| Objective observing the expression pattern of NF-κB, HMGB1 in the intestinal mucosal damage during acute necrotizing pancreatitis in rats at different time points, which is beneficial to investigate the correlation between NF-κB, HMGB1 and the intestinal mucosal damge during acute necrotizing pancreatitis.Methods 70 healthy SD rats are randomly divided into two group:the ANP group (the experimental group, A group), n=40; SO group (the sham operation group, S group), n=30;The two groups are respectively divided into five different time point groups:the 3h,6h,12h,24h,36h time point group, and the number of each time point for A group was 8 while the S group was 6. The rats of A group are all retrograde injected with 5% sodium taurocholate (0.1 ml/1 OOg,0.2ml/min) into the biliopancreatic duct to establish the ANP model, whlie the pancreas of the rats are turned up only in S group. The rats of A group and S group are continually forbidden to drink and eat after the model, and they are immediately injected with the 5% glucose and sodium chloride on the subcutaneous, repeated every 6h.The two groups rats are opened abdominal again at 3h,6h,12h,24h,36h time point, to observe the general situation in the abdominal cavity and the tissue of the pancreas and small intestine, to detect the concentration of AMY, Ca2+,DAO of the serum in the portal vein and the expression of NF-κBp65 and HMGB1 of the intestinal with ELISA and immunohistochemistry, to evaluate the pathological score of the pancreas and small intestine through light microscope after HE staining.Results (1) The general situation to the abdomen:pieces of hemorrhage and necrosis of pancreas, hemorrhagic ascites and saponification spot, gastric bowel bilges gas and expansion could be obviously found in A group, which changes gradually worsened over the time point. Those changes which are mentioned above could not be observed each time point in S group.(2) The concentrations of AMY in the serum of the blood:The concentrations of the AMY in the serum of the blood of A group are higher than the time point group with the corresponding in S group,,P<0.05. Compared with the concentrations of the AMY in the serum of the blood between the different time point group in the A group:①A3h lower than A6h,A12h,A24h,A36h,P<0.05;②A6h lower than A12h,A36h, P>0.05, A6h lower than A24h, P<0.05;③A12h lower than A24h, A36h, P>0.05;④A24h higher than A36h, P<0.05. While compared with the concentrations of the AMY in the serum of the blood between the different time point group in the S group, there is no significant difference, P>0.05.(3) The concentrations of Ca2+ in the serum of the blood:The concentrations of the Ca2+ in the serum of the blood of A group are lower than the time point group with the corresponding in S group,,P<0.05.Compared with the concentrations of the Ca2+ in the serum of the blood between the different time point group in the A group:①A3h higher than A6h,A12h,A24h,A36h,P<0.05;②A6h higher than A24,A12h,A36h, P<0.05;③A12h lower than A24h, A36h, P>0.05;④A24h higher than A36h, P>0.05. While compared with the concentrations of the Ca2+ in the serum of the blood between the different time point group in the S group, there is no significant difference, P>0.05.(4) Pancreas histopathology (light microscopy):①A3h:pancreas slight edema, acinar cells slightly swollen, spotting, a small amount of inflammatory cells infiltrated and the gland structure was normal;②A6h:pancreas edema, acinar cells significantly swollen, pieces of hemorrhage, more inflammatory cells infiltrated, while the gland structure profile was normal.③A12h:pancreatic obvious edema, acinar cells significantly swollen, pieces of hemorrhage and necrosis, a amount of inflammatory cells infiltrated, some of the normal gland structure disappeared.④A24h:the hemorrhage and necrosis of the pancreas tissue was significant, a large number of inflammatory cells infiltrated, most of the normal gland structure disappeared.⑤A36h:most of the pancreas with hemorrhage and necrosis, a large number of inflammatory cells infiltrated, the normal gland structure completely disappeared, which was unable to distinguish the normal acinar cells.⑥S group:the gland structure of the pancreas is normal, acinar cells integrity, no necrosis, hemorrhage and no inflammatory cells infiltrated.(5)Intestinal histopathology (light microscopy):①A3h:intestinal villis slight edema, no epithelial cells shed, the gland was normal and a small amount of inflammatory cells infiltrated;②A6h:intestinal villis edema, the space of the intestine subepithelial expanded, a part of epithelial cells shed, intestinal villis still neatly arranged and more inflammatory cells infiltrated;③A12h:intestinal villis obvious edema, the space of the intestine subepithelial expanded, some of the epithelial cells shed, intestinal villis irregularly arranged and a lot of inflammatory cells infiltrated;④A24h:intestinal villis obvious edema, the space of the intestine subepithelial expanded evidently, a large number of epithelial cell necrosis and shed, intestinal villis irregularly arranged and a lot of inflammatory cells infiltrated;⑤A36h:intestinal villis shorter, irregularly arranged and fusion in pairs, a large number of epithelial cells necrosis and focal ulcers, the space of the intestine subepithelial expanded evidently, a lot of inflammatory cells infiltrated.⑥S group:the basic morphology and the subcutaneous space of intestinal villis are normal, the columnar cells and goblet cells of intestinal villis are normal, without any significant inflammatory cells infiltrated.(6) The pathological score of the pancreas:the pathological score of the pancreas of the A group was higher than time point group with the corresponding in S group,,P <0.05.Compared with the pathological score of the pancreas between the different time point group in the A group:①A3h lower than A6h,A12h,A24h,A36h, P<0.05;②A6h lower than A12h,A24h,A36h, P<0.05;③A12h lower than A24h,A36h, P<0.05;④A24h lower than A36h, P<0.05. While compared with the pathological score of the pancreas between the different time point group in the S group, there is no significant difference, P>0.05.(7) The pathological score of the small intestine:Compared the pathological score of the small intestine of the S group with the A group:①A3h lower than S3h.P >0.05;②A6h,A12h,A24h,A36h higher than the S group, P<0.05. Compared with the pathological score of the small intestine between the different time point group in the A group:①A3h lower than A6h,A12h,A24h,A36h, p<0.05;②A6h lower than A12h,A24h,A36h, P<0.05;③A12h lower than A24h,A36h, P<0.05;④A24h lower than A36h, P>0.05.While compared with the pathological score of the small intestine between the different time point group in the S group, there is no significant difference, P>0.05.(8) The expression of NF-κBp65 in the intestinal mucosal cells:the expression of NF-KBp65 in the intestinal mucosal cells at 3h,6h,12h time point groups of the A group are higher than the time point groups with the corresponding in S group,,P <0.05. A24h,A36h lower than S24h,S36h, P>0.05.Compared with the expression of NF-κBp65 in the intestinal mucosal cells between the different time point group in the A group:①A3h higher than A6h, A12h, A24h, A36h, P<0.05;②A6h higher than A12h,A24h,A36h, P<0.05;③A12h higher than A24h,A36h, P<0.05;④A24h higher than A36h, P>0.05. While compared with the expression of NF-κBp65 in the intestinal mucosal cells between the different time point group in the S group, there is no significant difference, P>0.05(9) The expression of HMGB1 in the intestinal mucosal cells:①A3h higher than S3h, P>0.05;②A6h, A12h,A24h,A36h higher than S6h, S12h,S24h,S36h, P<0.05; Compared with the expression of HMGB1 in the intestinal mucosal cells between the different time point group in the A group:①A3h lower than A6h, A12h, A24h, A36h, P<0.05;②A6h lower than A24h, A36h, P<0.05, while A6h lower than A12h, P >0.05;③A12h lower than A24h,A36h, P<0.05;④A24h higher than A36h, P>0.05. While compared with the expression of HMGBl in the intestinal mucosal cells between the different time point group in the S group, there is no significant difference, P>0.05(10) The concentrations of NF-κBp65 in the small intestine tissues:A3h,A6h,A12h higher than the time point groups with the corresponding in S group, P<0.05. A24h,A36h higher than S24h,S36h, P>0.05.Compared with the concentrations of NF-KBp65 in the small intestine tissues between the different time point group in the A group:①A3h higher than A6h, A12h, A24h, A36h, P<0.05;②A6h higher than A12h,A24h,A36h, P<0.05;③A12h higher than A24h,A36h, P<0.05;④A24h higher than A36h, P>0.05. While compared with the concentrations of NF-KBp65 in the small intestine tissues between the different time point group in the S group, there is no significant differentce, P>0.05(11) The concentrations of HMGB1 in the small intestine tissues:A3h higher than S3h, P>0.05,the other time point groups in the A group were higher than the time point groups with the corresponding in S group, P<0.05. Compared with the concentrations of HMGB1 in the small intestine tissues between the different time point group in the A group:①A3h lower than A6h, A12h, A24h, A36h, P<0.05;②A6h lower than A24h, A36h, P<0.05, A6h lower than A12h, P>0.05;③A12h lower than A24h,A36h, P<0.05;④A24h than A36h, P>0.05. While compared with the concentrations of HMGB 1 in the small intestine tissues between the different time point group in the S group, there is no significant difference, P>0.05(12) The concentrations of DAO in the serum of the blood:A3h higher than S3h, P>0.05,while the other time point groups in A group were higher than the time point groups with the corresponding in S group, P<0.05.Compared with the concentrations of DAO in the serum of the blood between the different time point group in the A group: A3h lower than A6h,A12h,A24h,A36h, P<0.05;②A6h lower than A24h,A36h, P<0.05, while A6h lower than A12h, P>0.05;③A12h lower than A24h,A36h, P<0.05;④A24h lower than A36h, P>0.05.While compared with the concentrations of DAO in the serum of the blood between the different time point group in the S group, there is no significant difference, P>0.05.(13) correlation analysis:①There is positive correlation between the pathological score of the pancreas and the pathological score of the small intestine, and the correlation coefficient is 0.866, which is statistically significant difference, P< 0.05.②There is positive correlation between the concentrations of DAO in the serum of the blood and the pathological score of the small intestine, and the correlation coefficient is 0.78, which is statistically significant difference, P< 0.05.③There is negative correlation between the pathological score of the small intestine and the concentrations of NF-KBp65 of the small intestine, and the correlation coefficient is-0.893, which is statistically significant difference, P< 0.05.④There is positive correlation between the pathological score of the small intestine and the concentrations of HMGB1 of the small intestine, and the correlation coefficient is 0.798, which is statistically significant difference, P<0.05.⑤There is negative correlation between the concentrations of DAO in the serum of the blood and the concentrations of NF-κBp65 of the small intestine, and the correlation coefficient is-0.795, which is statistically significant difference, P<0.05.⑥There is positive correlation between the concentrations of DAO in the serum of the blood and the concentrations of HMGB1 of the small intestine, and the correlation coefficient is 0.870, which is statistically significant difference, P<0.05.Conclusions (1) there is intestinal mucosa damge at 6 hour after the model of ANP. (2) The highest expression of NF-κB in the intestinal mucosal at 3 hour after the model of ANP, decreasing gradually over the time point,which is negative correlation with the concentrations of DAO in the serum of the blood and the pathological score of the small intestine. (3) The expression of HMGB1 in the intestinal mucosal increases at 6 hour after the model of ANP, increasing gradually over the time point,which is positive correlation with the concentrations of DAO in the serum of the blood and the pathological score of the small intestine.(4) HMGB1 take part in the intestinal mucosa damge during ANP in rats. |
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