The Protective Effect And Mechanism Of Chenodeoxycholic Acid On Mucus Barrier Injury Of Caco-2 Cells

Objective: Obstructive jaundice(OJ)causes bile reduced in the intestinal lumen,increased release of endotoxin,imbalance between proliferation and apoptosis of intestinal epithelial cells,resulting in damage of the intestinal barrier.Intestinal mucus layer is the first barrier against invasion of harmful substances in the gut.Mucin2(MUC2)is main constituent.Our previous studies have confirmed the intestinal mucosal barrier injury in obstructive jaundice patients and rats,decreased protein expression of MUC2 in the intestine,and increased protein expression of MUC2 after internal and external drainage of bile,and internal drainage is superior to external drainage.Therefore,we speculated that bile acids can promote protein expression of MUC2 in intestinal epithelial cells and maintain intestinal mucosal barrier function.Cheodeoxycholic Acid(CDCA)is a primary bile acid that has the strongest stimulatory effect on mucin.Previous studies have shown that Notch1 signaling pathway is essential for the proliferation and differentiation of intestinal epithelial cells,and its activation will promote the proliferation of intestinal epithelial cells and restore intestinal barrier function.Therefore,we tried to use Notch as an entry point to observe the effect of CDCA on the protein and gene expression of MUC2 in Caco-2 cells mucus injury model and to explore its mechanism of action.Methods:Intestinal epithelial Caco-2 cell mucus injury model was established in vitro by lipopolysaccharide(LPS)induction.We examined the effect of different concentrations of CDCA on the expression of MUC2 and Notch proteins and genes in Caco-2 cell mucus injury model by Western Blot and Real-Time PCR.DAPT(specific inhibitor of Notch1)was used to reverse the effect.Results:1.LPS reduces the expression of MUC2 protein and gene,and mediates mucus barrier injury of Caco-2 cells.Western Blot results showed that the expression of MUC2 protein in LPS group was lower than that in DMSO group(MUC2/β-actin 0.24±0.06,0.39±0.04,P<0.01),and the difference was statistically significant.Real-Time PCR results are consistent with it.2.Different concentrations of CDCA increases the expression of MUC2 protein and gene,protects LPS-mediated mucus barrier injury of Caco-2 cells.Western Blot results showed that compared with LPS group,the expression of MUC2 protein in 50 μM、100 μM、150 μM CDCA group was significantly higher(MUC2/β-actin 0.87±0.07,1.06±0.06,1.36±0.06,0.24±0.06,P<0.01),and the highest was 150 μM CDCA group(P<0.01).Real-Time PCR results are consistent with it.3.CDCA protects mucus barrier injury of Caco-2 cells through Notch signaling pathway.Western Blot results showed that Notch1 protein expression was decreased in DAPT(20 μM)group compared with CDCA(150 μM)group(Notch1/β-actin 0.14±0.01,1.35±0.05,P<0.05),accompanied by decreased expression of MUC2 protein(MUC2/β-actin 0.28±0.03,1.36±0.06,P<0.05),and there was no statistical difference between the LPS group and the DAPT(20μM)group(P>0.05).Real-Time PCR results are consistent with it.Conclusion:1.LPS reduces the expression of MUC2 protein and gene,and mediates mucus barrier injury of Caco-2 cells.2.Different concentrations of CDCA increases the expression of MUC2 protein and gene,protects LPS-mediated mucus barrier injury of Caco-2 cells,and in a dose-dependent manner.3.CDCA protects the mucus barrier injury of Caco-2 cells by upregulating Notch signaling pathway.