Abstract:

OBJECTIVE: To investigate the effects of human cerebrospinal fluid (CSF) during development phase on migration and differentiation of fetal brain neural stem cells (NSCs).
METHODS: Fetal brain cells of gestational age of 16 weeks that were frozen in liquid nitrogen were obtained, resuscitated and incubated in DMEM/F12 medium containing epithium growth factor (EGF), basic fibroblast growth factor (bFGF), B27 and N2. The neurospheres cultured for 14 days were obtained. CSF was absorbed from the subarachnoid cavity and brain ventricle in the embryonic group. CSF was collected by lumbar puncture or ventricular puncture in the child group. The neurospheres cultured for 14 days were transplanted into the pure CSF in an incubator containing 5% CO2 at 37 ℃. Cellular migration and growth of neurospheres in CSF were observed. Effects of CSF on neural cell differentiation were identified by immunofluorescence.
RESULTS: Neural stem cells in the form of neurospheres derived from fetal brain were inoculated into the pure CSF, and cell migration were commonly observed besides few of neurospheres in child CSF culture at 6 hours following culture. Surrounding cells of neurospheres extended processes, forming cell cord that became cell webs after extension. Compared with the embryonic group, positive rate of glial fibrillary acidic protein was significantly increased in the children group (P < 0.01), but positive rates of nerve fiber and nestin were significantly decreased (P < 0.01). In addition, galactocerebroside-positive cells were only found in 3 baby CSF cultures.
CONCLUSION: There existed significant affections on both migration and differentiation of human neural stem cells when cultured in pure CSF with different developmental phase, suggesting that CSF is one of major niche factors for central neural system development.