摘要目的 探讨siRNA沉默细胞周期检测点激酶1(Chk1)基因对宫颈癌HeLa细胞放射敏感性的影响.方法 合成Chk1基因的小分子干扰RNA(si-Chk1),将si-NC或者si-Chk1分别转染到宫颈癌HeLa细胞中,0、2、4、6、8、10 Gy剂量的X射线照射细胞,RT-qPCR和Western blot检测转染后Chk1的mRNA水平和蛋白水平的表达,MTT方法检测细胞增殖能力,流式细胞仪检测细胞凋亡,克隆形成实验检测转染后HeLa细胞放射敏感性,Western blot检测转染后P21蛋白和抗凋亡基因Bcl-2蛋白的表达水平.结果 si-Chk1转染到HeLa细胞后,HeLa细胞Chk1基因的mRNA和蛋白表达水平均显著下降(t=2.12～5.86,P＜0.05),沉默Chk1的表达抑制了宫颈癌HeLa细胞的增殖(t=3.15 ～6.36,P＜0.05),同时降低宫颈癌HeLa细胞克隆形成能力(t=1.58～6.36,P ＜0.05),上调P21(t =4.35,P＜0.05)、下调Bcl-2蛋白(t=2.37,P＜0.05)的表达水平.结论 siRNA沉默Chk1表达能够增强HeLa细胞放射敏感性.

abstractsObjective To study the effect of checkpoint kinase 1 (Chk1) knock-down on radiosensitivity of human cervical carcinoma HeLa cells.Methods Chk1 was knock-down by using a specific siRNA of Chk1 gene.After transfection with si-CHK1 or its negative control si-NC,HeLa cells were irradiated with 0,2,4,6,8 and 10 Gy of X-rays.The expressions of Chk1 mRNA and protein in irradiated HeLa cells were determined by RT-qPCR and Western blot,respectively.Cell proliferation was measured by MTT assay,apoptosis was measured by flow cytometry assay,radiosensitivity was determined by colony formation,and the protein expressions of P21 and Bcl-2 were determined by Western blot.Results After transfection of si-Chk1,both mRNA and protein levels of Chk1 in HeLa cells were decreased (t =2.12-5.86,P ＜ 0.05).Chk1 knock-down decreased cell proliferation (t =3.15-6.57,P ＜ 0.05),reduced colony formation ability(t =1.58-6.36,P ＜ 0.05),increased the protein expression level of P21 (t =4.35,P ＜ 0.05) and decreased the protein level of Bcl-2(t =2.37,P ＜ 0.05) in the irradiated HeLa cells in comparison with nonirradiated control.Conclusions siRNA-mediated knock-down of Checkpoint kinase 1 enhances the radiosensitivity of HeLa cells.