Effect of glutamic acid on the nestin protein expression of neural stem cells from embryonic rat

Objective To investigate the effects of glutamic on the nestin protein expression of cultured neural stem cells(NSCs),and to provide a theoretical basis to further clarify the signaling pathways of proliferation and differentiation of NSCs and clinical nerve regeneration after injury.Methods The NSCs obtained from subgranular zone(SGZ) of mice's embryon were cultured in the serum-free culture medium,which was added basic fibroblast growth factor(bFGF) and epidermal growth factor(EGF).Then the fluorescence immunocytochemistry was adopted to detect the expression of NSCs specific factor Nestin,the proliferation of NSCs were also detected by 5-Bromo-2-deoxy Uridine(BrdU) method;The differentiation of NSCs were determined by the specific marker glial fibrillary acidic protein(GFAP),which was detected after7 days treatment by the cell culture medium containing 10% FBS.After 3 passages,the NSCs were assigned to six groups(group A,B,C,D,E,F).Group A was control group,and group B,C,D,E,F were treated by gradient concentrations of glutamic,the proliferation of each group was detectd and polymerase chain reaction(PCR) was performed to identify the proliferation result of NSCs.The group difference was determined by statistics.Results(1) Cell culture:the NSCs,isolated from the embryonic mice's cerebral cortex,were successfully cultured and could continuously proliferate and then formed the specific neurospheres.The results of immunofluorescence showed that both of the primary and the passaged neurospheres were all Nestin positive.The GFAP or BrdU positive were also detected in the differentiated neural stem cells.(2) Reverse transcription-polymerase chain reaction(RT-PCR) :The significant difference of Nestin mRNA expression existed in various groups(P 0.05),the extent of its expression showed decreasing gradually:Group F(2.21 ± 0.42),Group E(1.99 ± 0.37),Group D(1.58 ± 0.26),Group C(1.36 ±0.41),Group B(1.05 ± 0.34),Group A(0.83 ± 0.21).Conclutions Nestin mRNA in the glutamic group and control group were expressed,and its expression level increased with the higher concentration of glutamic.