目的观察胱硫醚γ裂解酶(CSE)siRNA转染人脐静脉内皮EAhy926细胞后CSE的表达及对H2S的影响。方法体外培养EAhy926细胞,经siRNA转染以沉默CSE基因表达,分为5组进行培养:正常对照组(A组):只加入转染试剂;阴性对照组(B组):转染Neg-siRNA;转染CSEsiRNA1组(C组);转染CSEsiRNA2组(D组);转染CSEsiRNA3组(E组)。采用Lipofectamine 2000介导转染EAhy926细胞,在荧光显微镜下观察细胞以检测转染效率;Western blot法检测CSE蛋白表达;收集细胞检测H2S含量。结果观察转染效率为60%左右;A组CSE蛋白表达为(153.41±23.43),B组24 h CSE蛋白表达为(150.22±17.56),而D组24 h CSE蛋白表达降至(59.43±21.81),差异有统计学意义(P〈0.01);并且A组H2S含量为(0.96±0.05),B组24 h H2S含量为(0.95±0.06),D组24 h H2S含量降至(0.54±0.08),差异有统计学意义(P〈0.01)。结论 CSE-siRNA能够有效地抑制EAhy926中CSE的表达,并且CSE-siRNA2沉默作用最强。
Objective To investigate the expression of cystathionine gamma lyase(CSE) after transfecting CSE small interfering RNA in human umbilical vein endothelial cell EAhy926 and the effect on H 2 S content.Methods EAhy926 cells were cultured in vitro,and were transfected by siRNA to knock down the expression of CSE.EAhy926 cells were divided into five groups: normal group(group A) : only to add the transfection reagents;negative control group(group B) : transfecting Neg-siRNA;transfecting CSEsiRNA1(group C);transfecting CSEsiRNA2(group D);transfecting CSEsiRNA3(group E).Transfecting EAhy926 cells with Lipofectamine 2000,detected the transfecting efficiency under flourence microscope,tested CSE protein expression by Western blot,collected the cells to test the content of H 2 S.Results The transfection efficiency was about 60%.The expression of CSE protein in group A was 153.41 ± 23.43 and 150.22 ± 17.56 in group B after 24 h,whereras decreased to 59.43 ± 21.81 in group D after 24 h,there was statistic difference among them(P 〈 0.01);and the content of H 2 S was 0.96 ± 0.05 in group A and 0.95 ± 0.06 in group B after 24 h,however decreased to 0.54 ± 0.08 in group D after 24 h,also had statistic difference among them(P 〈 0.01).Conclusion CSEsiRNA can inhibit the expression of CSE in the EAhy926 cells effectively,and CSEsiRNA2 shows the strongest gene silence effect.