015-04-15

    Doxorubicin (DOX) is known to induce serious cardiotoxicity, which is believed to be mediated by oxidative stress and complex interactions with iron. However, the relationship between iron and DOX-induced cardiotoxicity remains controversial and the role of iron chelation therapy to prevent cardiotoxicity is called into question. Firstly, we evaluated in vitro the effects of DOX in combination with dextran–iron on cell viability in cultured H9c2 cardiomyocytes and EMT-6 cancer cells. Secondly, we used an in vivo murine model of iron overloading (IO) in which male C57BL/6 mice received a daily intra-peritoneal injection of dextran–iron (15 mg/kg) for 3 weeks (D0–D20) and then (D21) a single sub-lethal intra-peritoneal injection of 6 mg/kg of DOX. While DOX significantly decreased cell viability in EMT-6 and H9c2, pretreatment with dextran–iron (125–1000 μg/mL) in combination with DOX, paradoxically limited cytotoxicity in H9c2 and increased it in EMT-6. In mice, IO alone resulted in cardiac hypertrophy (+ 22%) and up-regulation of brain natriuretic peptide and β-myosin heavy-chain (β-MHC) expression, as well as an increase in cardiac nitro-oxidative stress revealed by electron spin resonance spectroscopy. In DOX-treated mice, there was a significant decrease in left-ventricular ejection fraction (LVEF) and an up-regulation of cardiac β-MHC and atrial natriuretic peptide (ANP) expression. However, prior IO did not exacerbate the DOX-induced fall in LVEF and there was no increase in ANP expression. IO did not impair the capacity of DOX to decrease cancer cell viability and could even prevent some aspects of DOX cardiotoxicity in cardiomyocytes and in mice. - Highlights: • The effects of iron on cardiomyocytes were opposite to those on cancer cell lines. • In our model, iron overload did not potentiate anthracycline cardiotoxicity. • Chronic oxidative stress induced by iron could mitigate doxorubicin cardiotoxicity. • The role of iron in

  3. Antiamylase, Anticholinesterases, Antiglycation, and Glycation Reversing Potential of Bark and Leaf of Ceylon Cinnamon (Cinnamomum zeylanicum Blume) In Vitro.

    Science.gov (United States)

    Arachchige, Sirimal Premakumara Galbada; Abeysekera, Walimuni Prabhashini Kaushalya Mendis; Ratnasooriya, Wanigasekera Daya

    2017-01-01

    Ethanol (95%) and dichloromethane : methanol (DCM : M, 1 : 1 v/v) bark extracts (BEs) and leaf extracts (LEs) of authenticated Ceylon cinnamon (CC) were studied for antiamylase, antiglucosidase, anticholinesterases, and antiglycation and glycation reversing potential in bovine serum albumin- (BSA-) glucose and BSA-methylglyoxal models in vitro. Further, total proanthocyanidins (TP) were quantified. Results showed significant differences (p < 0.05) between bark and leaf extracts for the studied biological activities (except antiglucosidase) and TP. BEs showed significantly high (p < 0.05) activities for antiamylase (IC50: 214 ± 2-215 ± 10 μg/mL), antibutyrylcholinesterase (IC50: 26.62 ± 1.66-36.09 ± 0.83 μg/mL), and glycation reversing in BSA-glucose model (EC50: 94.33 ± 1.81-107.16 ± 3.95 μg/mL) compared to LEs. In contrast, glycation reversing in BSA-methylglyoxal (EC50: ethanol: 122.15 ± 6.01 μg/mL) and antiglycation in both BSA-glucose (IC50: ethanol: 15.22 ± 0.47 μg/mL) and BSA-methylglyoxal models (IC50: DCM : M: 278.29 ± 8.55 μg/mL) were significantly high (p < 0.05) in leaf. Compared to the reference drugs used some of the biological activities were significantly (p < 0.05) high (BEs: BChE inhibition and ethanol leaf: BSA-glucose mediated antiglycation), some were comparable (BEs: BSA-glucose mediated antiglycation), and some were moderate (BEs and LEs: antiamylase, AChE inhibition, and BSA-MGO mediated antiglycation; DCM : M leaf: BSA-glucose mediated antiglycation). TP were significantly high (p < 0.05) in BEs compared to LEs (BEs and LEs: 1097.90 ± 73.01-1381.53 ± 45.93 and 309.52 ± 2.81-434.24 ± 14.12 mg cyanidin equivalents/g extract, resp.). In conclusion, both bark and leaf of CC possess antidiabetic properties and thus may be useful in managing diabetes and its complications.

  4. Skin sensitizers in cosmetics and beyond: potential multiple mechanisms of action and importance of T-cell assays for in vitro screening.

    Science.gov (United States)

    Vukmanović, Stanislav; Sadrieh, Nakissa

    2017-05-01

    Allergic contact dermatitis (ACD) is a delayed-type hypersensitivity (DTH) reaction induced by repeated contact with sensitizers. The ability of a chemical to act as a sensitizer has most frequently been tested in animals. As the use of animals for these purposes is gradually and globally being phased out, there is a need for reliable in vitro surrogate assays. Currently proposed in vitro assays are designed to test four key events of the adverse outcome pathway (AOP) involving covalent modification of self-proteins by sensitizers (haptenation) and presentation of new antigens (hapten/carrier complexes) to the immune system. There appears to be imperfect alignment of in vitro assays with clinical and/or animal data, suggesting possibly additional mechanisms of ACD development. Indeed, studies on allergies to small drugs, small chemical-induced HLA-peptide exchange for vaccination purposes and cosmetic ingredient-induced exposure of autoantigens suggest a possibility of DTH response promotion by hapten/carrier-independent mechanisms. Therefore, there is a need for additional appropriate in vitro assays, in order to achieve maximal concordance between clinical and/or animal data and in vitro assays. In this paper, we will review evidence supporting the idea of diverse mechanisms of ACD development. We will also discuss the impact of these multiple mechanisms, on the AOP and on the in vitro assays that should be used for allergen detection. We will propose alloreactivity-like reactions, aided by computer modeling and biochemical tests of compound-HLA binding, as additional tools for better prediction of DTH reactions, resulting from exposure to ingredients in cosmetic products. The combination of the proposed tests, along with the existing assays, should further enhance animal-free assessment of sensitizing potential of individual chemicals.

  5. Investigation of anticancer potential of hypophyllanthin and phyllanthin against breast cancer by in vitro and in vivo methods

    Directory of Open Access Journals (Sweden)

    Madhukiran Parvathaneni

    2014-02-01

    Full Text Available Objective: To investigate the in vitro and in vivo anticancer activities of hypophyllanthin and phyllanthin isolated from Phyllanthus amarus Schum & Thonn against breast cancer. Methods: In vitro anticancer activity was evaluated against two cell lines (MCF-7 and MDAMB-231 using MTT assay. In vivo anticancer activity was tested using Sprague-Dawley rats with N-methyl-N-nitrosourea induced mammary cancer. Results: In vitro studies demonstrated a dose-dependent inhibitory effect on cell growth with IC50 values of (35.18依1.48 µg/mL (hypophyllanthin and (32.51依0.95 µg/mL (phyllanthin for MCF-7; (38.74 依1.24 (hypophyllanthin and (32.2依1.17 (phyllanthin for MDA-MB-231 breast cancer cell lines. Tumor weights per group at doses of 5 and 10 mg/kg/day for hypophyllanthin (12.82 and 12.06 g and phyllanthin (11.95 and 8.87 g treated groups were significantly (P<0.001 lower than untreated N-methyl-N-nitrosourea group (35.85. Conclusions: Results of the present research work indicated that the isolated lignan compounds, hypophyllanthin and phyllanthin showed significant anticancer activities against breast cancer, in vitro and in vivo.

  6. Osmotic potential of Zinnia elegans plant material affects the yield and morphology of tracheary elements produced in vitro

    NARCIS (Netherlands)

    Twumasi, P.; Schel, J.; Ieperen, van W.

    2010-01-01

    The Zinnia elegans cell suspension culture is excellent for xylogenesis studies at the cellular and molecular level, due to the high and synchronous in vitro differentiation of tracheary elements (TEs). The percentage TE differentiation (%TE) in the culture is, however, influenced by a number of

  7. In vitro regeneration of solanum aethiopicum L. (scarlet eggplant), an african vegetable crop with potential ornamental value

    Science.gov (United States)

    Successful in vitro regeneration of plantlets was obtained from shoot tips of five Solanum aethiopicum (African eggplants) accessions evaluated in two media, M1 and M2. The M1 medium consisted of Murashige and Skoog (MS) basal salt mixture supplemented with 20 g/L sucrose, 0.75 g/L MgCl2, and 2 g/L ...

  8. High frequency direct shoot organogenesis of leaf explants and a comparative evaluation of phytochemicals, antioxidant potential of wild vs. in vitro plant extracts of Lysimachia laxa.

    Science.gov (United States)

    Gupta, Sanjoy; Seal, Tapan; Mao, A A; Sarma, Soneswar

    2017-08-01

    The present studies were attempted to develop direct shoot organogenesis from in vitro grown leaf explants of Lysimachia laxa and comparative evaluation of phytochemical and antioxidant potential of in vitro raised and wild plants extracts. The fresh leaves of this species are used for deworming gastrointestinal worm infection in traditional medicine. Overexploitation of this species and poor regeneration has led to rapid decline in wild population, therefore, present investigation was attempted to develop an efficient rapid mass propagation protocol for this species. Our result showed significantly (P rooting induction (100%) with average root number of 11.70 and length 7.35 cm. All rooted plants were successfully acclimatized in greenhouse and transferred to field condition with a survival rate of 97%. The contents of phenolic and flavonoid were higher in in vitro raised plant in compared to wild plant extracts. Antioxidants assay showed high radical scavenging activity of IC50 1.61 ± 0.07 mg dry material and reducing power of 49.79 ± 0.11 mg/g ascorbic acid equivalent by aqueous methanol extracts of in vitro raised 3-months-old plants in compare to the wild plants. The present protocol is a viable option for pharmaceutical or nutraceutical industries for sustainable utilization of L. laxa with enhanced of phytochemical and antioxidant potency which is not reported elsewhere.

  9. Evaluation of the sensitizing potential of antibiotics in vitro using the human cell lines THP-1 and MUTZ-LC and primary monocyte‐derived dendritic cells

    Energy Technology Data Exchange (ETDEWEB)

    Sebastian, Katrin, E-mail: ksebastian@ukaachen.de [Department of Dermatology and Allergology, RWTH Aachen University Hospital, D-52074 Aachen (Germany); Ott, Hagen [Department of Dermatology and Allergology, RWTH Aachen University Hospital, D-52074 Aachen (Germany); Zwadlo-Klarwasser, Gabriele [IZKF (BIOMAT), RWTH Aachen University Hospital, D-52074 Aachen (Germany); Skazik-Voogt, Claudia; Marquardt, Yvonne; Czaja, Katharina; Merk, Hans F.; Baron, Jens Malte [Department of Dermatology and Allergology, RWTH Aachen University Hospital, D-52074 Aachen (Germany)

    2012-08-01

    Since the 7th amendment to the EU cosmetics directive foresees a complete ban on animal testing, alternative in vitro methods have been established to evaluate the sensitizing potential of small molecular weight compounds. To find out whether these novel in vitro assays are also capable to predict the sensitizing potential of small molecular weight drugs, model compounds such as beta-lactams and sulfonamides – which are the most frequent cause of adverse drug reactions – were co-incubated with THP-1, MUTZ-LC, or primary monocyte‐derived dendritic cells for 48 h and subsequent expression of selected marker genes (IL-8, IL-1β, CES1, NQO1, GCLM, PIR and TRIM16) was studied by real time PCR. Benzylpenicillin and phenoxymethylpenicillin were recognized as sensitizing compounds because they are capable to induce the mRNA expression of these genes in moDCs and, except for IL-8, in THP-1 cells but not in MUTZ-LC. Ampicillin stimulated the expression of some marker genes in moDCs and THP-1 cells. SMX did not affect the expression of these genes in THP-1, however, in moDCs, at least PIR was enhanced and there was an increase of the release of IL-8. These data reveal that novel in vitro DC based assays might play a role in the evaluation of the allergenic potential of novel drug compounds, but these systems seem to lack the ability to detect the sensitizing potential of prohaptens that require metabolic activation prior to sensitization and moDCs seem to be superior with regard to the sensitivity compared with THP-1 and MUTZ-3 cell lines. -- Highlights: ► We tested the sensitizing potential of small molecular weight drugs in vitro. ► In vitro assays were performed with moDCs and THP-1 cells. ► Beta-lactam antibiotics can be recognized as sensitizing compounds. ► They affect the expression of metabolic enzymes, cytokines and transcription factors. ► Sulfamethoxazole has no measurable effect on THP-1 cells and moDCs.

  10. Application of drag-reducing polymer solutions as test fluids for in vitro evaluation of potential blood damage in blood pumps.

    Science.gov (United States)

    Daly, Amanda R; Sobajima, Hideo; Olia, Salim E; Takatani, Setsuo; Kameneva, Marina V

    2010-01-01

    In vitro evaluation of the potential of a circulatory-assist device to damage blood cells has generally been performed using blood from various species. Problems with this approach include the variability of blood sensitivity to mechanical stress in different species, preparation of blood including the adjustment of hematocrit to a standard value, changes in the mechanical properties of blood that occur during storage, and necessity to pool blood samples to obtain an adequate amount of blood for in vitro circulating systems. We investigated whether the mechanical degradation of a drag-reducing polymer (DRP) solution resulting in the loss of drag-reducing ability can indicate the degree of shear-induced blood damage within blood pumps. DRP solution (polyethylene oxide, 4,500 kDa, 1,000 ppm) or porcine blood were driven through a turbulent flow system by a centrifugal pump, either the Bio-Pump BPX-80 (Medtronic, Inc.) or CentriMag (Levitronix LLC) at a constant pressure gradient of 300 mm Hg for 120 minutes. DRP mechanical degradation was evaluated by reduction of flow rate and solution viscosity. A proposed index of DRP mechanical degradation (PDI) is similar to the normalized index of hemolysis (NIH) typically used to quantify the results of in vitro testing of blood pumps. Results indicate that the mechanical degradation of DRP solutions may provide a sensitive standard method for the evaluation of potential blood trauma produced by blood pumps without the use of blood.

  11. The isolated chicken eye test as a suitable in vitro method for determining the eye irritation potential of household cleaning products.

    Science.gov (United States)

    Schutte, K; Prinsen, M K; McNamee, P M; Roggeband, R

    2009-08-01

    Eye irritation is an important endpoint in the safety evaluation of consumer products and their ingredients. Several in vitro methods have been developed and are used by different industry sectors to assess eye irritation. One such in vitro method in use for some time already is the isolated chicken eye test (ICE). This investigation focuses on assessing the ICE as a method to determine the eye irritation potential of household cleaning products, both for product safety assurance prior to marketing and for classification and labeling decisions. The ICE involves a single application of test substances onto the cornea of isolated chicken eyes. Endpoints are corneal swelling, corneal opacity and fluorescein retention. The ICE results were compared to historic LVET data in this study due to availability of such in vivo data and the ability to correlate LVET to human experience data on the outcome of accidental exposures to household cleaning products in general. The results of this study indicate that the ICE test is a useful in vitro method for evaluating the eye irritation/corrosion potential and establishing classification and labeling for household cleaning products. For new product formulations, it is best used as part of a weight-of-evidence approach and benchmarked against data from comparable formulations with known eye irritation/corrosion profiles and market experience.

  12. Efficacy of simple short-term in vitro assays for predicting the potential of metal oxide nanoparticles to cause pulmonary inflammation.

    Science.gov (United States)

    Lu, Senlin; Duffin, Rodger; Poland, Craig; Daly, Paul; Murphy, Fiona; Drost, Ellen; Macnee, William; Stone, Vicki; Donaldson, Ken

    2009-02-01

    There has been concern regarding risks from inhalation exposure to nanoparticles (NPs). The large number of particles requiring testing means that alternative approaches to animal testing are needed. We set out to determine whether short-term in vitro assays that assess intrinsic oxidative stress potential and membrane-damaging potency of a panel of metal oxide NPs can be used to predict their inflammogenic potency. For a panel of metal oxide NPs, we investigated intrinsic free radical generation, oxidative activity in an extracellular environment, cytotoxicity to lung epithelial cells, hemolysis, and inflammation potency in rat lungs. All exposures were carried out at equal surface area doses. Only nickel oxide (NiO) and alumina 2 caused significant lung inflammation when instilled into rat lungs at equal surface area, suggesting that these two had extra surface reactivity. We observed significant free radical generation with 4 of 13 metal oxides, only one of which was inflammogenic. Only 3 of 13 were significantly hemolytic, two of which were inflammogenic. Potency in generating free radicals in vitro did not predict inflammation, whereas alumina 2 had no free radical activity but was inflammogenic. The hemolysis assay was correct in predicting the proinflammatory potential of 12 of 13 of the particles examined. Using a battery of simple in vitro tests, it is possible to predict the inflammogenicity of metal oxide NPs, although some false-positive results are likely. More research using a larger panel is needed to confirm the efficacy and generality of this approach for metal oxide NPs.

  13. In Vitro characterization of Lactococcus lactis strains Isolated from Iranian Traditional Dairy Products as a Potential Probiotic

    OpenAIRE

    Fatemeh Nejati; Tobias Oelschlaeger

    2015-01-01

    Few studies have been reported regarding probiotic properties of Lactococcus lactis strains although they are extensively used as starter cultures in the production of dairy products. In this study 8 wild isolates of Lactococcus lactis were evaluated in vitro with regard to resistance to simulated gastric and intestinal juices, adherence ability to Caco-2 cells and HT29-MTX-E12 cell lines, anti-microbial activity, hydrophobicity and antibiotic susceptibility. The results revealed that all iso...

  14. Appraisal of Total Phenol, Flavonoid Contents, and Antioxidant Potential of Folkloric Lannea coromandelica Using In Vitro and In Vivo Assays

    Directory of Open Access Journals (Sweden)

    Tekeshwar Kumar

    2015-01-01

    Full Text Available The aim of this study was to determine the impending antioxidant properties of different extracts of crude methanolic extract (CME of leaves of Lannea coromandelica (L. coromandelica and its two ethyl acetate (EAF and aqueous (AqF subfractions by employing various established in vitro systems and estimation of total phenolic and flavonoid content. The results showed that extract and fractions possessed strong antioxidant activity in vitro and among them, EAF had the strongest antioxidant activity. EAF was confirmed for its highest phenolic content, total flavonoid contents, and total antioxidant capacity. The EAF was found to show remarkable scavenging activity on 2,2-diphenylpicrylhydrazyl (DPPH (EC50 63.9 ± 0.64 µg/mL, superoxide radical (EC50 8.2 ± 0.12 mg/mL, and Fe2+ chelating activity (EC50 6.2 ± 0.09 mg/mL. Based on our in vitro results, EAF was investigated for in vivo antioxidant assay. Intragastric administration of the EAF can significantly increase levels of superoxide dismutase (SOD, catalase (CAT, glutathione (GSH, and glutathione peroxidase (GSH-Px levels, and decrease malondialdehyde (MDA content in the liver and kidney of CCl4-intoxicated rats. These new evidences show that L. coromandelica bared antioxidant activity.

  15. Evaluation of the sensitizing potential of antibiotics in vitro using the human cell lines THP-1 and MUTZ-LC and primary monocyte-derived dendritic cells.

    Science.gov (United States)

    Sebastian, Katrin; Ott, Hagen; Zwadlo-Klarwasser, Gabriele; Skazik-Voogt, Claudia; Marquardt, Yvonne; Czaja, Katharina; Merk, Hans F; Baron, Jens Malte

    2012-08-01

    Since the 7th amendment to the EU cosmetics directive foresees a complete ban on animal testing, alternative in vitro methods have been established to evaluate the sensitizing potential of small molecular weight compounds. To find out whether these novel in vitro assays are also capable to predict the sensitizing potential of small molecular weight drugs, model compounds such as beta-lactams and sulfonamides - which are the most frequent cause of adverse drug reactions - were co-incubated with THP-1, MUTZ-LC, or primary monocyte-derived dendritic cells for 48 h and subsequent expression of selected marker genes (IL-8, IL-1β, CES1, NQO1, GCLM, PIR and TRIM16) was studied by real time PCR. Benzylpenicillin and phenoxymethylpenicillin were recognized as sensitizing compounds because they are capable to induce the mRNA expression of these genes in moDCs and, except for IL-8, in THP-1 cells but not in MUTZ-LC. Ampicillin stimulated the expression of some marker genes in moDCs and THP-1 cells. SMX did not affect the expression of these genes in THP-1, however, in moDCs, at least PIR was enhanced and there was an increase of the release of IL-8. These data reveal that novel in vitro DC based assays might play a role in the evaluation of the allergenic potential of novel drug compounds, but these systems seem to lack the ability to detect the sensitizing potential of prohaptens that require metabolic activation prior to sensitization and moDCs seem to be superior with regard to the sensitivity compared with THP-1 and MUTZ-3 cell lines. Copyright © 2012 Elsevier Inc. All rights reserved.

  16. Secukinumab, a novel anti–IL-17A antibody, shows low immunogenicity potential in human in vitro assays comparable to other marketed biotherapeutics with low clinical immunogenicity

    Science.gov (United States)

    Karle, Anette; Spindeldreher, Sebastian; Kolbinger, Frank

    2016-01-01

    ABSTRACT Secukinumab is a human monoclonal antibody that selectively targets interleukin-17A and has been demonstrated to be highly efficacious in the treatment of moderate to severe plaque psoriasis, starting at early time points, with a sustained effect and a favorable safety profile. Biotherapeutics—including monoclonal antibodies (mAbs)—can be immunogenic, leading to formation of anti-drug antibodies (ADAs) that can result in unwanted effects, including hypersensitivity reactions or compromised therapeutic efficacy. To gain insight into possible explanations for the clinically observed low immunogenicity of secukinumab, we evaluated its immunogenicity potential by applying 2 different in vitro assays: T-cell activation and major histocompatibility complex–associated peptide proteomics (MAPPs). For both assays, monocyte-derived dendritic cells (DCs) from healthy donors were exposed in vitro to biotherapeutic proteins. DCs naturally process proteins and present the derived peptides in the context of human leukocyte antigen (HLA)-class II. HLA-DR–associated biotherapeutic-derived peptides, representing potential T–cell epitopes, were identified in the MAPPs assay. In the T-cell assay, autologous CD4+ T cells were co-cultured with secukinumab-exposed DCs and T-cell activation was measured by proliferation and interleukin-2 secretion. In the MAPPs analysis and T-cell activation assays, secukinumab consistently showed relatively low numbers of potential T-cell epitopes and low T-cell response rates, respectively, comparable to other biotherapeutics with known low clinical immunogenicity. In contrast, biotherapeutics with elevated clinical immunogenicity rates showed increased numbers of potential T-cell epitopes and increased T-cell response rates in T-cell activation assays, indicating an approximate correlation between in vitro assay results and clinical immunogenicity incidence. PMID:26817498

  17. In vitro digestibility and starch content, predicted glycemic index and potential in vitro antidiabetic effect of lentil sprouts obtained by different germination techniques.

    Science.gov (United States)

    Świeca, Michał; Baraniak, Barbara; Gawlik-Dziki, Urszula

    2013-06-01

    The study focuses on changes in starch content and expected glycemic index (eGI) caused by different sprouting methods of lentil. On germination, a decrease was observed in total starch content (TS), α-amylase inhibitors activity (αAI) and eGI values. After elicitation, the highest TS content was determined in 3-day-old control sprouts (100.9 mg/gf.m.), whereas the lowest was in 4-day-old sprouts induced with 300 mM NaCl (57.8 mg/gf.m.). Resistant starch (RS) content was most effectively increased by induction with 600 mM mannitol. The highest eGI values were determined for 3-day-old sprouts induced with 300 mM NaCl, whereas the lowest were for 6-day-old sprouts induced with 100mM NaCl. In treated sprouts starch digestibility was connected with αAI activity and RS content. Sprouting conditions can modify starch content, its potential bioavailability and eGI values. Optimization of this process will allow for the maximum nutritional benefit. Copyright © 2012 Elsevier Ltd. All rights reserved.

  18. Targeted in vitro and in vivo gene transfer into T Lymphocytes: potential of direct inhibition of allo-immune activation

    Directory of Open Access Journals (Sweden)

    Mehra Mandeep R

    2006-11-01

    Full Text Available Abstract Background Successful inhibition of alloimmune activation in organ transplantation remains one of the key events in achieving a long-term graft survival. Since T lymphocytes are largely responsible for alloimmune activation, targeted gene transfer of gene of cyclin kinase inhibitor p21 into T cells might inhibit their aberrant proliferation. A number of strategies using either adenoviral or lentiviral vectors linked to mono or bispecific antibodies directed against T cell surface markers/cytokines did not yield the desired results. Therefore, this study was designed to test if a CD3promoter-p21 chimeric construct would in vitro and in vivo transfer p21 gene to T lymphocytes and result in inhibition of proliferation. CD3 promoter-p21 chimeric constructs were prepared with p21 in the sense and antisense orientation. For in vitro studies EL4-IL-2 thyoma cells were used and for in vivo studies CD3p21 sense and antisense plasmid DNA was injected intramuscularly in mice. Lymphocyte proliferation was quantified by 3H-thymidine uptake assay; IL-2 mRNA expression was studied by RT-PCR and using Real Time PCR assay, we monitored the CD3, p21, TNF-α and IFN-γ mRNA expression. Results Transfection of CD3p21 sense and antisense in mouse thyoma cell line (EL4-IL-2 resulted in modulation of mitogen-induced proliferation. The intramuscular injection of CD3p21 sense and antisense plasmid DNA into mice also modulated lymphocyte proliferation and mRNA expression of pro-inflammatory cytokines. Conclusion These results demonstrate a novel strategy of in vitro and in vivo transfer of p21 gene to T cells using CD3-promoter to achieve targeted inhibition of lymphocyte proliferation and immune activation.

  19. Anti-giardial therapeutic potential of dichloromethane extracts of Zingiber officinale and Curcuma longa in vitro and in vivo.

    Science.gov (United States)

    Dyab, Ahmad K; Yones, Doaa A; Ibraheim, Zedan Z; Hassan, Tasneem M

    2016-07-01

    Giardiosis is one of the common parasitic diarrhoea in humans, especially in children, worldwide. Many drugs are used for its treatment, but there is evidence of drug resistance, insufficient efficacy and unpleasant side effects. Natural products are good candidates for discovering more effective anti-giardial compounds. This study evaluated the activity of extracts of Zingiber officinale (ginger) and Curcuma longa (curcumin) against Giardia lamblia in vitro and in vivo. Giardia cyst suspension was prepared from children faecal specimens. For the in vitro experiment, 1, 10 and 50 mg⁄mL dichloromethane extracts of ginger and curcumin separately were incubated with Giardia cysts for 5, 10, 30 and 60 min. The viability was distinguished by 0.1 % eosin and a haemocytometer. For the in vivo experiments, Balb/c mice were infected with Giardia cyst suspension containing 10,000 cysts/mL. Infected mice were administered 10 and 20 mg kg(-1) day(-1) ginger and curcumin extracts separately for 7 days post-infection. The effectiveness of the extracts was evaluated by faecal cyst and intestinal trophozoite counts and histopathological examination of the small intestine. In vitro ginger extract had a higher significant effect on cyst viability than curcumin, in a dose- and time-dependent manner. In vivo ginger (more effective) and curcumin extracts significantly treated infected mice, and this was evidenced by the faecal cyst and intestinal trophozoite counts reduction, in addition to evident improvement of intestinal mucosal damages induced by Giardia infection. Z. officinale and C. longa extracts may represent effective and natural therapeutic alternatives with low side effects and without drug resistance in the treatment of giardiosis.

  20. Myogenic potential of whole bone marrow mesenchymal stem cells in vitro and in vivo for usage in urinary incontinence.

    Directory of Open Access Journals (Sweden)

    Monica Gunetti

    Full Text Available Urinary incontinence, defined as the complaint of any involuntary loss of urine, is a pathological condition, which affects 30% females and 15% males over 60, often following a progressive decrease of rhabdosphincter cells due to increasing age or secondary to damage to the pelvic floor musculature, connective tissue and/or nerves. Recently, stem cell therapy has been proposed as a source for cell replacement and for trophic support to the sphincter. To develop new therapeutic strategies for urinary incontinence, we studied the interaction between mesenchymal stem cells (MSCs and muscle cells in vitro; thereafter, aiming at a clinical usage, we analyzed the supporting role of MSCs for muscle cells in vitro and in in vivo xenotransplantation. MSCs can express markers of the myogenic cell lineages and give rise, under specific cell culture conditions, to myotube-like structures. Nevertheless, we failed to obtain mixed myotubes both in vitro and in vivo. For in vivo transplantation, we tested a new protocol to collect human MSCs from whole bone marrow, to get larger numbers of cells. MSCs, when transplanted into the pelvic muscles close to the external urethral sphincter, survived for a long time in absence of immunosuppression, and migrated into the muscle among fibers, and towards neuromuscular endplates. Moreover, they showed low levels of cycling cells, and did not infiltrate blood vessels. We never observed formation of cell masses suggestive of tumorigenesis. Those which remained close to the injection site showed an immature phenotype, whereas those in the muscle had more elongated morphologies. Therefore, MSCs are safe and can be easily transplanted without risk of side effects in the pelvic muscles. Further studies are needed to elucidate their integration into muscle fibers, and to promote their muscular transdifferentiation either before or after transplantation.

  1. Synthesis and in vitro biological evaluation of new pyrazole chalcones and heterocyclic diamides as potential anticancer agents

    Directory of Open Access Journals (Sweden)

    Sankappa Rai U.

    2015-05-01

    Full Text Available Synthesis and characterization of new heterocyclic pyrazole chalcones (4a–e and diamide (6a–e derivatives are described. Pyrazole chalcones were synthesized by the reaction of pyrazole aldehydes and suitable aromatic ketones. Diamides were synthesized by the reaction of phthalic acid and amines. Newly synthesized compounds were characterized by spectral studies and their biological activity was assessed in vitro using MCF-7 (human breast adenocarcinoma and HeLa (human cervical tumor cells cell lines. Few of the synthesized molecules inhibited the growth of the human breast cancer cell lines and human cervical tumor cell lines at low micromolar to nanomolar concentrations.

  2. In vitro and in vivo activity of iclaprim, a diaminopyrimidine compound and potential therapeutic alternative against Pneumocystis pneumonia.

    Science.gov (United States)

    Aliouat, E M; Dei-Cas, E; Gantois, N; Pottier, M; Pinçon, C; Hawser, S; Lier, A; Huang, D B

    2018-03-01

    Pneumocystis pneumonia is a serious complication that may affect immunosuppressed patients. The absence of reliable and safe therapeutic alternatives to trimethoprim-sulfamethoxazole (TMP/SMX) justifies the search for more effective and less toxic agents. In this study, the in vitro and in vivo anti-Pneumocystis jirovecii activity of iclaprim, a diaminopyrimidine compound that exerts its antimicrobial activity through the inhibition of dihydrofolate reductase (DHFR), as does TMP, was evaluated alone or in combination with SMX. The antimicrobial activity of iclaprim was tested in vitro using an efficient axenic culture system, and in vivo using P. carinii endotracheally inoculated corticosteroid-treated rats. Animals were orally administered iclaprim (5, 25, 50 mg/kg/day), iclaprim/SMX (5/25, 25/125, 50/250 mg/kg/day), TMP (50 mg/kg/day), or TMP/SMX (50/250 mg/kg/day) once a day for ten consecutive days. The in vitro maximum effect (E max ) and the drug concentrations needed to reach 50% of E max (EC 50 ) were determined, and the slope of the dose-response curve was estimated by the Hill equation (E max sigmoid model). The iclaprim EC 50 value was 20.3 μg/mL. This effect was enhanced when iclaprim was combined with SMX (EC 50 : 13.2/66 μg/mL) (p = 0.002). The TMP/SMX EC 50 value was 51.4/257 μg/mL. In vivo, the iclaprim/SMX combination resulted in 98.1% of inhibition compared to TMP/SMX, which resulted in 86.6% of inhibition (p = 0.048). Thus, overall, the iclaprim/SMX combination was more effective than TMP/SMX both in vitro and in vivo, suggesting that it could be an alternative therapy to the TMP/SMX combination for the treatment of Pneumocystis pneumonia.

  3. Human adipose-derived stem cells isolated from young and elderly women: their differentiation potential and scaffold interaction during in vitro osteoblastic differentiation.

    Science.gov (United States)

    de Girolamo, Laura; Lopa, Silvia; Arrigoni, Elena; Sartori, Matteo F; Baruffaldi Preis, Franz W; Brini, Anna T

    2009-01-01

    Several authors have demonstrated that adipose tissue contains multipotent cells capable of differentiation into several lineages, including bone, cartilage and fat. This study compared human adipose-derived stem cells (hASC) isolated from 26 female donors, under 35 and over 45 years old, showing differences in their cell numbers and proliferation, and evaluated their in vitro adipocytic and osteoblastic differentiation potential. The cellular yield of hASC from older donors was significantly greater than that from younger donors, whereas their clonogenic potential appeared slightly reduced. There were no significant discrepancies between hASC isolated from young and elderly women regarding their in vitro adipocytic differentiation, whereas the osteoblastic potential was significantly reduced by aging. We also assessed the influence of hydroxyapatite (HAP) and silicon carbide (SiC-PECVD) on hASC. Even when cultured on scaffolds, hASC from younger donors had better differentiation into osteoblast-like cells than hASC from older donors; their differentiation ability was up-regulated by the presence of HAP, whereas SiC-PECVD produced no significant effect on hASC osteoblastic differentiation. The large numbers of hASC resident in adipose tissue and their differentiation features suggest that they could be used for a successful bone regeneration process in vivo. We have shown that age does not seem to affect cell viability and in vitro adipocytic differentiation significantly, whereas it does affects osteoblastic differentiation, in the absence and presence of two-dimensional and three-dimensional scaffolds.

  4. The catechol-O-methyltransferase inhibitory potential of Z-vallesiachotamine by in silicoand in vitro approaches

    Directory of Open Access Journals (Sweden)

    Carolina dos Santos Passos

    Full Text Available AbstractZ-Vallesiachotamine is a monoterpene indole alkaloid that has a β-N-acrylate group in its structure. This class of compounds has already been described in different Psychotriaspecies. Our research group observed that E/Z-vallesiachotamine exhibits a multifunctional feature, being able to inhibit targets related to neurodegeneration, such as monoamine oxidase A, sirtuins 1 and 2, and butyrylcholinesterase enzymes. Aiming at better characterizing the multifunctional profile of this compound, its effect on cathecol-O-methyltransferase activity was investigated. The cathecol-O-methyltransferase activity was evaluated in vitro by a fluorescence-based method, using S-(5′-adenosyl-l-methionine as methyl donor and aesculetin as substrate. The assay optimization was performed varying the concentrations of methyl donor (S-(5′-adenosyl-l-methionine and enzyme. It was observed that the highest concentrations of both factors (2.25 U of the enzyme and 100 µM of S-(5′-adenosyl-l-methionine afforded the more reproducible results. The in vitro assay demonstrated that Z-vallesiachotamine was able to inhibit the cathecol-O-methyltransferase activity with an IC50 close to 200 µM. Molecular docking studies indicated that Z-vallesiachotamine can bind the catechol pocket of catechol-O-methyltransferase enzyme. The present work demonstrated for the first time the inhibitory properties of Z-vallesiachotamine on cathecol-O-methyltransferase enzyme, affording additional evidence regarding its multifunctional effects in targets related to neurodegenerative diseases.

  5. Determination of genetic toxicity and potential carcinogenicity in vitro--challenges post the Seventh Amendment to the European Cosmetics Directive.

    Science.gov (United States)

    Tweats, D J; Scott, A D; Westmoreland, C; Carmichael, P L

    2007-01-01

    Genetic toxicology and its role in the detection of carcinogens is currently undergoing a period of reappraisal. There is an increasing interest in developing alternatives to animal testing and the three R's of reduction, refinement and replacement are the basis for EU and national animal protection laws the Seventh Amendment to the EU Cosmetics Directive will ban the marketing of cosmetic/personal care products that contain ingredients that have been tested in animal models. Thus in vivo tests such as the bone marrow micronucleus test, which has a key role in current testing strategies for genotoxicity, will not be available for this class of products. The attrition rate for new, valuable and safe chemicals tested in an in vitro-only testing battery, using the in vitro tests currently established for genotoxicity screening, will greatly increase once this legislation is in place. In addition there has been an explosion of knowledge concerning the cellular and molecular events leading to carcinogenesis. This knowledge has not yet been fully factored into screening chemicals for properties that are not directly linked to mutation induction. Thus there is a pressing need for new, more accurate approaches to determine genotoxicity and carcinogenicity. However, a considerable challenge is presented for these new approaches to be universally accepted and new tests sufficiently validated by March 2009 when the animal testing and marketing bans associated with the Seventh Amendment are due to come into force. This commentary brings together ideas and approaches from several international workshops and meetings to consider these issues.

  6. Maintenance of potential spermatogonial stem cells in vitro by GDNF treatment in a chondrichthyan model (Scyliorhinus canicula L.).

    Science.gov (United States)

    Gautier, Aude; Bosseboeuf, Adrien; Auvray, Pierrick; Sourdaine, Pascal

    2014-10-01

    Previous work in dogfish, Scyliorhinus canicula, has identified the testicular germinative area as the spermatogonial stem cell niche. In the present study, an in vitro co-culture system of spermatogonia and somatic cells from the germinative area was developed. Long-term maintenance of spermatogonia has been successful, and addition of GDNF has promoted the development of clones of spermatogonia expressing stem cell characteristics such as alkaline phosphatase activity and has allowed maintenance of self-renewal in spermatogonia for at least 5 mo under culture conditions, notably by decreasing cell apoptosis. Furthermore, clones of spermatogonia expressed the receptor of GDNF, GFRalpha1, which is consistent with the effect of GDNF on cells despite the lack of identification of a GDNF sequence in the dogfish's transcriptome. However, a sequence homologous to artemin has been identified, and in silico analysis supports the hypothesis that artemin could replace GDNF in the germinative area in dogfish. This study, as the first report on long-term in vitro maintenance of spermatogonia in a chondrichthyan species, suggests that the GFRalpha1 signaling function in self-renewal of spermatogonial stem cells is probably conserved in gnathostomes. © 2014 by the Society for the Study of Reproduction, Inc.

  7. Procalcitonin Impairs Liver Cell Viability and Function In Vitro: A Potential New Mechanism of Liver Dysfunction and Failure during Sepsis?

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    Martin Sauer

    2017-01-01

    Full Text Available Purpose. Liver dysfunction and failure are severe complications of sepsis and result in poor outcome and increased mortality. The underlying pathologic mechanisms of hepatocyte dysfunction and necrosis during sepsis are only incompletely understood. Here, we investigated whether procalcitonin, a biomarker of sepsis, modulates liver cell function and viability. Materials and Methods. Employing a previously characterized and patented biosensor system evaluating hepatocyte toxicity in vitro, human hepatocellular carcinoma cells (HepG2/C3A were exposed to 0.01–50 ng/mL procalcitonin for 2×72 h and evaluated for proliferation, necrosis, metabolic activity, cellular integrity, microalbumin synthesis, and detoxification capacity. Acetaminophen served as positive control. For further standardization, procalcitonin effects were confirmed in a cellular toxicology assay panel employing L929 fibroblasts. Data were analyzed using ANOVA/Tukey’s test. Results. Already at concentrations as low as 0.25 ng/mL, procalcitonin induced HepG2/C3A necrosis (P<0.05 and reduced metabolic activity, cellular integrity, synthesis, and detoxification capacity (all P<0.001. Comparable effects were obtained employing L929 fibroblasts. Conclusion. We provide evidence for procalcitonin to directly impair function and viability of human hepatocytes and exert general cytotoxicity in vitro. Therapeutical targeting of procalcitonin could thus display a novel approach to reduce incidence of liver dysfunction and failure during sepsis and lower morbidity and mortality of septic patients.

  8. Mitochondrial Membrane Potential and Nuclear and Gene Expression Changes During Human Disc Cell Apoptosis: In Vitro and In Vivo Annulus Findings.

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    Gruber, Helen E; Hoelscher, Gretchen L; Bethea, Synthia; Hanley, Edward N

    2015-06-15

    A study using cultured human annulus cells and human annular tissue. To further explore and define mitochondrial mechanisms related to disc cell apoptosis in vitro and in vivo. Mitochondrial-dependent intrinsic signaling pathways are a well-recognized component of apoptosis (programmed cell death). Disc cell apoptosis is important because it is a major mechanism by which cell numbers decrease during disc degeneration. Our objective was to further explore and define mitochondrial mechanisms related to disc cell apoptosis. High-content screening techniques were used to study nuclear morphology and mitochondrial membrane potentials in cultured annulus cells. Gene expression in annulus tissue was studied with microarray analysis. Cultured cells showed significantly increased nuclear size (an indicator of apoptosis) with increasing Thompson grade (P potential (which results from the difference in electrical potential generated by the electrochemical gradient across the inner membrane of the mitochondrion) versus Thompson grade was identified in cultured human annulus cells in control conditions (r = 0.356, P potential was identified versus nontreated cells. Gene expression patterns in more degenerated Thompson grade III, IV, and V discs versus healthier grade I and II discs showed significant upregulation of a number of genes with well-recognized apoptosis roles in mitochondrial potential decline (ITM2B, beta-2-microglobulin, and cathepsin B, DAP, GAS1, and PDCD5) and TNF-α associations (cathepsin B, RAC1, and PPT1). Data presented here show the in vivo expression of apoptosis-related genes associated with the loss of mitochondrial membrane integrity and decreased mitochondrial membrane potential with increasing Thompson scores. These data, which mimic our novel, direct cell-based in vitro findings, stress the importance of mitochondrial changes related to apoptosis and TNF-α during human disc degeneration. N/A.

  9. Phenolic compounds, antioxidant activity and in vitro inhibitory potential against key enzymes relevant for hyperglycemia and hypertension of commonly used medicinal plants, herbs and spices in Latin America.

    Science.gov (United States)

    Ranilla, Lena Galvez; Kwon, Young-In; Apostolidis, Emmanouil; Shetty, Kalidas

    2010-06-01

    Traditionally used medicinal plants, herbs and spices in Latin America were investigated to determine their phenolic profiles, antioxidant activity and in vitro inhibitory potential against key enzymes relevant for hyperglycemia and hypertension. High phenolic and antioxidant activity-containing medicinal plants and spices such as Chancapiedra (Phyllantus niruri L.), Zarzaparrilla (Smilax officinalis), Yerba Mate (Ilex paraguayensis St-Hil), and Huacatay (Tagetes minuta) had the highest anti-hyperglycemia relevant in vitro alpha-glucosidase inhibitory activities with no effect on alpha-amylase. Molle (Schinus molle), Maca (Lepidium meyenii Walp), Caigua (Cyclanthera pedata) and ginger (Zingiber officinale) inhibited significantly the hypertension relevant angiotensin I-converting enzyme (ACE). All evaluated pepper (Capsicum) genus exhibited both anti-hyperglycemia and anti-hypertension potential. Major phenolic compounds in Matico (Piper angustifolium R.), Guascas (Galinsoga parviflora) and Huacatay were chlorogenic acid and hydroxycinnamic acid derivatives. Therefore, specific medicinal plants, herbs and spices from Latin America have potential for hyperglycemia and hypertension prevention associated with Type 2 diabetes. (c) 2010 Elsevier Ltd. All rights reserved.

  10. Evaluation of antiproliferative, anti-type 2 diabetes, and antihypertension potentials of ellagitannins from strawberries (Fragaria × ananassa Duch.) using in vitro models.

    Science.gov (United States)

    Pinto, Marcia da Silva; de Carvalho, Joao Ernesto; Lajolo, Franco Maria; Genovese, Maria Inés; Shetty, Kalidas

    2010-10-01

    Strawberries represent the main source of ellagic acid derivatives in the Brazilian diet, corresponding to more than 50% of all phenolic compounds found in the fruit. There is a particular interest in the determination of the ellagic acid content in fruits because of possible chemopreventive benefits. In the present study, the potential health benefits of purified ellagitannins from strawberries were evaluated in relation to the antiproliferative activity and in vitro inhibition of α-amylase, α-glucosidase, and angiotensin I-converting enzyme (ACE) relevant for potential management of hyperglycemia and hypertension. Therefore, a comparison among ellagic acid, purified ellagitannins, and a strawberry extract was done to evaluate the possible synergistic effects of phenolics. In relation to the antiproliferative activity, it was observed that ellagic acid had the highest percentage inhibition of cell proliferation. The strawberry extract had lower efficacy in inhibiting the cell proliferation, indicating that in the case of this fruit there is no synergism. Purified ellagitannins had high α-amylase and ACE inhibitory activities. However, these compounds had low α-glucosidase inhibitory activity. These results suggested that the ellagitannins and ellagic acid have good potential for the management of hyperglycemia and hypertension linked to type 2 diabetes. However, further studies with animal and human models are needed to advance the in vitro assay-based biochemical rationale from this study.

  11. Prediction of in vivo potential for metabolic activation of drugs into chemically reactive intermediate: correlation of in vitro and in vivo generation of reactive intermediates and in vitro glutathione conjugate formation in rats and humans.

    Science.gov (United States)

    Masubuchi, Noriko; Makino, Chie; Murayama, Nobuyuki

    2007-03-01

    The covalent binding of reactive intermediates to macromolecules might have potential involvement in severe adverse drug reactions. Thus, quantification of reactive metabolites is necessary during the early stage of drug discovery to avoid serious toxicity. In this study, the relationship between covalent binding and glutathione (GSH) conjugate formation in rat and human liver microsomes were investigated using 10 representative radioactive compounds that have been reported as hepatotoxic or having other toxicity derived from their reactive intermediates: acetaminophen, amodiaquine, carbamazepine, clozapine, diclofenac, furosemide, imipramine, indomethacin, isoniazid, and tienilic acid, all at a concentration of 10 microM. The GSH conjugate formation rate correlates well with the covalent binding of radioactivity (both rat and human, r2 = 0.93), which suggests that quantification of the GSH conjugate can be used to estimate covalent binding. To quantify the GSH-conjugation rate with non-radiolabeled compounds in vitro, the validation study for the determination of GSH conjugate formation using 35S-GSH by radio-HPLC was useful to predict metabolic activation. Following oral administration of 20 mg/kg of the radiolabeled compounds to rats, radioactivity that covalently bound to plasma and liver proteins was determined. The in vivo maximum covalent binding level in liver based on the free fraction of plasma area under the concentration curve (AUC) and in vitro covalent binding rate was found to correlate well (r2 = 0.79). Therefore, this model for in vitro covalent binding studies in human and rat and in vivo rat studies might be useful in predicting human metabolic activation of compounds.

  12. In Silico Screening and In Vitro Activity Measurement of Javamide Analogues as Potential p38 MAPK Inhibitors

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    Jae B. Park

    2017-12-01

    Full Text Available p38 Mitogen-activated protein kinase (p38 MAPK is a protein kinase critically involved in the progress of inflammation/stress-associated diseases. Our data suggested that javamide analogues may contain strong anti-inflammation activities, but there is little information about their effects on p38 MAPK. Therefore, in this paper, the effects of thirty javamide analogues on p38 MAPK were investigated using in silico screening and in vitro p38 MAPK assay methods. The javamide analogues were synthesized and their chemical structures were confirmed using nuclear magnetic resonance (NMR spectroscopic methods. Then, the javamide analogues were screened using an in silico modeling program. The screened analogues demonstrated a wide range of binding energy (ΔE; −20 to −39 and several analogues with ΔE; −34 to −39 showed strong binding affinity to p38 MAPK. In vitro p38 MAPK assay, the kinase was significantly inhibited by the analogues with great binding energy (ΔE; −34 to −39 and in silico scores (Avg. score; −27.5 to −29.3. Furthermore, the comparative analysis of both assays showed a positive correlation between the in silico scores and p38 MAPK inhibition. In fact, the javamide analogues with top five in silico scores (Avg. score; −27.5 to −29.3 were found to inhibit p38 MAPK by 27–31% (p < 0.05 better than those with less scores (ΔE < −27.0. Especially, javamide-II-O-ethyl ester with relatively high in silico score (Avg. score; −29.2 inhibited p38 MAPK (IC50 = 9.9 μM a little better than its methyl ester with best in silico score (Avg. score; −29.3. To support the ability to inhibit p38 MAPK, the treatment of javamide-II-ethyl and -methyl esters could suppress the production of IL-8 and MCP-1 protein significantly by 22–73% (p < 0.05 in the differentiated THP-1 cells, and the inhibition was slightly stronger by the ethyl ester than the methyl ester. Altogether, this study suggests that javamide-II-O-ethyl ester may

  13. IN VITRO MASS-SCREENING OF LACTIC ACID BACTERIA AS POTENTIAL BIOSORBENTS OF CESIUM AND STRONTIUM IONS

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    Hideki Kinoshita

    2015-04-01

    Full Text Available Many radionuclides were scattered by the explosion at the Fukushima Daiichi Nuclear Power Station. We examined whether lactic acid bacteria (LAB can sorb cesium ions (Cs+ and strontium ions (Sr2+ for radioprotection. Many strains showed biosorption to Cs+ and Sr2+ using an in vitro mass-screening although each strain showed different sorption. We selected MYU 111, MYU 758, and MYU 759 strains that showed especially high biosorption to Cs+ and/or Sr2+. MYU 111 was identified as Lactobacillus plantarum, and MYU 758 and 759 were Pediococcus pentosaceus. The selected strains tended to show higher biosorption when using the buffer method compared to the culture method. Further, they showed high biosorption at a low concentration of 1 ppb Cs+ and Sr2+ (max 28.8% and 97.7% sorption, respectively. This is the first study where lactic acid bacteria are shown to have biosorption of Cs+ and Sr2+.

  14. In vitro antiplasmodial activity and prophylactic potentials of extract and fractions of Trema orientalis (Linn.) stem bark.

    Science.gov (United States)

    Olanlokun, John Oludele; David, Oluwole Moses; Afolayan, Anthony Jide

    2017-08-15

    Trema orientalis (T. orientalis Linn) has been used in the management of malaria in the western part of Nigeria and despite its application in ethnomedicine, there is dearth of scientific evidence to justify the acclaimed prophylactic antimalarial usage of the plant. The aim of this study is to assess the in vitro antiplasmodial cell-free assay and chemopreventive efficacy of the methanol extract of the stem bark of T. orientalis and its fractions as a prophylactic regimen for malaria prevention. Also, the antimicrobial activities of the extract and the fractions were investigated. Vacuum liquid chromatography was used to obtain dichloromethane, ethylacetate and methanol fractions from the methanol extract of T. orientalis. The fractions were tested for their prophylactic and cell-free antimalarial activity using murine models and β-hematin formation assay respectively. Disc diffusion method was used to determine the antibacterial activity of the extract and its fractions against both Gram-positive and Gram-negative bacteria. In the prophylactic experiment, dichloromethane (DCMF), methanol fraction (MF) and extract (ME) (in this order) showed significant chemopreventive effects against P. berghei invasion of the red blood cells when compared with both Sulfadoxine-Pyrimethamine (SP) and untreated controls. Results of the in vitro study showed that the DCMF had the highest effect in preventing the formation of β-hematin when compared with other fractions. The DCMF also had the highest percentage inhibition of β-hematin formation when compared with chloroquine. The extract and fractions showed a concentration dependent antibacterial activity. Methanol extract had a pronounced inhibitory effect on Enterobacter cloaca ATCC 13047 and Enterococcus faecalis ATCC 29212. Serratia mercescens ATCC 9986 and Pseudomonas aeruginosa ATCC 19582 were the most susceptible bacteria. The results obtained showed that both extract and fractions of T. orientalis possessed

  15. Potential in vitro model for testing the effect of exposure to nanoparticles on the lung alveolar epithelial barrier

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    Raymond Derk

    2015-03-01

    Full Text Available Pulmonary barrier function plays a pivotal role in protection from inhaled particles. However, some nano-scaled particles, such as carbon nanotubes (CNT, have demonstrated the ability to penetrate this barrier in animal models, resulting in an unusual, rapid interstitial fibrosis. To delineate the underlying mechanism and specific bio-effect of inhaled nanoparticles in respiratory toxicity, models of lung epithelial barriers are required that allow accurate representation of in vivo systems; however, there is currently a lack of consistent methods to do so. Thus, this work demonstrates a well-characterized in vitro model of pulmonary barrier function using Calu-3 cells, and provides the experimental conditions required for achieving tight junction complexes in cell culture, with trans-epithelial electrical resistance measurement used as a biosensor for proper barrier formation and integrity. The effects of cell number and serum constituents have been examined and we found that changes in each of these parameters can greatly affect barrier formation. Our data demonstrate that use of 5.0 × 104 Calu-3 cells/well in the Transwell cell culture system, with 10% serum concentrations in culture media is optimal for assessing epithelial barrier function. In addition, we have utilized CNT exposure to analyze the dose-, time-, and nanoparticle property-dependent alterations of epithelial barrier permeability as a means to validate this model. Such high throughput in vitro cell models of the epithelium could be used to predict the interaction of other nanoparticles with lung epithelial barriers to mimic respiratory behavior in vivo, thus providing essential tools and bio-sensing techniques that can be uniformly employed.

  16. In vitro activity of potential old and new drugs against multidrug-resistant gram-negatives.

    Science.gov (United States)

    Rizek, Camila; Ferraz, Juliana Rosa; van der Heijden, Inneke Marie; Giudice, Mauro; Mostachio, Anna Karina; Paez, Jorge; Carrilho, Claudia; Levin, Anna Sara; Costa, Silvia F

    2015-02-01

    The aim of this study was to evaluate the in vitro susceptibility of MDR gram-negatives bacteria to old drugs such as polymyxin B, minocycline and fosfomycin and new drugs such as tigecycline. One hundred and fifty-three isolates from 4 Brazilian hospitals were evaluated. Forty-seven Acinetobacter baumannii resistant to carbapenens harboring adeB, blaOxA23, blaOxA51, blaOxA143 and blaIMP genes, 48 Stenotrophomonas maltophilia including isolates resistant to levofloxacin and/or trimethoprim-sulfamethoxazole harboring sul-1, sul-2 and qnrMR and 8 Serratia marcescens and 50 Klebsiella pneumoniae resistant to carbapenens harboring blaKPC-2 were tested to determine their minimum inhibitory concentrations (MICs) by microdilution to the following drugs: minocycline, ampicillin-sulbactam, tigecycline, and polymyxin B and by agar dilution to fosfomycin according with breakpoint criteria of CLSI and EUCAST (fosfomycin). In addition, EUCAST fosfomycin breakpoint for Pseudomonas spp. was applied for Acinetobacter spp and S. maltophilia, the FDA criteria for tigecycline was used for Acinetobacter spp and S. maltophilia and the Pseudomonas spp polymyxin B CLSI criterion was used for S. maltophilia. Tigecycline showed the best in vitro activity against the MDR gram-negative evaluated, followed by polymyxin B and fosfomycin. Polymyxin B resistance among K. pneumoniae was detected in 6 isolates, using the breakpoint of MIC > 8 ug/mL. Two of these isolates were resistant to tigecycline. Minocycline was tested only against S. maltophilia and A. baumannii and showed excellent activity against both. Fosfomycin seems to not be an option to treat infections due to the A. baumannii and S. maltophilia isolates according with EUCAST breakpoint, on the other hand, showed excellent activity against S. marcescens and K. pneumoniae. Copyright © 2014 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

  17. Transient receptor potential ankyrin 1 receptor activation in vitro and in vivo by pro-tussive agents: GRC 17536 as a promising anti-tussive therapeutic.

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    Indranil Mukhopadhyay

    Full Text Available Cough is a protective reflex action that helps clear the respiratory tract which is continuously exposed to airborne environmental irritants. However, chronic cough presents itself as a disease in its own right and despite its global occurrence; the molecular mechanisms responsible for cough are not completely understood. Transient receptor potential ankyrin1 (TRPA1 is robustly expressed in the neuronal as well as non-neuronal cells of the respiratory tract and is a sensor of a wide range of environmental irritants. It is fast getting acceptance as a key biological sensor of a variety of pro-tussive agents often implicated in miscellaneous chronic cough conditions. In the present study, we demonstrate in vitro direct functional activation of TRPA1 receptor by citric acid which is routinely used to evoke cough in preclinical and clinical studies. We also show for the first time that a potent and selective TRPA1 antagonist GRC 17536 inhibits citric acid induced cellular Ca(+2 influx in TRPA1 expressing cells and the citric acid induced cough response in guinea pigs. Hence our data provides a mechanistic link between TRPA1 receptor activation in vitro and cough response induced in vivo by citric acid. Furthermore, we also show evidence for TRPA1 activation in vitro by the TLR4, TLR7 and TLR8 ligands which are implicated in bacterial/respiratory virus pathogenesis often resulting in chronic cough. In conclusion, this study highlights the potential utility of TRPA1 antagonist such as GRC 17536 in the treatment of miscellaneous chronic cough conditions arising due to diverse causes but commonly driven via TRPA1.

  18. Transient receptor potential ankyrin 1 receptor activation in vitro and in vivo by pro-tussive agents: GRC 17536 as a promising anti-tussive therapeutic.

    Science.gov (United States)

    Mukhopadhyay, Indranil; Kulkarni, Abhay; Aranake, Sarika; Karnik, Pallavi; Shetty, Mahesh; Thorat, Sandeep; Ghosh, Indraneel; Wale, Dinesh; Bhosale, Vikram; Khairatkar-Joshi, Neelima

    2014-01-01

    Cough is a protective reflex action that helps clear the respiratory tract which is continuously exposed to airborne environmental irritants. However, chronic cough presents itself as a disease in its own right and despite its global occurrence; the molecular mechanisms responsible for cough are not completely understood. Transient receptor potential ankyrin1 (TRPA1) is robustly expressed in the neuronal as well as non-neuronal cells of the respiratory tract and is a sensor of a wide range of environmental irritants. It is fast getting acceptance as a key biological sensor of a variety of pro-tussive agents often implicated in miscellaneous chronic cough conditions. In the present study, we demonstrate in vitro direct functional activation of TRPA1 receptor by citric acid which is routinely used to evoke cough in preclinical and clinical studies. We also show for the first time that a potent and selective TRPA1 antagonist GRC 17536 inhibits citric acid induced cellular Ca(+2) influx in TRPA1 expressing cells and the citric acid induced cough response in guinea pigs. Hence our data provides a mechanistic link between TRPA1 receptor activation in vitro and cough response induced in vivo by citric acid. Furthermore, we also show evidence for TRPA1 activation in vitro by the TLR4, TLR7 and TLR8 ligands which are implicated in bacterial/respiratory virus pathogenesis often resulting in chronic cough. In conclusion, this study highlights the potential utility of TRPA1 antagonist such as GRC 17536 in the treatment of miscellaneous chronic cough conditions arising due to diverse causes but commonly driven via TRPA1.

  19. The pathogenic potential of Helicobacter cinaedi isolated from non-human sources: adherence, invasion and translocation ability in polarized intestinal epithelial Caco-2 cells in vitro.

    Science.gov (United States)

    Taniguchi, Takako; Yamazaki, Wataru; Saeki, Yuji; Takajo, Ichiro; Okayama, Akihiko; Hayashi, Tetsuya; Misawa, Naoaki

    2016-05-03

    Helicobacter cinaedi infection has been recognized as an increasingly important emerging disease in humans. Infection with H. cinaedi causes bacteremia, cellulitis and enteritis. H. cinaedi has been isolated from non-human sources, including dogs, cats and rodents; however, it remains unclear whether animal strains are pathogenic in humans and as zoonotic pathogens. In this study, H. cinaedi isolates were recovered from a dog and a hamster, and the ability of these isolates to adhere to, invade and translocate across polarized human intestinal epithelial Caco-2 cells was examined in vitro. To better understand the pathogenic potential of animal H. cinaedi isolates, these results were compared with those for a human strain that was isolated from a patient with bacteremia. The animal and human strains adhered to and invaded Caco-2 cells, but to a lesser degree than the C. jejuni 81-176 strain, which was used as a control. The integrity of tight junctions was monitored by measuring transepithelial electrical resistance (TER) with a membrane insert system. The TER values for all H. cinaedi strains did not change during the experimental periods compared with those of the controls; however, translocation of H. cinaedi from the apical side to the basolateral side was confirmed by cultivation and H. cinaedi-specific PCR, suggesting that the H. cinaedi strains translocated by transcellular route. This study demonstrated that H. cinaedi strains of animal origin might have a pathogenic potential in human epithelial cells as observed in a translocation assay in vitro with a human isolate.

  20. Nanoparticle Fullerene (C60) demonstrated stable binding with antibacterial potential towards probable targets of drug resistant Salmonella typhi - a computational perspective and in vitro investigation.

    Science.gov (United States)

    Skariyachan, Sinosh; Parveen, Asma; Garka, Shruti

    2017-12-01

    Salmonella typhi, a Gram negative bacterium, has become multidrug resistant (MDR) to wide classes of antibacterials which necessitate an alarming precaution. This study focuses on the binding potential and therapeutic insight of Nano-Fullerene C60 towards virulent targets of Salmonella typhi by computational prediction and preliminary in vitro assays. The clinical isolates of Salmonella typhi were collected and antibiotic susceptibility profiles were assessed. The drug targets of pathogen were selected by rigorous literature survey and gene network analysis by various metabolic network resources. Based on this study, 20 targets were screened and the 3D structures of few drug targets were retrieved from PDB and others were computationally predicted. The structures of nanoleads such as Fullerene C60, ZnO and CuO were retrieved from drug databases. The binding potential of these nanoleads towards all selected targets were predicted by molecular docking. The best docked conformations were screened and concept was investigated by preliminary bioassays. This study revealed that most of the isolates of Salmonella typhi were found to be MDR (p C60 showed better binding affinity towards the drug targets when compared to ZnO and CuO. The preliminary in vitro assays suggested that 100 μg/L Fullerene C60 posses significant inhibitory activities and absence of drug resistance to this nanoparticle. This study suggests that Fullerene C60 can be scaled up as probable lead molecules against the major drug targets of MDR Salmonella typhi.

  1. Improvement of In Vitro Osteogenic Potential through Differentiation of Induced Pluripotent Stem Cells from Human Exfoliated Dental Tissue towards Mesenchymal-Like Stem Cells

    Directory of Open Access Journals (Sweden)

    Felipe Augusto Andre Ishiy

    2015-01-01

    Full Text Available Constraints for the application of MSCs for bone reconstruction include restricted self-renewal and limited cell amounts. iPSC technology presents advantages over MSCs, providing homogeneous cellular populations with prolonged self-renewal and higher plasticity. However, it is unknown if the osteogenic potential of iPSCs differs from that of MSCs and if it depends on the iPSCs originating cellular source. Here, we compared the in vitro osteogenesis between stem cells from human deciduous teeth (SHED and MSC-like cells from iPSCs from SHED (iPS-SHED and from human dermal fibroblasts (iPS-FIB. MSC-like cells from iPS-SHED and iPS-FIB displayed fibroblast-like morphology, downregulation of pluripotency markers and upregulation of mesenchymal markers. Comparative in vitro osteogenesis analysis showed higher osteogenic potential in MSC-like cells from iPS-SHED followed by MSC-like cells from iPS-FIB and SHED. CD105 expression, reported to be inversely correlated with osteogenic potential in MSCs, did not display this pattern, considering that SHED presented lower CD105 expression. Higher osteogenic potential of MSC-like cells from iPS-SHED may be due to cellular homogeneity and/or to donor tissue epigenetic memory. Our findings strengthen the rationale for the use of iPSCs in bone bioengineering. Unveiling the molecular basis behind these differences is important for a thorough use of iPSCs in clinical scenarios.

  2. Metformin potentiates B-cell response to high glucose: an in vitro study on isolated perfused pancreas from normal rats.

    Science.gov (United States)

    Gregorio, F; Filipponi, P; Ambrosi, F; Cristallini, S; Marchetti, P; Calafiore, R; Navalesi, R; Brunetti, P

    1989-01-01

    This study investigated the effects of metformin on pancreatic A-B- and D-cell functions using the isolated perfused rat pancreas model. The lactate output rate following metformin infusion was also monitored. Metformin was infused at the low "therapeutic" concentration of 1.5 micrograms/ml and its effects were evaluated in three different glycaemic conditions: during a basal infusion of 4.44 mM glucose, during a moderate increase to 8.88 mM of glucose concentration, and finally during a higher 16.66 mM glycaemic stimulus. Basal insulin secretion and B-cell release during the lower hyperglycaemic stimulus were unaffected by metformin infusion. On the contrary, the drug significantly enhanced insulin response to 16.66 mM glucose, particularly by increasing the second phase of hormone release. Glucagon and somatostatin releases during metformin infusion were similar to the secretory pattern observed in the control experiments both in the basal condition and in the presence of the two different hyperglycaemic stimuli. Finally metformin did not modify the lactate output rate from perfused pancreas, irrespective of the different glycaemic conditions employed. Therefore our data suggest--at least in rats, in in vitro experiments but above all in the presence of markedly elevated hyperglycaemic conditions--that metformin may influence the glucose stimulatory effect on B-cell activity.

  3. Formulation and In-vitro Evaluation of Tretinoin Microemulsion as a Potential Carrier for Dermal Drug Delivery

    Science.gov (United States)

    Mortazavi, Seyed Alireza; Pishrochi, Sanaz; Jafari azar, Zahra

    2013-01-01

    In this study, tretinoin microemulsion has been formulated based on phase diagram studies by changing the amounts and proportions of inactive ingredients, such as surfactants, co-surfactants and oils. The effects of these variables have been determined on microemulsion formation, particle size of the dispersed phase and release profile of tretinoin from microemulsion through dialysis membrane. In released studies, static Franz diffusion cells mounted with dialysis membrane were used. Sampling was conducted every 3 h at room temperature over a period of 24 h. The amount of released drug was measured with UV-spectrophotometer and the percentage of drug released was calculated. Based on the results obtained, the oil phase concentration had a proportional effect on particle size which can consequently influence on drug release. The particle size and the amount of released drug were affected by the applied surfactants. The components of the optimized microemulsion formulation were 15% olive oil, 12% propylene glycol (as co-surfactant), 33% Tween®80 (as surfactant) and 40% distilled water, which was tested for viscosity and rheological behavior. The prepared tretinoin microemulsion showed pseudoplastic-thixotropic behavior. The profile of drug release follows zero order kinetics. The optimized tretinoin microemulsion showed enhanced in-vitro release profile compared to the commercial gels and creams. PMID:24523740

  4. Potential of plant essential oils and their components in animal agriculture – in vitro studies on antibacterial mode of action

    Directory of Open Access Journals (Sweden)

    Corliss A. O'bryan

    2015-09-01

    Full Text Available The broad field of agriculture is currently undergoing major changes in practices, with new catch phrases including organic and sustainable. Consumers are more aware than ever before of the food that they eat and they want food free of toxic chemicals, antibiotics and the like. The antimicrobial activity of essential oils and their components has been recognized for several years. Recent research has demonstrated that many of these essential oils have beneficial effects for livestock, including reduction of foodborne pathogens in these animals. Essential oils as natural antimicrobials offer the opportunity to help maintain the safety of our food supply and minimize consumers’ concerns about consumption of synthetic chemicals. Numerous studies have been made into the mode of action of essential oils and the resulting elucidation of bacterial cell targets have contributed to new perspectives on countering antimicrobial resistance and pathogenicity of these bacteria. In this review, after a brief discussion of the uses essential oils in agriculture as antimicrobials, we give an overview of the current knowledge about the antibacterial mode of action of essential oils and their constituents as determined in vitro.

  5. Rethinking In Vitro Embryo Culture: New Developments in Culture Platforms and Potential to Improve Assisted Reproductive Technologies1

    Science.gov (United States)

    Smith, Gary D.; Takayama, Shuichi; Swain, Jason E.

    2011-01-01

    ABSTRACT The preponderance of research toward improving embryo development in vitro has focused on manipulation of the chemical soluble environment, including altering basic salt composition, energy substrate concentration, amino acid makeup, and the effect of various growth factors or addition or subtraction of other supplements. In contrast, relatively little work has been done examining the physical requirements of preimplantation embryos and the role culture platforms or devices can play in influencing embryo development within the laboratory. The goal of this review is not to reevaluate the soluble composition of past and current embryo culture media, but rather to consider how other controlled and precise factors such as time, space, mechanical interactions, gradient diffusions, cell movement, and surface interactions might influence embryo development. Novel culture platforms are being developed as a result of interdisciplinary collaborations between biologists and biomedical, material, chemical, and mechanical engineers. These approaches are looking beyond the soluble media composition and examining issues such as media volume and embryo spacing. Furthermore, methods that permit precise and regulated dynamic embryo culture with fluid flow and embryo movement are now available, and novel culture surfaces are being developed and tested. While several factors remain to be investigated to optimize the efficiency of embryo production, manipulation of the embryo culture microenvironment through novel devices and platforms may offer a pathway toward improving embryo development within the laboratory of the future. PMID:21998170

  6. Technological Potential of Lactobacillus Strains Isolated from Fermented Green Olives: In Vitro Studies with Emphasis on Oleuropein-Degrading Capability

    Directory of Open Access Journals (Sweden)

    Massimo Iorizzo

    2016-01-01

    Full Text Available Technological properties of two strains of Lactobacillus plantarum (B3 and B11 and one of Lactobacillus pentosus (B4, previously isolated from natural fermented green olives, have been studied in vitro. Acidifying ability, salt, temperature, and pH tolerances of all strains were found in the range reported for similar strains produced in Italy and optimal growth conditions were found to be 6.0–8.0 pH, 15–30°C temperature, and less than 6% NaCl. Moreover, all strains showed very good tolerance to common olive phenol content (0.3% total phenol and high oleuropein-degrading capability. It was found that medium composition affected the bacterial oleuropein degradation. B11 strain grown in a nutrient-rich medium showed a lower oleuropein-degrading action than when it was cultivated in nutrient-poor medium. Furthermore, enzymatic activity assays revealed that oleuropein depletion did not correspond to an increase of hydroxytyrosol, evidencing that bacterial strains could efficiently degrade oleuropein via a mechanism different from hydrolysis.

  7. Rethinking in vitro embryo culture: new developments in culture platforms and potential to improve assisted reproductive technologies.

    Science.gov (United States)

    Smith, Gary D; Takayama, Shuichi; Swain, Jason E

    2012-03-01

    The preponderance of research toward improving embryo development in vitro has focused on manipulation of the chemical soluble environment, including altering basic salt composition, energy substrate concentration, amino acid makeup, and the effect of various growth factors or addition or subtraction of other supplements. In contrast, relatively little work has been done examining the physical requirements of preimplantation embryos and the role culture platforms or devices can play in influencing embryo development within the laboratory. The goal of this review is not to reevaluate the soluble composition of past and current embryo culture media, but rather to consider how other controlled and precise factors such as time, space, mechanical interactions, gradient diffusions, cell movement, and surface interactions might influence embryo development. Novel culture platforms are being developed as a result of interdisciplinary collaborations between biologists and biomedical, material, chemical, and mechanical engineers. These approaches are looking beyond the soluble media composition and examining issues such as media volume and embryo spacing. Furthermore, methods that permit precise and regulated dynamic embryo culture with fluid flow and embryo movement are now available, and novel culture surfaces are being developed and tested. While several factors remain to be investigated to optimize the efficiency of embryo production, manipulation of the embryo culture microenvironment through novel devices and platforms may offer a pathway toward improving embryo development within the laboratory of the future.

  8. In Silico, In Vitro, and In Vivo Studies Indicate the Potential Use of Bolaamphiphiles for Therapeutic siRNAs Delivery.

    Science.gov (United States)

    Kim, Taejin; Afonin, Kirill A; Viard, Mathias; Koyfman, Alexey Y; Sparks, Selene; Heldman, Eliahu; Grinberg, Sarina; Linder, Charles; Blumenthal, Robert P; Shapiro, Bruce A

    2013-03-19

    Specific small interfering RNAs (siRNAs) designed to silence different oncogenic pathways can be used for cancer therapy. However, non-modified naked siRNAs have short half-lives in blood serum and encounter difficulties in crossing biological membranes due to their negative charge. These obstacles can be overcome by using siRNAs complexed with bolaamphiphiles, consisting of two positively charged head groups that flank an internal hydrophobic chain. Bolaamphiphiles have relatively low toxicities, long persistence in the blood stream, and most importantly, in aqueous conditions can form poly-cationic micelles thus, becoming amenable to association with siRNAs. Herein, two different bolaamphiphiles with acetylcholine head groups attached to an alkyl chain in two distinct configurations are compared for their abilities to complex with siRNAs and deliver them into cells inducing gene silencing. Our explicit solvent molecular dynamics (MD) simulations showed that bolaamphiphiles associate with siRNAs due to electrostatic, hydrogen bonding, and hydrophobic interactions. These in silico studies are supported by various in vitro and in cell culture experimental techniques as well as by some in vivo studies. Results demonstrate that depending on the application, the extent of siRNA chemical protection, delivery efficiency, and further intracellular release can be varied by simply changing the type of bolaamphiphile used.Molecular Therapy-Nucleic Acids (2013) 2, e80; doi:10.1038/mtna.2013.5; published online 19 March 2013.

  9. Evaluation of in vitro antimicrobial potential and GC–MS analysis of Camellia sinensis and Terminalia arjuna

    Directory of Open Access Journals (Sweden)

    Divya Gupta

    2017-03-01

    Full Text Available Traditionally, Camellia sinensis and Terminalia arjuna are being used widely to cure various diseases like cardiovascular diseases, cancer etc. In the present study, extracts of these plants were evaluated for their antimicrobial activities against some human pathogenic bacteria viz. E. coli, P. aeruginosa, S. aureus and fungus C. albicans. In-vitro inhibition of these pathogenic microorganisms produced inhibition zone ranging from 9 to 18 mm. MIC values of these plant extracts ranged from 6.25 to 12.5 mg/ml. MBC of C. sinensis for E. coli, P. aeruginosa and S. aureus was found to be 50 and 12.5 mg/ml, respectively. In case of T. arjuna, the MBC of all the tested microorganisms was found to be 25 mg/ml. The MFC of C. sinensis and T. arjuna against C. albicans was observed to be 50 and 25 mg/ml, respectively. GC–MS analysis of C. sinensis and T. arjuna extract identified 13 and 21 compounds, respectively.

  10. High FSH decreases the developmental potential of mouse oocytes and resulting fertilized embryos, but does not influence offspring physiology and behavior in vitro or in vivo.

    Science.gov (United States)

    Li, Min; Zhao, Yue; Zhao, Cui H; Yan, Jie; Yan, Ying L; Rong, Li; Liu, Ping; Feng, Huai-Liang; Yu, Yang; Qiao, Jie

    2013-05-01

    Do different concentrations of FSH in the assisted reproductive technology (ART) procedure in vitro or in vivo affect the developmental competence of oocytes, the embryos and the offspring conceived from these embryos? Improper FSH treatment (200 IU/l in vitro, 10 IU/ml in vivo and 200 IU/ml in vivo) impairs the development competence of oocyte and embryo, but does not influence offspring physiology and behavior. Exogenous FSH has been widely used in the field of ART. However, the effects of different concentrations of FSH on the developmental competence of oocytes, embryos and the offspring conceived from these embryos, are still unknown. In a prospective study, a total of 45 mice at 8-10 weeks of age were primed in vivo with different dosages of FSH (9 mice in the 10 IU/ml, 10 mice in the 50 IU/ml, 10 mice in the 100 IU/ml and 16 mice in the 200 IU/ml groups). Fresh MII oocytes were retrieved from ovaries: this was designated as in vivo group. Thirty six mice at 8-10 weeks of age were sacrificed by cervical dislocation to obtain ovaries without FSH treatment (9 mice in the 0 IU/l, 9 mice in the 50 IU/l, 8 mice in the 100 IU/l and 10 mice in the 200 IU/l groups), and then the immature oocytes were collected from these ovaries and cultured in vitro matured medium supplemented with 0, 50, 100 and 200 IU/l FSH: this was designated as in vitro group. Spindle assembly of matured MII oocytes was stained via an immunofluorescence method and the oocytes ratio of normal spindle was analyzed. The developmental competence of the resulting fertilized embryos in the pre- and post-implantation stages was examined in in vitro and in vivo groups. Furthermore, physiological index, including reproductive potential and body weight, of the offspring was investigated by mating experiments and behavior index, including learning, memory, probing and intelligence, was tested by Morris water maze in in vitro and in vivo groups. In the in vitro groups, the oocyte maturation competence

  11. Paradoxically, iron overload does not potentiate doxorubicin-induced cardiotoxicity in vitro in cardiomyocytes and in vivo in mice.

    Science.gov (United States)

    Guenancia, Charles; Li, Na; Hachet, Olivier; Rigal, Eve; Cottin, Yves; Dutartre, Patrick; Rochette, Luc; Vergely, Catherine

    2015-04-15

    Doxorubicin (DOX) is known to induce serious cardiotoxicity, which is believed to be mediated by oxidative stress and complex interactions with iron. However, the relationship between iron and DOX-induced cardiotoxicity remains controversial and the role of iron chelation therapy to prevent cardiotoxicity is called into question. Firstly, we evaluated in vitro the effects of DOX in combination with dextran-iron on cell viability in cultured H9c2 cardiomyocytes and EMT-6 cancer cells. Secondly, we used an in vivo murine model of iron overloading (IO) in which male C57BL/6 mice received a daily intra-peritoneal injection of dextran-iron (15mg/kg) for 3weeks (D0-D20) and then (D21) a single sub-lethal intra-peritoneal injection of 6mg/kg of DOX. While DOX significantly decreased cell viability in EMT-6 and H9c2, pretreatment with dextran-iron (125-1000μg/mL) in combination with DOX, paradoxically limited cytotoxicity in H9c2 and increased it in EMT-6. In mice, IO alone resulted in cardiac hypertrophy (+22%) and up-regulation of brain natriuretic peptide and β-myosin heavy-chain (β-MHC) expression, as well as an increase in cardiac nitro-oxidative stress revealed by electron spin resonance spectroscopy. In DOX-treated mice, there was a significant decrease in left-ventricular ejection fraction (LVEF) and an up-regulation of cardiac β-MHC and atrial natriuretic peptide (ANP) expression. However, prior IO did not exacerbate the DOX-induced fall in LVEF and there was no increase in ANP expression. IO did not impair the capacity of DOX to decrease cancer cell viability and could even prevent some aspects of DOX cardiotoxicity in cardiomyocytes and in mice. Copyright © 2015 Elsevier Inc. All rights reserved.

  12. In-vitro cytocidal effect of water on bladder cancer cells: The potential role for intraperitoneal lavage during radical cystectomy.

    Science.gov (United States)

    Taoka, Rikiya; Williams, Stephen B; Ho, Philip L; Kamat, Ashish M

    2015-01-01

    We investigate the cytocidal effect of water on bladder cancer cells. Intraperitoneal lavage with sterile water is sometimes used during radical cystectomy to lyse cancer cells that might have escaped the surgical specimen. The efficacy of this approach at the cellular level is unknown. Three bladder cancer cell lines of varying grade, RT4, TCCSUP and T24 were exposed to sterile water, and morphological changes were closely observed under microscopy. Changes of cell membrane integrity, cell viability, and cell number of re-incubated cells after water exposure were measured to determine water induced hypotonic shock. The low-grade RT4 cells started swelling immediately upon exposure to water followed by rupture within 3 minutes. The higher grade TCCSUP and T24 cells demonstrated limited hypotonic swelling with significantly less cell rupture after 10 minutes. The damage to cell membrane of RT4 cells was evident at 1 minute; only 10.0% of cells were intact at 10 minutes. On the other hand, 41.9% and 77.8% of TCCSUP and T24 cells were intact at 10 minutes, respectively. Percentage of viable cells at 10 minutes was 2.1 ± 2.3%, 2.3 ± 0.4%, and 16.1 ± 0.6% for RT4, TCCSUP, and T24, respectively. Cytocidal effect of hypotonic shock can be achieved, to varying degrees, by exposing bladder cancer cells to water for at least 10 minutes. This in vitro study may have bearing on the effects seen with intraperitoneal lavage using sterile water during radical cystectomy.

  13. Stromal cell-derived factor-1β potentiates bone morphogenetic protein-2-stimulated osteoinduction of genetically engineered bone marrow-derived mesenchymal stem cells in vitro.

    Science.gov (United States)

    Herberg, Samuel; Fulzele, Sadanand; Yang, Nianlan; Shi, Xingming; Hess, Matthew; Periyasamy-Thandavan, Sudharsan; Hamrick, Mark W; Isales, Carlos M; Hill, William D

    2013-01-01

    Skeletal injuries are among the most prevalent clinical problems and bone marrow-derived mesenchymal stem/stromal cells (BMSCs) have successfully been used for the treatment thereof. Stromal cell-derived factor-1 (SDF-1; CXCL12) is a member of the CXC chemokine family with multiple splice variants. The two most abundant variants, SDF-1α and SDF-1β, share identical amino acid sequences, except for four additional amino acids at the C-terminus of SDF-1β, which may mediate surface stabilization via glycosaminoglycans and protect SDF-1β from proteolytic cleavage, rendering it twice as potent as SDF-1α. Increasing evidence suggests that SDF-1 is involved in bone formation through regulation of recruitment, engraftment, proliferation, and differentiation of stem/progenitor cells. The underlying molecular mechanisms, however, have not yet been fully elucidated. In this study, we tested the hypothesis that SDF-1β can potentiate bone morphogenetic protein-2 (BMP-2)-stimulated osteogenic differentiation and chemotaxis of BMSCs in vitro. Utilizing retrovirus-mediated gene transfer to generate novel Tet-Off-SDF-1β BMSCs, we found that conditional SDF-1β expression is tightly regulated by doxycycline in a dose-dependent and temporal fashion, leading to significantly increased SDF-1β mRNA and protein levels. In addition, SDF-1β was found to enhance BMP-2-stimulated mineralization, mRNA and protein expression of key osteogenic markers, and regulate BMP-2 signal transduction via extracellular signal-regulated kinases 1/2 (Erk1/2) phosphorylation in genetically engineered BMSCs in vitro. We also showed that SDF-1β promotes the migratory response of CXC chemokine receptor 4 (CXCR4)-expressing BMSCs in vitro. Taken together, these data support that SDF-1β can play an important role in BMP-2-stimulated osteogenic differentiation of BMSCs and may exert its biological activity in both an autocrine and paracrine fashion.

  14. Nickel subsulfide is genotoxic in vitro but shows no mutagenic potential in respiratory tract tissues of BigBlue rats and Muta Mouse mice in vivo after inhalation.

    Science.gov (United States)

    Mayer, C; Klein, R G; Wesch, H; Schmezer, P

    1998-12-03

    Carcinogenic nickel compounds are known to induce promutagenic DNA lesions such as DNA strand breaks and DNA adducts in cultured mammalian cells. In standard mutation assays, in contrast, they were found to be either inactive or weakly active. In our in vitro mutation studies in a lacI transgenic embryonic fibroblast cell line, nickel subsulfide (Ni3S2) increased mutation frequency up to 4. 5-fold. We subsequently applied the comet assay and transgenic rodent mutation assays to investigate the DNA damaging effect and mutagenic potential of nickel subsulfide in target cells of carcinogenesis. A 2-h in vitro treatment of freshly isolated mouse nasal mucosa and lung cells with nickel subsulfide clearly induced DNA fragmentation in a concentration dependent manner. The strong effect was not seen in the same cell types following inhalative treatment of mice and rats, leading only in the mouse nasal mucosa to high DNA damage. When the same inhalative treatment was applied to lacZ and lacI transgenic mice and rats, the spontaneous mutation frequency of these target genes in the respiratory tissues was not increased. These results support a recently proposed non-genotoxic model of nickel carcinogenesis, which acts through gene silencing via DNA methylation and chromatin condensation. This model may also explain our in vitro mutation data in the lacI transgenic cell line, in which nickel subsulfide increased mutation frequency, but in about one-third of the mutants, molecular analysis did not reveal any DNA sequence change in the coding region of the lacI gene despite of the phenotypic loss of its function. Copyright 1998 Elsevier Science B.V.

  15. Standardization, chemical profiling, in vitro cytotoxic effects, in vivo anti-carcinogenic potential and biosafety profile of Indian propolis.

    Science.gov (United States)

    Kapare, Harshad; Lohidasan, Sathiyanarayanan; Sinnathambi, Arulmozhi; Mahadik, Kakasaheb

    2017-12-04

    Propolis from apiculture is known for wide range of medicinal properties owing to its vast chemical constituents including polyphenols, flavonoids and anticancer agent Caffeic acid phenethyl ester (CAPE). The objective of the study was to extract and standardize Indian propolis (IP) with respect to selected markers by newly developed High performance liquid chromatography (HPLC) method, to evaluate in vitro and in vivo anticancer activity and biosafety of Indian propolis. IP was extracted, optimized and standardized using a newly developed and validated HPLC method for simultaneous estimation of caffeic acid, apigenin, quercetin and CAPE. The standardised ethanolic extract of IP (EEIP) was screened for in vitro cytotoxicity using sulforhodamine B (SRB) assay, in vivo anti-carcinogenic effect against Dalton's Lymphoma ascites (DLA) cells, hemolytic effect and pesticide analysis. The EEIP was found to contain more amount of total flavonoids (23.61+ 0.0452 mg equivalent of quercetin/g), total polyphenolics (34.82 + 0.0785 mg equivalent of gallic acid/g) and all selected markers except caffeic acid compared to all other extracts. EEIP showed better anti-cancer potential than CAPE on MCF-7 and HT-29 cell line and significant (p carcinogenic effects against DLA in comparison with 5-fluorouracil. EEIP was found to be non-hemolytic. From in vitro cytotoxicity, in vivo anti-carcinogenicity and biosafety studies it can be concluded that the standardized EEIP is safe and can be considered for further development as a biomedicine. Copyright © 2017 Transdisciplinary University, Bangalore and World Ayurveda Foundation. Published by Elsevier B.V. All rights reserved.

  16. Lysergic acid diethylamide (LSD) is a partial agonist of D2 dopaminergic receptors and it potentiates dopamine-mediated prolactin secretion in lactotrophs in vitro.

    Science.gov (United States)

    Giacomelli, S; Palmery, M; Romanelli, L; Cheng, C Y; Silvestrini, B

    1998-01-01

    The hallucinogenic effects of lysergic acid diethylamide (LSD) have mainly been attributed to the interaction of this drug with the serotoninergic system, but it seems more likely that they are the result of the complex interactions of the drug with both the serotoninergic and dopaminergic systems. The aim of the present study was to investigate the functional actions of LSD at dopaminergic receptors using prolactin secretion by primary cultures of rat pituitary cells as a model. LSD produced a dose-dependent inhibition of prolactin secretion in vitro with an IC50 at 1.7x10(-9) M. This action was antagonized by spiperone but not by SKF83566 or cyproheptadine, which indicates that LSD has a specific effect on D2 dopaminergic receptors. The maximum inhibition of prolactin secretion achieved by LSD was lower than that by dopamine (60% versus 80%). Moreover, the fact that LSD at 10(-8)-10(-6) M antagonized the inhibitory effect of dopamine (10(-7) M) and bromocriptine (10(-11) M) suggests that LSD acts as a partial agonist at D2 receptors on lactotrophs in vitro. Interestingly, LSD at 10(-13)-10(-10) M, the concentrations which are 10-1000-fold lower than those required to induce direct inhibition on pituitary prolactin secretion, potentiated the dopamine (10(-10)-2.5x10(-9) M)-mediated prolactin secretion by pituitary cells in vitro. These results suggest that LSD not only interacts with dopaminergic receptors but also has a unique capacity for modulating dopaminergic transmission. These findings may offer new insights into the hallucinogenic effect of LSD.

  17. Study on the Cytotoxic, Genotoxic and Clastogenic Potential of Attalea phalerata Mart. ex Spreng. Oil Pulp In Vitro and In Vivo Experimental Models.

    Science.gov (United States)

    Freitas de Lima, Fernando; Lima Tolouei Menegati, Sara Emilia; Karenina Traesel, Giseli; Souza de Araújo, Flávio Henrique; Honaiser Lescano, Caroline; Moraes Peixoto, Sara; Mao Silva, Felipe Ariel; Heredia Vieira, Silvia Cristina; do Carmo Vieira, Maria; Oesterreich, Silvia Aparecida

    2016-01-01

    Attalea phalerata Mart. ex Spreng. (Arecaceae), popularly known as "bacuri", is used in Brazilian folk medicine. Its oil is used orally to relieve pulmonary congestion and joint pain. In topical applications, it is applied as an effective hair tonic and anti-dandruff. The in natura pulp and its nuts are used as food because of its nutritional value. Despite its use in folk medicine, there is a lack of data regarding its in vivo/in vitro cytotoxic/genotoxic and clastogenic effects. Therefore, in this study, we evaluated the cytotoxic, genotoxic and clastogenic effects of Attalea phalerata Mart. ex Spreng. oil (APMO) in vitro and in vivo. For the analysis of cytotoxic potential, the Artemia salina and MTT (3-(4,5-dimethizzol-zyl)-2,5-diphenyltetrazolium bromide) assays were performed. Possible cytotoxic, genotoxic and clastogenic effects of APMO intake were determined by performing the comet and micronucleus assays. Male and female Wistar rats were orally treated with doses of 125, 250, 500 or 1000 mg.kg-1 of the APMO daily for 28 consecutive days (four weeks). The results showed that the APMO did not induce cell death in the experiments of Artemia salina and MTT, indicating that it has no cytotoxicity. The APMO did not cause significant damage to the DNA of the rats in the four doses used when compared to the negative control group (saline + Tween® 80). The APMO did not present any significant increase in micronucleated polychromatic erythrocytes (MNPCEs) for the four tested doses. When compared to the positive control group, all groups (comet and micronucleus tests) were statistically different. These data suggest that the administration of Attalea phalerata Mart oil. ex Spreng does not cause cytotoxicity, genotoxicity and clastogenicity in experimental models in vitro and in vivo following oral administration in this study.

  18. Thalidomide inhibits alternative activation of macrophages in vivo and in vitro: a potential mechanism of anti-asthmatic effect of thalidomide.

    Science.gov (United States)

    Lee, Hyun Seung; Kwon, Hyouk-Soo; Park, Da-Eun; Woo, Yeon Duk; Kim, Hye Young; Kim, Hang-Rae; Cho, Sang-Heon; Min, Kyung-Up; Kang, Hye-Ryun; Chang, Yoon-Seok

    2015-01-01

    Thalidomide is known to have anti-inflammatory and immunomodulatory actions. However, the effect and the anti-asthmatic mechanism of thalidomide in the pathogenesis of asthmatic airways are not fully understood. This study is designed to determine the effect and the potential mechanism of thalidomide in the pathogenesis of asthmatic airways using animal model of allergic asthma. Six-week-old female BALB/C mice were sensitized with alum plus ovalbumin (OVA) and were exposed to OVA via intranasal route for 3 days for challenge. Thalidomide 200 mg/kg was given via gavage twice a day from a day before the challenge and airway hyperresponsivenss (AHR), airway inflammatory cells, and cytokines in bronchoalveolar lavage fluids (BALF) were evaluated. The expression levels of pro-inflammatory cytokines and other mediators were evaluated using ELISA, real time (RT)-qPCR, and flow cytometry. CRL-2456, alveolar macrophage cell line, was used to test the direct effect of thalidomide on the activation of macrophages in vitro. The mice with thalidomide treatment showed significantly reduced levels of allergen-induced BALF and lung inflammation, AHR, and the expression of a number of pro-inflammatory cytokines and mediators including Th2 related, IL-17 cytokines, and altered levels of allergen-specific IgG1/IgG2a. Of interesting note, thalidomide treatment significantly reduced expression levels of allergen- or Th2 cytokine-stimulated alternative activation of macrophages in vivo and in vitro. These studies highlight a potential use of thalidomide in the treatment of allergic diseases including asthma. This study further identified a novel inhibitory effect of thalidomide on alternative activation of macrophages as a potential mechanism of anti-asthmatic effect of thalidomide.

  19. Screening of in vitro cytotoxicity, antioxidant potential and bioactivity of nano- and micro-ZrO2 and -TiO2 particles.

    Science.gov (United States)

    Karunakaran, Gopalu; Suriyaprabha, Rangaraj; Manivasakan, Palanisamy; Yuvakkumar, Rathinam; Rajendran, Venkatachalam; Kannan, Narayanasamy

    2013-07-01

    Nanometal oxides are used in tissue engineering and implants. The increased use of nanoparticles suggests the need to study their adverse effects on biological systems. The present investigation explores in vitro cytotoxicity, antioxidant potential, and bioactivity of nano- and micro-particles such as zirconia (ZrO2) and titania (TiO2) on biological systems such as National Institute of Health (NIH) 3T3 mouse embryonic fibroblasts cell line, di(phenyl)-(2,4,6-trinitrophenyl) iminoazanium (DPPH) and simulated body fluid (SBF). The cell line viability % indicated that nano ZrO2 and TiO2 were less toxic than microparticles up to 200µgml(-1). DPPH assay revealed that the free radical scavenging potential of tested particles were higher for nano ZrO2 (76.9%) and nano TiO2 (73.3%) at 100mg than that for micron size particles. Calcium deposition percentage of micro- and nano-ZrO2 particles, after SBF study, showed 0.066% and 0.094% respectively, whereas for micro- and nano-TiO2, it was 0.251% and 0.615% respectively. FTIR results showed a good bioactivity through hydroxyapatite formation. The present investigation clearly shows that nanoparticles possess good antioxidant potential and better biocompatibility under in vitro conditions which are dose and size dependent. Hence, cytotoxicity itself is not promising evaluation method for toxicity rather than particles individual characterisation using antioxidant and bioactivity analysis. Copyright © 2013 Elsevier Inc. All rights reserved.

  20. Fertilization in vitro with spermatozoa from different mice increased variation in the developmental potential of embryos compared to artificial parthenogenetic activation.

    Science.gov (United States)

    Miao, De-Qiang; Liang, Bo; Wang, Jun-Zuo; Wang, Hui-Li; Cui, Wei; Liu, Yong; Tan, Jing-He

    2009-03-01

    Although successful embryo development is dependent upon genetic and epigenetic contributions from both the male and female, the male potential to adversely affect embryo development has been scarcely studied. It is unclear whether the sperm variation among different males would affect the outcome of oocyte evaluation by embryo development following fertilization. In the present study, variation in the developmental potential of mouse embryos was first compared between in vitro fertilization with epididymal spermatozoa from different males and Sr(2+) parthenogenetic activation using oocytes of different qualities, and then the effect of male on fertilization and embryo development was examined using randomly chosen oocytes and spermatozoa from cauda epididymidis, vas deferens or electro-ejaculates. Rates of fertilization and blastocyst formation were significantly higher with spermatozoa from cauda epididymidis or vas deferens than with ejaculated spermatozoa. Rates of embryonic development differed significantly between different males, but not between different ejaculates of the same male. Analysis of standard errors of means and coefficients of variance indicated that as long as multiple males were involved, the variation in oocyte fertilization/activation and blastocyst formation was always higher after fertilization than after Sr(2+) parthenogenetic activation whether spermatozoa were collected from epididymidis, vas deferens or ejaculates and regardless of oocyte qualities. It is concluded that (1) epididymal mouse spermatozoa fertilize more oocytes than ejaculated spermatozoa under identical experimental conditions; (2) like farm animals, the mice also show a remarkable male effect on the developmental potential of in vitro produced embryos although they are supposed to be less genetically diverse; (3) parthenogenetic activation is recommended for assessment of oocyte quality to exclude the effect of male.

  1. Thalidomide inhibits alternative activation of macrophages in vivo and in vitro: a potential mechanism of anti-asthmatic effect of thalidomide.

    Directory of Open Access Journals (Sweden)

    Hyun Seung Lee

    Full Text Available Thalidomide is known to have anti-inflammatory and immunomodulatory actions. However, the effect and the anti-asthmatic mechanism of thalidomide in the pathogenesis of asthmatic airways are not fully understood.This study is designed to determine the effect and the potential mechanism of thalidomide in the pathogenesis of asthmatic airways using animal model of allergic asthma.Six-week-old female BALB/C mice were sensitized with alum plus ovalbumin (OVA and were exposed to OVA via intranasal route for 3 days for challenge. Thalidomide 200 mg/kg was given via gavage twice a day from a day before the challenge and airway hyperresponsivenss (AHR, airway inflammatory cells, and cytokines in bronchoalveolar lavage fluids (BALF were evaluated. The expression levels of pro-inflammatory cytokines and other mediators were evaluated using ELISA, real time (RT-qPCR, and flow cytometry. CRL-2456, alveolar macrophage cell line, was used to test the direct effect of thalidomide on the activation of macrophages in vitro.The mice with thalidomide treatment showed significantly reduced levels of allergen-induced BALF and lung inflammation, AHR, and the expression of a number of pro-inflammatory cytokines and mediators including Th2 related, IL-17 cytokines, and altered levels of allergen-specific IgG1/IgG2a. Of interesting note, thalidomide treatment significantly reduced expression levels of allergen- or Th2 cytokine-stimulated alternative activation of macrophages in vivo and in vitro.These studies highlight a potential use of thalidomide in the treatment of allergic diseases including asthma. This study further identified a novel inhibitory effect of thalidomide on alternative activation of macrophages as a potential mechanism of anti-asthmatic effect of thalidomide.

  2. Thalidomide Inhibits Alternative Activation of Macrophages In Vivo and In Vitro: A Potential Mechanism of Anti-Asthmatic Effect of Thalidomide

    Science.gov (United States)

    Park, Da-Eun; Woo, Yeon Duk; Kim, Hye Young; Kim, Hang-Rae; Cho, Sang-Heon; Min, Kyung-Up; Kang, Hye-Ryun; Chang, Yoon-Seok

    2015-01-01

    Background Thalidomide is known to have anti-inflammatory and immunomodulatory actions. However, the effect and the anti-asthmatic mechanism of thalidomide in the pathogenesis of asthmatic airways are not fully understood. Objective This study is designed to determine the effect and the potential mechanism of thalidomide in the pathogenesis of asthmatic airways using animal model of allergic asthma. Methods Six-week-old female BALB/C mice were sensitized with alum plus ovalbumin (OVA) and were exposed to OVA via intranasal route for 3 days for challenge. Thalidomide 200 mg/kg was given via gavage twice a day from a day before the challenge and airway hyperresponsivenss (AHR), airway inflammatory cells, and cytokines in bronchoalveolar lavage fluids (BALF) were evaluated. The expression levels of pro-inflammatory cytokines and other mediators were evaluated using ELISA, real time (RT)-qPCR, and flow cytometry. CRL-2456, alveolar macrophage cell line, was used to test the direct effect of thalidomide on the activation of macrophages in vitro. Results The mice with thalidomide treatment showed significantly reduced levels of allergen-induced BALF and lung inflammation, AHR, and the expression of a number of pro-inflammatory cytokines and mediators including Th2 related, IL-17 cytokines, and altered levels of allergen-specific IgG1/IgG2a. Of interesting note, thalidomide treatment significantly reduced expression levels of allergen- or Th2 cytokine-stimulated alternative activation of macrophages in vivo and in vitro. Conclusion These studies highlight a potential use of thalidomide in the treatment of allergic diseases including asthma. This study further identified a novel inhibitory effect of thalidomide on alternative activation of macrophages as a potential mechanism of anti-asthmatic effect of thalidomide. PMID:25905462

  3. In-vitro evaluation of the P-glycoprotein interactions of a series of potentially CNS-active Amaryllidaceae alkaloids

    DEFF Research Database (Denmark)

    Eriksson, André Huss; Rønsted, Nina; Jäger, Anna Katharina

    2012-01-01

    Drug compounds interacting with the blood-brain barrier efflux transporter P-glycoprotein (P-gp) might have limited access to brain tissue. The aim of the present study was to evaluate whether nine potentially CNS-active Amaryllidaceae alkaloids of the crinine, lycorine and galanthamine types int...

  4. In vitro ecology of Seiridium cardinale and allied species: the effect of solute stress and water potential on fungal growth

    Directory of Open Access Journals (Sweden)

    Elena TURCO

    2011-05-01

    Full Text Available Normal 0 14 false false false IT ZH-TW X-NONE MicrosoftInternetExplorer4 Defining the potential implications of global climate change on Mediterranean forest ecosystems requires a basic knowledge on the ecology of fungal pathogens under conditions that would stress host plants. The Mediterranean cypress (Cupressus sempervirens-Seiridium spp. pathosystem represents an important case study. In the last century, epidemics of cypress canker have killed historical plantations and the decades-long host resistance will probably break down in the future as a result of both host and pathogen adaptation to increasing temperature and decreasing summer precipitation. In this study, the effect of osmotic water stress on mycelial growth of Seiridium cardinale, S. unicorne and S. cupressi in culture was examined and compared to that of Diplodia cupressi, which is a pathogen of cypress known to be favoured by host water stress. Growth responses were evaluated on potato sucrose agar amended with KCl or NaCl to give water potentials in the range of -0.34 to -15 MPa. Mycelial growth decreased with decreasing water potential and ceased at -15 MPa, although the mycelium remained alive. Histochemical analysis conducted on S. cardinale grown at -12 MPa revealed melanization and thickening of hyphal walls, in addition to abundance of lipid-rich organelles. These results suggest that the three Seiridium spp. might survive drying cycles in cypress wood, but their tolerance is different. Successful survival strategies may partly result from changes in mycelium structure. Furthermore, S. unicorne was positively stimulated by a water potential of -3 MPa, suggesting that it may have high adaptive potential for life in a drier Mediterranean ecosystem, which is predicted to occur under scenarios of global warming. Normal 0 14 MicrosoftInternetExplorer4

  5. The Combined Use of in Silico, in Vitro, and in Vivo Analyses to Assess Anti-cancerous Potential of a Bioactive Compound from Cyanobacterium Nostoc sp. MGL001

    Directory of Open Access Journals (Sweden)

    Niveshika

    2017-11-01

    Full Text Available Escalating incidences of cancer, especially in developed and developing countries, demand evaluation of potential unexplored natural drug resources. Here, anticancer potential of 9-Ethyliminomethyl-12-(morpholin-4-ylmethoxy-5,8,13,16-tetraaza -hexacene-2,3-dicarboxylic acid (EMTAHDCA isolated from fresh water cyanobacterium Nostoc sp. MGL001 was screened through in silico, in vitro, and in vivo studies. For in silico analysis, EMTAHDCA was selected as ligand and 11 cancer related proteins (Protein Data Bank ID: 1BIX, 1NOW, 1TE6, 2RCW, 2UVL, 2VCJ, 3CRY, 3HQU, 3NMQ, 5P21, and 4B7P which are common targets of various anticancer drugs were selected as receptors. The results obtained from in silico analysis showed that EMTAHDCA has strong binding affinity for all the 11 target protein receptors. The ability of EMTAHDCA to bind active sites of cancer protein targets indicated that it is functionally similar to commercially available anticancer drugs. For assessing cellular metabolic activities, in vitro studies were performed by using calorimetric assay viz. 3-(4,5-dimethylthiazol-2-yl-2,5 diphenyltetrazolium bromide (MTT. Results showed that EMTAHDCA induced significant cytotoxic response against Dalton's lymphoma ascites (DLA cells in a dose and time dependent manner with an inhibitory concentration (IC50 value of 372.4 ng/mL after 24 h of incubation. However, in case of normal bone marrow cells, the EMTAHDCA did not induce cytotoxicity as the IC50 value was not obtained even with higher dose of 1,000 ng/mL EMTAHDCA. Further, in vivo studies revealed that the median life span/survival days of tumor bearing mice treated with EMTAHDCA increased significantly with a fold change of ~1.9 and 1.81 corresponding to doses of 5 and 10 mg/kg body weight (B.W. of EMTAHDCA respectively, as compared to the DL group. Our results suggest that 5 mg/kg B.W. is effective since the dose of 10 mg/kg B.W. did not show any significant difference as compared to 5 mg/kg B

  6. Evaluation of the Potential Nephroprotective and Antimicrobial Effect of Camellia sinensis Leaves versus Hibiscus sabdariffa (In Vivo and In Vitro Studies

    Directory of Open Access Journals (Sweden)

    Doa’a Anwar Ibrahim

    2014-01-01

    Full Text Available Green tea and hibiscus are widely consumed as traditional beverages in Yemen and some regional countries. They are relatively cheap and the belief is that they improve health state and cure many diseases. The aim of this study was to evaluate the potential protective and antibacterial activity of these two famous plants in vitro through measuring their antibacterial activity and in vivo through measuring nonenzymatic kidney markers dysfunction after induction of nephrotoxicity by gentamicin. Gram positive bacteria like MRSA (methicillin resistant Staphylococcus aureus were isolated from hospitalized patients’ different sources (pus and wound and Gram negative bacteria including E. coli and P. aeruginosa were used in vitro study. In addition, the efficacy of these plants was assessed in vivo through measuring nonenzymatic kidney markers including S. creatinine and S. urea. Green tea was shown antimicrobial activity against MRSA with inhibition zone 19.67 ± 0.33 mm and MIC 1.25 ± 0.00 mg/mL compared with standard reference (vancomycin 18.00 ± 0.00 mg/mL. Hibiscus did not exhibit a similar effect. Both Hibiscus- and green tea-treated groups had nephroprotective effects as they reduced the elevation in nonenzymatic kidney markers. We conclude that green tea has dual effects: antimicrobial and nephroprotective.

  7. Photophysical Characterization and in Vitro Phototoxicity Evaluation of 5,10,15,20-Tetra(quinolin-2-ylporphyrin as a Potential Sensitizer for Photodynamic Therapy

    Directory of Open Access Journals (Sweden)

    Letícia D. Costa

    2016-03-01

    Full Text Available Photodynamic therapy (PDT is a selective and minimally invasive therapeutic approach, involving the combination of a light-sensitive compound, called a photosensitizer (PS, visible light and molecular oxygen. The interaction of these per se harmless agents results in the production of reactive species. This triggers a series of cellular events that culminate in the selective destruction of cancer cells, inside which the photosensitizer preferentially accumulates. The search for ideal PDT photosensitizers has been a very active field of research, with a special focus on porphyrins and porphyrin-related macrocycle molecules. The present study describes the photophysical characterization and in vitro phototoxicity evaluation of 5,10,15,20-tetra(quinolin-2-ylporphyrin (2-TQP as a potential PDT photosensitizer. Molar absorption coefficients were determined from the corresponding absorption spectrum, the fluorescence quantum yield was calculated using 5,10,15,20-tetraphenylporphyrin (TPP as a standard and the quantum yield of singlet oxygen generation was determined by direct phosphorescence measurements. Toxicity evaluations (in the presence and absence of irradiation were performed against HT29 colorectal adenocarcinoma cancer cells. The results from this preliminary study show that the hydrophobic 2-TQP fulfills several critical requirements for a good PDT photosensitizer, namely a high quantum yield of singlet oxygen generation (Φ∆ 0.62, absence of dark toxicity and significant in vitro phototoxicity for concentrations in the micromolar range.

  8. Effect of contact angle, zeta potential and particles size on the in vitro studies of Al2O3 and SiO2 nanoparticles.

    Science.gov (United States)

    Karunakaran, Gopalu; Suriyaprabha, Rangaraj; Rajendran, Venkatachalam; Kannan, Narayanasamy

    2015-02-01

    Currently, nanometal oxides find their role in different biological applications such as tissue engineering, implant and bone replacement materials. Owing to the increased use of nanoparticles, it is necessary to understand their release and toxicity in the biological system. In this regard, three independent studies such as in vitro cytotoxicity, antioxidant activity and biocompatibility of nano- and micrometal oxide particles such as alumina (Al2O3) and silica (SiO2) are evaluated. It is evident from cell viability study that nanoAl2O3 and SiO2 particles are less toxic when compared with microAl2O3 and SiO2 to NIH 3T3 cell lines up to 200 µg/ml. Antioxidant properties of micro- and nanoAl2O3 in terms of radical scavenging percentage for micro- and nanoAl2O3 are 59.1% and 72.1%, respectively, at 100 mg. Similarly, the radical scavenging percentage of nano- and bulk SiO2 are 81.0% and 67.2%, respectively. The present study reveals that the cellular behaviour, interaction and biocompatibility of metal oxides differ with dose, particle size, contact angle and zeta potential. The present study opens up a new strategy to analyse in vitro nanotoxicity.

  9. Biomonitoring of the genotoxic effects and oxidative potentials of commercial edible dung beetles (Onitis sp.), grasshopper (Caelifera sp.) and mole crickets (Gryllotalpa sp.) in vitro.

    Science.gov (United States)

    Koc, Kubra; Incekara, Umit; Turkez, Hasan

    2014-09-01

    In this investigation, the genotoxic and oxidative effects of water soluble extracts of dung beetles, flying grasshopper and mole crickets have been assessed on cultured human blood cells. The extracts were added to the culture tubes at 12 different concentrations (0-2000 ppm). Micronucleus test was used to monitor the DNA and the chromosomal damage produced by aqueous extracts in vitro. In addition, to assess the oxidative effects, total antioxidant capacity (TAC) and total oxidant status (TOS) levels were also measured. Our results indicated that these extracts did not show genotoxic effects at the tested concentrations. However, the extracts caused dose-dependent alterations in both TAC and TOS levels. Based on the findings, it was concluded that the studied insects can be consumed safely, but it is necessary to consider the cellular damages which are likely to appear depending on oxidative stress at higher concentrations. It has also been suggested that this in vitro approach for oxidative and genotoxicity assessments may be useful to evaluate the potential health risks of edible insects. © The Author(s) 2014.

  10. Potential of Microbispora sp. V2 as biocontrol agent against Sclerotium rolfsii, the causative agent of southern blight of Zea mays L (Baby corn)--in vitro studies.

    Science.gov (United States)

    Patil, N N; Waghmode, M S; Gaikwad, P S; Gajbhiye, M H; Gunjal, A B; Nawani, N N; Kapadnis, B P

    2014-11-01

    The study was undertaken with the aim of exploring novel and beneficial agro activities of rare actinomycetes like Microbispora sp. V2. The antagonistic activity of Microbispora sp. V2 was evaluated as a biocontrol agents against Sclerotium rolfsii, a soil-borne fungal plant pathogen. The methodology performed for evaluation of biocontrol agent was in vitro evaluation assay which comprised of three tests viz., cellophane overlay technique, seed germination test and Thiram (fungicide) tolerance of Microbispora sp. V2. The isolate was found to inhibit the fungal pathogen Sclerotium rolfsii to 91.43% in cellophane assay. In seed germination assay, Microbispora sp. V2 treated seeds resulted in 25.75% increased germination efficiency, as compared to seeds infected by Sclerotium rolfsii. The isolate Microbispora sp. V2 could tolerate 1000 microg mL(-1) of Thiram (fungicide). The in vitro assay studies proved that Microbispora sp. V2 can be used as antifungal antagonist and thus posses' great potential as biocontrol agent against southern blight caused by Sclerotium rolfsii in Zea mays L (Baby corn) which causes large economical losses.

  11. Total Phenol Content and In Vitro Antioxidant Potential of Helicanthus elastica (Desr. Danser-A Less-explored Indian Mango Mistletoe

    Directory of Open Access Journals (Sweden)

    Koppala Narayana Sunil Kumar

    2014-10-01

    Full Text Available Natural products are an important source of antioxidant molecules like tannins, phenolic compounds, flavonoids, etc., Helicanthus elastica (Desr. Danser (Loranthaceae is one such plant belonging to the category of mistletoe, and grows commonly on the mango trees in India. In the present study, an attempt has been made to assess the antioxidant properties of the plant. Ethanol extract of H. elastica growing on mango tree was studied using different in vitro models. Shade-dried whole plant material was extracted with ethanol by cold percolation. Fifty milligrams of the alcohol extract of H. elastica was weighed and dissolved in 10 ml of methanol. The resultant 5 mg/ml solution was suitably diluted to obtain different concentrations. Total phenol content, reducing power assay, and scavenging of free radicals like nitric oxide, hydroxyl, hydrogen peroxide, and 1,1-diphenyl-2-picrylhydrazyl were studied by standardized in vitro chemical methods using ascorbic acid as the standard. The total phenol content of the plant was found to be 1.89% w/w. The extract showed good reducing power as well as scavenging of free radicals (nitric oxide, hydroxyl, superoxide anion, and hydrogen peroxide at concentrations ranging from 5 to 100 μg/ml. The study revealed the antioxidant potential of H. elastica.

  12. Evaluation of the Potential Nephroprotective and Antimicrobial Effect of Camellia sinensis Leaves versus Hibiscus sabdariffa (In Vivo and In Vitro Studies)

    Science.gov (United States)

    Anwar Ibrahim, Doa'a; Noman Albadani, Rowida

    2014-01-01

    Green tea and hibiscus are widely consumed as traditional beverages in Yemen and some regional countries. They are relatively cheap and the belief is that they improve health state and cure many diseases. The aim of this study was to evaluate the potential protective and antibacterial activity of these two famous plants in vitro through measuring their antibacterial activity and in vivo through measuring nonenzymatic kidney markers dysfunction after induction of nephrotoxicity by gentamicin. Gram positive bacteria like MRSA (methicillin resistant Staphylococcus aureus) were isolated from hospitalized patients' different sources (pus and wound) and Gram negative bacteria including E. coli and P. aeruginosa were used in vitro study. In addition, the efficacy of these plants was assessed in vivo through measuring nonenzymatic kidney markers including S. creatinine and S. urea. Green tea was shown antimicrobial activity against MRSA with inhibition zone 19.67 ± 0.33 mm and MIC 1.25 ± 0.00 mg/mL compared with standard reference (vancomycin) 18.00 ± 0.00 mg/mL. Hibiscus did not exhibit a similar effect. Both Hibiscus- and green tea-treated groups had nephroprotective effects as they reduced the elevation in nonenzymatic kidney markers. We conclude that green tea has dual effects: antimicrobial and nephroprotective. PMID:24949007

  13. The potential role of polyphenols in the modulation of skin cell viability by Aspalathus linearis and Cyclopia spp. herbal tea extracts in vitro.

    Science.gov (United States)

    Magcwebeba, Tandeka Unathi; Riedel, Sylvia; Swanevelder, Sonja; Swart, Pieter; De Beer, Dalene; Joubert, Elizabeth; Andreas Gelderblom, Wentzel Christoffel

    2016-11-01

    The relationship between polyphenol constituents, antioxidant properties of aqueous and methanol extracts of green tea (Camellia sinensis), the herbal teas, rooibos (Aspalathus linearis) and honeybush (Cyclopia spp.), against skin cell viability was investigated in vitro. The effect of extracts, characterised in terms of polyphenol content and antioxidant properties, on cell viability of premalignant, normal and malignant skin cells was determined. Phenolic composition, particularly high levels of potent antioxidants, of rooibos and green tea methanol extracts was associated with a strong reduction in cell viability specifically targeting premalignant cells. In contrast, the aqueous extracts of Cyclopia spp. were more effective in reducing cell viability. This correlated with a relatively high flavanol/proanthocyanidin content and ABTS radical cation scavenging capacity. The major green tea flavanol (epigallocatechin gallate) and rooibos dihydrochalcone (aspalathin) exhibited differential effects against cell viability, while the major honeybush xanthone (mangiferin) and flavanone (hesperidin) lacked any effect presumably due to a cytoprotective effect. The underlying mechanisms against skin cell viability are likely to involve mitochondrial dysfunction resulting from polyphenol-iron interactions. The polyphenol constituents and antioxidant parameters of herbal tea extracts are useful tools to predict their activity against skin cell survival in vitro and potential chemopreventive effects in vivo. © 2016 Royal Pharmaceutical Society.

  14. Exploring the antioxidant potentiality of two food by-products into a topical cream: stability, in vitro and in vivo evaluation.

    Science.gov (United States)

    Rodrigues, F; Sarmento, B; Amaral, M Helena; Oliveira, M Beatriz P P

    2016-01-01

    Coffee silverskin (CS), a food by-product of the coffee roasting industry, has been studied as an active ingredient for skin care products due to its high potential of antioxidant activity and low cytotoxicity. Another food waste used as ingredient with promising characteristics is obtained from Medicago sativa (MS), which antioxidants and isoflavones content is high. The aim of this study is to evaluate and characterize a new body formulation containing two food by-products extracts. Different parameters (such as pH, rheological behavior, color, antioxidant content and microbiological analysis) of a body cream formulation containing by-products (CSMS) and a formulation without extracts (F) were evaluated under a stability study during 180 days at different temperatures. Moreover, the in vitro cell toxicity and the in vivo skin safety and protective effects were also assessed. Formulation showed stable physical properties and antioxidant activity during 180 days of storage. In vitro toxicity was screened in two skin cell lines (fibroblasts and keratinocytes) and any toxicity was reported. The in vivo test carried out showed that, with respect to irritant effects, CSMS formulation can be regarded as safe for topical application and the skin hydratation improved after 30 days of its use. Also, considering the consumer acceptance, more than 90% of volunteers classified it as very pleasant. CSMS formulation is stable and safe for topical use as no adverse and/or side effects were observed during the application period of testing, improving skin protective properties.

  15. Comparison of in vitro erosion potentials between beverages available in the United Kingdom and the United States

    Science.gov (United States)

    Murrell, Sarah; Marshall, Teresa A.; Moynihan, Paula J.; Qian, Fang; Wefel, James S.

    2009-01-01

    Summary Objective Our objective was to compare the physiochemical properties and erosion potentials between beverages available in the UK and the US. Methods The physiochemical properties (pH, titratable acidity and fluoride concentration) and erosion potential on enamel surfaces of beverages available in the UK were compared to similar beverages from the US. Enamel windows were exposed to beverages for 25 hours. Teeth were sectioned through the windows, and lesion depths were defined as the average distance between the original tooth structure and the base of demineralization. Results The pH was lower in UK apple juice, orange juice, Diet Pepsi® and Sprite Zero® (pbeverages (pconcentrations were lower in UK apple juice, orange juice, Coke®, and Diet Coke®, Sprite® and Sprite Zero® (pbeverages available in the UK and the US. PMID:19962418

  16. In vitro screening of inhibition of PPAR-γ activity as a first step in identification of potential breast carcinogens

    DEFF Research Database (Denmark)

    Kopp, Tine Iskov; Lundqvist, J.; Petersen, R. K.

    2015-01-01

    measured in the H295R steroidogenesis assay after incubation with the chemicals. Ethylene glycol, ethyl acetate, and dimethyl sulphoxide inhibited PPAR-γ transactivation in a dose-dependent manner. The inhibitory effect on PPAR-γ was specific for PPAR-γ since the AB domain of PPAR-γ was required...... for the inhibitory effect. In the second step, ethylene glycol significantly increased production of oestradiol by 19% (p ethyl acetate inhibited production of testosterone (p ... followed by a well-established steroidogenesis assay for production of sex hormones in exposed H295 R cells may provide a screening tool for potential breast carcinogens. This initial screening thus identified ethylene glycol and possibly ethyl acetate as potential breast carcinogens....

  17. Transdominant Rev and Protease Mutant Proteins of HIV-SIV as Potential Antiviral Agents in Vitro and in Vivo (AIDS)

    Science.gov (United States)

    1993-10-30

    1577- 1580. 7. Sarver, N., E.M. Cantin , P.S. Chang, J.A. Zaia, P.A. Landne, D.A. Stephens, and J.J Rossi. 1990. Ribozymes as potential anti-HIV-1...activities. Nature (London) 334: 585-591. 16 Chang, P.S., E.M. Cantin , J.A. Zaia, P.A. Landne, D.A. Stephens, N. Sarver, and J.J. Rossi. 1990. Clinical

  18. Mitochondrial network in glioma's invadopodia displays an activated state both in situ and in vitro: potential functional implications.

    Science.gov (United States)

    Arismendi-Morillo, Gabriel; Hoa, Neil T; Ge, Lisheng; Jadus, Martin R

    2012-12-01

    Gliomas are typically characterized by their infiltrative nature, and the prognosis can be linked to the invasive nature of the tumoral cells. Glioblastoma multiforme are very invasive cancers and this contributes to their lethality. The invadopodia are considered essential for their motility. Human glioma cell invadopodia were examined with transmission electron and immunofluorescent microscopy. By electron microscopy, in situ gliomas (fibrillary astrocytoma, anaplastic astrocytoma, glioblastoma multiforme, pilocytic astrocytoma) show mitochondria with a dense matrix condensed configuration, indicating an active state. The mitochondria were frequently in close contact with an extended smooth endoplasmic reticulum displaying an endoplasmic reticulum subfraction associated with mitochondria. Mitochondria were seen within the filopodia that were penetrating into the extracellular matrix. The activated mitochondria and smooth endoplasmic reticulum were also detected within the invadopdia, which was associated microblood vessels. Fluorescent microscopy confirmed that D54 and U251 glioma cells growing in vitro also contained filopodia with mitochondria. The U251 glioma cells' filopodia that penetrated through 1.2-μm pores of transwell chambers also contained mitocondria, suggesting that the mitochondria are actively involved in the invasion process. Migration and invasion of tumor cells requires an increase in cellular motility and involves formation of lamellipodia, protrusions of the plasma membrane, and individual filopodia [ 1 ]. Gliomas are typically characterized by their infiltrative nature, resulting in a poorly demarcated interface between tumor and normal brain tissue. Their poor prognosis can be linked to the invasive nature of these cells. The motility of these tumor cells is correlated with the presence of invadopodia [ 2 ], and, consequently, more insight is necessary into their structural and molecular aspects. Evidence of robust invadopodia activity in

  19. Antioxidant potential, in vitro cytotoxicity and apoptotic effect induced by crude organic extract of Anthracophyllum lateritium against RD sarcoma cells.

    Science.gov (United States)

    Fernando, Dilusha M; Wijesundera, Ravi L C; Soysa, Preethi; de Silva, Dilip; Nanayakkara, Chandrika M

    2015-11-06

    Macrofungi have an established history of use in traditional oriental medicine. Anthracophyllum lateritium is a terrestrial macrofungus found in the dry zone forest reserves in Sri Lanka. Yet there are no scientific reports on bioactive properties of this species. Hence, the current study was aimed at determining the antioxidant potential, in vitro antiproliferative activity and apoptotic effect induced by crude methanolic extract of A. lateritium against RD sarcoma cell line. The crude extract of A. lateritium was dissolved in methanol (MEFCA) and antioxidant activity was evaluated using in vitro assays: inhibition of DPPH (1,1-diphenyl-2-picrylhydrazyl) radical scavenging, ferric ion reducing power and 2-deoxy-D-ribose degradation assay. Total phenol and flavonoid contents of MEFCA were assayed using folin Ciocalteu method and aluminium chloride colorimetric method. In vitro cytotoxicity was determined using MTT assay against RD cells after 24 h exposure to MEFCA. Ethidium bromide/ acridine orange staining, DNA fragmentation and protein synthesis experiments were used to study the apoptotic features and antiproliferative activities of the treated cells. Glutathione assay and griess nitrite assay were used to analyze the reduced glutathione content and liberation of nitric oxide from apoptotic cells. MEFCA showed promising antioxidant activity with EC50 values of 8.00 ± 0.35 μg/mL for DPPH scavenging and 83.33 ± 0.45 μg/mL for 2-deoxy-D-ribose degradation assay. The phenolic content was 265.15 ± 0.46 of (w/w) % of Gallic acid equivalents and flavonoid content was 173.01 ± 0.35 of (w/w) % of Epigallocatechingallate. A. lateritium showed strong in vitro cytotoxic activity with an EC50 of 18.80 ± 4.83 μg/mL for MTT assay against RD cells. Ethidium bromide/acridine orange staining and DNA fragmentation indicated the apoptotic features of treated cells. Protein levels showed a dose dependent decrease supporting the fact that A. lateritium

  20. Tumour cell derived effects on monocyte/macrophage polarization and function and modulatory potential of Viscum album lipophilic extract in vitro.

    Science.gov (United States)

    Estko, Myriam; Baumgartner, Stephan; Urech, Konrad; Kunz, Matthias; Regueiro, Ursula; Heusser, Peter; Weissenstein, Ulrike

    2015-04-24

    Macrophages are highly versatile cells that play an important role in tumour microenvironment. Tumour associated macrophages (TAMs) have been linked to both, good or bad prognosis of several cancer types depending on their number, composition and polarization. Viscum album lipophilic extract (VALE) contains several pentacyclic triterpenes known to modulate the activity of monocytes and other immune cells and to exhibit anticancer properties. In our in vitro study, we investigated the effect of tumour cell lines on macrophage polarization and monocyte chemotactic transmigration and examined the modulatory potential of VALE and its predominant triterpene oleanolic acid (OA). Human peripheral blood monocytes were differentiated into monocyte derived macrophages (MDM) using M-CSF and polarized into M1 by IFN-γ and LPS and into M2 macrophages by IL-4 and IL-13 or by co-culture with two different tumour cell lines. Polarized macrophages were subsequently treated with VALE or OA. Phenotypic markers and cytokines were assessed by flow cytometry and immunoanalysis. Migration of human peripheral blood monocytes induced by monocyte chemotactic protein-1 (MCP-1) or supernatants of different tumour cell lines under the influence of VALE or OA was measured in a chemotaxis transmigration assay. In vitro polarized M1 and M2 type macrophages revealed specific phenotypic patterns and tumour cell co-cultured MDM displayed ambiguous phenotypes with M1 as well as M2 associated markers. VALE and OA showed modest influence on cell surface marker profile and cytokine expression of tumour cell co-cultured macrophages. All tumour cell supernatants markedly enhanced the migratory activity of monocytes. VALE and OA significantly inhibited MCP-1 induced monocyte transmigration, whereas monocyte migration initiated by tumour cell derived supernatants was not affected. In our study we reconfirmed that co-culture with different tumour cell lines can result in a mixed macrophage phenotype with M1

  1. Enhancing antioxidant activity, microbial and sensory quality of mango (Mangifera indica L.) juice by γ-irradiation and its in vitro radioprotective potential.

    Science.gov (United States)

    Naresh, Kondapalli; Varakumar, Sadineni; Variyar, Prasad Shekhar; Sharma, Arun; Reddy, Obulam Vijaya Sarathi

    2015-07-01

    Gamma irradiation is an effective method currently being used for microbial decontamination and insect disinfestations of foods. In the present study, mango (Mangifera indica L.) juice was irradiated at doses of 0, 1.0, 3.0 and 5.0 kGy and microbial load, total polyphenols, flavonoids, ascorbic acid content, antioxidant activities, colour and sensory properties were evaluated immediately after irradiation and also during storage. Microbiological assay of the fresh and stored mango juice showed better quality after γ-irradiation. The total polyphenols and flavonoids were significantly (p mango juice without any adverse changes in the sensory qualities. Significant in vitro plasmid DNA protection was observed in the presence of mango juice against radiation induced damage, even at the dose of 5 kGy. This study confirmed the potential of γ-irradiation as a method for microbial decontamination and improving the quality of the mango juice without compromising on the sensory attributes.

  2. Antagonistic potentiality of Trichoderma harzianum towards seed-borne fungal pathogens of winter wheat cv. Protiva in vitro and in vivo.

    Science.gov (United States)

    Hasan, M M; Rahman, S M E; Kim, Gwang-Hee; Abdallah, Elgorban; Oh, Deog-Hwan

    2012-05-01

    The antagonistic effect of Trichoderma harzianum on a range of seed-borne fungal pathogens of wheat (viz. Fusarium graminearum, Bipolaris sorokiniana, Aspergillus spp., and Penicillium spp.) was assessed. The potential of T. harzianum as a biocontrol agent was tested in vitro and under field conditions. Coculture of the pathogens and Trichoderma under laboratory conditions clearly showed dominance of T. harzianum. Under natural conditions, biocontrol effects were also obtained against the test fungi. One month after sowing, field emergence (plant stand) was increased by 15.93% over that obtained with the control treatment, and seedling infection was reduced significantly. Leaf blight severity was decreased from 22 to 11 at the heading stage, 35 to 31 at the flowering stage, and 86 to 74 at the grain filling stage. At harvest, the number of tillers per plant was increased by 50%, the yield was increased by 31.58%, and the 1,000-seed weight was increased by 21%.

  3. Quartz-Containing Ceramic Dusts: In vitro screening of the cytotoxic, genotoxic and pro-inflammatory potential of 5 factory samples

    Science.gov (United States)

    Ziemann, C.; Jackson, P.; Brown, R.; Attik, G.; Rihn, B. H.; Creutzenberg, O.

    2009-02-01

    Inhalation of some respirable crystalline silica (MMAD RF>TG>Ti>BR>TC>Al2O3. DNA-damage was maximal for BR and TI followed by DQ12>TG>TC>RF>Al2O3. All dusts induced PGE2-liberation (DQ12>BR>TC>TG>Ti>RF>Al2O3) at 50μg/cm2 (4h), but TNF-a mRNA (10μg/cm2, 24h) was only increased by DQ12, TG (quartz-dependently), and TC. In conclusion, these in vitro tests were an adequate approach to screen the toxic potential of quartz-containing ceramic dusts, but the quartz-content was too low to differentiate the various quartz-varieties.

  4. Effects of a very low dose rate of chronic ionizing radiation on the division potential of human embryonic lung fibroblasts in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Croute, F.; Vidal, S.; Soleilhavoup, J.P.; Vincent, C.; Serre, G.; Planel, H.

    1986-01-01

    Among the various parameters that are supposed to play a role in aging at the cellular level, the ''free radical theory'' involves biochemical modifications that can be induced by radiation. Human embryonic lung fibroblasts were serially subcultivated at low density under chronic low dose rate irradiation (40 mrad/day) and in a normal environment. Irradiation increases cell attachment and the population doubling/day throughout their entire in vitro lifespan. Consequently, the doubling potential reached by irradiated cells was higher than that of control cultures. Finally, the total number of cells produced under chronic irradiation was 8-14 times higher than in a normal environment. These results are discussed with respect to the increased enzymatic activities (superoxidismutase, catalase, glutathion-reductase, G6PD) found in some irradiated organisms.

  5. Assessment of the antidiabetic potential of selected medicinal plants using in vitro bioassays of muscle glucose transport and liver glucose production

    DEFF Research Database (Denmark)

    Beidokhti, M N; Sanchez Villavicencio, M L; Eid, H M

    2016-01-01

    by skeletal muscle cells are the principal contributors to the associated hyperglycemic state. The aim of this study was to assess the antidiabetic potential of five medicinal plant extracts using in vitro cell based assays targeting glucose uptake in C2C12 skeletal muscle cells [1] and glucose-6-phosphatase...... activity (G6Pase) [2] in rat hepatoma H4IIE. Cells were treated for 18h with maximal non-toxic concentrations (50 µg/mL) of the ethanolic extract of Psidium guajava (leaf and bark), Phyllanthus niruri (aerial parts), Eugenia jambolana (dried fruit) and Rhizophora mucronata (bark), which were determined...... by the lactate dehydrogenase (LDH) cytotoxicity assay. None of the extracts were able to reduce G6Pase activity. In contrast, one plant extract (P. guajava leaf extract) was found to significantly increase deoxyglucose uptake in C2C12 muscle cells (161%, p ≤0.001), to levels higher that of the positive control...

  6. The 10 basic requirements for a scientific paper reporting antioxidant, antimutagenic or anticarcinogenic potential of test substances in in vitro experiments and animal studies in vivo

    DEFF Research Database (Denmark)

    Verhagen, H.; Aruoma, O.I.; van Delft, J.H.M.

    2003-01-01

    , provided they can be justified on scientific grounds. The 10 basic requirements for a scientific paper reporting antioxidant, antimutagenic or anticarcinogenic potential of test substances in in vitro experiments and animal studies in vivo concern the following areas: (1) Hypothesis-driven study design; (2......There is increasing evidence that chemicals/test substances cannot only have adverse effects, but that there are many substances that can (also) have a beneficial effect on health. As this journal regularly publishes papers in this area and has every intention in continuing to do so in the near...... future, it has become essential that studies reported in this journal reflect an adequate level of scientific scrutiny. Therefore a set of essential characteristics of studies has been defined. These basic requirements are default properties rather than non-negotiables: deviations are possible and useful...

  7. Loss of Local Astrocyte Support Disrupts Action Potential Propagation and Glutamate Release Synchrony from Unmyelinated Hippocampal Axon Terminals In Vitro.

    Science.gov (United States)

    Sobieski, Courtney; Jiang, Xiaoping; Crawford, Devon C; Mennerick, Steven

    2015-08-05

    Neuron-astrocyte interactions are critical for proper CNS development and function. Astrocytes secrete factors that are pivotal for synaptic development and function, neuronal metabolism, and neuronal survival. Our understanding of this relationship, however, remains incomplete due to technical hurdles that have prevented the removal of astrocytes from neuronal circuits without changing other important conditions. Here we overcame this obstacle by growing solitary rat hippocampal neurons on microcultures that were comprised of either an astrocyte bed (+astrocyte) or a collagen bed (-astrocyte) within the same culture dish. -Astrocyte autaptic evoked EPSCs, but not IPSCs, displayed an altered temporal profile, which included increased synaptic delay, increased time to peak, and severe glutamate release asynchrony, distinct from previously described quantal asynchrony. Although we observed minimal alteration of the somatically recorded action potential waveform, action potential propagation was altered. We observed a longer latency between somatic initiation and arrival at distal locations, which likely explains asynchronous EPSC peaks, and we observed broadening of the axonal spike, which likely underlies changes to evoked EPSC onset. No apparent changes in axon structure were observed, suggesting altered axonal excitability. In conclusion, we propose that local astrocyte support has an unappreciated role in maintaining glutamate release synchrony by disturbing axonal signal propagation. Certain glial cell types (oligodendrocytes, Schwann cells) facilitate the propagation of neuronal electrical signals, but a role for astrocytes has not been identified despite many other functions of astrocytes in supporting and modulating neuronal signaling. Under identical global conditions, we cultured neurons with or without local astrocyte support. Without local astrocytes, glutamate transmission was desynchronized by an alteration of the waveform and arrival time of axonal

  8. In vitro antioxidant and antiproliferative potential of medicinal plants used in traditional Indian medicine to treat cancer.

    Science.gov (United States)

    Baskar, Arul-Albert; Al Numair, Khalid S; Alsaif, Mohammed A; Ignacimuthu, Savarimuthu

    2012-01-01

    The goal of this study was to evaluate the antioxidant and antiproliferative activities of 10 traditional medicinal plants, Asclepias curassavica, Ophiorrhiza mungos Linn., Cynodon dactylon (L.) Pers, Costus speciosus (J. Koenig.) Smith Costaceae, Achyranthes aspera L., Amaranthus tristis Roxb., Blepharis maderaspatensis L., Merremia emerginata Hall.f., Aegle marmelos Corr., and Tabernaemontana heyneana Wall., used in the traditional Indian system of medicine as a cure for cancer. The present study focuses on the anticancer potential of traditional medicinal plants to induce apoptosis in cancer cell lines. Plants were sequentially extracted with hexane, ethyl acetate, and methanol. The extract was concentrated to yield the crude extract, which was tested for antioxidant activity using 1,1-diphenyl-2-picrylhydrazyl, nitric oxide and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assays on four cancer cell lines and a normal cell line. The anticancer potential of cytotoxic extracts was determined by the Annexin-fluorescein isothiocyanate-conjugated assay in human colon adenocarcinoma cell lines (COLO 320 DM). All the tested extracts showed significant antioxidant and antiproliferative activities in a concentration- and time-dependant manner in the following descending order: A. curassavica > C. dactylon > C. speciosus root > A. tristis > M. emarginata > O. mungos > T. Heyneana > B. maderaspatensis > A. marmelos > A. aspera. The results of the present study support the need of further studies to isolate potential anticancer drug with cancer cell-specific cytotoxicity. Additionally, the study supports the anticancer property of medicinal plants used in the traditional Indian medicine system and further evaluation of the selected medicinal plants for an effective anticancer drug with minimal side effects.

  9. In silico and In vitro evaluation of the anti-inflammatory potential of Centratherum punctatum Cass-A.

    Science.gov (United States)

    Shankaran, Krithika S; Ganai, Shabir Ahmad; K P, Arun; P, Brindha; Mahadevan, Vijayalakshmi

    2017-03-01

    Centratherum punctatum Cass., a herb belonging to the family Asteraceae has been traditionally used as a curative against diverse disorders like inflammation, tumor, depression, and hypertension. Though the medicinal properties of this plant have been attributed to the presence of flavonoids, glucosides, alkaloids, Vitamin C, etc., the molecular constituents of this plant and of the flavonoids that contribute to its medicinal activity have not been explored yet. This work attempts to evaluate the potential of Centratherum punctatum extract as an anti-inflammatory agent. Ethanolic extracts of Centratherum punctatum analyzed by High Performance Thin Layer Chromatography (HPTLC) and Liquid Chromatography-Mass Spectrometry (LC-MS/MS) identified the presence of the flavones kaempferol, glycoside Isorhamnetin-3-O-rutinoside, and kaempferol-3-glucoside. The plant extract exhibited anti-oxidant property as confirmed by DPPH assay and IC50 value of 271.6 μg/mL during inhibition of protein denaturation, 186.8 μg/mL during RBC membrane stabilization, and 278.2 μg/mL for proteinase inhibition. Membrane stabilizing functions of flavones and flavones glycosides validated the anti-inflammatory potential of the extract. In silico evaluation using a rigorous molecular docking protocol with receptors of Cox2, TNF-α, Interleukin 1β convertase, and Histamine H1 predicted high binding affinity of the isoflavones and isoflavone glycosides of Centratherum punctatum Cass. The interactions have also been shown to compare well with that of known drugs valdecoxib through Gln178, His342, and Gly340, desloratadine (through Lys191 and Thr194) and belnacasin (through Asp288 and Gly287) proven to function through the anti-inflammatory pathway. This work establishes the anti-inflammatory potential of Centratherum punctatum Cass. extract as an alternative to existing therapeutic approach to inflammation through a systematic in silico approach supplementing the findings.

  10. The skin-depigmenting potential of Paeonia lactiflora root extract and paeoniflorin: in vitro evaluation using reconstructed pigmented human epidermis.

    Science.gov (United States)

    Qiu, J; Chen, M; Liu, J; Huang, X; Chen, J; Zhou, L; Ma, J; Sextius, P; Pena, A-M; Cai, Z; Jeulin, S

    2016-10-01

    The roots of the herb Paeonia lactiflora ('White Peony') are used in association with other herbs in traditional clinical cosmetic practice in China as oral treatment for skin pigmentary disorders, such as brown or dark pigmentary spots. However, the skin-depigmenting potential of Paeonia lactiflora root extract and its main ingredient paeoniflorin has been scarcely investigated by topical application. The purpose of this study was to evaluate the efficacy of Paeonia lactiflora root extract and paeoniflorin as skin whitening agent in cosmetic application. Paeonia lactiflora root extract (containing 53.25% of paeoniflorin) and paeoniflorin (97% purity) were applied topically on reconstructed pigmented human epidermis model, a three-dimensional (3D) human skin equivalent, showing morphological and functional characteristics similar to those of in vivo human skin. Two specific methods were used for quantifying melanin inside the reconstructed pigmented epidermis: Fontana-Masson staining (2D quantification) and multiphoton microscopy (3D quantification). Compared to vehicle (dimethyl sulfoxide DMSO), a significant decrease in 2D and 3D melanin content was observed after topical application on reconstructed pigmented epidermis of Paeonia lactiflora extract at 300 μg mL(-1) (-28% and -27%, respectively) and paeoniflorin at 120 μg mL(-1) /250 μM (-30% and -23%, respectively), which is in the same order of magnitude as the positive reference 4-n-butylresorcinol at 83 μg mL(-1) /500 μM (-26% and -40%, respectively). These results demonstrate, for the first time, the depigmenting potential of paeoniflorin and thus the potential interest of using Paeonia lactiflora root extracts containing paeoniflorin in cosmetic or dermatological applications for reducing the severity of some hyperpigmented skin disorders. © 2016 Society of Cosmetic Scientists and the Société Française de Cosmétologie.

  11. In vitro toxicity of particulate matter (PM collected at different sites in the Netherlands is associated with PM composition, size fraction and oxidative potential - the RAPTES project

    Directory of Open Access Journals (Sweden)

    Lebret Erik

    2011-09-01

    Full Text Available Abstract Background Ambient particulate matter (PM exposure is associated with respiratory and cardiovascular morbidity and mortality. To what extent such effects are different for PM obtained from different sources or locations is still unclear. This study investigated the in vitro toxicity of ambient PM collected at different sites in the Netherlands in relation to PM composition and oxidative potential. Method PM was sampled at eight sites: three traffic sites, an underground train station, as well as a harbor, farm, steelworks, and urban background location. Coarse (2.5-10 μm, fine (2. Following overnight incubation, MTT-reduction activity (a measure of metabolic activity and the release of pro-inflammatory markers (Tumor Necrosis Factor-alpha, TNF-α; Interleukin-6, IL-6; Macrophage Inflammatory Protein-2, MIP-2 were measured. The oxidative potential and the endotoxin content of each PM sample were determined in a DTT- and LAL-assay respectively. Multiple linear regression was used to assess the relationship between the cellular responses and PM characteristics: concentration, site, size fraction, oxidative potential and endotoxin content. Results Most PM samples induced a concentration-dependent decrease in MTT-reduction activity and an increase in pro-inflammatory markers with the exception of the urban background and stop & go traffic samples. Fine and qUF samples of traffic locations, characterized by a high concentration of elemental and organic carbon, induced the highest pro-inflammatory activity. The pro-inflammatory response to coarse samples was associated with the endotoxin level, which was found to increase dramatically during a three-day sample concentration procedure in the laboratory. The underground samples, characterized by a high content of transition metals, showed the largest decrease in MTT-reduction activity. PM size fraction was not related to MTT-reduction activity, whereas there was a statistically significant

  12. The quaternary lidocaine derivative, QX-314, exerts biphasic effects on transient receptor potential vanilloid subtype 1 channels in vitro

    DEFF Research Database (Denmark)

    Rivera-Acevedo, Ricardo E; Pless, Stephan Alexander; Ahern, Christopher A

    2011-01-01

    BACKGROUND: Transient receptor potential vanilloid subfamily member 1 (TRPV1) channels are important integrators of noxious stimuli with pronounced expression in nociceptive neurons. The experimental local anesthetic, QX-314, a quaternary (i.e., permanently charged) lidocaine derivative, recently...... concentrations (less than 1 mM), QX-314 potently inhibited capsaicin-evoked TRPV1 currents with an IC₅₀ of 8.0 ± 0.6 μM. CONCLUSIONS: The results of this study show that the quaternary lidocaine derivative QX-314 exerts biphasic effects on TRPV1 channels, inhibiting capsaicin-evoked TRPV1 currents at lower...

  13. In vivo and in vitro immunomodulatory potential of swertiamarin isolated from Enicostema axillare (Lam.) A. Raynal that acts as an anti-inflammatory agent.

    Science.gov (United States)

    Saravanan, S; Pandikumar, P; Prakash Babu, N; Hairul Islam, V I; Thirugnanasambantham, K; Gabriel Paulraj, M; Balakrishna, K; Ignacimuthu, S

    2014-10-01

    Swertiamarin is a secoiridoid glycoside found in Enicostema axillare (Lam) A. Raynal, a medicinal plant used as a depurative in the Indian system of traditional medicine. The present study evaluated the immunomodulatory activity of isolated swertiamarin. In vivo immunomodulatory activity of swertiamarin (2, 5, and 10 mg/kg b.w.) was evaluated in a model of sheep red blood cells (SRBC) by assessing its effect on organ weight, hemagglutinating antibody titer (HA), plaque-forming cells (PFC), quantitative hemolysis of SRBC, and delayed type hypersensitivity (DTH). In vitro immunomodulatory potential was studied on isolated splenocytes, neutrophils, and peritoneal macrophages. In silico immunomodulatory effects were evaluated by docking of swertiamarin on proinflammatory cytokines to confirm its potential. In in vivo studies, the animals treated with swertiamarin showed a significant (P ≤ 0.05) increase in antibody titer, plaque-forming cells, and also in weight of the thymus and spleen. A decreased response to DTH reaction was recorded with the treatment of swertiamarin. In in vitro studies, treatment with swertiamarin modulated the messenger RNA (mRNA) and protein expression of IFN-γ, IL-10, and IL-4 significantly (P ≤ 0.05) and also favored Th2-mediated response on concanavalin A (Con A)-induced splenocytes. The compound inhibited the release of free radicals significantly (P ≤ 0.05) in phytohemagglutinin (PHA)-induced neutrophils and also ameliorated the mRNA and protein expression of proinflammatory cytokines (TNF-α, IL-1β, and IL-6) in lipopolysaccharide (LPS)-induced macrophages. In in silico, the best docked pose of swertiamarin with the target proteins (TNF-α, IL-1β, and IL-6) was confirmed that swertiamarin acted as an anti-inflammatory mediator.

  14. Phenolic profiling and therapeutic potential of local flora of Azad Kashmir; In vitro enzyme inhibition and antioxidant

    Directory of Open Access Journals (Sweden)

    Raza Muhammad Asam

    2017-12-01

    Full Text Available The current study supports the phytochemical screening, evaluation of antioxidant and enzyme inhibition potential and correlations between antioxidant activities and phenolics of Rumex dentatus (Family: Polygonaceae, Mentha spicata (Family: Lamiaceae, Withania somnifera (Family: Solanaceae, Nerium indicum (Family: Apocynaceae and Artemisia scoparia (Family: Asteraceae. The herbal materials were extracted in ethanol (90% and partitioned between several solvents based on polarities. Total phenols were determined with FC method and ranged 21.33 ± 1.53 - 355.67 ± 6.03 mg GAE/ mg of the extract. Antioxidant activities (DPPH, total iron reducing capacity, phosphomolybdate assay & FRAP and enzyme inhibition potential (Protease, AChE & BChE were performed by the standard protocols. The results showed that all extracts exhibited significant DPPH activity ranging from 12.67 ± 2.08 - 92.67 ± 1.53%. The extracts that were active in DPPH activity also potrayed marvelous FRAP, total iron reducing and phosphomolybdate values. Correlation studies of antioxidant activities and the content of phenolic compounds in plant materials exhibited positive correlation between them. The outcome of enzyme inhibition activity exhibited that about 80% of the fractions under surveillance plants intimated more than 50% inhibition. Isolation of bioactive compounds from these plants is in progress.

  15. Apoptosis induced by low-intensity ultrasound in vitro: Alteration of protein profile and potential molecular mechanism

    Science.gov (United States)

    Feng, Yi; Wan, Mingxi

    2017-03-01

    To analyze the potential mechanism related to the apoptosis induced by low intensity focused ultrasound, comparative proteomic method was introduced in the study. After ultrasound irradiation (3.0 W/cm2, 1 minute, 6 hours incubation post-irradiation), the human SMMC-7721 hepatocarcinoma cells were stained by trypan blue to detect the morphologic changes, and then the percentage of early apoptosis were tested by the flow cytometry with double staining of FITC-labelled Annexin V/Propidium iodide. Two-dimensional SDS polyacrylamide gel electrophoresis was used to get the protein profile and some proteins differently expressed after ultrasound irradiation were identified by MALDI-TOF mass spectrometry. It's proved early apoptosis of cells were induced by low intentisy focused ultrasound. After ultrasound irradiation, the expressing characteristics of several proteins changed, in which protein p53 and heat shock proteins are associated with apoptosis initiation. It is suggested that the low-intensity ultrasound-induced apoptotic cancer therapy has the potential application via understanding its relevant molecular signaling and key proteins. Moreover, the comparative proteomic method is proved to be useful to supply information about the protein expression to analyze the metabolic processes related to bio-effects of biomedical ultrasound.

  16. Potassium Iodide Potentiates Antimicrobial Photodynamic Inactivation Mediated by Rose Bengal in In Vitro and In Vivo Studies.

    Science.gov (United States)

    Wen, Xiang; Zhang, Xiaoshen; Szewczyk, Grzegorz; El-Hussein, Ahmed; Huang, Ying-Ying; Sarna, Tadeusz; Hamblin, Michael R

    2017-07-01

    Rose bengal (RB) is a halogenated xanthene dye that has been used to mediate antimicrobial photodynamic inactivation for several years. While RB is highly active against Gram-positive bacteria, it is largely inactive in killing Gram-negative bacteria. We have discovered that addition of the nontoxic salt potassium iodide (100 mM) potentiates green light (540-nm)-mediated killing by up to 6 extra logs with the Gram-negative bacteria Escherichia coli and Pseudomonas aeruginosa, the Gram-positive bacterium methicillin-resistant Staphylococcus aureus, and the fungal yeast Candida albicans The mechanism is proposed to be singlet oxygen addition to iodide anion to form peroxyiodide, which decomposes into radicals and, finally, forms hydrogen peroxide and molecular iodine. The effects of these different bactericidal species can be teased apart by comparing the levels of killing achieved in three different scenarios: (i) cells, RB, and KI are mixed together and then illuminated with green light; (ii) cells and RB are centrifuged, and then KI is added and the mixture is illuminated with green light; and (iii) RB and KI are illuminated with green light, and then cells are added after illumination with the light. We also showed that KI could potentiate RB photodynamic therapy in a mouse model of skin abrasions infected with bioluminescent P. aeruginosa. Copyright © 2017 American Society for Microbiology.

  17. Caries-preventive potential of an adhesive patch after thermomechanical loading--a microbial-based in vitro study.

    Science.gov (United States)

    Schmidlin, Patrick R; Klück, Ilja; Zimmermann, Jörg; Roulet, Jean-François; Seemann, Rainer

    2006-02-01

    To assess the enamel-protective potential of a newly devised adhesive patch for smooth enamel sealing. Approximal surfaces of 30 extracted molars were divided into three areas: the buccal thirds were treated with a flowable composite (Tetric Flow, Ivoclar Vivadent) and served as negative control sites, the lingual thirds were left untreated and served as positive control sites, and the middle thirds served as the test areas. This was sealed with either 1. a twofold application of an unfilled resin (Heliobond, Ivoclar Vivadent), 2. an adhesive prototype patch (Ivoclar Vivadent), or 3. an adhesive patch in combination with a flowable composite. After thermomechanical loading and demineralization in a microbial-based artificial caries chamber, demineralization depth was assessed using a confocal laser scanning microscope. Negative control sites treated with the flowable composite showed no signs of demineralization. Areas treated with the patch showed no signs of demineralization, irrespective of whether it was used in combination with a flowable composite or directly bonded to the enamel. Caries-like lesions in untreated sites showed a mean depth of 134.3 +/- 35.9 microm. Demineralization depth at sites treated with the unfilled resin was 76.2 +/- 26.5 microm (p = 0.023). Under the conditions of the present study, the adhesive patch under investigation completely protected the underlying enamel from demineralization. This merits further study to assess its potential as an interproximal sealant.

  18. The broad-spectrum antimycobacterial activities of phenothiazines, InVitro: somewhere in all of this there may be patentable potentials.

    NARCIS (Netherlands)

    Ingen, J. van

    2011-01-01

    The phenothiazines are neuroleptic drugs that have long been known to have antimycobacterial activity, in vitro. Of the various commercially available phenothiazines, thioridazine, chlorpromazine and trifluoperazine are most active against mycobacteria, in vitro. Their MICs for Mycobacterium

  19. Quartz-Containing Ceramic Dusts: In vitro screening of the cytotoxic, genotoxic and pro-inflammatory potential of 5 factory samples

    Energy Technology Data Exchange (ETDEWEB)

    Ziemann, C; Creutzenberg, O [Fraunhofer Institute of Toxicology and Experimental Medicine, Hannover (Germany); Jackson, P [CERAM Research Ltd., Stoke-on-Trent (United Kingdom); Brown, R [TOXSERVICES, Stretton (United Kingdom); Attik, G; Rihn, B H, E-mail: christina.ziemann@item.fraunhofer.d [Nancy-University, Faculte de Pharmacie, Nancy (France)

    2009-02-01

    Inhalation of some respirable crystalline silica (MMAD < approx. 4 mum) leads to inflammatory and malignant diseases. Comprehensive physicochemical/biological data and suitable in vitro/in vivo methods may distinguish between more or less harmful quartz-varieties. Within the European Collective Research Project SILICERAM an in vitro screening battery was established to evaluate cytotoxicity (LDH-release, MTT-assay), genotoxicity (Comet-assay) and pro-inflammatory potential (PGE{sub 2}-liberation, TNF-a mRNA expression) of 5 respirable quartz-containing dusts from ceramic plants: brickwork (BR: 7.8% quartz), tableware granulate/cast (TG/TC: 5.8%/3.1%), tiles (TI: 8.1%), refractory (RF: 3.7%). DQ12 (87% a-quartz) and Al{sub 2}O{sub 3} were used as particulate positive and negative controls, respectively. Primary rat alveolar macrophages and the macrophage cell line NR8383 served as model systems. Aluminium lactate was used as inhibitor of biologically active silica, enabling differentiation of silica- and non-specific toxicity. At 200mug/cm{sup 2} (2h) the dusts did not alter significantly LDH-release (except TC), whereas the MTT-assay demonstrated the mainly quartz-independent rank order: DQ12>RF>TG>Ti>BR>TC>Al{sub 2}O{sub 3}. DNA-damage was maximal for BR and TI followed by DQ12>TG>TC>RF>Al{sub 2}O{sub 3}. All dusts induced PGE{sub 2}-liberation (DQ12>BR>TC>TG>Ti>RF>Al{sub 2}O{sub 3}) at 50mug/cm{sup 2} (4h), but TNF-a mRNA (10mug/cm{sup 2}, 24h) was only increased by DQ12, TG (quartz-dependently), and TC. In conclusion, these in vitro tests were an adequate approach to screen the toxic potential of quartz-containing ceramic dusts, but the quartz-content was too low to differentiate the various quartz-varieties.

  20. In-vitro evaluation of bioactive compounds, anti-oxidant, lipid peroxidation and lipoxygenase inhibitory potential of Citrus karna L. peel extract.

    Science.gov (United States)

    Singh, Jagdeep; Sood, Shailja; Muthuraman, Arunachalam

    2014-01-01

    Many medicinal plants have been studied for their antioxidant and their pharmacological activity. Citrus species were well documented as potential antioxidant based therapy for cancer, inflammation, heart disease. Citrus seeds and peels have been shown to possess high antioxidant activity. Therefore, the present study to explore the antioxidant and lipid peroxidation & lipoxygenase inhibitory action of Citrus karna peel extracts were undertaken. Extraction was performed with different solvents of increasing polarity and yield was calculated. Peel extracts were also analyzed for the presence of phenols, flavonoids, vitamin C, and carotenoids. Then the Citrus karna peel extracts were evaluated for the antioxidant and lipid peroxidation & lipoxygenase inhibitory action In-Vitro. In further, the quantification of hesperidin and naringin was carried out by HPLC-DAD method. The results indicated the presence of phenols, flavonoids, vitamin C, carotenoids, hesperidin and naringin in Citrus karna peel extracts with maximum yield of (3.91% w/w). Citrus karna peel extracts were also found to have potential antioxidant and lipid peroxidation & lipoxygenase inhibitory action. Therefore, Citrus karna peel extracts could be used for the future therapeutic medicine due to presence of potential bioactive compounds.

  1. Decreased proliferative, migrative and neuro-differentiative potential of postnatal rat enteric neural crest-derived cells during culture in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Yu, Hui [Department of Pediatric Surgery, the Second Affiliated Hospital, Xi’an Jiaotong University, No 157, Xi Wu Road, Xi’an 710004, Shaanxi (China); Institute of Neurobiology, Environment and Genes Related to Diseases Key Laboratory of Chinese Ministry of Education, Xi’an Jiaotong University, No 96, Yan Ta Xi Road, Xi’an 710061, Shaanxi (China); Pan, Wei-Kang; Zheng, Bai-Jun; Wang, Huai-Jie [Department of Pediatric Surgery, the Second Affiliated Hospital, Xi’an Jiaotong University, No 157, Xi Wu Road, Xi’an 710004, Shaanxi (China); Chen, Xin-Lin; Liu, Yong [Institute of Neurobiology, Environment and Genes Related to Diseases Key Laboratory of Chinese Ministry of Education, Xi’an Jiaotong University, No 96, Yan Ta Xi Road, Xi’an 710061, Shaanxi (China); Gao, Ya, E-mail: ygao@mail.xjtu.edu.cn [Department of Pediatric Surgery, the Second Affiliated Hospital, Xi’an Jiaotong University, No 157, Xi Wu Road, Xi’an 710004, Shaanxi (China)

    2016-05-01

    A growing body of evidence supports the potential use of enteric neural crest-derived cells (ENCCs) as a cell replacement therapy for Hirschsprung's disease. Based on previous observations of robust propagation of primary ENCCs, as opposed to their progeny, it is suggested that their therapeutic potential after in vitro expansion may be restricted. We therefore examined the growth and differentiation activities and phenotypic characteristics of continuous ENCC cultures. ENCCs were isolated from the intestines of postnatal rats and were identified using an immunocytochemical approach. During continuous ENCC culture expansion, proliferation, migration, apoptosis, and differentiation potentials were monitored. The Cell Counting Kit-8 was used for assessment of ENCC vitality, Transwell inserts for cell migration, immunocytochemistry for cell counts and identification, and flow cytometry for apoptosis. Over six continuous generations, ENCC proliferation potency was reduced and with prolonged culture, the ratio of migratory ENCCs was decreased. The percentage of apoptosis showed an upward trend with prolonged intragenerational culture, but showed a downward trend with prolonged culture of combined generations. Furthermore, the percentage of peripherin{sup +} cells decreased whilst the percentage of GFAP{sup +} cells increased with age. The results demonstrated that alterations in ENCC growth characteristics occur with increased culture time, which may partially account for the poor results of proposed cell therapies. - Highlights: • Differences were identified between primary and daughter ENCCs. • Daughter ENCCs had reduced proliferation, migration and differentiation. • Daughter ENCCs also had increased apoptosis. • These altered characteristics warrant further investigation.

  2. Combining in vitro, in vivo and in silico approaches to evaluate nutraceutical potentials and chemical fingerprints of Moltkia aurea and Moltkia coerulea.

    Science.gov (United States)

    Zengin, Gokhan; Ceylan, Ramazan; Katanić, Jelena; Mollica, Adriano; Aktumsek, Abdurrahman; Boroja, Tatjana; Matić, Sanja; Mihailović, Vladimir; Stanić, Snežana; Aumeeruddy-Elalfi, Zaahira; Yilmaz, Mustafa Abdullah; Mahomoodally, Mohamad Fawzi

    2017-09-01

    Methanolic extracts of Moltkia aurea Boiss. (MA) and Moltkia coerulea (Willd.) Lehm. (MC) were investigated for their antioxidant capacity and enzymatic inhibitory potential against acetylcholinesterase, butyrylcholinesterase, α-amylase, α-glucosidase, and tyrosinase in vitro. MA and MC were also explored for their antimicrobial effect, as well as for their possible genotoxic/antigenotoxic potential on Drosophila melanogaster in vivo. The total bioactive components (phenolic (TPC) and flavonoid contents (TFC)) were determined and liquid chromatography-tandem mass spectrometry (LC-MS/MS) metabolite profiling of MA and MC appraised. The plausible docking poses of bioactive compounds to key enzymes were further studied using molecular modelling approach. MA proved to be a better antioxidant with higher TPC and TFC compared to MC. Protocatechuic acid, rutin, hesperidin and malic acid were the most abundant in these extracts. Both MA and MC exhibited antigenotoxic potential with a %R in DNA damage of 60.90 and 53.14% respectively. The docking studies revealed that rutin, hesperidin, and rosmarinic acid have the best scores for all the enzymes tested. MA and MC were found to be rich in phytochemicals with potent antioxidant, antimicrobial, and antigenotoxic activities that can be further studied for the management of neurodegenerative complications, diabetes, and hyperpigmentation. Copyright © 2017 Elsevier Ltd. All rights reserved.

  3. A well-refined in vitro model derived from human embryonic stem cell for screening phytochemicals with midbrain dopaminergic differentiation-boosting potential for improving Parkinson's disease.

    Science.gov (United States)

    Hsieh, Wen-Ting; Chiang, Been-Huang

    2014-07-09

    Stimulation of endogenous neurogenesis is a potential approach to compensate for loss of dopaminergic neurons of substantia nigra compacta nigra (SNpc) in patients with Parkinson's disease (PD). This objective was to establish an in vitro model by differentiating pluripotent human embryonic stem cells (hESCs) into midbrain dopaminergic (mDA) neurons for screening phytochemicals with mDA neurogenesis-boosting potentials. Consequently, a five-stage differentiation process was developed. The derived cells expressed many mDA markers including tyrosine hydroxylase (TH), β-III tubulin, and dopamine transporter (DAT). The voltage-gated ion channels and dopamine release were also examined for verifying neuron function, and the dopamine receptor agonists bromocriptine and 7-hydroxy-2-(dipropylamino)tetralin (7-OH-DPAT) were used to validate our model. Then, several potential phytochemicals including green tea catechins and ginsenosides were tested using the model. Finally, ginsenoside Rb1 was identified as the most potent phytochemical which is capable of upregulating neurotrophin expression and inducing mDA differentiation.

  4. Blockade of Aquaporin 1 Inhibits Proliferation, Motility, and Metastatic Potential of Mesothelioma In Vitro but not in an In Vivo Model

    Directory of Open Access Journals (Sweden)

    Sonja Klebe

    2015-01-01

    Full Text Available Background. Malignant mesothelioma (MM is an aggressive tumor of the serosal membranes, mostly the pleura. It is related to asbestos exposure and has a poor prognosis. MM has a long latency period, and incidence is predicted to remain stable or increase until 2020. Currently, no biomarkers for a specific targeted therapy are available. Previously, we observed that expression of aquaporin 1 (AQP1 was an indicator of prognosis in two independent cohorts. Here we determine whether AQP1 inhibition has therapeutic potential in the treatment of MM. Methods. Functional studies were performed with H226 cells and primary MM cells harvested from pleural effusions. AQP1 expression and mesothelial phenotype was determined by immunohistochemistry. AQP1 function was inhibited by a pharmacological blocker (AqB050 or AQP1-specific siRNA. Cell proliferation, migration, and anchorage-independent cell growth were assessed. A nude mouse heterotopic xenograft model of MM was utilised for the in vivo studies. Results. Inhibition of AQP1 significantly decreases cell proliferation, metastatic potential, and motility without inducing nonspecific cytotoxicity or increasing apoptosis. In vivo blockade of AQP1 had no biologically significant effect on growth of established tumours. Conclusions. Targeted blockade of AQP1 restricts MM growth and migration in vitro. Further work is warranted to fully evaluate treatment potential in vivo.

  5. Production and Characterization of Glass-Ceramic Materials for Potential Use in Dental Applications: Thermal and Mechanical Properties, Microstructure, and In Vitro Bioactivity

    Directory of Open Access Journals (Sweden)

    Francesco Baino

    2017-12-01

    Full Text Available Multicomponent silicate glasses and their corresponding glass-ceramic derivatives were prepared and tested for potential applications in dentistry. The glasses were produced via a melting-quenching process, ground and sieved to obtain fine-grained powders that were pressed in the form of small cylinders and thermally treated to obtain sintered glass-ceramic samples. X-ray diffraction investigations were carried out on the materials before and after sintering to detect the presence of crystalline phases. Thermal analyses, mechanical characterizations (assessment of bending strength, Young’s modulus, Vickers hardness, fracture toughness, and in vitro bioactivity tests in simulated body fluid were performed. On the basis of the acquired results, different potential applications in the dental field were discussed for the proposed glass-ceramics. The use of such materials can be suggested for either restorative dentistry or dental implantology, mainly depending on their peculiar bioactive and mechanical properties. At the end of the work, the feasibility of a novel full-ceramic bilayered implant was explored and discussed. This implant, comprising a highly bioactive layer expected to promote osteointegration and another one mimicking the features of tooth enamel, can have an interesting potential for whole tooth substitution.

  6. Polarity directed optimization of phytochemical and in vitro biological potential of an indigenous folklore: Quercus dilatata Lindl. ex Royle.

    Science.gov (United States)

    Ahmed, Madiha; Fatima, Humaira; Qasim, Muhammad; Gul, Bilquees; Ihsan-Ul-Haq

    2017-08-03

    Plants have served either as a natural templates for the development of new chemicals or a phytomedicine since antiquity. Therefore, the present study was aimed to appraise the polarity directed antioxidant, cytotoxic, protein kinase inhibitory, antileishmanial and glucose modulatory attributes of a Himalayan medicinal plant- Quercus dilatata. Total phenolic and flavonoid contents were determined colorimetrically and various polyphenols were identified by RP-HPLC analysis. Brine shrimp lethality, SRB and MTT assays were employed to test cytotoxicity against Artemia salina and human cancer cell lines respectively. Antileishmanial activity was determined using standard MTT protocol. Glucose modulation was assessed by α-amylase inhibition assay while disc diffusion assay was used to establish protein kinase inhibitory and antifungal spectrum. Among 14 extracts of aerial parts, distilled water-acetone extract demonstrated maximum extract recovery (10.52% w/w), phenolic content (21.37 ± 0.21 μg GAE/mg dry weight (DW)), total antioxidant capacity (4.81 ± 0.98 μg AAE/mg DW) and reducing power potential (20.03 ± 2.4 μg/mg DW). On the other hand, Distilled water extract proficiently extracted flavonoid content (4.78 ± 0.51 μg QE/mg DW). RP-HPLC analysis revealed the presence of significant amounts of phenolic metabolites (0.049 to 15.336 μg/mg extract) including, pyrocatechol, gallic acid, catechin, chlorogenic acid, p-coumaric acid, ferulic acid and quercetin. Highest free radical scavenging capacity was found in Methanol-Ethyl acetate extract (IC 50 8.1 ± 0.5 μg/ml). In the brine shrimp toxicity assay, most of the tested extracts (57%) showed high cytotoxicity. Among these, Chloroform-Methanol extract had highest cytotoxicity against THP-1 cell line (IC 50 3.88 ± 0.53 μg/ml). About 50% of the extracts were found to be moderately antiproliferative against Hep G2 cell line. Methanol extract exhibited considerable protein kinase inhibitory

  7. Potentiation of antimicrobial photodynamic inactivation mediated by a cationic fullerene by added iodide: in vitro and in vivo studies.

    Science.gov (United States)

    Zhang, Yunsong; Dai, Tianhong; Wang, Min; Vecchio, Daniela; Chiang, Long Y; Hamblin, Michael R

    2015-03-01

    Antimicrobial photodynamic inactivation with fullerenes bearing cationic charges may overcome resistant microbes. We synthesized C60-fullerene (LC16) bearing decaquaternary chain and deca-tertiary-amino groups that facilitates electron-transfer reactions via the photoexcited fullerene. Addition of the harmless salt, potassium iodide (10 mM) potentiated the ultraviolet A (UVA) or white light-mediated killing of Gram-negative bacteria Acinetobacter baumannii, Gram-positive methicillin-resistant Staphylococcus aureus and fungal yeast Candida albicans by 1-2+ logs. Mouse model infected with bioluminescent Acinetobacter baumannii gave increased loss of bioluminescence when iodide (10 mM) was combined with LC16 and UVA/white light. The mechanism may involve photoinduced electron reduction of (1)(C60>)* or (3)(C60>)* by iodide producing I· or I2 followed by subsequent intermolecular electron-transfer events of (C60>)-· to produce reactive radicals.

  8. A novel class of potential prion drugs: preliminary in vitro and in vivo data for multilayer coated gold nanoparticles.

    Science.gov (United States)

    Ai Tran, Hoang Ngoc; Sousa, Fernanda; Moda, Fabio; Mandal, Subhra; Chanana, Munish; Vimercati, Chiara; Morbin, Michela; Krol, Silke; Tagliavini, Fabrizio; Legname, Giuseppe

    2010-12-01

    Gold nanoparticles coated with oppositely charged polyelectrolytes, such as polyallylamine hydrochloride and polystyrenesulfonate, were examined for potential inhibition of prion protein aggregation and prion (PrPSc) conversion and replication. Different coatings, finishing with a positive or negative layer, were tested, and different numbers of layers were investigated for their ability to interact and reduce the accumulation of PrPSc in scrapie prion infected ScGT1 and ScN2a cells. The particles efficiently hampered the accumulation of PrPSc in ScN2a cells and showed curing effects on ScGT1 cells with a nanoparticle concentration in the picomolar range. Finally, incubation periods of prion-infected mice treated with nanomolar concentrations of gold nanoparticles were significantly longer compared to untreated controls.

  9. Purvalanol A, olomoucine II and roscovitine inhibit ABCB1 transporter and synergistically potentiate cytotoxic effects of daunorubicin in vitro.

    Directory of Open Access Journals (Sweden)

    Daniela Cihalova

    Full Text Available Cyclin-dependent kinase inhibitors (CDKi have high potential applicability in anticancer therapy, but various aspects of their pharmacokinetics, especially their interactions with drug efflux transporters, have not yet been evaluated in detail. Thus, we investigated interactions of five CDKi (purvalanol A, olomoucine II, roscovitine, flavopiridol and SNS-032 with the ABCB1 transporter. Four of the compounds inhibited efflux of two ABCB1 substrates, Hoechst 33342 and daunorubicin, in MDCKII-ABCB1 cells: Olomoucine II most strongly, followed by roscovitine, purvalanol A, and flavopiridol. SNS-032 inhibited ABCB1-mediated efflux of Hoechst 33342 but not daunorubicin. In addition, purvalanol A, SNS-032 and flavopiridol lowered the stimulated ATPase activity in ABCB1 membrane preparations, while olomoucine II and roscovitine not only inhibited the stimulated ATPase but also significantly activated the basal ABCB1 ATPase, suggesting that these two CDKi are ABCB1 substrates. We further revealed that the strongest ABCB1 inhibitors (purvalanol A, olomoucine II and roscovitine synergistically potentiate the antiproliferative effect of daunorubicin, a commonly used anticancer drug and ABCB1 substrate, in MDCKII-ABCB1 cells as well as in human carcinoma HCT-8 and HepG2 cells. We suggest that this pronounced synergism is at least partly caused by (i CDKi-mediated inhibition of ABCB1 transporter leading to increased intracellular retention of daunorubicin and (ii native cytotoxic activity of the CDKi. Our results indicate that co-administration of the tested CDKi with anticancer drugs that are ABCB1 substrates may allow significant dose reduction in the treatment of ABCB1-expressing tumors.

  10. A novel class of potential prion drugs: preliminary in vitro and in vivo data for multilayer coated gold nanoparticles

    Science.gov (United States)

    Ai Tran, Hoang Ngoc; Sousa, Fernanda; Moda, Fabio; Mandal, Subhra; Chanana, Munish; Vimercati, Chiara; Morbin, Michela; Krol, Silke; Tagliavini, Fabrizio; Legname, Giuseppe

    2010-12-01

    Gold nanoparticles coated with oppositely charged polyelectrolytes, such as polyallylamine hydrochloride and polystyrenesulfonate, were examined for potential inhibition of prion protein aggregation and prion (PrPSc) conversion and replication. Different coatings, finishing with a positive or negative layer, were tested, and different numbers of layers were investigated for their ability to interact and reduce the accumulation of PrPSc in scrapie prion infected ScGT1 and ScN2a cells. The particles efficiently hampered the accumulation of PrPSc in ScN2a cells and showed curing effects on ScGT1 cells with a nanoparticle concentration in the picomolar range. Finally, incubation periods of prion-infected mice treated with nanomolar concentrations of gold nanoparticles were significantly longer compared to untreated controls.Gold nanoparticles coated with oppositely charged polyelectrolytes, such as polyallylamine hydrochloride and polystyrenesulfonate, were examined for potential inhibition of prion protein aggregation and prion (PrPSc) conversion and replication. Different coatings, finishing with a positive or negative layer, were tested, and different numbers of layers were investigated for their ability to interact and reduce the accumulation of PrPSc in scrapie prion infected ScGT1 and ScN2a cells. The particles efficiently hampered the accumulation of PrPSc in ScN2a cells and showed curing effects on ScGT1 cells with a nanoparticle concentration in the picomolar range. Finally, incubation periods of prion-infected mice treated with nanomolar concentrations of gold nanoparticles were significantly longer compared to untreated controls. Electronic supplementary information (ESI) available: HRTEM image of a 2A coated nanogold particle; Western blot