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目的研究热休克蛋白70(Hsp70)在苯并(a)芘(BaP)所致DNA损伤修复中的保护作用。方法转染人支气管上皮细胞(16HBE)含有Hsp70基因(hsp70)的质粒,以增高Hsp70的表达,以正常培养和转染载体质粒细胞作为对照,分别为Hsp70过表达组(16HBE/hsp70)、对照组(16HBE)和转染对照组(16HBE/pcDNA)。用16μmol BaP染毒上述3组细胞2 h,恢复不同的时间(0,2,4,8,24 h),碱性单细胞电泳(SCGE)检测DNA修复。转染被不同浓度(0,10,20,30,40μmol)BaP二氢二醇环氧化物(BPDE)损伤的荧光素酶报告基因质粒,检测不同组细胞对质粒的修复程度。以Olive尾距(OTMs)和相对荧光素酶活力评价DNA修复能力的差异。结果转染增高了Hsp70表达量约80%。3组细胞OTMs值均随恢复时间延长而降低。相对于对照组和转染对照组,Hsp70过表达组的OTMs值在恢复前2 h降低更迅速,且在恢复的各个时间点均小于对照组,差异均有统计学意义(P<0.01)。Hsp70过表达组的荧光素酶活力在恢复的各个时间点始终大于对照组和转染对照组,且在10μmol时,P<0.01,在20μmol时,P<0.05。结论Hsp70能促进细胞对BaP造成DNA损伤的修复
Objective To study the protective effect of heat shock protein 70 (Hsp70) on DNA damage induced by benzo [a] pyrene (BaP). Methods The plasmid containing Hsp70 gene (hsp70) was transfected into human bronchial epithelial cells (16HBE) to increase the expression of Hsp70. The normal cultured and transfected plasmid vector was used as control group, which were respectively Hsp70 overexpressing group (16HBE / hsp70) Group (16HBE) and transfection control group (16HBE / pcDNA). The above three groups of cells were treated with 16μmol BaP for 2 h, and then recovered for different time (0, 2, 4, 8, 24 h). DNA repair was detected by alkaline single cell electrophoresis (SCGE). The plasmids were transfected with luciferase reporter plasmids damaged by BaP dihydropyrane epoxide (BPDE) at different concentrations (0, 10, 20, 30, 40μmol). Differences in DNA repair capacity were evaluated using Olive tail length (OTMs) and relative luciferase activity. Results Transfection increased Hsp70 expression by about 80%. The OTMs of the three groups decreased with the prolongation of recovery time. Compared with the control group and the transfected control group, the OTMs of Hsp70 overexpression group decreased more rapidly 2 hours before recovery and were lower than those of the control group at various time points (P <0.01). The luciferase activity of Hsp70 overexpression group was always higher than that of control group and transfected control group at each time points of recovery, and P <0.01 at 10μmol, P <0.05 at 20μmol. Conclusion Hsp70 can promote the repair of DNA damage caused by BaP in cells