摘要
目的研究镉对人肝癌细胞SMMC-7721增殖及凋亡的影响及其作用机制。方法体外培养的人肝癌细胞SMMC-7721分别以0、40、80、160μmol/L的CdCl2处理0～48 h,应用四甲基偶氮唑蓝(MTT)法检测细胞相对存活率,流式细胞仪检测细胞凋亡百分率,分光光度法检测半胱天冬酶Caspase-3酶活性的变化,逆转录聚合酶链反应(RT-PCR)和蛋白印迹(western blot)法测Caspase-3 mRNA和蛋白表达的变化。结果随着镉处理浓度的增加,氯化镉对细胞活力的抑制作用明显增强,40、80、160μmol/L氯化镉处理SMMC-7721细胞,其抑制率分别为33.94%、48.04%和68.54%,均明显高于0μmol/L氯化镉处理组,差异均有统计学意义(P<0.01);40、80、160μmol/L镉处理组细胞的凋亡百分率分别为(25.77±4.66)%、(34.97±9.25)%和(55.14±5.67)%,与0μmol/L镉处理组比较明显升高,差异均有统计学意义(P<0.01);镉处理SMMC-7721细胞24和48 h,Caspase-3相对酶活性均随着镉浓度的增加而明显增强;40μmol/L镉处理SMMC-7721细胞,12、24和48 h可明显上调细胞中Caspase-3基因及蛋白的表达,Caspase-3基因mRNA表达水平分别为对照组的4.1、3.7和3.9倍,Caspase-3蛋白表达分别为对照组的5.8、6.0和7.2倍。结论镉能明显抑制人肝癌细胞增殖,诱导细胞凋亡,Caspase-3基因在镉诱导SMMC-7721细胞凋亡过程中起着重要的作用。
Objective To study the effection on proliferation and apoptosis of SMMC-7721 cells induced by cadmium and its mechanism.Methods SMMC-7721 cells were incubated with 0,40,80,160 μmol/L CdCl2 for 0-48 hours.Cell viability was measured by methyl thiazolyl tetrazolium assay（MTT）.The occurrence of apoptosis was determined by flowcytometry.Absorption spectrometry was adopted to measure the relative activity of Caspase-3.The expression of Caspase-3 gene in SMMC-7721 cells after treatment with cadmium was determined by the methods of reverse transcription polymerase chain reaction（RT-PCR） and western-blot analysis.Results Cell viability decreased in dose-dependent manner with the dose increase of cadmium.The inhibition ratio of cell proliferation incubated with 40,80,160 μmol/L CdCl2 was 33.94%,48.04%,and 68.54%,respectively.Compared with 0 μmol/L cadmium,the inhibition ratio increased obviously（P0.01）.The apoptosis rate of SMMC-7721 cell incubated with 40,80,160 μmol/L CdCl2 was 25.77±4.66%,34.97±9.25%,and 55.14±5.67%,respectively.Compared with 0 μmol/L cadmium,the apoptosis rate increased obviously in the incubated cells with 40,80,160 μmol/L cadmium（P0.01 for all）.With the dose increase of cadmium,we found that the enzyme activity of Caspase-3 increased obviously.RT-PCR analysis revealed that the incubated cell with 40μmol/L cadmium for 12,24 and 48 hours,the expression of Caspase-3 mRNA was 4.1,3.7,and 3.9 times higher compared with that of the controls.Western blot analysis revealed that in the incubated cells with 40 μmol/L cadmium for 12,24,and 48 hours,the expressin of Caspase-3 was 5.8,6.0,and 7.2 times higher compared to that of the controls,respectively.Conclusion CdCl2 can obviously restrain the proliferation and induce apoptosis of SMMC-7721 cell.The results suggest that Caspase-3 could play an important role in cadmium-induced cell apoptosis.
出处
《中国公共卫生》
CAS
CSCD
北大核心
2012年第12期1599-1601,共3页
Chinese Journal of Public Health
基金
辽宁省博士启动基金资助项目(20091082)
