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Research Containing: Cell Proliferation

Floating culture promotes the maintenance of hematopoietic stem cells

by on 9 June 2015in Biology & Biotechnology No comment

In this study we examined the effect of the specific gravity of culture medium on the frequency of hematopoietic stem cell (HSC) maintenance. We used a newly developed high-specific-gravity media. Bone marrow cells were isolated and cultured, and HSC activity was evaluated. The number of hematopoietic progenitor/stem cells was markedly higher in the medium with high specific gravity. In high-specific-gravity media, cells did not precipitate, maintenance of HSCs was increased, and there was a concomitant accumulation of beta-catenin. This novel technique for maintaining HSC populations provides an important new tool for studies in regenerative medicine. (c) 2007 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

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<Go to ISI>://WOS:000250102900017

An integrated microfluidic culture device for quantitative analysis of human embryonic stem cells

by on 9 June 2015in Biology & Biotechnology No comment

We have successfully designed and fabricated an integrated microfluidic platform, the hESC-mu Chip, which is capable of reproducible and quantitative culture and analysis of individual hESC colonies in a semi-automated fashion. In this device, a serpentine microchannel allows pre-screening of dissociated hESC clusters, and six individually addressable cell culture chambers enable parallel hESC culture, as well as multiparameter analyses in sequence. In order to quantitatively monitor hESC proliferation and pluripotency status in real time, knock-in hESC lines with EGFP driven by the endogenous OCT4 promoter were constructed. On-chip immunoassays of several pluripotency markers were carried out to confirm that the hESC colonies maintained their pluripotency. For the first time, our studies demonstrated well characterized hESC culture and analysis in a microfluidic setting, as well as a proof-of-concept demonstration of parallel/multiparameter/real-time/automated examination of self-renewal and differentiation in the same device.

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<Go to ISI>://WOS:000263465900008

Proliferation, osteogenic differentiation, and distribution of rat bone marrow stromal cells in nonwoven fabrics by different culture methods

by on 9 June 2015in Biology & Biotechnology No comment

The proliferation, osteogenic differentiation, and distribution patterns of stromal cells from rat bone marrow were investigated in a three-dimensional nonwoven fabric of polyethylene terephthalate fiber by the static, agitated, and stirred culture methods; stirring speeds were 10, 50, and 100 rpm in the stirred culture method. The culture method affected the time profile of proliferation and osteogenic differentiation of cells or their distribution in the fabric. The extent of cell proliferation and osteogenic differentiation became higher in order of the stirred at 100 rpm = the stirred at 50 rpm > the stirred at 10 rpm > the agitated > the static methods. In addition, the cells were more uniformly proliferated in the fabric by the stirred culture method with time than they were proliferated in the fabric by other methods. The alkaline phosphatase (ALP) activity and calcium content were higher for cells cultured by the stirred culture method than those cultured by other methods. The total ALP activity, calcium content, and bone mineral density were higher for every stirred method than those for other methods. However, the distribution uniformity of cells differentiated was low irrespective of the culture method. It is concluded that the extent of proliferation and differentiation of cells or their distribution uniformity in the nonwoven fabrics was influenced by the culture method.

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<Go to ISI>://WOS:000253960900011

Morphofunctional status and osteogenic differentiation potential of human mesenchymal stromal precursor cells during in vitro modeling of microgravity effects

by on 9 June 2015in Biology & Biotechnology No comment

We studied the effects of long-term (20-day) simulated microgravity (clinostatic exposure) and osteogenic differentiation stimuli on cultured mesenchymal stromal precursor cells isolated from human bone marrow. Clinostatic exposure significantly reduced proliferative activity of mesenchymal stem cells in comparison with the static and dynamic control, increased the number of large flat cells in the culture, and stimulated migration activity of cells. Phenotypic studies of surface antigens (CD90, CD54, CD106, CD105, CD34, CD45, class 1 HLA) during clinostatic exposure of mesenchymal stem cell cultures showed differences in their expression between experimental and control groups. Studies of osteogenesis of precursor cell showed that cell differentiation potential can be directed towards osteogenesis by a combination of clinostatic exposure and differentiation stimuli. The results confirm gravity sensitivity of human bone marrow precursor cells and open new vistas for understanding of the mechanisms of bone tissue loss in humans under conditions of space mission.

Related URLs:
http://ovidsp.ovid.com/ovidweb.cgi?T=JS&CSC=Y&NEWS=N&PAGE=fulltext&D=emed8&AN=2008092502
http://sfxhosted.exlibrisgroup.com/mayo?sid=OVID:embase&id=pmid:&id=doi:10.1007%2Fs10517-007-0387-1&issn=0007-4888&isbn=&volume=144&issue=4&spage=608&pages=608-613&date=2007&title=Bulletin+of+Experimental+Biology+and+Medicine&atitle=Morphofunctional+status+and+osteogenic+differentiation+potential+of+human+mesenchymal+stromal+precursor+cells+during+in+vitro+modeling+of+microgravity+effects&aulast=Gershovich&pid=%3Cauthor%3EGershovich+J.G.%3C%2Fauthor%3E&%3CAN%3E2008092502%3C%2FAN%3E

Dynamic Three-Dimensional Culture Methods Enhance Mesenchymal Stem Cell Properties and Increase Therapeutic Potential

by on 9 June 2015in Biology & Biotechnology No comment

Mesenchymal stemcells (MSCs) are capable of self-renewal and differentiation along the osteogenic, chondrogenic, and adipogenic lineages and have potential applications in a range of therapies. MSCs can be cultured as monolayers on tissue culture plastic, but there are indications that they lose cell-specific properties with time in vitro and so poorly reflect in vivo MSC behavior. We developed dynamic three-dimensional (3D) techniques for in vitro MSC culture using spinner flasks and a rotating wall vessel bioreactor. We characterized the two methods for dynamic 3D MSC culture and compared the properties of these cultures with monolayer MSCs. Our results showed that under optimal conditions, MSCs form compact cellular spheroids and remain viable in dynamic 3D culture. We demonstrated altered cell size and surface antigen expression together with enhanced osteogenic and adipogenic differentiation potential in MSCs from dynamic 3D conditions. By microarray analysis of monolayer and spinner flask MSCs, we identified many differences in gene expression, including those confirming widespread changes to the cellular architecture and extracellular matrix. The upregulation of interleukin 24 in dynamic 3D cultures was shown to selectively impair the viability of prostate cancer cells cultured in medium conditioned by dynamic 3D MSCs. Overall, this work suggests a novel therapeutic application for dynamic 3D MSCs and demonstrates that these methods are a viable alternative to monolayer techniques and may prove beneficial for retaining MSC properties in vitro.

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<Go to ISI>://WOS:000280228300018

Computational modeling for the optimization of a cardiogenic 3D bioprocess of encapsulated embryonic stem cells

by on 9 June 2015in Biology & Biotechnology No comment

We present a computational fluid dynamics (CFD)-based model aimed at the identification of optimized culture conditions promoting efficient cardiogenesis of hydrogel-bead-encapsulated embryonic stem cells (ESCs) within a rotating bioreactor. The numerical approach, integrating diffusion, convection, and multiphase fluid dynamics calculations, allowed to evaluate (i) the microgravity motion of the floating beads, (ii) the O-2 delivery to the cells, also (iii) taking into account the cellularO(2) consumption, as a function of different rotation speeds of the breeding chamber. According to our results, a 25rpm rotation (i) enhances an adequate mixing of the cell carriers, avoiding sedimentation and excessive packing, also maintaining a quite homogeneous distribution of the suspended beads and (ii) imparts a proper cellular O-2 supply, providing cells close to a normoxia condition. The bioreactor working conditions derived from the numerical analysis allowed the attainment of in vitro long-term cell viability maintenance, supporting efficient large-scale generation of ESC-derived cardiomyocytes (ESC-DCs) through a chemical-based conditioning bioprocess. In conclusion, we demonstrated the feasibility of using CFD-based tools, as a reliable and cost-effective strategy to assist the design of a 3D cardiogenic bioprocess.

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<Go to ISI>://WOS:000300230600020

Spaceflight enhances cell aggregation and random budding in Candida albicans

by on 9 June 2015in Biology & Biotechnology No comment

This study presents the first global transcriptional profiling and phenotypic characterization of the major human opportunistic fungal pathogen, Candida albicans, grown in spaceflight conditions. Microarray analysis revealed that C. albicans subjected to short-term spaceflight culture differentially regulated 452 genes compared to synchronous ground controls, which represented 8.3% of the analyzed ORFs. Spaceflight-cultured C. albicans-induced genes involved in cell aggregation (similar to flocculation), which was validated by microscopic and flow cytometry analysis. We also observed enhanced random budding of spaceflight-cultured cells as opposed to bipolar budding patterns for ground samples, in accordance with the gene expression data. Furthermore, genes involved in antifungal agent and stress resistance were differentially regulated in spaceflight, including induction of ABC transporters and members of the major facilitator family, downregulation of ergosterol-encoding genes, and upregulation of genes involved in oxidative stress resistance. Finally, downregulation of genes involved in actin cytoskeleton was observed. Interestingly, the transcriptional regulator Cap1 and over 30% of the Cap1 regulon was differentially expressed in spaceflight-cultured C. albicans. A potential role for Cap1 in the spaceflight response of C. albicans is suggested, as this regulator is involved in random budding, cell aggregation, and oxidative stress resistance; all related to observed spaceflight-associated changes of C. albicans. While culture of C. albicans in microgravity potentiates a global change in gene expression that could induce a virulence-related phenotype, no increased virulence in a murine intraperitoneal (i.p.) infection model was observed under the conditions of this study. Collectively, our data represent an important basis for the assessment of the risk that commensal flora could play during human spaceflight missions. Furthermore, since the low fluid-shear environment of microgravity is relevant to physical forces encountered by pathogens during the infection process, insights gained from this study could identify novel infectious disease mechanisms, with downstream benefits for the general public.

Related URLs:
http://www.ncbi.nlm.nih.gov/pubmed/24324620

Induction of vascular endothelial phenotype and cellular proliferation from human cord blood stem cells cultured in simulated microgravity

by on 9 June 2015in Biology & Biotechnology No comment

Recent studies have demonstrated that stem cells derived from adult hematopoietic tissues are capable of trans-differentiation into non-hematopoietic cells, and that the culture in microgravity (μ g) may modulate the proliferation and differentiation. We investigated the application of μ g to human umbilical cord blood stem cells (CBSC) in the induction of vascular endothelial phenotype expression and cellular proliferation. CD34+ mononuclear cells were isolated from waste human umbilical cord blood samples and cultured in simulated μ g for 14 days. The cells were seeded in rotary wall vessels (RWV) with or without microcarrier beads (MCB) and vascular endothelial growth factor was added during culture. Controls consisted of culture in I G. The cell cultures in RWV were examined by inverted microscopy. Cell counts, endothelial cell and leukocyte markers per-formed by flow-cytometry and FACS scan were assayed at days 1, 4, 7 and at the termination of the experiments. Culture in RWV revealed significantly increased cellular proliferation with three-dimensional (3D) tissue-like aggregates. At day 4, CD34+ cells cultured in RWV bioreactor without MCB developed vascular tubular assemblies and exhibited endothelial phenotypic markers. These data suggest that CD34+ human umbilical cord blood progenitors are capable of trans-differentiation into vascular endothelial cell phenotype and assemble into 3D tissue structures. Culture of CBSC in simulated μ g may be potentially beneficial in the fields of stem cell biology and somatic cell therapy. © 2005 Elsevier Ltd. All rights reserved.

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<Go to ISI>://WOS:000229063800018