Role Of Wnt/β-catenin Pathway In HS Inhibition Of Hepatic Ischemia/Reperfusion Injury In Rats

Objective The effects of saturated hydrogen physiological saline preconditioning on hepatic ischemia-reperfusion injury in rats were observed by establishing a model of hepatic partial ischemia-reperfusion injury in SD rats.Through pathological examination,determination of liver function,liver tissue oxidative stress index and Wnt/β-catenin pathway-related protein expression level,the Wnt/β-catenin pathway was studied from the molecular level to relieve liver ischemia in rats.The role of perfusion injury provides a theoretical basis for the clinical application of hydrogen in reducing hepatic ischemia-reperfusion injury.Methods The 40 male Sprague-Dawley rats were randomly divided into 5 groups,such as Sham group,I/R1+NS,I/R1+HS group,I/R2+NS group and I/R2+HS group.Except the Sham group,the ischemia time of I/R1 group and I/R2 group was 20 min and 60 min,respectively,and the reperfusion time was 6 h.The NS group received intraperitoneal injection of normal saline 15 minutes before operation.The HS group received intraperitoneal injection of saturated hydrogen normal saline 15 minutes before operation at a dose of 10 ml/Kg.The hepatic ischemia-reperfusion model was established by the widely recognized Nauta method at home and abroad,blocking 70% of the liver blood supply.After 6 hours of reperfusion in the liver of each group,the samples were taken to detect serum ALT and AST content.The content of MDA and MPO in liver tissue was detected by colorimetry.The pathologic of liver tissue were observed by microscope.The apoptosis of hepatocytes was detected by TUNEL.Western blot was used to detect the expression of β-catenin and DKK-1 protein.Results 1.The liver ischemic was 20 min,60min,and the blood flow was reperfused for 6h.The levels of ALT and AST in the control group and the experimental group were significantly higher than Sham group(P<0.05).The longer the blood time,the higher the serum ALT and AST levels(P<0.05).Compared with the control group,it had lower serum ALT and AST levels(P<0.05),while the experimental group 2 had serum ALT,The AST reduction was more pronounced(P < 0.01).2.There was no significant change in the levels of MDA and MPO in the sham operation group after reperfusion for 6 hours in the 70% hepatic ischemia model of SD rats.Compared with the sham group,the MDA and MPO contents in the liver tissue of the control group and the experimental group were significantly increased,and the difference was statistically significant(P<0.01).Compared with the control group,the MDA and MPO levels were significantly higher in the experimental group(P<0.01).3.The Sham group have showed normal liver structure.In the I/R1+NS group,the structure of the hepatic lobule was visible.The structure was basically normal,the cells were arranged neatly,and some hepatocyte edema was observed.The apoptosis and necrosis of the cells was observed.In the I/R1+HS group,the structure of the hepatic lobule was clearly visible,the liver cells were arranged neatly,and few cell edema and inflammatory cell infiltration were observed,and no apoptotic or necrotic cells were observed.I/R2+NS group: The structure of hepatic lobule was obviously changed,the cell line arrangement was disordered,the cell edema was obvious,the swelling was spherical,and obvious vacuolization and punctate necrosis were observed.A large number of inflammatory cells gathered and infiltrated around the portal area,and some of the portal area were visible.The fragments were necrotic,some hepatocytes shrank,cytoplasmic acidophilus,and nuclear condensation showed hepatocyte apoptosis.Compared with the I/R2+NS group,the hepatic lobule structure of the I/R2+HS group was clearly clear,the hepatocytes were arranged neatly,the edema was relieved,some were vacuolated,scattered punctate necrosis,and inflammatory cells in the portal area were clustered.The infiltration was significantly improved,a small number of hepatocytes shrink,cytoplasmic acidophilus,and nuclear condensation showed hepatocyte apoptosis.4.TUNEL test showed that the number of positive cells in the control group increased significantly,and the apoptosis index of hepatocytes increased significantly(P<0.01).The positive cells of the experimental group were corresponding to the ischemic time.The number decreased,especially in the experimental group 2,the hepatocyte apoptosis index decreased(P<0.01).5.The expression of DKK-1 was up-regulated in the control group.Compared with the control group,the expression of DKK-1 was decreased in the corresponding ischemia time group(P<0.05).Compared with the sham group,the expression of β-catenin protein was down-regulated in the control group.After HS pretreatment,the expression of β-catenin protein was up-regulated in the experimental group compared with the control group,but it still lower than the sham operation group.Conclusions 1.Saturated hydrogen physiological saline pretreatment significantly reduced liver ischemia-reperfusion injury in rats.2.Saturated hydrogen normal saline may act through the Wnt/β-catenin signaling pathway,inhibitting the degradation of β-catenin protein and the expression of DKK-1 protein,and it can enter the nucleus to regulate cell apoptosis to hepatic ischemia.