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Cell Culture Experiments (cell + culture_experiment)
Selected AbstractsEvidence of occult HCV genotypes in haemophilic individuals with unapparent HCV mixed infectionsHAEMOPHILIA, Issue 4 2008C. PARODI Summary., Individuals with haemophilia who received non heat-treated factor concentrates were likely to undergo multiple exposures to the hepatitis C virus (HCV). Therefore, HCV mixed-genotype infections might be more frequent in these patients than in the general population. Their prevalence is extremely variable in similar groups of patients tested by different assays due to the fact that currently available genotyping techniques are not suitable to detect multiple HCV genotypes in a viral population. As an HCV viral reservoir, the peripheral blood mononuclear cell (PBMC) might harbor viral variants distinct from the genotypes detected in plasma. We investigated the presence of HCV genotypes in a group of chronically infected haemophilic patients in the PBMC compartment using a non-stimulated cell culture system that allows the detection of the HCV genome in culture supernatants. We compared them to the HCV genotypes found in plasma samples. Cell culture experiments performed with PBMC demonstrated the presence of additional HCV genotypes that were undetected in the corresponding plasma samples with the same genotyping technique. Although mixed infections at HCV genotype level became evident in 5.6% of the patients (16/288), the culture methodology increased the number of HCV infections with multiple genotypes to 62.5% (10/16) (P < 0.0001). Once more, the role of mononuclear cells as HCV viral reservoirs is emphasized. Considering minor strains could influence the outcome of treatment, detection of covert HCV mixed-genotype infections might be essential for choosing the adequate therapeutic regimen. [source] Gelatin-based photopolymers for bone replacement materialsJOURNAL OF POLYMER SCIENCE (IN TWO SECTIONS), Issue 24 2009Monika Schuster Abstract Gelatin-based monomers were considered as suitable base component for the 3D structuring of potential bone replacement materials by stereolithographic techniques. Different methacrylate-based gelatin derivatives were prepared, whereas a polyethylene glycol modified derivative GP4M turned out to have the highest tolerance toward other monomers. These are essential as they allow the tuning of the photoreactivity and the mechanical properties. Cell culture experiments with osteoblast- and endothelial-like cells confirmed negligible cytotoxicity of these monomers. Finally, we were able to show the possibility of producing arbitrary cellular structures with these gelatin-containing formulations using stereolithography. © 2009 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem, 2009 [source] Development of Electrospun Three-arm Star Poly(, -caprolactone) Meshes for Tissue Engineering ApplicationsMACROMOLECULAR BIOSCIENCE, Issue 8 2010Dario Puppi Abstract We have developed three-dimensional electrospun microfibrous meshes of a novel star branched three-arm poly(, -caprolactone) (*PCL) as potential scaffolds for tissue engineering applications. The processing conditions required to obtain uniform fibers were optimized by studying their influence on fiber morphology and size. Polymer molecular weight and solution feed rate influenced both the mesh microstructure and the tensile properties of the developed mats. Electrospun samples were also tested for their mechanical properties in wet conditions, showing higher yield strength and strain in comparison to that observed in dry conditions. Cell culture experiments employing MC3T3-E1 osteoblast like cells showed good cell viability adhesion and collagen production on the *PCL scaffolds. [source] H2/NH3 Plasma-Grafting of PEEK-WC-PU Membrane to Improve their cyto-Compatibility with HepatocytesPLASMA PROCESSES AND POLYMERS, Issue S1 2009Stefania Laera Abstract Plasma treatments in H2 and NH3 RF (13.56 MHz) glow discharges have been used for modifying the surface of PEEK-WC-PU membranes. Water contact angle (WCA) and X-Ray photoelectron spectroscopy (XPS) analyses were performed to study the compositional changes of PEEK-WC-PU membranes after grafting. Cell culture experiments with human hepatocytes clearly show that grafting N-containing groups improves the cyto-compatibility of the membranes. [source] Three-dimensional fibrous PLGA/HAp composite scaffold for BMP-2 deliveryBIOTECHNOLOGY & BIOENGINEERING, Issue 1 2008Hemin Nie Abstract A protein loaded three-dimensional scaffold can be used for protein delivery and bone tissue regeneration. The main objective of this project was to develop recombinant human bone morphogenetic protein-2 (rhBMP-2) loaded poly(D,L -lactide-co-glycolide)/hydroxylapatite (PLGA/HAp) composite fibrous scaffolds through a promising fabrication technique, electrospinning. In vitro release of BMP-2 from these scaffolds, and the attachment ability and viability of marrow derived messenchymal stem cells (MSCs) in the presence of the scaffolds were investigated. The PLGA/HAp composite scaffolds developed in this study exhibit good morphology and it was observed that HAp nanoparticles were homogeneously dispersed inside PLGA matrix within the scaffold. The composite scaffolds allowed sustained (2,8 weeks) release of BMP-2 whose release rate was accelerated with increasing HAp content. It was also shown that BMP-2 protein successfully maintained its integrity and natural conformations after undergoing the process of electrospinning. Cell culture experiments showed that the encapsulation of HAp could enhance cell attachment to scaffolds and lower cytotoxicity. Biotechnol. Bioeng. 2008;99: 223,234. © 2007 Wiley Periodicals, Inc. [source] Preparation and properties of ionically cross-linked chitosan nanoparticlesPOLYMERS FOR ADVANCED TECHNOLOGIES, Issue 7 2009Hui Liu Abstract Chitosan nanoparticles were fabricated by a method of tripolyphosphate (TPP) cross-linking. The influence of fabrication conditions on the physical properties and drug loading and release properties was investigated by transmission electron microscopy (TEM), dynamic light scattering (DLS), and UV,vis spectroscopy. The nanoparticles could be prepared only within a zone of appropriate chitosan and TPP concentrations. The particle size and surface zeta potential can be manipulated by variation of the fabrication conditions such as chitosan/TPP ratio and concentration, solution pH and salt addition. TEM observation revealed a core,shell structure for the as-prepared nanoparticles, but a filled structure for the ciprofloxacin (CH) loaded particles. Results show that the chitosan nanoparticles were rather stable and no cytotoxicity of the chitosan nanoparticles was found in an in vitro cell culture experiment. Loading and release of CH can be modulated by the environmental factors such as solution pH and medium quality. Copyright © 2008 John Wiley & Sons, Ltd. [source] Drosophila multiplexin (Dmp) modulates motor axon pathfinding accuracyDEVELOPMENT GROWTH & DIFFERENTIATION, Issue 5 2009Frauke Meyer Multiplexins are multidomain collagens typically composed of an N-terminal thrombospondin-related domain, an interrupted triple helix and a C-terminal endostatin domain. They feature a clear regulatory function in the development of different tissues, which is chiefly conveyed by the endostatin domain. This domain can be found in proteolytically released monomeric and trimeric versions, and their diverse and opposed effects on the migratory behavior of epithelial and endothelial cell types have been demonstrated in cell culture experiments. The only Drosophila multiplexin displays specific features of both vertebrate multiplexins, collagens XV and XVIII. We characterized the Drosophila multiplexin (dmp) gene and found that three main isoforms are expressed from it, one of which is the monomeric endostatin version. Generation of dmp deletion alleles revealed that Dmp plays a role in motor axon pathfinding, as the mutants exhibit ventral bypass defects of the intersegmental nerve b (ISNb) similar to other motor axon guidance mutants. Transgenic overexpression of monomeric endostatin as well as of full-length Dmp, but not trimeric endostatin, were able to rescue these defects. In contrast, trimeric endostatin increased axon pathfinding accuracy in wild type background. We conclude that Dmp plays a modulating role in motor axon pathfinding and may be part of a buffering system that functions to avoid innervation errors. [source] Functional analysis of murine CBF1 during Drosophila developmentDEVELOPMENTAL DYNAMICS, Issue 4 2006Markus Kaspar Abstract Transcription factors of the CSL family are the main mediators of the Notch signalling pathway. The CSL factor in Drosophila is called Suppressor of Hairless (Su(H)) and it has been shown that it acts as a transcriptional repressor in the absence of a Notch signal and as a transcriptional activator in its presence in several developmental contexts. Furthermore, recent data suggest that Su(H) can also activate and maintain transcription of some target genes in a Notch-independent manner. However, although it has been shown that the mammalian CSL ortholog, CBF1, acts as a repressor of transcription in cell culture experiments, so far in vivo evidence for such a function has been lacking. Moreover, it is not known whether CBF1 can activate transcription in a Notch-independent manner, just like Su(H). Here we have investigated these questions by introducing murine CBF1 (mCBF1) and asked whether it can functionally replace Su(H) during Drosophila development. We found that this is indeed the case. We show that mCBF1 can act as a repressor of transcription and can activate and maintain the expression of some target genes in a Notch-independent manner. Our results, therefore, indicate that CBF1 can exert these functions also in its normal context, that is during mammalian development. Developmental Dynamics 235:918,927, 2006. © 2006 Wiley-Liss, Inc. [source] Tailoring Cell Behavior on Polymers by the Incorporation of Titanium Doped Phosphate Glass Filler,ADVANCED ENGINEERING MATERIALS, Issue 7 2010Wojciech Chrzanowski Abstract Understanding tissue response to materials, to enable modulation and guided tissue regeneration is one of the main challenges in biomaterials science. Nowadays polymers, glasses, and metals dominate as biomaterials. Often native properties of those materials are not sufficient and there is a need to combine them, so as to modify and adjust their properties to the application. The primary aim of this study was to improve cell response to polymer (PLDL) using phosphate glass as filler (titanium doped phosphate glass). As a control ,-tricalcium phosphate (TCP) filler was used. Various concentrations of the filler were used (10,40 vol%). Wetting behavior, , -potentials, mechanical and thermal properties, and human cells response to the materials were evaluated. Results showed that with increase in glass filler loading wettability improved, , -potentials dropped, and increase in stiffness of materials was observed. Importantly cell culture experiments showed more developed and well spread cells on the samples with glass content up to 20 vol%. Cells responded much more positively to the glass filled samples than to TCP filled. However, expression of osteocalcin and osteopontin, proteins that indicate formation of the mineralized structures was positive for all the samples including pure PLDL. It was concluded that due to improved wetting behavior, lower , -potentials, and specific chemistry of the glass filler it was possible to alter cells response, improve bioactivity of the polymer, and vary mechanical properties. [source] Inhibition of prostaglandin synthesis and actions by genistein in human prostate cancer cells and by soy isoflavones in prostate cancer patientsINTERNATIONAL JOURNAL OF CANCER, Issue 9 2009Srilatha Swami Abstract Soy and its constituent isoflavone genistein inhibit the development and progression of prostate cancer (PCa). Our study in both cultured cells and PCa patients reveals a novel pathway for the actions of genistein, namely the inhibition of the synthesis and biological actions of prostaglandins (PGs), known stimulators of PCa growth. In the cell culture experiments, genistein decreased cyclooxygenase-2 (COX-2) mRNA and protein expression in both human PCa cell lines (LNCaP and PC-3) and primary prostate epithelial cells and increased 15-hydroxyprostaglandin dehydrogenase (15-PGDH) mRNA levels in primary prostate cells. As a result genistein significantly reduced the secretion of PGE2 by these cells. EP4 and FP PG receptor mRNA were also reduced by genistein, providing an additional mechanism for the suppression of PG biological effects. Further, the growth stimulatory effects of both exogenous PGs and endogenous PGs derived from precursor arachidonic acid were attenuated by genistein. We also performed a pilot randomised double blind clinical study in which placebo or soy isoflavone supplements were given to PCa patients in the neo-adjuvant setting for 2 weeks before prostatectomy. Gene expression changes were measured in the prostatectomy specimens. In PCa patients ingesting isoflavones, we observed significant decreases in prostate COX-2 mRNA and increases in p21 mRNA. There were significant correlations between COX-2 mRNA suppression, p21 mRNA stimulation and serum isoflavone levels. We propose that the inhibition of the PG pathway contributes to the beneficial effect of soy isoflavones in PCa chemoprevention and/or treatment. © 2008 Wiley-Liss, Inc. [source] Physically crosslinked composite hydrogels of PVA with natural macromolecules: Structure, mechanical properties, and endothelial cell compatibilityJOURNAL OF BIOMEDICAL MATERIALS RESEARCH, Issue 2 2009Y. Liu Abstract Polyvinyl alcohol (PVA) hydrogels have been considered potentially suitable for applications as engineered blood vessels because of their structure and mechanical properties. However, PVA's hydrophilicity hinders its capacity to act as a substrate for cell attachment. As a remedy, PVA was blended with chitosan, gelatin, or starch, and hydrogels were formed by subjecting the solutions to freeze-thaw cycles followed by coagulation bath immersion. The structure-property relationships for these hydrogels were examined by measurement of their swelling, rehydration, degradation, and mechanical properties. For the case of pure PVA hydrogels, the equilibrium swelling ratio was used to predict the effect of freeze thaw cycles and coagulation bath on average molecular weights between crosslinks and on mesh size. For all hydrogels, trends for the reswelling ratio, which is indicative of the crosslinked polymer fraction, were consistent with relative tensile properties. The coagulation bath treatment increased the degradation resistance of the hydrogels significantly. The suitability of each hydrogel for cell attachment and proliferation was examined by protein adsorption and bovine vascular endothelial cell culture experiments. Protein adsorption and cell proliferation was highest on the PVA/gelatin hydrogels. This study demonstrates that the potential of PVA hydrogels for artificial blood vessel applications can be improved by the addition of natural polymers, and that freeze-thawing and coagulation bath treatment can be utilized for fine adjustment of the physical characteristics. © 2009 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2009 [source] Gene expression study of Saccharomyces cerevisiae under changing growth conditionsJOURNAL OF CHEMICAL TECHNOLOGY & BIOTECHNOLOGY, Issue 8 2009Pengcheng Fu Abstract BACKGROUND: DNA microarrays technology has been used to obtain expression profiles of thousands of genes at the same time for a given organism at relatively low costs. While gene expression approaches are being developed which allow holistic analysis of complex biological processes, there exist very few illustrative examples on the integration of large scale modeling and high throughput time course experiments to upgrade the information contents on yeast biology. RESULTS:Saccharomyces cerevisiae cell culture experiments with perturbed growth conditions were designed so that the metabolic states would be shifted from one to another. Microarrays were used to explore changes in gene expression across the entire yeast genome during the perturbation experiments. Changes in transcript abundance in these growth periods were investigated to study the cellular response to different glucose and oxygen supply. Computational results and experimental observations representing the three characteristic metabolic states were compared on the S. cerevisiae metabolic pathways, as well as the visualization platform provided by the metabolic phenotypic phase plane (PhPP) for the gene regulation on cell metabolism and adaptation of cells to environmental changes. CONCLUSIONS: The integrated expression study described reveals that S. cerevisiae cells respond to environmental changes mainly by down-regulating a number of genes to alter the cell metabolism so that the cells adapt to the variations in their growth conditions. Copyright © 2009 Society of Chemical Industry [source] Potent anti-amyloidogenic and fibril-destabilizing effects of polyphenols in vitro: implications for the prevention and therapeutics of Alzheimer's diseaseJOURNAL OF NEUROCHEMISTRY, Issue 1 2003Kenjiro Ono Abstract Cerebral deposition of amyloid ,-peptide (A,) in the brain is an invariant feature of Alzheimer's disease (AD). A consistent protective effect of wine consumption on AD has been documented by epidemiological studies. In the present study, we used fluorescence spectroscopy with thioflavin T and electron microscopy to examine the effects of wine-related polyphenols (myricetin, morin, quercetin, kaempferol (+)-catechin and (,)-epicatechin) on the formation, extension, and destabilization of ,-amyloid fibrils (fA,) at pH 7.5 at 37°C in vitro. All examined polyphenols dose-dependently inhibited formation of fA, from fresh A,(1,40) and A,(1,42), as well as their extension. Moreover, these polyphenols dose-dependently destabilized preformed fA,s. The overall activity of the molecules examined was in the order of: myricetin = morin = quercetin > kaempferol > (+)-catechin = (,)-epicatechin. The effective concentrations (EC50) of myricetin, morin and quercetin for the formation, extension and destabilization of fA,s were in the order of 0.1,1 µm. In cell culture experiments, myricetin-treated fA, were suggested to be less toxic than intact fA,, as demonstrated by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide assay. Although the mechanisms by which these polyphenols inhibit fA, formation from A,, and destabilize pre-formed fA,in vitro are still unclear, polyphenols could be a key molecule for the development of preventives and therapeutics for AD. [source] Nitric oxide synthase type-II is synthesized by human gingival tissue and cultured human gingival fibroblastsJOURNAL OF PERIODONTAL RESEARCH, Issue 4 2000H. K. Kendall Nitric oxide is known to be an important inflammatory mediator, and is implicated in the pathophysiology of a range of inflammatory disorders. The aim of this study was to determine the localization and distribution of endothelial NOS (NOSII) in human gingival tissue, and to ascertain if human gingival fibroblasts express NOS-II when stimulated with interferon gamma (IFN-,) and bacterial lipopolysaccharide (LPS). The distribution of NOS-II in inflamed and non-inflamed specimens of human gingivae was studied using a monoclonal antibody against nitric oxide synthase II. Cultures of fibroblasts derived from healthy human gingivae were used for the cell culture experiments. The results from immunohistochemical staining of the tissues indicated an upregulation of NOS-II expression in inflamed compared to non-inflamed gingival tissue. Fibroblasts and inflammatory cells within the inflamed connective tissue were positively stained for NOS-II. In addition, basal keratinocytes also stained strongly for NOS-II, in both healthy and inflamed tissue sections. When cultured human gingival fibroblasts were stimulated by INF-, and Porphyromonas gingivalis LPS, NOS-II was more strongly expressed than when the cells were exposed to LPS or IFN-, alone. These data suggest that, as for other inflammatory diseases, NO plays a role in the pathophysiology of periodontitis. [source] Continuous perfusion microfluidic cell culture array for high-throughput cell-based assaysBIOTECHNOLOGY & BIOENGINEERING, Issue 1 2005Paul J. Hung Abstract We present for the first time a microfluidic cell culture array for long-term cellular monitoring. The 10 × 10 array could potentially assay 100 different cell-based experiments in parallel. The device was designed to integrate the processes used in typical cell culture experiments on a single self-contained microfluidic system. Major functions include repeated cell growth/passage cycles, reagent introduction, and real-time optical analysis. The single unit of the array consists of a circular microfluidic chamber, multiple narrow perfusion channels surrounding the main chamber, and four ports for fluidic access. Human carcinoma (HeLa) cells were cultured inside the device with continuous perfusion of medium at 37°C. The observed doubling time was 1.4 ± 0.1 days with a peak cell density of ,2.5*105 cells/cm2. Cell assay was demonstrated by monitoring the fluorescence localization of calcein AM from 1 min to 10 days after reagent introduction. Confluent cell cultures were passaged within the microfluidic chambers using trypsin and successfully regrown, suggesting a stable culture environment suitable for continuous operation. The cell culture array could offer a platform for a wide range of assays with applications in drug screening, bioinformatics, and quantitative cell biology. © 2004 Wiley Periodicals, Inc. [source] | ||||||||||||||||