Inducement Of Salvianolic Acid B On Proliferation And Differentiation Of Neural Stem Cells From Cerebra Of Mouse Embryos

Objective: To establish culture procedures of neural stem cells from cerebra of mouse embryoes, observe their growth and characteristics and investigate the effects of salvianolic acid B on their proliferation and differentiation ability.Methods: Neural stem cells were isolated from cerebra of mouse embryoes (embryonic and fetal day, ED13.5). Harvested cells were cultured in serum-free media containing basic fibroblast growth factor (bFGF) and B27. Observe the morphological characteristic of neural stem cells under optics. The growth kinetics of neural stem cells was measured by MTT method. Cell cycles were measured by flow cytometre staining with PI. Neural stem cells and their offspring induced by serum were identified with RT-PCR and Immunocytochemistry. Then, by using MTT assay, select the drugs efficacious concentrations in logarithmic growth phases of cell. Effect of salvianolic acid B on proliferation was measured with MTT assay, neurospheres count, and flow cytometre. Study their effect on differentiation of neural stem cells by immunofluorescence and RT-PCR.Results: (1) In this study, a culture model of neural stem cells was successfully set up. In this model, primary cells from E13.5 mouse cerebra were dissected out, and cultured as floating neurospheres. The number of cells did not increase almost in 2 days, and then entered logarithmic growth phases about 3 days, the doubling time of cells at the logarithm growth period was about 24 hours. The 90% of primary culture cells stayed in the G0/G1 phases. Primary cells and passages expressed nestin mRNA and protein in cytoplasm. After induced by serum for 4 hours, neural stem cells began to express neuron-specific gene tau and glial cells-specific gene GFAP. Typical...