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Research Containing: Cell Adhesion

The Importance of Caveolin-1 as Key-Regulator of Three-Dimensional Growth in Thyroid Cancer Cells Cultured under Real and Simulated Microgravity Conditions

by cfynanon 22 August 2016in Biology & Biotechnology No comment

We recently demonstrated that the CAV1 gene was down-regulated, when poorly differentiated thyroid FTC-133 cancer cells formed spheroids under simulated microgravity conditions. Here, we present evidence that the caveolin-1 protein is involved in the inhibition of spheroid formation, when confluent monolayers are exposed to microgravity. The evidence is based on proteins detected in cells and their supernatants of the recent spaceflight experiment: "NanoRacks-CellBox-Thyroid Cancer". The culture supernatant had been collected in a special container adjacent to the flight hardware incubation chamber and stored at low temperature until it was analyzed by Multi-Analyte Profiling (MAP) technology, while the cells remaining in the incubation chamber were fixed by RNAlater and examined by mass spectrometry. The soluble proteins identified by MAP were investigated in regard to their mutual interactions and their influence on proteins, which were associated with the cells secreting the soluble proteins and had been identified in a preceding study. A Pathway Studio v.11 analysis of the soluble and cell-associated proteins together with protein kinase C alpha (PRKCA) suggests that caveolin-1 is involved, when plasminogen enriched in the extracellular space is not activated and the vascular cellular adhesion molecule (VCAM-1) mediated cell-cell adhesion is simultaneously strengthened and activated PRKCA is recruited in caveolae, while the thyroid cancer cells do not form spheroids.

Related URLs:
http://www.ncbi.nlm.nih.gov/pubmed/26633361

Genetic and Apoptotic Changes in Lungs of Mice Flown on the STS-135 Mission in Space.

by cfynanon 22 August 2016in Biology & Biotechnology No comment

AIM: The goal of the study was to evaluate changes in lung status due to spaceflight stressors that include radiation above levels found on Earth.;MATERIALS AND METHODS: Within hours after return from a 13-day mission in space onboard the Space Shuttle Atlantis, C57BL/6 mice (FLT group) were euthanized; mice housed on the ground in similar animal enclosure modules served as controls (AEM group). Lung tissue was collected to evaluate the expression of genes related to extracellular matrix (ECM)/adhesion and stem cell signaling. Pathway analysis was also performed. In addition, immunohistochemistry for stem cell antigen-1 (SCA-1), the terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay for apoptosis, and staining for histological characteristics were performed.;RESULTS: There were 18/168 genes significantly modulated in lungs from the FLT group (p<0.05 vs. AEM); 17 of these were up-regulated and one was down-regulated. The greatest effect, namely a 5.14-fold increase, was observed on Spock1 (also known as Spark/osteonectin), encoding a multi-functional protein that has anti-adhesive effects, inhibits cell proliferation and regulates activity of certain growth factors. Additional genes with increased expression were cadherin 3 (Cdh3), collagen, type V, alpha 1 (Col5a1), integrin alpha 5 (Itga5), laminin, gamma 1 (Lamc1), matrix metallopeptidase 14 (Mmp14), neural cell adhesion molecule 1 (Ncam1), transforming growth factor, beta induced (Tgfbi), thrombospondin 1 (Thbs1), Thbs2, versican (Vcan), fibroblast growth factor receptor 1 (Fgfr1), frizzled homolog 6 (Fzd6), nicastrin (Ncstn), nuclear factor of activated T-cells, cytoplasmic, calcineurin-dependent 4 (Nfatc4), notch gene homolog 4 (Notch4) and vang-like 2 (Vangl2). The down-regulated gene was Mmp13. Staining for SCA-1 protein showed strong signal intensity in bronchiolar epithelial cells of FLT mice (p<0.05 vs. AEM). TUNEL positivity was also significantly higher in the FLT mice (p<0.05 vs. AEM), but no consistent histological differences were noted. CONCLUSION: The results demonstrate that spaceflight-related stress had a significant impact on lung integrity, indicative of tissue injury and remodeling. Related URLs:
http://www.ncbi.nlm.nih.gov/pubmed/26130787

A biaxial rotating bioreactor for the culture of fetal mesenchymal stem cells for bone tissue engineering

by cfynanon 9 June 2015in Biology & Biotechnology No comment

The generation of effective tissue engineered bone grafts requires efficient exchange of nutrients and mechanical stimulus. Bioreactors provide a manner in which this can be achieved. We have recently developed a biaxial rotating bioreactor with efficient fluidics through in-silico modeling. Here we investigated its performance for generation of highly osteogenic bone graft using polycaprolactone-tricalcium phosphate (PCL-TCP) scaffolds seeded with human fetal mesenchymal stem cell (hfMSC). hfMSC scaffolds were cultured in either bioreactor or static cultures, with assessment of cellular viability, proliferation and osteogenic differentiation it) vitro and also after transplantation into immunodeficient mice. Compared to static culture, bioreactor-cultured hfMSC scaffolds reached cellular confluence earlier (day 7 vs. day 28), with greater cellularity (2x, p < 0.01), and maintained high cellular viability in the core, which was 2000 Inn from the surface. In addition, bioreactor culture was associated with greater osteogenic induction, ALP expression (1.5x P < 0.01), calcium deposition (5.5x, p < 0.001) and bony nodule formation on SEM, and in-vivo ectopic bone formation in immunodeficient mice (3.2x, p < 0.001) compared with static-cultured scaffolds. The use of biaxial bioreactor here allowed the maintenance of cellular viability beyond the limits of conventional diffusion, with increased proliferation and osteogenic differentiation both in vitro and in vivo, suggesting its utility for bone tissue engineering applications. (C) 2009 Elsevier Ltd. All rights reserved.

Related URLs:
<Go to ISI>://WOS:000264953900005

Effects of simulated microgravity on embryonic stem cells

by cfynanon 9 June 2015in Biology & Biotechnology No comment

There have been many studies on the biological effects of simulated microgravity (SMG) on differentiated cells or adult stem cells. However, there has been no systematic study on the effects of SMG on embryonic stem (ES) cells. In this study, we investigated various effects (including cell proliferation, cell cycle distribution, cell differentiation, cell adhesion, apoptosis, genomic integrity and DNA damage repair) of SMG on mouse embryonic stem (mES) cells. Mouse ES cells cultured under SMG condition had a significantly reduced total cell number compared with cells cultured under 1 g gravity (1G) condition. However, there was no significant difference in cell cycle distribution between SMG and 1G culture conditions, indicating that cell proliferation was not impaired significantly by SMG and was not a major factor contributing to the total cell number reduction. In contrast, a lower adhesion rate cultured under SMG condition contributed to the lower cell number in SMG. Our results also revealed that SMG alone could not induce DNA damage in mES cells while it could affect the repair of radiation-induced DNA lesions of mES cells. Taken together, mES cells were sensitive to SMG and the major alterations in cellular events were cell number expansion, adhesion rate decrease, increased apoptosis and delayed DNA repair progression, which are distinct from the responses of other types of cells to SMG.

Related URLs:
http://ovidsp.ovid.com/ovidweb.cgi?T=JS&CSC=Y&NEWS=N&PAGE=fulltext&D=emed10&AN=2011698685
http://sfxhosted.exlibrisgroup.com/mayo?sid=OVID:embase&id=pmid:&id=doi:10.1371%2Fjournal.pone.0029214&issn=1932-6203&isbn=&volume=6&issue=12&spage=e29214&pages=&date=2011&title=PLoS+ONE&atitle=Effects+of+simulated+microgravity+on+embryonic+stem+cells&aulast=Wang&pid=%3Cauthor%3EWang+Y.%3C%2Fauthor%3E&%3CAN%3E2011698685%3C%2FAN%3E

Formation and differentiation of three-dimensional rat marrow stromal cell culture on microcarriers in a rotating-wall vessel

by cfynanon 9 June 2015in Biology & Biotechnology No comment

Using a high aspect ratio vessel (HARV), this study investigated the formation of 3-D rat marrow stromal cell culture on microcarriers and the expression of bone-related biochemical markers under conditions of simulated microgravity. In addition, it calculated the shear stresses imparted on the surface of microcarriers of different densities by the medium fluid in an HARV. Secondary rat marrow stromal cells were cultured on two types of microcarriers, Cytodex-3 beads and modified bioactive glass particles. Examination of cellular morphology by scanning electron microscopy revealed the presence of three-dimensional multicellular aggregates consisting of multiple cell-covered Cytodex-3 microcarriers bridged together. Mineralization was observed in the aggregates. Spherical cell-bead aggregates were observed in an HARV, while cell-bead assemblies were mostly loosely packed in a chainlike or branched structure in a cell bag. The expressions of alkaline phosphatase activity, collagen type I, and osteopontin were shown via the use of histochemical staining, immunolabeling, and confocal scanning electron microscopy. Using a numerical approach, it was found that at a given rotational speed and for a given culture medium, a larger density difference between the microcarrier and the culture medium (e.g., a modified bioactive glass particle) imparted a higher maximum shear stress on the microcarrier.

Related URLs:
<Go to ISI>://WOS:000072925300005

[Cytoskeleton structures and adhesion properties of human stromal precursors under conditions of simulated microgravity]

by cfynanon 9 June 2015in Biology & Biotechnology No comment

Cytoskeletal alterations occur is several cell types including lymphocytes, glial cells, and osteoblasts, during spaceflight and under simulated microgravity (SMG). One of the potential mechanisms of cytoskeletal gravisensitivity of a cell is disruption of extracellular matrix and integrin interactions. Therefore, we investigated the effects of SMG on F-actin cytoskeleton structure, vinculin focal adhesions, expression of some integrin subtypes and cellular adhesion molecules in mesenchymal stem cells (hMSCs) derived from human bone marrow in vitro. Simulated microgravity was produced by RPM (manufactured by Dutch Space, The Netherlands). Culture flasks with MSCs were settled on the inner platform of RPM for 30 minutes, 6, 24, 48 and 120 h. The results have shown that actin cytoskeleton is very fast reorganized even after 30 minutes of simulated microgravity. The number of cells with disrupted actin cytoskeleton steadily increased with the increasing exposure time. However these changes were reversible because the cells partly recovered their F-actin structure after 120-hours-exposure. In addition, we observed vinculin redistribution inside the cells after 6 hours and subsequent terms of exposures. This process was accompanied with increasing of fluorescence intensity of cells. The expression of integrin-alpha2 increased after exposure in RPM. Also we observed decrease in the number of VCAM-1-positive cells and changes in the expression of ICAM-1. Thus, our findings indicate that SMG leads to reversible microfilament alterations of hMSCs and alters adhesion properties of this type of cells.

Related URLs:
http://ovidsp.ovid.com/ovidweb.cgi?T=JS&CSC=Y&NEWS=N&PAGE=fulltext&D=medl&AN=20058807
http://sfxhosted.exlibrisgroup.com/mayo?sid=OVID:medline&id=pmid:20058807&id=doi:&issn=0041-3771&isbn=&volume=51&issue=11&spage=896&pages=896-904&date=2009&title=Tsitologiia&atitle=%5BCytoskeleton+structures+and+adhesion+properties+of+human+stromal+precursors+under+conditions+of+simulated+microgravity%5D.&aulast=Gershovich&pid=%3Cauthor%3EGershovich+PM%3C%2Fauthor%3E&%3CAN%3E20058807%3C%2FAN%3E

Attachment to Cytodex Beads Enhances Differentiation of Human Retinal Progenitors in 3-D Bioreactor Culture

by cfynanon 9 June 2015in Biology & Biotechnology No comment

Retinal degenerations are the leading cause of genetically inherited blindness. One of the strategies currently being tested for the treatment is cell/tissue transplantation. As such stem cells and tissue engineered constructs are of great importance. This report describes the growth of multipotential human retinal progenitors (cell line) in a 3-D bioreactor culture vessel with (adhesive substrate) laminin coated collagen 1/cytodex beads and without adhesive substrate (beadless culture). The study demonstrates that progenitors are capable of growth and differentiation in the bioreactor with or without beads. The presence of adhesive substrate accelerates and enhances photoreceptor differentiation in the bioreactor, reflected by significantly higher level expressions of several photoreceptor specific proteins; N acetyl transferase (AaNat), rhodopsin and cone transducin GNB3. Both monomeric and dimeric forms of rhodopsin are expressed in cells attached to beads, whereas, only the monomeric form is expressed in beadless culture. Similarly, a different isomeric form of tyrosine hydroxylase (a doublet) is expressed in cell bead attached cultures. Co-culturing retinal progenitors with retinal pigment epithelium (RPE) in cell bead cultures further stabilizes the photoreceptor phenotype and rhodopsin expression. Most of the retinal neuronal phenotypes are confirmed by an expression of specific proteins. The adhesive substrate in the form of collagen 1, laminin coated cytodex beads, could be just an effector for stabilization or a positive signal, modulating extracellular matrix (ECM) molecules and/or neurotrophins. In the future, the bioreactor culture system could be utilized to grow retina-like structures from ciliary epithelium by incorporating biodegradable substrates.

Related URLs:
<Go to ISI>://WOS:000301640300005

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

by cfynanon 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.

Related URLs:
<Go to ISI>://WOS:000229063800018

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