预诱导HSP90α对HepG2细胞热应激的影响

Effects of pre-induction of HSP90αon the heat stress in HepG2 cells

目的探讨预诱导热休克蛋白90α(HSP90α)后,细胞热应激反应的变化,并探索诱导细胞热习服的策略。方法将人肝细胞癌细胞系HepG2培养于42℃细胞培养箱制作热应激模型,用不损伤细胞活性浓度的HSP90抑制剂17-DMAG诱导HSP90α表达增多,通过流式细胞术检测细胞活性氧(ROS)含量和凋亡,CCK-8检测细胞活性,Western blot检测蛋白变化。结果热应激0.5~4 h后,细胞内ROS含量逐渐升高(F=332.22,P<0.01);在37℃恢复至24 h后,热应激1~4 h组细胞内γH2AX较0 h组显著增多,同时细胞活性也随热应激时间延长而降低(F=59.97,P<0.01),热应激4 h组活性只有0 h组的(71.39±2.55)%;使用5 nmol/L 17-DMAG预处理24 h诱导HSP90α表达增多后,能显著减少热应激后的γH2AX和细胞凋亡率(F=62.79,P<0.01),使热应激4 h组细胞凋亡率从(26.06±2.48)%降至(15.93±1.67)%。结论通过诱导HSP90α表达增多能减少细胞热应激后的DNA损伤和凋亡,有利于细胞快速达到热习服。

Objective To investigate the responses to heat stress in cells with pre-induction of HSP90α,and explore the strategy to induce thermal acclimation of cells.Methods Human hepatocellular carcinoma cell line HepG2 was cultured at 42℃to create a heat stress model.Expression of HSP90αwas pre-induced by 17-DMAG,an HSP90 inhibitor,at a concentration that does not damage cell viability.Reactive oxygen species(ROS)and apoptosis were detected by flow cytometry.Cell viability was detected by CCK-8,and protein changes were detected by Western blot.Results After heat shock for 0.5-4 h,the content of intracellular ROS gradually increased(F=332.22,P<0.01).After recovery to 24 h at 37℃,moreγH2AX was observed in the 1-4 h heat shock group.Meanwhile,cell viabilities were attenuated as heat shock prolonged(F=31.94,P<0.01),and 4 h heat shock decreased cell viability to(71.39±2.55)%of control group.After the expression of HSP90αwas pre-induced by 5 nmol/L 17-DMAG for 24 h,heat shock for 4 h at 42℃followed by a recovery at 37℃to 24 h,γH2AX was decreased compared with cells without HSP90αpre-induction;and the apoptosis rate of cells were also decreased,from(26.06±2.48)%to(15.93±1.67)%(F=62.79,P<0.01).Conclusion Pre-induction of HSP90αcan reduce cells’DNA damage and apoptosis after heat stress,help⁃ing cells obtain rapid heat acclimation.