Study On The Expression And Importance Of Autophagy In Severe Acute Pancreatitis

Objective: To explore the relationship between autophagy and SAP(severe acute pancreatitis)and provide a new target for the treatment of severe acute pancreatitis by studying the expression and change of autophagy in the experimental severe acute pancreatitis.Methods: 48 healthy male SD rats were randomly divided into two groups,the control group(SO)and acute necrotizing pancreatitis(ANP)group,each group had 24 rats.Acute necrotizing pancreatitis model was induced by retrograde biliopancreatic duct injection of 5% taurocholic cholate.In SO group,pancreas were flipped only after laparotomy.Each group of rats were randomly divided into three subgroups,according to three time points,3h,6h and 12 h.And each group had 8 rats.Collecting blood at each point of time(3h,6h,12h)after establishing models respectively and testing serum levels of amylase.Fraction of pancreatic tissue were fixed by 4% formaldehyde and 2.5% glutaraldehyde respectively for HE staining and electron microscopy.And some were used to test the expression of LC3 B,Beclin1 and P62 by immunohistochemistry.Results:1.Pancreatic tissue pathology observation and rating:(1)General observation: There was no structural abnormality of pancreas in SO group,and no ascites as well.we could see tumescent pancreatic tissue at 3h in ANP group.In the 6h ANP,the pancreatic tissue swelled obviously,with some scattered petechiae and mild ascites.We could see splinter hemorrhagic necrosis in pancreatic tissue and omental around,with hemorrhagic ascites.(2)Pancreatic pathology rating: According to Schmidt criteria:(1)Between the same time of SO group and ANP group,there was a remarkable difference(P<0.05);(2)In 12 h ANP group,the degree of injury increased severely,and the rating scores were significantly different when compared with 3h ANP group(P<0.05).2.Serum amylase:(1)In SO group,there was no significant difference between the three points of time(P>0.05);(2)Values increased obviously in ANP group when compared with SO group,and there was a significant difference between them(P<0.05).3.Electron microscope performance evaluation of pancreatic tissue :(1)In the SO group,pancreatic cells nuclear membrane was integrity and the structure of the organelles was normal,and there was a small amout of autophagy in this group.(2)In the ANP group: the autophagosome with double membrane of shape “C”was visible at 3h;in the 6h ANP group,the mitochondrion swelled obviously and the number of autolysosome and autophagy increased significantly,and some autophagosome was merging with lysosomes.the number of autolysosome and autophagy in 12 h ANP group increased significantly,and we could see some lysosomes were processing endogenous substrates in cells.4.LC3 B,Beclin1 and P62 pancreatic tissue immunohistochemical staining score :(1)LC3B was little expression at each point of time in SO group,and there was no significant difference(P>0.05);compared with the same time point in SO group,LC3 B expression was increased obviously in ANP group(P<0.05),and the expression was significantly increased with time expansion(P <0.05);(2)In SO group,Beclin1 was little expression,and the expression at each time point was no significantly different(P>0.05);compared with the SO group,Beclin1 expression was significantly increased(P<0.05),especially at 6h and 12 h time point when compared with 3h ANP group respectively(P<0.05);(3)P62 was little expression at each point in time of SO group,and there existed no significant difference(P>0.05);compared with the SO group,the P62 expression in ANP was extremely increased(P<0.05),but there was no significant difference between each time point of ANP group(P>0.05).Conclusion:1.Using the method of retrograde biliopancreatic duct injection of 5% sodium taurocholate can successfully reproduce ANP animal model.2.Compared with SO group,autophagy expression in ANP group is increased;and the expression of autophagy increases as pathological damage aggravated;3.Compared with SO group,the P62 expression in ANP is extremely increases,which suggests that the function of autophagy degradation in ANP group is weakened.4.Autophagy dysfunction may be an important factor for tissue injury in severe acute pancreatitis.