Biological Characteristics Of Mesenchymal Stromal Cell From SD Rat Among Different Ages

Because of bone marrow mesenchymal stem cells (Mesenchymal Stromal Cells, MSCs) have the ability of multiple differentiation, and easily be separated, in recent years, with the rise of genetic engineering, MSCs was considered as target cells for gene therapy to treat diseases which was difficult to cure, e.g. bone defects, degenerative disease, heart failure, etc. And the majority of the patients is elderly people, maybe it was concerned with the change of the growth and differentiation of the MSCs in aging body, so, it becomes even more important to discuss the biology characteristics of mesenchymal stem cells, and whether its function will change with age. This experiment based on comparative analysis on biological characteristics of MSCs between different ages of SD rats, to discuss the effects of the ages of MSCs. Objects:To study the biological characteristics of bone marrow mesenchymal stem cells of SD rat from different ages, and investigate the influence of age on the biological characteristics. Methods:Isolated MSCs of the three ages form the SD rat:young group(3weeks)、mild group(3months)、old group(14months). Use the L-DMEM with10%FBS to culture them in vitro amplification. The cultured EPCs were identified by Wright’s Giemsa staining for cellular morphology, Flow cytometry to identify cell surface antigen. Using the MTT to compare the proliferation of bone marrow mesenchymal stem cells of SD rat from different ages. Alizarin red semi-quantitative method is used to measure the deposition of calcium salts, so, find the difference among the MSCs of the three ages. Oil Red O quantification is used to compare the adipogenic capacity. By way of detecting the Alkaline Phosphatase (ALP) of MSCs, we can find the influence of age on MSCs osteogenesis ability.Result:(1) Identified by flow cytometry of cell surface antigens CD29and CD44positive, CD34and CD45negative, it can be proved that the adherent culture method can get the bone marrow mesenchymal stem cells.(2) Observed the cells under an inverted microscope, we can find that the primary MSCs of three ages show a colony-like growth. Once digest cells by trypsin, the cell won’t be the colony growth anymore, they are spindle growth. When they are in third generation, their form begin to become stable.(3) The MTT results show:when they are in third and sixth generation, No significant differences in proliferation, but when they grow up to ninth, twelfth and fifteenth, the young group is significantly better than the others.(4) The result of osteogenic differentiation and adipogenic differentiation display that the calcium nodules and lipid vacuoles which inducted by young group are more and better than the mild group, and the mild group is superior to the older group.(5) ALP test results showed that osteogenic ability of MSCs of young group is better than the middle-aged group, the middle-aged group is superior to the older group.Conclusion:The MSCs from the young, mild and old SD rats are all have strong proliferation, osteogenic differentiation and adipogenic differentiation, but the young group is the best among these, and the mild group is better than the old group.