氯倍他索对大鼠神经干细胞自发分化的影响

Effects of clobetasol on spontaneous differentiation of rat neural stem cells

目的:研究氯倍他索(Clo)对神经干细胞自发分化的影响,为Clo治疗中枢性神经损伤提供医学理论基础。方法:利用无血清悬浮法分离培养及鉴定大鼠NSCs;选取第三代NSCs,以含10%胎牛血清完全培养基贴壁诱导分化7 d,培养基中各加入同体积二甲基亚砜、2. 5、5μmol/L的Clo;免疫荧光染色和Western Blot方法检测NSCs分化后轴突膜蛋白(GAP-43)、酸性胶质蛋白(GFAP)、髓磷脂碱性蛋白(MBP)表达情况,进一步探讨Clo的作用机制。结果:分离培养的NSCs高表达Nestin; Clo作用组表达GAP-43和MBP较对照组明显提高(P <0. 05),GFAP表达水平低于对照组(P <0. 05),Clo作用组表达音猬因子(SHH)和锌指蛋白(Gli 1)均高于对照组(P <0.05)。结论:Clo可在体外通过SHH信号通路促进NSCs向神经元细胞和少突胶质细胞分化,与此同时Clo可抑制NSCs向星形胶质细胞分化。

Objective： To study the effects of clobetasol（ Clo） on the spontaneous differentiation of rat neural stem cells,provide a theoretical basis for Clo to treat central nerve injury.Methods： The serum-free suspension method was used to isolate and culture the rat NSCs.The third-generation NSCs were selected and adhered to the 10% FBS serumcontaining medium to induce differentiation for 7 days.Clobetasol 0（ Same volume of Dimethyl sulfoxide）,2.5 and 5μmol/L were added to the medium.Immunofluorescent staining and Western Blot methods were used to detect the expression of growth associated protein-43（ GAP-43）,glial fibrillary acidic protein（ GFAP） and myelin basic protein（ MBP）after differentiation of NSCs,and Nestin was used to identify NSCs.Results： Nestin was highly expressed in NSCs.The expression of GAP-43 and MBP in Clo treated group was significantly higher than those in control group（ P〈0.05）; the expression level of GFAP was lower than that in control group（ P〈0.05） and the expression of sonic hedeahog（ SHH）and Gli 1 were higher than those of control group（ P〈0.05）.Conclusion： Clo could promote the differentiation of NSCs into neurons and oligodendrocytes through the SHH signal pathway in vitro,and meanwhile while inhibit the differentiation of NSCs into astrocytes.