大鼠胚胎神经干细胞和PEP-1-SOD1融合蛋白联合培养及鉴定

摘要:目的　探讨神经干细胞（NSCs）和PEP-1-SOD1联合培养对NSCs生长分化的影响。方法　孕14 d的SD大鼠进行NSCs的分离培养及鉴定后，NSCs和细胞转导肽PEP-1介导铜，锌-超氧化物歧化酶（Cu，Zn-SOD，SOD1）形成的PEP-1-SOD1融合蛋白联合培养。实验共分为4组，空白组:正常培养的NSCs，不予处理。低浓度组:NSCs在0.5 μmol/L PEP-1-SOD1的无血清培养基中培养。中浓度组:NSCs在2.5 μmol/L PEP-1-SOD1的无血清培养基中培养。高浓度组:NSCs在4.5 μmol/L PEP-1-SOD1的无血清培养基中培养。MTT法和免疫组化检测联合培养后对细胞增殖影响及细胞分化标志蛋白表达。结果　与不同浓度PEP-1-SOD1联合培养48 h后NSCs细胞增殖较空白组均出现明显的增殖活跃，差异有统计学意义（P＜0.05）；中、高浓度组较空白组及低浓度组差异有统计学意义（P＜0.05）；但中、高浓度2组之间差异无统计学意义（P＞0.05）。结论　2.5 μmol/L的 PEP-1-SOD1为比较适宜的NSCs培养浓度，培养48 h最有利于NSCs增殖。

Co-culture and identification of neural stem cells and PEP-1-SOD1 from rat embryo

Abstract: Objective This work was to investigate the effect of co-culture on the in vitro growth and differentiation of neural stem cells and PEP-1-SOD1. Methods After the cultivation of neural stem cells in pregnant SD rats and identification, NSCs were co-cultured with PEP-1-SOD1 fusion protein, which was formed by cell penetrating peptide (CPP) interacting with copper, zinc -superoxide dismutase (SOD1), to observe the growth and differentiation of NSCs. The experiment was divided into 4 groups: the control group with the normal cultured NSCs without treatment, the low concentration group with NSCs in serum-free medium containing 0.5 μmol/L PEP-1-SOD1, the medium concentration group with 2.5 μmol/L PEP-1-SOD1 in medium, and the high concentration group with 4.5 μmol/L PEP-1-SOD1 in medium. MMT tests were conducted for the effect of co-culture on cell proliferation, and immunohistochemistry tests were conducted for the effect of co-culture on the expression of cell differentiation marker protein. Results Compared to the control group, the proliferation of NSCs was significantly improved after the co-culture with PEP-1-SOD1 at different concentrations for 48h (P<0.05). Compared to the control group and the low concentration group, cell proliferation was enhanced significantly (P<0.05) after the co-culture with 2.5 and 4.5 μmol/L PEP-1-SOD1, but there was no statistical difference between the control group and the low concentration group (P>0.05). Conclusion 2.5 μmol/L PEP-1-SOD1 is the suitable concentration for the co-culture with NSCs, and 48h is the most favorable to the proliferation of NSCs.