Identification of proliferation and differentiation capability of hippocampal neural stem/progenitor cells by adherent culture / 中国组织工程研究

ABSTRACT

BACKGROUND: In vitro isolation and culture of neural stem/progenitor cells will provide a good cell model for the study of neurodevelopment, neurological diseases, and neural transplantation. OBJECTIVE: To study the highly effective method for isolation and expansion of hippocampal neural stem/progenitor cells from newborn mice, and to identify the proliferation and differentiation of hippocampal neural stem/progenitor cells using improved adherent culture method. METHODS: Neural stem/progenitor cells were isolated from the hippocampus of newborn C57BL/6 mice and were expanded for several passages. Combination of polylysine and laminin were used for adherent culture to promote cell attachment. Morphological observation and immunofluorescence cytochemical staining were conducted to detect the expression of neural progenitor-specific marker protein Nestin and proliferation index Ki-67. After 7 days of induction and differentiation, the expression of GFAP, DCX, Tuj1 and S100β was detected by immunofluorescence. RESULTS AND CONCLUSION: About 82％ of the cultured neural stem/progenitor cells expressed Nestin, and about 49％ expressed Ki-67. A small number of cells were DCX-positive neurons after induction and differentiation, while most of the cells were positive for GFAP. The ratio of neurons to astrocytes was 11.7 identified by Tuj1 and S100β double staining. The neural stem/progenitor cells derived from the hippocampus were efficiently isolated and cultured. The cell proliferation and differentiation abilities were effectively identified after adherent culture, which can provide sufficient cell sources for further experimental research.