| Purpose: This study aims to investigate the effects of reversine on hepatic fibrosis and inflammation mechanisms in hepatic fibrosis rat model.Methods: 1.25 healthy male Sprague-Dawley rats were randomly divided into control group(5 rats)and model group(20 rats).Rats in the model group were intraperitoneally injected with thioacetamide solution to establish the hepatic fibrosis rat model.During 4 weeks,2 rats died,then after 4 weeks,randomly selected 3 rats in the model group for liver biopsy to observe that if the hepatic fibrosis model was successful.The remaining 15 rats in model group were randomly divided into hepatic fibrosis group,low concentration group(intraperitoneally injected with reversine 50ug/kg),and high concentration group(intraperitoneally injected with reversine 200ug/kg),while the rats in the control group and hepatic fibrosis group without treatment.After 3 weeks,all rats were sacrificed to collect liver and blood.2.By measuring the body weight and liver wet weight of each rat to calculate the liver index.Testing the serum of liver function.3.Observe the liver pathological morphology by HE staining and Masson staining,and analysis of collagen area.4.Immunohistochemistry was used to detect the expression of ?-SMA and TGF-?1 protein in liver of rats in each group.5.The m RNA expression levels of IL-1?,IL-6,IL-17 A,NLRP3,PDGFB,RELA and TNF-? in liver tissue of each group were detected by q RT-PCR.Results:1.The liver index was(3.2 ± 0.002)% in hepatic fibrosis group and(2.3 ± 0.002)% in control group.There was a statistically significant difference between the two groups(P < 0.001).The liver index of rats in low concentration group and high concentration group were(2.6 ± 0.004)% and(2.8 ± 0.002)%,respectively,and there was no significant difference compared with the liver fibrosis group(P> 0.05).The serum AST in control group was(86±3.61)U/L,and the serum AST in hepatic fibrosis group was(151.7±22.19)U/L.There was a significant difference in serum AST between the two groups(P<0.001).After treatment by reversine,the AST levels in low concentration group(P=0.002)and high concentration group(P<0.001)were reduced.There was no significant difference in serum TBIL,ALT and ALB levels(P>0.05).2.After intraperitoneally injected with thioacetamide,the hepatic lobule structure was disordered and fiber septum was formed.Occasionally,pseudolobules were observed.Infiltration with many inflammatory cells in the portal area,spotted necrosis and deposition of collagen fibers also could seen.This phenomenon is more serious in hepatic fibrosis group rats.By comparing the liver collagen area of each group,the liver collagen area in hepatic fibrosis group was 7.61±1.19,which was higher than the control group(P=0.003).The liver collagen area in low concentration and high concentration groups were 5.41±0.65 and 2.34±0.50,respectively,which were lower than those in liver fibrosis group(P=0.036,P=0.006,respectively).3.By analyzing the IOD of ?-SMA and TGF-?1 proteins in the liver of rats in each group,we found that the IOD of ?-SMA and TGF-?1 proteins in hepatic fibrosis group were 9141.04±782.54 and 48398.67±3897.37,respectively.And the IOD of ?-SMA and TGF-?1 proteins in control group were 630.68±244.96 and 5852.37±703.33,which were lower than those in hepatic fibrosis group(P<0.001).The IOD of ?-SMA protein in high concentration group decreased when comparing with hepatic fibrosis group(P<0.001).The IOD of TGF-?1 protein in low concentration group and high concentration group were 28038.3±7391.04 and 18131.24±3204.93 respectively,which were less than the hepatic fibrosis group.4.q RT-PCR detected the expression of IL-1?,IL-6,IL-17 A,NLRP3,PDGFB,RELA and TNF-? m RNA in liver tissue of rats in each group.In hepatic fibrosis group,therelative expression levels of IL-1?,IL-17 A,NLRP3,PDGFB,and RELA m RNA were 1.16±0.08,1.8±0.17,1.84±0.13,3.21±0.11,and 1.53±0.07,respectively.All the expression were increased when comparing those on control group(P=0.028,P=0.001,P<0.001,P<0.001,P<0.001,respectively).While the expression levels of IL-6 and TNF-? m RNA were 1.19±0.15 and 0.9±0.01(P>0.05).Compared with the hepatic fibrosis group,the relative expression levels of IL-1?m RNA in low concentration group and high concentration group were 0.75±0.03 and 0.89±0.07(P=0.001,P=0.011).And the relative expression of IL-17 A m RNA in low concentration group was 0.76±0.27(P=0.004).The m RNA expression of NLRP3 and PDGFB in the liver of low concentration group were 1.25±0.04 and 1.94±0.09,respectively.The relative expression levels of NLRP3 and PDGFB m RNA in high concentration group were 1.02±0.07 and 1.23±0.2,respectively,which were lower than those in hepatic fibrosis group(P<0.01).The relative expression levels of RELA m RNA in low concentration and high concentration groups were 1.27±0.08 and 0.85±0.07,respectively,which significantly different from those in the hepatic fibrosis group(P=0.013,P<0.001,respectively).Conclusions: 1.Reversine can inhibit the expression of ?-SMA and TGF-?1 and reduce the formation of collagen fibers in hepatic fibrosis rats,so that it can lessen the degree of liver fibrosis.2.Reversine can improve liver function in rats with hepatic fibrosis.3.Reversine may reduce inflammation by inhibiting the expression of IL-1?,NLRP3,PDGFB,and RELA m RNA,so as to reduce the degree of hepatic fibrosis 4.Low concentration of reversine can inhibit the m RNA expression of IL-17 A in hepatic fibrosis rats.However,high concentration of reversine has no effect on its expression. |
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