ECHS1经线粒体途径调控HepG2细胞凋亡

ECHS1 Involved in Antagonising Apoptosis in HepG2 Cells via the Mitochondrial Pathway

目的:观察烯脂酰辅酶A水合酶短链1(enoyl-CoA hydratase short chain 1,ECHS1)对HepG2细胞凋亡的影响。方法:构建ECHS1基因的干扰质粒,转染HepG2细胞后通过嘌呤霉素筛选稳定干扰ECHS1的细胞株(HepG2-siECHS1),利用Western blot验证其干扰效率;利用流式细胞仪及TUNEL方法检测ECHS1干扰后HepG2细胞凋亡情况,再采用Western blot检测凋亡相关蛋白表达水平。结果:成功构建了ECHS1的干扰质粒;Western blot验证了ECHS1干扰后HepG2细胞株中ECHS1表达低于空白对照组(未转染HepG2细胞)及阴性对照组(空载体pU6转染HepG2细胞)(P<0.05);流式细胞仪术显示HepG2-siECHS1组细胞株凋亡率(14.98±1.07)%,阴性对照组(10.55±0.19)%,两者差异有统计学意义(P<0.05);TUNEL实验显示HepG2-siECHS1组细胞株凋亡率(6.13±0.12)%,阴性对照组(2.89±0.21)%,两者差异有统计学意义(P<0.05);与阴性对照组比较,ECHS1干扰后p53、促凋亡蛋白Bax及Bid均表达增加。结论:ECHS1通过线粒体途径拮抗HepG2细胞凋亡。

Objective： To investigate the impact of enoyl-CoA hydratase short chain 1 （ECHS1） on apoptosis of HepG2 cells. Methods： The ECHS1 interference plasmids were constructed and transfected into HepG2 ceils. The stable interference cell lines were screened via puromycin. Interference efficacy was detected by Western blot. Flow cytometry and TUNEL assays studied the apoptosis of HepG2 cells. Apoptosis-related proteins were detected by Western blot. Results： The HepG2 cells with stable ECHS1 gene interference were successfully established. The expression level of ECHS1 protein in HepG2 cells after transfection with ECHS1 siRNA was significantly lower than that in the blank control cells （HepG2 ceils without transfection） and the negative control cells （HepG2 cells transfected with pU6 vector） （P 〈 0.05 ）. Apoptosis ratio of the ECHS1 siRNA group was significant higher than that of negative control group by both Flow cytometry and TUNEL assays （ P 〈 0.05 ）. Expressions of p53, Bax and Bid in ECHS1 siRNA group were higher than those in negative control group. Conclusion： ECHS1 may antagonise the apoptosis of HepG2 cells via the mitochondrial pathway.