The Pathogenic Effect And Mechanism Of PKM2 In Acute Liver Injury Induced By LPS/D-Gal

Objective: Pyruvate kinase(PK)is the key enzyme in the last step of glycolysis.Pyruvate kinase isozyme type M2(PKM2)plays an important role in the regulation of tumor cell proliferation.Recent studies have found that PKM2 also plays an important role in the pathophysiological process of inflammatory response.This study investigate the potential role and mechanism of PKM2 in acute liver injury induced by lipopolysaccharides(LPS)/D-galactosamine(D-Gal)in mice.Method: Male BALB/c mice were injected with LPS(10 ?g/kg)/D-Gal(700 mg/kg)to establish an acute liver injury model.In order to clarify the change of PKM2 in LPS/D-Gal model,we sacrificed mice at different time points(0h?2h?4h and 6h after LPS/D-Gal injection)and used immunoblot analysis to determine the expression of PKM2 in liver tissue.The colorimetric method was used to detect the change of pyruvate kinase activity and pyruvate content in liver tissue.To explore the role of PKM2 in acute liver injury induced by LPS/D-Gal in mice,pretreatment with PKM2 agonist ML265,which acts to inhibit PKM2 nuclear translocation,also known as PKM2 nuclear translocation inhibitor(intraperitoneal injection 30 min before LPS/D-Gal injection)and post-treatment(intraperitoneal injection 2 hours after LPS/D-Gal injection),the ALT and AST activities were measured by the Lai method,the level of IL-6 and TNF-? in plasma were determined by ELISA.The liver sections were stained with hematoxylin and eosin for histopathological evaluation and hepatic apoptosis was determined by TUNEL analysis.To further investigate the effects of glucose metabolism in LPS/D-Gal-induced liver injury,we used ethyl pyruvate as a pre-and post-treatment to observe liver damage and Caspase-3 activity in the same manner as described above.Results:(1)In the LPS/D-Gal-induced liver injury,the level of PKM2 in the nucleus up-regulated,pyruvate kinase activity decreased,and pyruvate content decreased.(2)Pretreatment with PKM2 nuclear translocation inhibitor ML265 suppressed LPS/D-Gal-induced elevation of plasma aminotransferases and IL-6 levels.ML265 alleviated LPS/D-Gal-induced histological abnormalities in the liver.Compared with the model group,ML265 suppressed LPS/D-Gal-induced up-regulation of PKM2 in the nucleus.Increased pyruvate kinase activity and pyruvate content.ML265 pretreatment suppressed mortality in LPS/D-Gal exposed mice.(3)ML265 pretreatment restrained the apoptosis signal activated by LPS/D-Gal,such as TNF-?,Caspase-3,8,9 activity and TUNEL-positive apoptotic bodies decreased.(4)ML265 post-treatment alleviated liver injury caused by LPS/D-Gal,while Caspase-3,8,9 activity inhibited,TUNEL-positive apoptotic bodies reduced.Increased pyruvate kinase activity and pyruvate content.The analysis of KEGG indicated that AKT,NF-kB,c-Jun and FOXO1 A was up-regulated in the apoptosis pathway.(5)Ethyl pyruvate treatment reduced the levels of plasma aminotransferases in acute liver injury induced by LPS/D-Gal.EP alleviated LPS/D-Gal-induced histological abnormalities in the liver.EP suppressed LPS/D-Gal-induced Caspase-3 activity.Conclusion: The results of this study indicate that the accumulation of PKM2 in the nucleus plays an important role in LPS/D-Gal-induced acute liver injury,and its mechanism is related to the recovery of pyruvate content and inhibition of apoptotic signaling pathway.This suggests that inhibition of PKM2 and supplementation of pyruvate may have potential applications in the prevention and treatment of inflammatory liver diseases.