Isolation of rat embryonic cortical neural stem cells and induction of differentiation

Neural stem cells (NSCs) play an essential role in the development of embryonic nervous system and have the capacity for self-renewal in the adulthood which may be important for normal functions of the CNS, such as learning, memory and response to injury. These cells exist at different stages of development in different areas of the CNS. The purpose of this study was isolation and confirmation of stem cell and induction of differentiation of the cells isolated from the 15 day old embryonic rat cortex. This was an experimental study on 15 day old embryonic rat cortices (n=6). 15 days after plug formation in female rats, the animals were transported to the laboratory. Under anesthesia and sterile condition, embryos were removed from the uterus. The meninges were separated by use of a stereoscope and the cortices of the embryos were dissected in HBSS buffer. Then the cortices were cut into small pieces and cultured in DMEM-F12 medium containing bFGF, EGF and B-27 supplement. The medium changed every day to keep the cells in an undifferentiated and proliferative state. DFN medium (DMEM-F12 and supplements N2) without growth factors was used for induction of differentiation. Immunocytochemical technique was used for confirmation of stem cells and detection of various neural cell types. Immunocytochemical evaluation revealed that, these cells were neural stem cells (nestin positive) and had the potential to differentiate in to the neuronal (expression of Beta tubulin III), oligodendrocyte (expression of OSP marker) or astrocyte (expression of GFAP marker). This is a reliable method for isolation of embryonic neural stem cells and considering their embryonic origin; they can be used to investigate the effect of various agents on the process of CNS development. Also they might be effective in the treatment of neurodegenerative lesions.