Regulatory Effect And Mechanism Of High Mobility Group Protein B1 On Autophagy And Apoptosis Of Lymphocytes In Severe Acute Pancreatitis

Objective: To investigate the role of HMGB1 in autophagy and apoptosis of spleen lymphocytes in mice with severe acute pancreatitis(SAP)and its molecular mechanism.Methods: 50 adult male SPF BALB/C mice were randomly divided into control group,mild acute pancreatitis(MAP)group,SAPgroup,SAP+ anti-HMGB1 antibody group(hereinafter abbreviated as SAP+ Ab group)and SAP+ glycyrrhizin(GL)group.The control group was given intraperitoneal injection of normal saline,The MAPgroup was induced by intraperitoneal injection of caerulein,the SAPgroup was induced by intraperitoneal injection of caerulein and lipopolysaccharide(LPS),and the SAP+Ab group and SAP+GL group were given intraperitoneal injection of anti-HMGB1 antibody and glycyrrhizin(GL)respectively after successful SAPmodel.12 hours after the establishment of the model,the pancreatic tissue was taken for HE staining and the pancreatic injury was graded.The levels of serum amylase and lipase were determined by ultraviolet spectrophotometer.The expression levels of HMGB1,Caspase-3 and LC3-II in serum were detected by ELISA.The apoptosis rate and autophagy rate of spleen lymphocytes were detected by flow cytometry(FCM).The mRNA and protein expression levels of factors such as HMGB1,Caspase-3,LC3-II,receptor for advanced glycation end produts(RAGE),phosphorylated p38 mitogen activated protein kinase(p-p38 MAPK)in spleen lymphocytes were detected by RT-q PCR and Western blot.Results:(1)Compared with the control group,the pancreatic histopathological score and HMGB1 expression level in MAP,SAP,SAP+ Ab and SAP+ GL groups were increased(P<0.05);Compared with MAPgroup,the pancreatic histopathological score and HMGB1 expression in SAP,SAP+ Ab and SAP+ GL groups were increased(P<0.05);Compared with SAPgroup,the pancreatic histopathological score and HMGB1 expression level of mice in SAP+ Ab and SAP+ GL groups were significantly decreased(P<0.05).(2)Compared with the control group,the apoptosis rate and autophagy rate of spleen lymphocytes in MAPgroup were only slightly increased(P>0.05),the apoptosis rate and autophagy rate of spleen lymphocytes in SAPgroup,SAP+ Ab group and SAP+ GL group was significantly increased(P<0.05);compared with MAPgroup,the apoptosis rate and autophagy rate of spleen lymphocytes in SAPgroup,SAP+ Ab group and SAP+ GL group were significantly increased(P<0.05);compared with SAPgroup,the apoptosis rate and autophagy rate of spleen lymphocytes in SAP+ Ab group and SAP+ GL group were significantly decreased(P<0.05).The expression level of HMGB1 in SAPgroup was positively correlated with the autophagy rate and apoptosis rate of spleen lymphocytes(P<0.05).(3)Compared with the control group,the expression of Caspase-3,LC3-II mRNA and protein in MAPgroup was only slightly increased(P>0.05),while the expression of Caspase-3,LC3-II mRNA and protein in SAPgroup,SAP+ Ab group and SAP+ GL group was significantly increased(P<0.05);compared with MAPgroup,the expression of Caspase-3,LC3-II in SAPgroup,SAP+Ab group and SAP+ GL group was significantly increased(P<0.05);compared with SAPgroup,the expressions of Caspase-3,LC3-II mRNA and protein in SAP+ Ab group and SAP+ GL group were significantly decreased(P<0.05).The expression levels of HMGB1 mRNA,HMGB1 protein and serum HMGB1 in spleen lymphocytes in SAPgroup were positively correlated with the expression levels of LC3-II and Caspase-3 mRNA in spleen lymphocytes,LC3-II and Caspase-3 protein in spleen lymphocytes,LC3-II and Caspase-3 in serum,respectively.(4)Compared with the control group,the expression levels of p-p38 MAPK protein,RAGE mRNA and protein in MAPgroup were only slightly increased(P>0.05),while the expression levels in SAPgroup,SAP+ Ab group and SAP+ GL group were significantly increased(P<0.05);compared with MAPgroup,the expression levels of p-p38 MAPK protein,RAGE mRNA and protein in SAPgroup,SAP+ Ab group and SAP+ GL group were significantly increased(P<0.05);Compared with SAPgroup,the expression levels of p-p38 MAPK protein,RAGE mRNA and protein in SAP+ Ab group and SAP+ GL group were significantly decreased(P<0.05).In SAPgroup,the expression levels of HMGB1 mRNA and HMGB1 protein in spleen lymphocytes were positively correlated with the expression levels of RAGE mRNA,RAGE and p-p38 MAPK protein in spleen lymphocytes,respectively.Conclusion:(1)In acute pancreatitis,especially in SAPmice,the expression of HMGB1 in spleen lymphocytes was significantly up-regulated,and the autophagy and apoptosis of lymphocytes were increased.(2)The high expression of HMGB1 activates RAGE/p38 MAPK signal pathway,which leads to the up-regulation of LC3-II and Caspase-3 expression,which may be an important reason for the increase of autophagy and apoptosis of spleen lymphocytes in SAPmice.Inhibition of HMGB1 expression can reverse the above effects and improve pancreatic injury.HMGB1 may be a potential target for SAPtherapy.