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目的研究去血清损伤中PKC信号通路的改变,同时阐明神经节苷脂对大鼠嗜铬细胞瘤细胞株(PC12)细胞去血清损伤的保护作用及其作用机制。方法采用去血清损伤24h作为损伤模型,MTT测定细胞存活率,Hoechst 33258/PI观察细胞损伤后的死亡类型及坏死、凋亡的细胞数;Western blotting检测PCl2细胞在损伤后细胞浆和细胞膜PKC蛋白的变化。结果去血清损伤时间依赖性对PC12细胞有损伤,多数细胞呈凋亡征象,而神经节苷脂对去血清损伤有明显的保护作用;去血清损伤时,PKC信号通路被活化,主要表现为PC12细胞胞浆中的PKC蛋白减少,胞膜上的PKC蛋白逐渐增加,实现PKC蛋白转位;而神经节苷脂可以抑制这种转位,从而起到保护作用。结论神经节苷脂对去血清损伤有保护作用,其保护作用部分是由于神经节苷脂抑制PKC信号通路的活化。
Objective To study the changes of PKC signaling pathway in serum-deprived rats and to elucidate the protective effect of gangliosides on serum-depleted rat pheochromocytoma cell line (PC12) and its possible mechanism. Methods The decellularized serum was harvested for 24 h. The cell viability was determined by MTT assay. The type of cell death and the number of necrotic and apoptotic cells were observed by Hoechst 33258 / PI. The expression of PKC protein in cytoplasm and membrane of PC12 cells was detected by Western blotting The change. Results Time-dependent degeneration of serum could damage PC12 cells, and most of the cells showed signs of apoptosis, while gangliosides had a protective effect on serum-deprivation injury. PKC signaling pathway was activated when de-serum was damaged, which was mainly PC12 PKC protein in the cytoplasm decreased, the membrane protein PKC gradually increased, to achieve PKC protein translocation; and ganglioside can inhibit this translocation, which play a protective effect. Conclusion Gangliosides have a protective effect on de-seruming injury, and its protective effect is partly due to inhibition of PKC signaling by gangliosides.