Therapeutic effects of cFLIP gene silencing on acute necrotizing pancreatitis rats and its mechanism

Abstract

Objective To investigate the therapeutic effects of cFLIP-siRNA tail venous injection on acute necrotizing pancreatitis (ANP) rats, and explore the related mechanism. Methods Three pairs of cFLIP siRNAs (cFLIP-siRNA) and negative control(siRNA-NC) were designed and synthesized. The most effective siRNA to inhibit the expression of cFLIP gene was selected for the experiment. Thirty SD rats were treated with retrograde pancreaticobiliary duct injection of 5% taurocholic acid sodium salt solution to establish ANP rat model. They were randomly divided into ANP group, siRNA-NC group and cFLIP-siRNA group with 10 rats in each group using random number method. The same volume of normal saline, siRNA-NC and cFLIP-siRNA were injected into tail vein, respectively. The rats were sacrificed after 24 h and the blood samples were colelcted. The levels of serum amylase and endotoxin were measured. The pancreatic tissues were collected and routine pathological examination was performed. The expressions of cFLIP-S and caspase-8 protein in pancreatic tissues were detected by immunohistochemistry and Western blotting. Results The levels of serum amylase in ANP group, siRNA-NC group and cFLIP-siRNA group were (1 286±209), (1 297±305) and (552±256)U/L, the level of serum endotoxin were (136±32), (128±56) and (46±23)ng/L, respectively. The pancreatic pathological scores were (4.97±1.16), (4.92±2.03) and (1.67±1.98). The expression level of cFLIP-S protein were (8.56±2.72), (9.12±2.45) and (3.82±1.46), respectively, which were significantly lower in cFLIP-siRNA group than in ANP group and siRNA-NC group (P<0.05), and the difference was statistically significant (P<0.05). The expression level of caspase-8 protein in the three groups were (2.25±1.24), (2.41±1.14) and (5.56±1.79), respectively, which were significantly higher in cFLIP-siRNA group than in ANP group and siRNA-NC group, and the difference was statistically significant (P<0.05). However, there was no significant difference between ANP group and siRNA-NC group. Conclusions Inhibition of cFLIP gene expression can reduce pancreatic injury in ANP rats. The possible mechanism was that the up-regulation of caspase-8 can inhibit pancreatic cell necrosis and promote apoptosis. Key words: Pancreatitis, acute necrotizing; RNA, small interfering; Apoptosis; cFLIP genes