| ObjectiveIn this study, by regulating the expression of liver heme oxygenase-1(HO-1), to observe the expression of hepatic bile salt transport protein by HO-1, Explore the rat biliary epithelial cell in ischemia and reperfusion injury (IRI) and its possible mechanismsMethods1. To establish the models of70%part of the liver ischemia-reperfusion injury by the method of operation which Fei X had reported.2. The use of chemical synthesis of Ad-HO-1, Ad-HO-1-of siRNA, empty adenoviruses and saline, into the donor liver by Genital dorsal vein. laparotomy after24hours, establish rat liver ischemia-reperfusion injury model. The animals were randomly divided into four groups:NaCl group, Empty adenovirus group, Amplification of HO-1adenovirus group, Silencing HO-1adenovirus group. To Collect specimens on1hour,1day,1week and2weeks after operation. To detect the levels of ALT,AST,TB,ALP in serum after ischemia reperfusion injury of liver. The function of liver was also evaluated. The expressions of IL-6,IL-10,HGF and TNF-a in the liver were detected. The mRNA and protein expressions of salt transporter proteins(Sodium taurocholate cotransporting polypeptide, Bile salt expot pump,Multidrug resistance-associated protein-2,sodium taurocholate cotransporting polypeptide and the HO-1mRNA expressions of liver tissue of levels in1d were measured. The livere and biliary bile duct were detected by immunofluorescence double staining and Chloro-acetate esterase staining. Observe the morphological changes in the structure of the size of the bile duct and the surrounding liver tissue.Results1.128cases of ischemia-reperfusion injury models of liver were Successfully established and partial liver ischemia time was45minutes.2. Compared with the same time points, serum ALT, AST, TB, and the ALP in HO-1groups was significantly lower than in NaCl and Adenovirus groups (p<0.05). serum ALT, AST, TB, and the ALP in siRNA groups was significantly higher than in NaCl and Adenovirus groups (p<0.05). HO-1groups was significantly lower than the siRNA groups (p<0.01).3. Compared with the same time points, siRNA groups serum HGF and IL-10was significantly lower than in NaCl and Adenovirus groups (p<0.05); HO-1groups serum HGF and IL-10was significantly higher than in NaCl and Adenovirus groups (p<0.05). HO-1groups was significantly higher than the siRNA groups (p<0.01). serum IL-6and TNF-a in G3was significantly lower than in NaCl and Adenovirus groups (p<0.05). serum IL-6and TNF-a in siRNA groups was significantly higher than in NaCl and Adenovirus groups (p<0.05). HO-1groups was significantly lower than the siRNA groups (p<0.01).4. Compared with the same time points, HO-1groups of HO-1, Ntcp, Bsep, Mrp2mRNA expression level was significantly higher than in NaCl and Adenovirus groups (p<0.05); siRNA groups of HO-1, Ntcp, Bsep, Mrp2mRNA expression level was significantly lower than in NaCl and Adenovirus groups (p<0.05); HO-1groups was significantly higher than the siRNA groups (p<0.01).5. siRNA groups Ntcp protein expression levels were significantly lower than the NaCl and Adenovirus groups (p<0.05). HO-1groups Ntcp protein expression levels were significantly higher than the NaCl and Adenovirus groups (p<0.05). HO-1groups was significantly higher than the siRNA groups (p<0.01).6. Compared with the same time points, Immunofluorescence double staining and chloro-acetate esterase staining in siRNA groups can be seen:The size of the bile duct epithelial cells is fuzzy, part of the bile duct basement membrane shedding, liver cells around loose, with degeneration, serious injury performance. The HO-1groups was less severe injury, size of the bile duct epithelial cells is clear, the continuity of the basement membrane is better, around the bile duct liver cells closely arranged, no degenerationConclusions:1. Rat liver warm ischemia reperfusion injury model is a stable, reliable, simple, the model is workable.2. Ad-HO-1can effectively increase HO-1gene expression in rat liver; Ad-HO-1-siRNA can effectively reduce the expression of HO-1gene in rat liver.3. The liver function damaged by Ischemia and reperfusion. Ad-HO-1can effectively improve liver function. Reduce the proinflammatory cytokine IL-6, TNF-a expression, an increase of protective cytokines of HGF, IL-10expression. Reduce the size of the bile duct ischemia and reperfusion injury.4. The mechanism of HO-1protect biliary ischemia-reperfusion injury in rats, may be induced by bile salt transporter protein high expression. Bile salt transporter protein expression can contribute to the transport and excretion of bile, reduce the strong cytotoxic bile formation, ultimately reduce the role of bile salts of the bile duct. |
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