| Objective: Severe acute pancreatitis(SAP)is a very serious disease with rapid onset,high mortality,and frequent complications.Serotonin(5-HT),as a neurotransmitter,in the signal transduction system of its synthesis,release,binding to receptors,and reuptake,etc.,any abnormality in any link may lead to changes in gastrointestinal motility and secretion function.and the production of visceral hypersensitivity.However,the role of5-HT in SAP intestinal mucosal barrier damage remains unclear.The aim of our study was to investigate the function of 5-HT on the intestinal tight junction barrier in a SAP rat model through the 5-HT signaling pathway,and to seek new ideas for improving the intestinal mucosal barrier damage caused by SAP.Methods: 30 SPF healthy male Wistar rats were randomly divided into sham operation group(SO)(n=10),severe acute pancreatitis group(SAP)(n=10)and p-chlorophenylacetic acid intervention group(SAP+ p-chlorophenylalanine [PCPA])(n=10).Rat SAP model was established by retrograde cholangiopancreatography and retrograde injection of 5%sodium taurocholate into the pancreatic duct.The intervention group received intraperitoneal injection of PCPA(200 mg/kg)24 hours before SAP model induction,and rats in each group were killed 24 h after SAP induction.At the end of the experiment,rats in each group were killed after anesthesia,and intestinal tract,pancreatic tissue and inferior vena cava blood was collected for the follow-up experiment.Serum amylase and lipase levels were measured by automatic biochemical analyzer,and the concentrations and contents of tumor necrosis factor-α(TNF-α),interleukin-6(IL-6)and 5-HT in ileum were also measured.He staining was used to evaluate the pathological changes of pancreas and intestines of rats in each group,and pathological scores were made.The expression of intestinal tight junction proteins Occludin,Claudin-1 and zonulacluden-1(ZO-1)was detected by Westernblot.The expression of tryptophan hydroxylase TPH1 was detected by q-PCR and immunohistochemistry)and serotonin reuptake transporter(SERT);Immunofluorescence method was used to detect the distribution of TJ protein in intestine.Results:(1)the results of biochemistry and ELISA detection.Compared with SO group,the levels of serum lipase,amylase,TNF-α and IL-6 in SAP group and SAP+PCPA group were significantly higher,while the levels of lipase,amylase,TNF-α and IL-6 in SAP+PCPA group were significantly lower than those in SAP group.At the same time,the intestinal 5-HT content in SAP and SAP+PCPA groups was significantly higher than that in SAP group,while the 5-HT content in SAP+PCPA group was significantly lower than that in SAP+PCPA group.Besides,there was no significant difference in serum 5-HT level among the three groups.(2)HE staining.No pathological changes were found in intestinal epithelium and pancreas in SO group.The pathological scores of pancreatic and intestinal injury in SAP group and SAP+PCPA group were significantly higher than those in SO group,however those in SAP+PCPA group were significantly lower than those in SAP group.(3)the expression of TJ protein in ileum of rats in each group was detected by Westernblotting.The expressions of occludin,Claudin-1 and ZO-1 in SAP group and SAP+PCPA group were significantly inferior to SO group.Moreover,the expression of occludin,Claudin-1 and ZO-1 in SAP+PCPA group was significantly lower than that in SAP group.These results reflect the damage of intestinal mucosal barrier in rats.Although the injury of intestinal mucosal barrier could not be reversed in SAP+PCPA group,it was significantly improved.(4)the expression of TPH1 and SERT was detected by q-PCR and immunohistochemistry.Compared with SO group,the expression of TPH1 in SAP group and SAP+PCPA group was significantly increased,however the expression of SERT was significantly decreased.Besides,the expression of TPH1 in SAP+PCPA group was significantly lower than that in SAP group,while the expression of SERT was significantly increased in SAP+PCPA group.These results showed that the expression of TPH1 increased and the expression of SERT decreased in SAP rats,which further explored the reason for the increase of 5-HT in SAP rats,which could be significantly improved in SAP+PCPA rats.(5)the expression of TJ protein in intestinal tissue was detected by immunofluorescence staining.ZO-1(green),latent protein(green)and compact protein-1(green)were observed at the junction of intestinal epithelial cells.Compared with SO group,the expression of TJ protein in SAP and SAP+PCPA groups was significantly decreased,and the expression of TJ protein in SAP+PCPA group was significantly higher than that in SAP group.Combined with the pathological results,it was determined that the degree of intestinal injury was related to the expression of TJ protein,which was consistent with the results of westernblotting..Conclusion: This study confirmed that in the SAP rat model,the activity of TPH1 in intestinal epithelial cells was inhibited by PCPA,reducing the endogenous release of5-HT in the rat intestine can reduce the damage of intestinal TJ protein and protect the intestinal mucosal barrier of SAP patients.It is suggested that the 5-HT signaling pathway provides new ideas in improving the intestinal mucosal barrier damage caused by SAP. |
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