富含半胱氨酸蛋白61对子宫内膜癌细胞增殖及细胞周期的影响及其机制研究

Effect of cysteine-rich protein 61 on proliferation and cell cycle of endometrial cancer cells and its mechanism

目的研究富含半胱氨酸蛋白61(CYR61)对子宫内膜癌(EC)细胞株HEC-1B增殖及细胞周期的影响,并探讨其机制。方法取对数生长期HEC-1B细胞,随机分为Control组(不处理)、pcDNA3.1-NC组(转染pcDNA3.1-NC)、pcDNA3.1 CYR61组(转染pcDNA3.1 CYR61)、pcDNA3.1 CYR61+LiCl(Wnt/βcatenin通路激活剂)组(转染pcDNA3.1 CYR61,并加入LiCl使其终浓度为60 mmol/L)。转染48 h后采用MTT法检测增殖抑制率,PI染色法检测细胞周期占比,Western blotting检测细胞β-catenin、Caspase-3、Cyclin D1蛋白的表达。结果与Control组、pcDNA3.1-NC组比较,pcDNA3.1 CYR61组CYR61、β-catenin、Caspase-3、Cyclin D1蛋白相对表达量、G_(0)/G_(1)期占比升高(P<0.05),S、G_(2)/M期占比降低(P<0.05);与pcDNA3.1 CYR61组比较,pcDNA3.1 CYR61+LiCl组CYR61、β-catenin、Caspase-3、Cyclin D1蛋白相对表达量、G_(0)/G_(1)期占比降低(P<0.05),S、G_(2)/M期占比升高(P<0.05)。4组24 h、48 h、72 h增殖抑制率比较,采用重复测量设计的方差分析,结果:①不同时间点增殖抑制率有差异(F=25.416,P=0.000);②3组增殖抑制率有差异(F=46.962,P=0.000),pcDNA3.1 CYR61组增殖抑制率较高,增殖抑制效果好;③3组增殖抑制率变化趋势有差异(F=37.541,P=0.000)。结论上调CYR61可抑制EC细胞株HEC-1B增殖,阻滞其细胞周期,其作用机制可能与抑制Wnt/β-catenin信号通路有关。

Objective To study the effect of cysteine-rich protein 61(CYR61)on the proliferation and cell cycle of endometrial cancer(EC)cell line HEC-1B,and to explore the possible mechanism.Methods Log phase HEC-1B cells were taken and randomly divided into control group(no treatment),pcDNA3.1-NC group(transfected with pcDNA3.1-NC),pcDNA3.1 CYR61 group(transfected with pcDNA3.1 CYR61),pcDNA3.1 CYR61+LiCl[Wnt/β-catenin(β-catenin)pathway activator]group(pcDNA3.1 CYR61 was transfected,and LiCl was added to make the final concentration 60 mmol/L).At 48h after transfection,the proliferation inhibition rate was detected by MTT experiment.The cycle proportion was detected by PI staining method.The protein expression level ofβ-catenin,aspartate-specific caspase-3(Caspase-3),and Cyclin D1(Cyclin D1)in different groups were detected by Western blotting method.Results Compared with control group and pcDNA3.1-NC group,the proportion of S,G_(2)/M phase,and the protein expression ofβ-catenin,Caspase-3,Cyclin D1 in the pcDNA3.1 CYR61 group were decreased,and the CYR61 protein expression and the proportion of G_(0)/G_(1) phase was increased(P<0.05).Compared with pcDNA3.1 CYR61 group,the proportion of S,G_(2)/M phase,and the protein expression ofβ-catenin,Caspase-3,Cyclin D1 in the pcDNA3.1 CYR61+LiCl group were increased,and the CYR61 protein expression,the proportion of G_(0)/G_(1) phase was decreased(P<0.05).The proliferation inhibition rates of pcDNA3.1-NC group,pcDNA3.1 CYR61 group,and pcDNA3.1 CYR61+LiCl group were different(F=46.962,P=0.000).The pcDNA3.1 CYR61 group had higher proliferation inhibition rate and better proliferation inhibition effect.There were differences in the change trend of proliferation inhibition rate among pcDNA3.1-NC group,pcDNA3.1 CYR61 group,and pcDNA3.1 CYR61+LiCl group(F=37.541,P=0.000).Conclusion Up-regulation of CYR61 can inhibit the proliferation of EC cell line HEC-1B and block its cell cycle,which may be achieved by inhibiting the Wnt/β-catenin signaling pathway.