RNA干扰Caspase-3基因抑制大鼠肝脏缺血再灌注损伤

目的 观察RNA干扰(RNAi)Caspase-3基因对大鼠肝脏缺血再灌注损伤(IRI)的保护作用.方法 构建针对大鼠Caspase-3基因的短发夹状RNA(shRNA)真核表达载体.肝脏IRI前48 h经门静脉注射磷酸盐缓冲液(PBS)或Caspase-3 shRNA,实验随机分为3组,假手术组、PBS组和shRNA组.阻断大鼠70%入肝血流40 min,再灌注6,12,24 h,3,5,7 d检测血清谷丙转氨酶(ALT)和谷草转氨酶(AST)水平,肝组织Caspase-3 mRNA的表达,细胞凋亡情况,丙二醛(MDA)以及超氧化物歧化酶(SOD)的含量.结果 与PBS组比较,shRNA组再灌注6,12,24 h血清ALT和AST水平显著降低(P<0.05),shRNA组大鼠肝组织的Caspase-3 mRNA水平、肝细胞凋亡指数(25.21%±3.18%vs 35.24%±2.33%,P<0.05)和肝组织中MDA含量[(96.3±12.8)nmol/mg vs(133.5±12.4)nmol/mg,(P<0.05)]显著降低,SOD活性显著升高[(22.5±3.4)U/mg vs(12.2±3.1)U/mg,(P<0.05)].结论 RNA干扰Caspase-3基因可以抑制细胞凋亡的发生,保护肝脏缺血再灌注损伤.

Inhibition of liver ischemic reperfusion injury through silencing Cuspuse-3 gene by RNA interference in rats

Objective To investigate the protective effect of silencing Caspase-3 gene on liver is-chemic reperfusion injury. Methods We constructed the eukaryotic expression vector of small hairpin RNA targeting rat Caspase-3 gene. PBS or Caspase-3 shRNA was injected through portal vein 48 h before operation. The rats was divided into the sham-operated group, PBS group and shRNA group. The left lateral and median lobes of the liver were rendered ischemic 40 min resulting in a segmental (70%) hepatic ischemia. The serum ALT and AST levels were detected 6,12, 24 h, 3, 5 and 7d after reperfusion. The mRNA expression of Caspase-3, apoptosis index, concentration of the malondialde-hyde (MDA) and superoxide dismutase (SOD) in the liver tissue were determined. Results Compared with PBSgroup and blank vector group, the ALT and AST levels were significantly de-creased in shRNA group 6, 12 and 24 h after reperfusion (P<0.05). Caspase-3 mRNA level, apopto-sis index (25.21±3.18% vs 35.24±2.33%, P＜0.05) and concentration of MDA in the liver tissue (96.3±12.8 nmol/mg vs 133.5±12.4 nmol/mg, P<0.05) were significantly decreased while the SOD activity were significantly increased (22.5±3.4 U/rag vs 12.2±3.1 U/mg, P＜0.05). Conclusion Caspase-3 gene silencing can protect the liver ischemia/reperfusion injury.