Sodium Butyrate Ameliorates Visceral Hypersensitivity Via Down-regulating IRAK1

Background:Irritable bowel syndrome(IBS)has been regarded as one of the most common digestive diseases which hampered not only the living quality of patients but also the development of social economy.Many studies have shown that the status of visceral hypersensitivity could be the clinical feature of partial IBS patients,which is possibly related to the damage of intestinal epithelial barrier.The intestinal epithelial barrier contributes to sustaining intestinal homeostasis and preventing toxic factors from infiltrating into body.Subsequently,the low-grade intestinal epithelial inflammation in some IBS patients is considered to be an important pathogenic factor.Sodium butyrate,mainly produced by intestinal Clostridium,could sustain symbiosis with the host and participate in intestinal mucosal physiological function.Pre'vious studies demonstrated that sodium butyrate could suppress intestinal inflammation.A recent study shows that the abundance of butyrate-producing flora was declined in IBS patients' feces,indicating the role of butyrate in the pathogenesis of IBS.Interleukin 1 receptor associated kinase(IRAK1)as an important signal molecule in toll-like receptors signaling pathways,could promote intestinal inflammation through regulating related factors.The present study was designed to investigate relationship between expression of IRAK-1 in colon and visceral hypersensitivity via clinical and animal samples.Furthermore,we demonstrated the mechanism of butyric acid in treatment of IBS patients via vitro experiment.Objectives1.To investigate the correlation between abdominal pain and intestinal epithelial expression of IRAK1.2.To investigate the function of sodium butyrate to the expression of epithelial IRAK1 and p-IRAK1 and visceral hypersensitivity in IBS animal model.3.To investigate the function of sodium butyrate to the expression of IRAK1 and p-IRAK1 in vitro experiment.Methods:1.Clinical studiesIBS-D patients(assessed according to the Rome ? criteria)were recruited.All the participants were asked to score the frequency and severity of their abdominal symptoms by a validated questionnaire.The intestinal epithelial biopsy specimens were processed for IRAKI immunohistochemistry to analyse the correlation between abdominal pain and intestinal IRAK1 expression.2.Animal studiesIntracolonic 2,4,6-trinitrobenzenesulfonic acid(TNBS)in 50%ethanol was used to produce IBS model.Sodium butyrate group was treated with sodium butyrate(100nmol)intracolonically after TNBS treatment.The 28m day was the end point.Colorectal distension test was used to evaluate the visceral hypersensitivity.Mucosal samples from mice were collected.Mucosal IRAKI and p-IRAK1 levels were measured by specific ELISA kit and immunohistochemistry to evaluate the correlation between visceral hypersensitivity and protein expression.3.Cell studiesHT-29 cells were cultured under appropriate condition.After divided into three groups,IL-33(10ng/ml)stimulated HT-29 cells,with or without sodium butyrate(5mmol/L)pretreatment.Total protein extracts were collected after 2 h,using western blot method to test IRAK1 and p-IRAK1 protein expression.Results:1.Clinical studiesThe expression of IRAK-1 in colon epithelium of IBS-D patients was higher compared with normal group.The expression of IRAK-1 had a positive correlation with abdominal pain score and frequency score.2.Animal studiesThe expression of IRAKI and p-IRAKI were both up-regulated in colon epithelium of model mice compared with control group,and had negative correlation with visceral pain threshold.Compared with control group,the expression of IRAKI and p-IRAKI was down-regulated in sodium butyrate treated group and the visceral pain threshold of treated group increased.3.Cell studiesIL-33 stimulation could increase the protein expression of IRAKI and p-IRAK1 in HT-29 cells,and sodium butyrate pretreatment could diminish this effect.Conclusion:1.Intestinal epithelial IRAKI might facilitate abdominal pain in IBS-D patients.2.Intestinal epithelial IRAKI might facilitate visceral hypersensitivity in IBS animal model.Sodium butyrate may function through down-regulating IRAKI and p-IRAK1.3.HT-29 cells could resist up-regulation of IRAKI and p-IRAK1 by IL-33 stimulation under sodium butyrate pretreatment.