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目的研究热休克预处理对过氧化氢诱导的细胞基因突变损伤是否具有保护效应。方法将V79细胞分为热休克处理组(热处理组)、过氧化氢损伤组(损伤组)和热休克预处理后过氧化氢损伤组(保护组)。采用6-巯基鸟嘌呤(6-TG)筛选次黄嘌呤磷酸核糖转移酶(HPRT)基因突变细胞并形成细胞克隆,应用Gi- emsa染色法计数突变细胞克隆数,计算各组细胞的相对克隆形成率及基因突变率。结果(1)温度>42℃,1h的热休克处理会诱导明显的HPRT基因突变,细胞相对克隆形成率降低,基因突变率与37℃处理1h组比较,差异具有统计学意义(P<0.01);温度≤42℃热休克处理1 h,不会诱导明显的HPRT基因突变,细胞相对克隆形成率高,基因突变率与37℃处理1 h组比较,差异无统计学意义(P>0.05)。(2)过氧化氢浓度≥0.5 mmol/L时,各亚组HPRT基因突变率与0 mmol/L亚组比较,差异均有统计学意义(P<0.05或0.01)。(3)细胞在接受热休克预处理后再行过氧化氢处理,过氧化氢浓度≤1 mmol/L的细胞没有发生明显的HPRT基因突变,与0 mmol/L亚组比较,差异无统计学意义(P>0.05);而过氧化氢浓度超过1 mmol/L时,细胞发生明显的HPRT基因突变,与0 mmol/L亚组比较,差异均有统计学意义(P<0.05或0.01)。(4)损伤组与保护组相应浓度的亚组(除0,4 mmol/L组)比较,差异均有统汁学意义(P<0.01)。结论热休克预处理对过氧化氢诱导的HPRT基因突变损伤细胞具有明显的保护作用。
Objective To investigate whether heat shock pretreatment has a protective effect against hydrogen peroxide-induced cellular gene mutation injury. Methods V79 cells were divided into heat shock group (heat treatment group), hydrogen peroxide injury group (injury group) and heat shock pretreatment hydrogen peroxide injury group (protection group). The 6th-thioguanine (6-TG) was used to screen the mutant cells of hypoxanthine phosphoribosyltransferase (HPRT) gene and the cell clones were formed. The number of mutant cells was counted by Giemsa staining and the relative clonality of each group was calculated Rate and mutation rate. Results (1) Heat shock treatment at temperature> 42 ℃ for 1 h could induce significant HPRT gene mutation and decrease the relative clone formation rate. The gene mutation rate was significantly higher than that at 37 ℃ for 1 h (P <0.01) ; Heat shock treatment at temperature below 42 ℃ for 1 h did not induce obvious HPRT gene mutation, and the relative clone formation rate was high. The mutation rate was not significantly different from that at 37 ℃ for 1 h (P> 0.05). (2) When the concentration of hydrogen peroxide was> 0.5 mmol / L, the mutation rate of HPRT gene in each subgroup was significantly different from that in 0 mmol / L subgroup (P <0.05 or 0.01). (3) Cells were treated with hydrogen peroxide after heat shock preconditioning, and no obvious HPRT gene mutation occurred in cells with hydrogen peroxide concentration ≤1 mmol / L. There was no significant difference between the two groups (P> 0.05). When the concentration of hydrogen peroxide exceeded 1 mmol / L, the gene mutation of HPRT gene was significantly different from that in 0 mmol / L subgroup (P <0.05 or 0.01). (4) The differences between the injury group and the protection group were significant (P <0.01) compared with the corresponding concentration of the protection group (except 0,4 mmol / L group). Conclusion Heat shock pretreatment has a significant protective effect on hydrogen peroxide-induced HPRT gene mutation in injured cells.