海马神经干细胞不同传代方法的比较

海马神经干细胞不同传代方法的比较

李梅;王湘臻;徐铁军

【期刊名称】《中国组织工程研究》

【年(卷),期】2011(015)006

【摘要】BACKGROUND: Neural stem cells (NSCs) cultured in vitro can form sphere due to its proliferation characteristics during suspension culture. How to isolate cell spheres into single cells is a problem facing us during subculture.OBJECTIVE: To establish an ideal subculture method for harvesting NSCs from the rat hippocampus to obtain a large number of NSCs for investigation.METHODS: NSCs were isolated from neonatal 1-day rat hippocampus. After a growth of typically 5-6 days, primary neurospheres were passaged by mechanical dissociation, trypsin, TrypLE or Accutase digestion. Each method was used to dissociate neurospheres for three passages every 7 days. The ratio of cell viability and neurosphere numbers were estimated on day 1 and day 4 respectively after passage. This experiment was repeated three times.RESULTS AND CONCLUSION: Three enzymatic dissociation each obtained single-cell suspension; while mechanical dissociation resulted in mixtures of single cells and small neurospheres. The cell viability after Accutase treatment was significantly higher than trypsin (P ＜ 0.01) and TrypLE digestion (P ＜0.05). Simultaneously, more newly formed neurospheres were generated by Accutase treatment as compared with