Discoidin Domain Receptor-2 Participate In Alcoholic Liver Fibrosis And Hepatic Sinusoidal Capillarization Through Mediating The Interaction Between Collagen And Hepatic Stellate Cells

Objective By establishing the alcoholic liver fibrosis rat model, to observe the initiation of liver fibrosis, hepatic sinusoidal capillarization, by using histological and electron microscopic methods. Investigate the expression of gene and protein of matrix metalloproteinase2 and discoidin domain receptor 2 in liver tissues of rats as the model went on to explore the possible effect of them in alcoholic liver fibrosis and hepatic sinusoidal capillarization, thus to reveal the further pathogenesy of alcoholic liver fibrosis and to provide the experimental basis for the afterward treatment of alcoholic liver fibrosis targeting discoidin domain receptor 2.Methods After fed normally for a week, fifty adult male Wistar rats were divided into three groups randomly, 38 for alcoholic liver fibrosis model group (ALF model), 6 for normal control group (N Group) and another 6 for pyrazol control group (P Group). The rats of the ALF model group were given olive oil diet plus the intragastric administration of alcohol and pyrazol mixture. The alcohol was administrated to rat in distillate spirit form, the quantity and concentration of which increased every two weeks from 4 g/kg·d, 30% to 9.6g/kg·d, 60% with pyrazol (25mg/kg·d) dissolved. The rats in N Group were given equal normal sodium and the rats of P Group were infused with equal pyrazol dissolved in equal normal sodium. To get liver samples, the animals in model group were sacrificed at the end of 4^th (M1 group), 8^th (M2 group), 12^th (M3 group) and 16^th (M4 group) week, while the rats in N Group and P Group were both sacrificed at the end of 16^th week. One part of the liver samples taken were used to observe the pathologic changes of liver with the methods of HE stains and Masson stains; another part were used to survey the development of hepatic sinusoidal capillarization with the methods of reticular fiber strains, immunohistochemistry stains of collagen typeⅠ,Ⅳand laminin and electron microscope observing; the rest part were used to detect the expression of gene and protein of MMP2 and DDR2 in the liver tissues with the methods of Real Time Fluorescence Quantitation-PCR and Western blot to make a dependability analysis with the index of hepatic sinusoidal capillarization.Results The severity of liver fibrosis in the ALF model group was aggravated with increased duration of alcoholic feeding: fatty degeneration, ballooning degeneration, inflammation and fibrosis could be viewed as the modeling extended. The deposition of collagen fibers and reticular fibers increased gradually (P < 0.05). Under electron microscope, it could be observed that the lost of fenestrae in the sinusoidal endothelial cells (SECs) and the basal membrane forming, suggesting the development of hepatic sinusoidal capillarization in modeling 12^th week. The expression of mRNA and protein of MMP2 and DDR2 increased gradually as the aggravation of fibrosis. The correlation analysis reveled that there was a remarkable positive correlation between the expressions of MMP2 and the index of hepatic sinusoidal capillarization. Meanwhile, the expressions of DDR2 have an obviously positive correlation with the expressions of MMP2 and the index of hepatic sinusoidal capillarization. No markedly abnormal changes appeared in C Group and P Group and little MMP2 and DDR2 expressed.Conclusions We successfully built the rat model of alcoholic liver fibrosis through olive oil diet plus the intragastric administration of alcohol and pyrazol mixture, during which we could observe the phenomenon of hepatic sinusoidal capillarization. The expression of mRNA and protein of MMP2 and DDR2 increased gradually as the modeling went on and the expressions of DDR2 have an obviously positive correlation with the expressions of MMP2 and the index of hepatic sinusoidal capillarization, suggesting that DDR2 may participate in alcoholic liver fibrosis and hepatic sinusoidal capillarization through mediating the interaction between collagen and hepatic stellate cells by MMP2.