| Stem cells display a broad potential (often multipotential) for giving rise to diverse differentiated progenies, together with their extensive capacity for self-renewal. With an orderly chain of highly regulated processes involving cell proliferation, migration, differentiation, and maturation leading to the production and sustenance of most cell lineages in adult organisms. The stem cell is the earliest cell type on this chain. In addition to the hematopoietic and intestinal stem cells, considered for many years as paradigms of stem cells, adult organisms contain several other classes of stem cells, such as neural stem cells, epithelial stem cells, mesenchymal stem cells. In vivo and in vitro studies have identified the bone marrow mesenchymal stem cell as the source of multipotent stem cells that gives rise to progenitors for several mesenchymal tissues, including bone, cartilage, tendon, adipose, muscle , and hematopoietic-supporting stroma. hMSCS have a vast repertoire in the body tissues of umbilical blood, peripheral blood, bone and trabecular bone , pancreas, adipose and synovium tissue, what's more, experiments on the differntiation of hMSCS into other embryonic layers have also been carried out. The objective of this study is to analyze gene expression difference between hMSCS and hUVECS, to discuss the feasibility of induced endothelial differentiation from MSCS through gene transfection in vitro. At first,0.1% collagenase â… and 0.25% pancreatin were mixed in proportion of 1:1 to digest human umbilical vein. With this method, a large number of endothelial cells of high purity could be harvested quickly, which provided a reliable sample source for gene expression analysis with fewer subcultures and minor cell gene variation. Via the combined way of density gradient centrifugation and adherence screening, high homogeneous mesenchymal stem cells were secured. They may be kept in a homogeneous status of undifferentiation and high plasticity through high density cell plating and low ratio subculture.Immunocytochemistry, immunofluorescence and FACS, from the protein level, evaluated the purity of hMSCS and hUVECS and detected the cellular immunological basis of hMSCS to differentiate into endothelial cells. Transmission electronic microscope compared ultrastructural difference between hMSCS and hUVECS.Gene expression difference between them were analyzed in cDNA microarrays, which from mRNA level revealed the molecular basis of endothelial differentiation from hMSCS and provided important molecular reference for our experiment on the induced endothelial differentiation from hMSCS through VEGF165 transfection.We then transfected hMSCS with eukaryon expression plasmid pcDNA3.1/GS containing the target gene VEGF165 by liposome transfecting method, and had further detection. The results were as follows:1. The hUVECS acquired through collagenase-pancreatin mixture digestion were of high purity and large number.2. Primary hMSCS expressed a marker of early endothelial differentation and endothelial progenitor cells, KDR/Flk-1, which was associated with the early development of endothelial cells.3. There was no significant gene expression difference between hMSCS and hUVECS in 84% genes observed in the expression profile chips. In addition, hMSCS also express other early endothelial differentiation markers: ICAM-1 (Intercellular adhesion molecule 1 ), VCAM-1 (Vascular cell adhesion molecule 1), EDF-1 (Endothelial differentiation factor 1), MDG1 (Microvascular endothelial differentiation gene 1), EDG-2(Endothelial differentiation lysophosphatidic acid G-protein-coupled receptor, 2), et al. 4. hUVECS highly expressed 344 genes including CXCRF,VEGF, CTNNB1, Ang-2 et al,many of which were related to signal molecules and proteins. VEGF controlled their expression and biological activity. 5. Transfected with eukaryon expression plasmid pcDNA3.1/GS, which was inserted VEGF165, hMSCS were morphologically changed. The cell body was shortened, processes increased, and transformed into polyang...
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