热休克反应对氧化应激所致nucleolin裂解的影响

观察热休克反应对氧化应激（0.5 mmol/L H2O2）诱导细胞凋亡发生时 nucleolin(C23)裂解的影响并探讨其机制。方法： 采用caspase-3 活性定量分析及免疫印迹技术检测氧化应激诱导的原代心肌细胞、RAW 264.7巨噬细胞 及K562白血病细胞凋亡及C23的裂解；通过热休克反应观察热休克反应对C23裂解的影响。结果： 0.5 mmol/L H2O2处理细胞2 h caspase-3活性显著升高，12 h达高峰。免疫印迹结果显示上述各种细胞的C23蛋白均发生了裂解（有80 kD裂解片断出现），而且最早出现裂解片段的时间都在H2O2处理30 min到1 h左右；同时热休克反应通过诱导HSP70，HSP25等应激蛋白表达而显著减少氧化应激所致C23的裂解。结论： 氧化应激诱导凋亡发生的同时也引起C23的裂解；热休克反应显著抑制氧化应激所致C23的裂解，其机制与热休克反应诱导多种热休克蛋白表达有关。

Effect of heat shock response on the cleavage of nucleolin induced by oxidative stress

Objective To observe the cleavage of nucleolin (C23) during apoptosis induced by oxidative stress and to clarify the effect of heat shock response (HSR) on the cleavage of nucleolin and its possible molecular mechanism. Methods We added 0.5 mmol/L peroxide hydrogen (H 2O 2) into cultured cells to mimic oxidative stress. Apoptosis and cleavage of C23 were detected using caspase-3 colorimetric assay and Western blotting respectively. HSR was performed to observe the effect of HSR on cleavage of C23 induced by oxidative stress, and over-expressions of HSP70 and HSP25 were detected by Western blotting.Results Activity of caspase-3 increased significantly after 2 hours of 0.5 mmol/L H 2O 2 treatment, and reached the peak after 12 hours. The cleavage of C23 appeared 30 minutes to 1 hour after the treatment of H 2O 2 as indicated by a cleaved fragmentation of 80 kD, which was significantly inhibited by HSR. Moreover, HSR could induce HSP70 and HSP25 over-expressions. Conclusion Oxidative stress can induce the activation of caspase -3, cleavage of C23, and apoptosis. HSR can significantly inhibit the cleavage of C23 induced by oxidative stress, which is related to the over-expressions of HSP70, HSP25, and other stress proteins.