葡萄糖调节蛋白75对缺糖诱导的凋亡相关基因Bax和NF-κB改变的影响

Effect of Grp75 on the alteration of apoptosis related gene Bax and NF-κB induced by glucose deprivation

目的通过已获得稳定表达葡萄糖调节蛋白（Grp75）的PC12细胞株,检测Grp75对缺糖诱导的细胞凋亡过程中Bax和NF-κB的影响。方法Grp75过表达组和对照组细胞无糖培养6h、12h2、4h和48h后,进行相关的实验。免疫印迹法检测缺糖状态下两组细胞中Grp75的表达水平和NF-κB的活性;应用半定量RT-PCR和免疫印迹法比较Bax表达水平的变化;免疫细胞化学通过构象特异性的Bax 6A7抗体检测Bax的活化。结果Bax活化和NF-κB活性的下降在缺糖诱导的PC12细胞凋亡过程中发挥了重要的作用。而Grp75通过阻止Bax的活化和NF-κB活性的下降抑制缺糖诱导的凋亡。缺糖状态下Grp75过表达组和对照组细胞中Bax表达水平均未发生改变。结论Bax活化和NF-κB活性下降与缺糖诱导的PC12细胞凋亡关系密切,Grp75通过阻止Bax的活化和维持NF-κB的活性保护PC12细胞。

Objective To study the effect of glucose regulated protein 75（Grp75） on the alteration of Bax and NF-kB induced by glucose deprivation through the stably transfected PC12 ceils with Grp75. Methods The ceils of Grp75-overexpressing group and control group incubated in glucose-free DMEM medium for indicated time （6, 12, 24 and 48hours）. The expression level of Grp75, Bax and the activity of NF-kB were determined by Western blotting, and the expression level of Bax was determined by semi-quantitative RT-PCR and Western blotting. Immunocytochemistry was performed using a conformation specific anti-Bax （6A7） antibody to detect the activation of Bax. Results The activation of Bax and the decline of NF-kB activity played important roles in the apoptosis of PC12 cells induced by glucose deprivation. Grp75 inhibited the apoptosis induced by glucose deprivation through inhibition of the activation of Bax and the decline of NF-kB activity. There was no change in Bax expression level under glucose deprivation in two groups. Conclusion The activation of Bax and the decline of NF-kB activity were associated with apoptosis of PCI2 cells induced by glucose deprivation, and Grp75 provided protection to PC12 cells through inhibition of activation of Bax and maintaining activation of NF-kB.