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Chkl的高表达可能是肿瘤对化疗药物的敏感性降低的重要因素之一,本研究的目的是观察siRNA干扰Chk1对人乳腺癌耐药细胞株MCF-7/adr(耐阿霉素)生长及细胞周期的影响,探讨Chk1在乳腺癌细胞耐药中的作用机制。采用RNAi技术抑制MCF-7/adr细胞中Chk1的表达。Westernblot检测转染前后细胞内Chk1蛋白表达情况,经阿霉素作用后,流式细胞术(FCM)检测其细胞周期分布及细胞凋亡率,MTT法检测细胞增殖。Western blot结果显示,Chk1 siRNA转染24h后,MCF-7/adr细胞中Chk1蛋白表达下降了67%,明显低于对照组和空载体转染组(P<0.05)。FCM法检测结果显示,同时,抑制Chk1的表达可解除阿霉素引起的G_2/M期阻滞;使阿霉素诱导的细胞凋亡率由转染前的(5.54±0.15)%上升到(22.24±0.13)%(P<0.05);在阿霉素浓度为0.4mg/L、4mg/L时,细胞的增殖活性分别下降13%、34%。提示siRNA干扰Chk1能够通过调控MCF-7/adr细胞周期及增殖从而增强乳腺癌细胞对阿霉素的敏感性,为临床上克服乳腺癌化疗耐药提供了新的作用靶点。
The high expression of Chkl may be one of the important factors that reduce the sensitivity of tumor to chemotherapeutic drugs. The purpose of this study is to observe the effect of siRNA on the growth of human breast cancer cell line MCF-7 / adr (resistant to doxorubicin) Cell cycle, explore the mechanism of Chk1 in breast cancer cell resistance. The RNAi technique was used to inhibit the expression of Chk1 in MCF-7 / adr cells. Western blot was used to detect the expression of Chk1 protein in the cells before and after transfection. After treated with doxorubicin, the cell cycle distribution and apoptosis rate were detected by flow cytometry (FCM) and the cell proliferation was detected by MTT assay. Western blot results showed that the expression of Chk1 protein in MCF-7 / adr cells decreased by 67% after transfected with Chk1 siRNA for 24 hours, which was significantly lower than that in control group and empty vector transfected group (P <0.05). FCM assay showed that, at the same time, the inhibition of Chk1 expression could relieve the G 2 / M arrest induced by doxorubicin; the apoptosis rate induced by doxorubicin increased from (5.54 ± 0.15)% before transfection to 22.24 ± 0.13)% (P <0.05). When adriamycin concentration was 0.4mg / L and 4mg / L, the cell proliferation activity decreased by 13% and 34% respectively. These results suggest that siRNA knockdown of Chk1 can enhance the sensitivity of breast cancer cells to doxorubicin by regulating the cell cycle and proliferation of MCF-7 / adr cells, which may provide a new target for overcoming chemoresistance in breast cancer.