雌激素对人肝癌细胞增殖和凋亡的影响

Effect of estrogen on proliferation and apoptosis of human hepatocellular carcinoma cells

目的探究雌激素对人肝癌细胞株增殖和凋亡的影响。方法采用普通聚合酶链反应(PCR)技术、Western印迹实验检测肝癌细胞株中雌激素受体(ER)α、ERβ的mRNA和蛋白质的表达情况;细胞增殖实验检测雌激素对肝癌细胞增殖的影响;流式细胞术检测雌激素对肝癌细胞凋亡率的影响;Tunel染色法检测雌激素对肝癌细胞晚期凋亡率的影响;雌激素处理肝癌细胞株后,基因芯片技术检测表达增加的凋亡基因。结果ERα和ERβ在肝癌细胞株SMMC7721、HepG2、Bel-7404、huh7、MHCC-97L、MHCC-97H、PLC、LM3、SK-HEP-1及Bel-7402中均有表达;不同浓度的雌激素处理肝癌细胞不同时间后,可显著抑制肝癌细胞增殖,且抑制效果存在时间和剂量依赖性;与0.00μmol/L组相比,不同浓度的雌激素(1.25、2.50、5.00、10.00、20.00、40.00、80.00和160.00μmol/L)处理HepG2、SMMC-7721细胞24 h、48 h后,HepG2和SMMC-7721细胞的细胞凋亡率增加,存在剂量和时间的依赖性;Tunel结果显示,与0.00μmol/L组相比,10.00μmol/L雌激素处理HepG2、SMMC-7721细胞48 h后,细胞晚期凋亡率均显著增加;20.00μmol/L雌激素处理SMMC-7721细胞48 h后,凋亡基因BAK1、GADD45A、HRK、LTA、TNFRSF-10A均表达增加。结论雌激素对人肝癌细胞具有抑制生长、诱导凋亡的作用。

Objective To investigate the effect of estrogen on proliferation and apoptosis of human hepatocellular carcinoma cell lines.Methods Conventional PCR and Western blot were used to detect mRNA and protein expression of estrogen receptor(ER)α,ERβin hepatocellular carcinoma cell lines.Cell proliferation assay(MTT assay)was used to detect the effect of estrogen on the proliferation of hepatocellular carcinoma cells.Flow cytometry(FCM)was used to detect the effect of estrogen on the apoptosis rate of hepatocellular carcinoma cells.Tunel staining was used to detect the effect of estrogen on the late-stage apoptosis rate of hepatocellular carcinoma cells.After estrogen treatment,RT2 Profiler PCR arrays was used to detect the increased expression of apoptotic genes.Results ERαand ERβwere expressed in hepatocellular carcinoma cell lines with SMMC7721,HepG2,Bel-7404,huh7,MHCC-97L,MHCC-97H,PLC,LM3,SK-HEP-1 and Bel-7402.MTT results showed that different concentrations of estrogen(1.25,2.50,5.00,10.00,20.00,40.00,80.00 and 160.00μmol/L)could significantly inhibit the proliferation of hepatocellular carcinoma cells after treatment for different time(24 h,48 h,72 h),and the inhibition effect was time-and dose-dependent.FCM results showed that compared with the blank group,after treatment with different concentrations of estrogen(1.25,2.50,5.00,10.00,20.00,40.00,80.00 and 160.00μmol/L)for different durations(24 h,48 h and 72 h),apoptosis of the HepG2 and SMMC-7721 cell lines were significantly increased,and there was a dose-and time-dependence.Tunel results showed that compared with the blank group,after treatment with estrogen(10.00μmol/L)for 48 h,late-stage apoptosis of the HepG2 and SMMC-7721 cell lines were significantly increased.After treatment SMMC-7721 cells with estrogen(20.00μmol/L)for 48 h,the expressions of BAK1,GADD45A,HRK,LTA and TNFRSF-10A were increased.Conclusions Estrogen could inhibit proliferation and induce apoptosis of human hepatocellular carcinoma cells.