Role And Mechanism Of PP2A In DCD Liver Injury Caused By Prolonged Cold Ischemia

Objective Liver transplantation is the best choice for end-stage liver disease(ESLD),and shortage of donor livers promotes usage of DCD livers.Currently,SCS is still the most convenient and economical method for liver preservation in vitro.Prolonged cold ischemia damage liver function severely leading to poor prognosis.To this end,we intend to explore the mechanism of prolonged cold ischemia induced DCD liver damage,and verify the role of PP2 A in this process.Methods First,establish rat DCD model to obtain livers,and perform SCS for 12 h and 24 h respectively.Liver function was evaluated by an isolated perfusion rat liver(IPRL)system.Expression of PP2 A,Drp1,CHOP and apoptosis marker protein caspases was detected by WB.Afterwards,AAV8 virus and pharmacological inhibitors were used to interfere PP2 A,mitochondrial division and endoplasmic reticulum stress(ERS)respectively to investigate the relationship of PP2 A,mitochondria and ER.Results(1)RNA sequencing results showed that the expression of apoptosis regulation pathway in DCD liver tissue increased,and interference of PP2 A decreased the gene expression of apoptosis regulation pathway and stress response pathway in liver tissue.(2)Prolonged cold storage aggravates DCD liver injury,which is manifested by the significantly increased ALT and AST in the perfusion solution in vitro,the liver pathological injury score of liver slices increased,and the BRL cell apoptosis rate increased.Protein detection showed that the expressions of PP2 A,Drp1 and CHOP increased,and the expression of apoptosis marker proteins caspases and Bax increased.Mitochondrial damage is aggravated,and the release of cytochrome c into the cytoplasm is increased.(3)Interfering with PP2 A expression alleviates DCD liver damage.(4)Inhibition of the mitochondrial division related protein Drp1 reduces the level of ERS,while inhibition of ERS does not affect Drp1 expression.Conclusion During cold ischemia,up-regulation of PP2 A expression induce mitochondrial fragmentation,leading to release of apoptotic factor including Cyt c as well as ERS,resulting in hepatocytes apoptosis.