Effects Of TIMP-1/2 SiRNA On PDGF/ERK In Hepatic Fibrosis Rats

Objective To investigate the effect of TIMP-1- and TIMP-2-targeted small interfering RNA(siRNA) on PDGF/ERK signaling pathway in hepatic stellate cells in rats with hepatic fibrosis induced by CCL4.Methods A total of 50 healthy, male, Sprague-Dawley rats were randomly divided into five groups(n = 10 each): normal group, model group, negative control group, TIMP-1 siRNA group and TIMP-2 si RNA group. The model of hepatic fibrosis was constructed by a classical method, injecting CCL4. In addition to the normal group, other groups were given subcutaneous injection of 40% CCL4. While the plasmid of TIMP-1 siRNA, TIMP-2 siRNA and negative control were injected into rats from the tail vein. The same dose of 0.9% sodium chloride injection was injected into the rats in normal and model group. Hepatic stellate cells(HSCs) were isolated from all the rats by pronase-collagenase method followed by density centrifugation after 12 weeks. Expression levels of TIMP-1, TIMP-2, platelet-derived growth factor(PDGF) and its receptor were determined by real-time polymerase chain reaction. Expression levels of PDGF and its receptor and phosphorylation of extracellular signal regulated kinase(p-ERK-1/2) were tested by western blotting analyses.Result The results of real-time PCR showed that the expression of TIMP-1, TIMP-2, PDGF-BB and PDGFRβ in the TIMP-1/2 si RNA groups were significantly lower than those in the model group and negative control group(P<0.05). The results of Western-blot showed that compared with the normal group, the other groups of PDGF-BB, PDGFRβ and p-ERK1/2 expression were significantly increased in HSCs of liver fibrosis rats(P<0.05). However, compared to the model and negative control group, the expression of PDGF-BB and its receptor and p-ERK1/2 in the TIMP-1 and-2 siRNA groups were obviously decreased(P<0.05). There were no significant difference of PDGF-BB and its receptor between in TIMP-1 and TIMP-2 siRNA treatment groups(P>0.05).Conclusions TIMP-1 and TIMP-2 targeted siRNA can effectively down-regulate the upstream cytokines of PDGF-BB and its receptor expression, then reduce the expression of p-ERK-1/2, which inhibit the proliferation and activation of HSCs and ultimately alleviate the liver fibrosis.