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目的:探讨多柔比星(doxorubicin)对乳腺癌MCF-7细胞表达DNA损伤修复相关蛋白乳腺癌易感蛋白1(breast cancer-associated protein 1,BRCA1)和多聚腺苷二磷酸核糖聚合酶-1[poly (ADP-ribose) polymerase-1,PARP-1]的影响。方法:蛋白质印迹法检测不同浓度的多柔比星干预并恢复不同时间后,MCF-7细胞表达BRCA1和PARP-1的水平及其PARP-1活性的动态变化。FCM法检测多柔比星和PARP-1抑制剂3-氨基苯酰胺(3-aminobenzamide,3-ABA)单独或联合干预后MCF-7细胞及SKBR3.0细胞(BRCA1突变型乳腺癌细胞)的凋亡情况。结果:随着多柔比星浓度的提高,PARP-1的活性产物聚腺苷二磷酸核糖[poly(ADP-ribose),PAR]逐渐增加(P<0.01),然而其全长表达(相对分子质量为1.13×105的片段)略有降低,断裂(相对分子质量为8.9×104的片段)有所增加;而BRCA1表达却呈现先增加后减少的趋势(P<0.01)。PARP-1活性随着恢复时间的延长逐渐升高(P<0.01),但全长PARP-1表达变化不大,断裂略有减弱;BRCA1表达却逐渐减少(P<0.01)。3-ABA能抑制PARP-1活性(P<0.01),诱导PARP-1断裂,但不影响BRCA1表达。多柔比星及3-ABA都能诱导MCF-7细胞凋亡,与对照组相比较,其差异均有统计学意义(P<0.05);多柔比星和3-ABA两者联合作用时能进一步增加BRCA1野生型乳腺癌细胞MCF-7的凋亡,与多柔比星及3-ABA单独使用相比较,差异具有统计学意义(P<0.05);多柔比星和3-ABA两者联合作用时能诱导BRCA1突变型乳腺癌细胞SKBR3.0发生凋亡,与两者联合作用MCF-7细胞的影响相比较,差异具有统计学意义(P<0.05)。结论:多柔比星干预能影响DNA损伤修复蛋白PARP-1的活性及BRCA1的表达。多柔比星和PARP-1抑制剂3-ABA都能诱导MCF-7细胞凋亡,两者联合应用能增加这种凋亡诱导效应。
Objective: To investigate the effects of doxorubicin on the expression of breast cancer-associated protein 1 (BRCA1) and poly (ADP-ribose) polymerase- 1 [poly (ADP-ribose) polymerase-1, PARP-1]. Methods: Western blotting was used to detect the changes of BRCA1 and PARP-1 expression and PARP-1 activity in MCF-7 cells treated with different concentrations of doxorubicin. MCF-7 cells and SKBR3.0 cells (BRCA1 mutant breast cancer cells) were treated with 3-aminobenzamide (3-ABA) and doxorubicin and PARP-1 inhibitor alone or in combination with FCM Apoptosis. RESULTS: With the increase of doxorubicin concentration, the poly (ADP-ribose) (PAR) activity of PARP-1 increased gradually (P <0.01) The fragment with a mass of 1.13 × 105 decreased slightly, and the fragment with a molecular weight of 8.9 × 104 increased, while the expression of BRCA1 increased first and then decreased (P <0.01). The activity of PARP-1 increased gradually with the prolongation of recovery time (P <0.01). However, the expression of PARP-1 did not change much and the fragmentation slightly decreased. The expression of BRCA1 decreased gradually (P <0.01). 3-ABA inhibited PARP-1 activity (P <0.01), induced PARP-1 cleavage but did not affect BRCA1 expression. Doxorubicin and 3-ABA could induce the apoptosis of MCF-7 cells, compared with the control group, the difference was statistically significant (P <0.05); when combined with doxorubicin and 3-ABA (P <0.05). Compared with doxorubicin and 3-ABA alone, the apoptosis of MCF-7 cells in BRCA1 wild-type breast cancer cells was further increased Could induce the apoptosis of SKBR3.0 in BRCA1 mutant breast cancer cells in combination with MCF-7 cells. The difference was statistically significant (P <0.05). Conclusion: Doxorubicin intervention can affect DNA damage repair protein PARP-1 activity and BRCA1 expression. Doxorubicin and PARP-1 inhibitor 3-ABA can induce apoptosis in MCF-7 cells, a combination of both can increase this apoptosis-inducing effect.