结肠癌中ASPM对细胞增殖、侵袭和凋亡的影响

Effects of ASPM on proliferation,invasiveness and apoptosis of colon adenocarcinoma cells

目的探讨头小畸形相关基因(abnormal spindle-like microcephaly associated,ASPM)对结肠癌细胞SW116和HCT116增殖、侵袭和凋亡的影响以及对β-catenin的调节作用。方法挑选ASPM含量较高的结肠癌细胞株HCT116和SW116。体外培养结肠癌细胞HCT116和SW116,设计siRNA-ASPM序列,转染细胞,分为阴性对照组(negative control,NC)组、siRNA1组和siRNA2组。采用CCK8法检测细胞增殖情况,Transwell实验检测细胞侵袭能力,Annexin V-FITC/PI细胞凋亡检测试剂盒检测细胞的凋亡率。qRT-PCR和Western blot法检测HCT116各组细胞中ASPM和β-catenin mRNA和蛋白的表达。挽救实验探究加入LiCl处理后,结肠癌细胞中ASPM对β-catenin表达的影响。结果siRNA-ASPM转染HCT116和SW116细胞后,siRNA组中ASPM表达量下降(P<0.05)。与NC组相比,siRNA1组、siRNA2组细胞的增殖和侵袭能力均降低,凋亡率升高;siRNA1组、siRNA2组中ASPM和β-catenin蛋白表达明显降低(P<0.05)。挽救实验结果证实,与siRNA-ASPM组相比,LiCl能部分恢复β-catenin mRNA的表达。结论沉默ASPM表达可抑制结肠癌细胞活力,降低细胞侵袭能力并诱导细胞凋亡,其机制可能与抑制Wnt/β-catenin信号通路有关。

Purpose To study the effects of(abnormal spindle-like microcephaly associated,ASPM)on proliferation,invasiveness and apoptosis of colon adenocarcinoma cells HCT116 and SW116,and on the expression ofβ-catenin.Methods Colon cancer cell lines HCT116 and SW116 with high expression of ASPM were selected.HCT116 and SW116 cells were cultured in vitro,and siRNA-ASPM sequence was designed.The transfected cells were divided into negative control(NC)group,siRNA1 group and siRNA2 group.CCK8 assay was used to detect the proliferation of cells.Transwell assay was used to detect cell invasiveness.Annexin V-FITC/PI was used to detected the apoptosis of cells.The expression of ASPM andβ-catenin was detected by qRT-PCR and Western blot.Rescue experiment was used to explore the effect of ASPM expression on the expression ofβ-catenin after treatmented with LiCl.Results The expression of ASPM mRNA and protein in siRNA group was significantly lower than those in NC group after transfection(P<0.05).Compared with NC group,the proliferation and invasiveness of the cells in siRNA1 and siRNA2 group decreased,and the apoptotic rate increased,respectively(P<0.05).The expression of ASPM andβ-catenin decreased significantly in siRNA1 and siRNA2 group(P<0.05).The rescue experiment confirmed that compared with siRNA-ASPM group,β-catenin mRNA expression was increased in LiCl supplementation group.Conclusion ASPM gene silencing can inhibit the proliferation,invasion of colonic cells,which may be related to down-regulation ofβ-catenin level.