Isolation, culture and identification of gut neural crest stem cells of postnatal mouse in vitro

AIM: To establish a simplified culture system to isolate and passage gut neural crest stem cells (GNCSCs) from postnatal mouse, observe their proliferation and differentiation, and determine the expression of GNCSCs and their derivate antigens-p75,GFAP,Peripherin and α-Actin. METHODS: The guts of postnatal 1-,5-day mice were dissociated and triturated into single cells which were seeded into serum-free DMEM/F12 medium to observe the formation of neurospheres. After 10% fetal bovine serum was added into the medium, the differentiation of neurospheres was identified with immunocytochemistry and immunofluorescence staining. RESULTS: A minority of gut cells formed neurospheres in the simplified serum-free medium. The staining of neurospheres and their derivatives indicated neurospheres could generate neurons, glia cells and smooth muscle cells under induction. CONCLUSION: GNCSCs, with the capacities of self-renew and proliferation and expressing p75, exist in the postnatal mouse gut. They can be isolated and cultured in vitro, and can differentiate into three types of cells that are assential for gut nerve system.