Explore The Ubiquitin Ligase Parkin Mediated Programmed Cell Death Process And Mechanism Of Action In Hepatic Ischemia-reperfusion Injury

There are still many problems in liver surgery.For example,surgery related or postoperative liver ischemia-reperfusion injury is always an unavoidable problem in liver surgery.How to avoid or reduce liver ischemia-reperfusion injury is a research focus on liver surgery.With the gradual deep understanding of ischemia-reperfusion,especially the rapid development of molecular biology and genetics in recent years,to study the pathways and mechanisms of hepatocyte injury after hepatic ischemia-reperfusion and how to reduce these injuries are important topics in the study of hepatic ischemia-reperfusion.An important pathway for hepatic ischemia-reperfusion injury is achieved by apoptosis/autophagy,but so far,how apoptosis/autophagy coordinated to mediate cell death,and how apoptosis/autophagy balances ischemia-reperfusion injury is still not fully understood.In recent years,many studies have suggested that autophagy plays a significant role in ischemia-reperfusion.Autophagy is regulated by many factors.It is now found that the ubiquitin-proteasome system widely present in the body play a very important role in regulation of ischemia-reperfusion injury.As a newly discovered member of the E3 ubiquitin ligase family,Parkinson-related protein(Parkin)is thought to precisely regulate some important physiological or pathophysiological processes in the body,such as apoptosis,inflammatory response,cell differentiation,etc.Mainly concentrated in the nervous system,but Parkin protein plays a role in regulating liver function,especially in hepatic ischemia-reperfusion.There is no relevant research.Based on this,the purpose of this study is to investigate the role of Parkin protein in liver ischemia-reperfusion injury and to explore its mechanism and molecular pathway.MethodsPart ⅠThe rat liver ischemia-reperfusion model was used to detect the expression of Parkin m RNA and protein in the liver before and after ischemia-reperfusion.The similarities and differences of hepatocyte autophagy before and after ischemia-reperfusion were observed,and Parkin were explored.RNAi plasmid targeting Parkin was used to reduce the expression of Parkin protein and the changes of ischemia-reperfusion injury,hepatocyte autophagy and injury were detected.From the above test results,the intrinsic relationship between hepatocyte injury/autophagy and Parkin after hepatic cell ischemia-reperfusion was studied.Part Ⅱ In addition to autophagy,other regulatory factors such as apoptosis,DNA damage/repair,and cell cycle have important impact on hepatocyte ischemia-reperfusion injury.In this part,we constructed a rat model of hepatic ischemia-reperfusion,and used the RNAi plasmid to reduce the expression of Parkin and then detected hepatocyte apoptosis and cell cycle changes.From the above results,the intrinsic link between apoptosis,DNA damage repair,cell cycle and Parkin expression after ischemia-reperfusion was confirmed.ResultsPart Ⅰ After the ischemia-reperfusion injury of hepatocytes,the Parkin expression was up-regulated,and the level of autophagy was significantly increased.When the expression of Parkin was decreased,the level of autophagy decreased.This part of the results showed that Parkin is autophagy related.There is a significant positive correlation between Parkin expression and autophagy in the regulatory genes of HIRI.Part Ⅱ In the liver after ischemia-reperfusion injury,extensive apoptosis of hepatocytes can be observed.Down-regulation of Parkin by RNAi aggravates hepatocyte apoptosis,down-regulation of Parkin resulted in a decrease in G2/M distribution.In the ischemia-reperfusion group,the anti-apoptotic protein BCL-2,DNA repair protein Ku70,and cyclin Chk1 were down-regulated after Parkin down-regulating.The results of this part indicate that Parkin is not only closely related to autophagy,but also closely related to other factors that cause hepatic ischemia-reperfusion,such as apoptosis,cell cycle and DNA damage repair.ConclusionParkin has a protective effect on hepatocyte injury by effectively inducing autophagy after ischemia-reperfusion;and Parkin may also be reduced by reducing liver ischemia-reperfusion injury.Inhibition of apoptosis,promotion of DNA repair,and cell cycle arrest in G2/M phase contribute to the survival of hepatocytes,suggesting that Parkin is a protective gene for hepatic ischemia-reperfusion injury.