麦冬皂苷D对氧糖剥夺/复氧后新生鼠大脑皮层离体星形胶质细胞HIF-1 α-VEGF通路的作用

Effects of ophiopogonin D on the HIF-1α-VEGF path of neonatal rat cerebral cortex astrocytes in vitro after oxygen glucose deprivation and reperfusion

目的:探讨麦冬皂苷D对氧糖剥夺/复氧(OGD/R)后星形胶质细胞HIF-1α-VEGF通路的作用。方法:分离培养和鉴定新生大鼠星形胶质细胞,建立星形胶质细胞OGD/R模型;设正常组、模型组、麦冬皂苷D给药组;在OGD4h/R2h、4h、6h后用ELISA检测星形胶质细胞细胞培养液中VEGF蛋白表达量,Western Blot检测星形胶质细胞胞核内HIF-1α蛋白的含量,免疫荧光检测星形胶质细胞及其在OGD/R后细胞的光密度、面密度和阳性细胞数量。结果:模型组VEGF、HIF-1α蛋白含量高于正常组(P<0.05,P<0.01),麦冬皂苷D给药组高于模型组(P<0.05,P<0.01),且随再灌注时间延长,VEGF、HIF-1α蛋白含量呈增加趋势。模型组星形胶质细胞的面密度、光密度、阳性细胞个数较正常组均明显降低(P<0.01),麦冬皂苷D给药组星形胶质细胞的面密度、光密度、阳性细胞个数较模型组均显著升高(P<0.05,P<0.01)。结论:麦冬皂苷D能够促进OGD/R后新生鼠离体星形胶质细胞的增殖;麦冬皂苷D能够明显上调OGD/R后新生鼠离体星形胶质细胞VEGF、HIF-1α的蛋白含量;麦冬皂苷对OGD/R后神经损伤的保护及修复作用可能是通过星形胶质细胞HIF-1α-VEGF通路实现的。

Objective： To discuss the effects of ophiopogonin D on the HIF-1α -VEGF path of neonatal rat cerebral cortex astrocytes in vitro after oxygen glucose deprivation and repeffusion. Methods： Isolated ,cultured and identified neonatal rat astrocytes, established the OGD/R model of neonatal rat astrocytes in vitro; set up normal group, model group, and ophiopogonin D treatment group, after oxygen glucose deprivation 4h and reperfusion 2h, 4h and 6h, analyzed separately the expression levels of VEGF protein in the culture supernatant of astrocytes by ELISA, HIF-1α protein in the nucleus of astrocytes by Western Blot, and the optical density, areal density and the number of positive cells of OGD/R astrocytes by double-labelling immunofluorescence. Results： The content of VEGF and HIF-1α in model group was higher than the normal （P〈0.05, P〈0.01）, while ophiopogonin D treatment group was higher than the model （P〈0.05, P〈0.01）, following reperfusion time lasted, both the expression levels of VEGF and HIF-1α appeared upward trend. Corresponding to this, all of the optical density, areal density and the number of positive cells of astrocytes in model group obviously decreased compared with the normal （P〈0.01）, and ophiopogonin D treatment group remarkably increased compared with the model, differences of which had statistical significance. Conclusion： ophiopogonin D could promote the proliferation of neonatal rat cerebral cortex astrocytes in vitro after OGD/R. Ophiopogonin D could raise up the expression levels of VEGF and HIF- 1α of neonatal rat cerebral cortex astrocytes in vitro after OGD/R; the protective and adjective effects of ophiopogonin D on nerve injury after oxygen glucose deprivation and reperfusion achieved potentially though HIF-1α -VEGF path of neonatal rat cerebral cortex astrocytes in vitro.