The Expression Of Insulin-Like Growth Factor-1 And The Intervention Effect Of Traditional Chinese Herbal Drug In Rat With Liver Fibrosis

Objective:Liver fibrosis is a common step of advanced liver diseases caused by diversified reasons. Such stage is characterized by destruction of hepatic lobules and formation of false liver acinus. Despite the mechanism of liver fibrosis has not been fully illustrated in the past decades, researchers have pointed out that activation of quiescent stage hepatic stellate cells(HSC) and accumulation of extracellular matrix(ECM) mainly account for the occurrence of fibrosis. Many cytokines and signaling pathway are involved in activating HSC. Insulin-like growth factor-1(IGF-1) signaling is strongly correlate with growth, differentiation, proliferation, and survival of cell. The objective of this research is to investigate the expression of IGF-1 in fibrosis rat and the effect of Traditional Chinese Herbal Drug on its signaling pathway.Methods:In vivo: Healthy Sprague-Dawley rats were divided into six groups randomly noted: control group, model group, GFK treatment group, DS treatment group, GFK control group, DS control group. The experimental model of fibrosis was established by subcutaneous injection of CCl4(in a vehicle of olive oil). Control group were disposed with normal saline and drug control groups were disposed with GFK or DS. Four weeks after CCl4 disposition, treatment groups were given GFK and DS respectively by subcutaneous injection according to the weight. The whole process of intervention lasts for 4 weeks, all the rats were sacrificed at the end of 8th week. Liver samples were collected then for the use of section making and molecular experiment.General and pathological observation: Conventional HE staining was used to observe morphological changes. General status of mentality and physical were observed.Detection of extracellular matrix: mRNA level of Collagen I and Collagen Ⅲ were measured to access the matrix.Marker of the activation of hepatic stellate cell: Expression of α-SMA were detected by immunohistochemistry.Expression of IGF-1 and relevant genes of IGF-1 in rats’ liver: Expression of IGF-1 were detected by immunohistochemistry. The expression of genes associated with IGF-1 signaling pathway(PI3K, cyclin D1) were detected.Correlation analysis: Correlation between IGF-1 and PI3 K,cyclin D1,CollagenⅠ,CollagenⅢ,α-SMA was done by Pearson method. Result:1. General and pathological observation changes: Model animals were stay on a level that acted significantly lower than normal group, liver volume increased obviously, envelop tension, greasy feeling. Liver fibrosis was definitely observed in all model rats characterized by destruction of hepatic lobules, disordered arrangement of liver cells steatosis and necrosis of cells in both portal area and hepatic lobules in contrast with control group. While alleviation of tissue reaction were observed in drug groups. There was no significant different between control groups.2. Detection of extracellular matrix: In model group, expressions of Collagen Ι, Collagen Ⅲ were up-regulating compared to normal(P<0.05); GanFuKang attenuated the up-regulation of Collagen Ι, Collagen Ⅲ while DS attenuated the up-regulation of Collagen Ι. There was significant difference between two drugs(P<0.05). There was no significant different between control groups.3. Assessment of the activation of hepatic stellate cell:Immunohistochemistry shows expression of α-SMA in modle was up-regulating compared to normal. Evident concentration of α-SMA was found in fibrotic band areas around the periportal. The decline of both IGF-1 can be confirmed in treatment(P<0.05) and there was significant difference between two treatment group. There was no significant different between control groups.4. Expression of IGF-1 and relevant genes of IGF-1 in rats’ liver: Expression of IGF-1 in modle was up-regulating compared to normal by Immunohistochemistry assessment, IGF-1 was concentrated extracellularly. The treatment of GanFuKang and DS alleviated the positive expression. In model group, mRNA level of PI3 K, cyclin D1 was up-regulation compared to normal(P<0.05); treatment of GanFuKang attenuated the up-regulation of PI3 K and cyclin D1, so did DS. Significant difference between two treatment groups was observed. There was no significant different between control groups.5. Correlation analysis: Positive correlations were found between IGF-1 and PI3 K,cyclin D1,CollagenⅠ,CollagenⅢ,α-SMA.Conclusion:1. IGF-1 may be conducive to stimulation of HSC and deposition of ECM in liver in the CCl4-induced hepatic fibrosis rats.2. GFK and DS weaken liver fibrosis by inhibition of IGF/PI3K/cyclin D1 pathway and significant difference in anti-fibrosis function between GFK and DS was found in our study.