| Objective:In this study,The Yangyin Yiqi Runchang Decotion(YYRD),a clinical prescription from a famous traditional Chinese medicine(TCM)professor Li Qingsheng in Yunnan Province,was selected to intervene in slow transit constipation(STC)model rats.Then we observed the effects of YYRD on the intestinal flora,short-chain fatty acids,interstitial cells of Cajal(ICCs),5-hydroxytryptamine and vasoactive intestinal peptide in STC model rats.The study was designed to explore whether YYRD could alleviate STC via improving intestinal flora composition and gastrointestinal motility,and to preliminarily explain the molecular mechanism of the laxative effect of YYRD,in order to provide experimental basis and theoretical support for clinical application.Method: 1.The STC model rats was replicated with compound diphenoxylate tablets.Whether the model was successfully replicated was evaluated by observing changes in the intestinal transit rate,fecal water content and colon pathology.2.The rats were gavaged with compound diphenoxylate for 7 days.Since the 8th day of the experiment,the TCM intervention was started by intragastric administration while continuing to replicate STC model rats.Rats in the positive control group were gavaged with Live Combined Bifidobacterium and Lactobacillus Tablets(LCBL).After 7 days of drug intervention,all rats were sacrificed,and samples were collected to detect experimental indicators.3.Tested indicators.(1)Evaluation indicators of drug efficacy: The intestinal transit rate of rats was detected by the carbon powder propulsion trial,and the fecal water content of rats was detected by the drying method.(2)Indicators related to intestinal motility: The concentrations of 5-hydroxytryptamine(5-HT)and vasoactive intestinal peptide(VIP)in rat plasma were detected by ELISA.The expression of interstitial cells of Cajal(ICCs)in colon tissue was detected by immunohistochemistry.(3)Indicators related to intestinal flora: The composition of the intestinal flora of rats was detected by 16 S rDNA sequencing and the concentration of short-chain fatty acids(SCFAs)was detected by gas chromatography.Result: 1.YYRD stimulated intestinal transit rate and increased fecal water content.(1)Effect of YYRD on the intestinal transit rate of rats: Compared with the Control group,the model rat's intestinal transit rate decreased significantly(P<0.01).Compared with the model group,the intestinal transit rate of rats in all drug intervention groups increased,and the differences were statistically significant.(2)Effect of YYRD on the fecal water content of rats: Compared with the control group,the fecal water content of rats in model group decreased.Compared with the model group,the fecal water content of rats in all drug intervention groups increased.(3)Effect of YYRD on the pathology of rat colon tissue: Compared with the control group,the mucosa of the colon tissue of the model group was partially shed,the muscle layer was slightly thinned,and the number of goblet cells was reduced.Compared with the model group,the mucosal detachment of colon tissue and the thickness of muscular layer were improved in all drug groups,and the number of goblet cells increased.2.YYRD promoted intestinal motility via increasing the expression of ICCs in rat colon tissue and regulating the concentration of 5-HT and VIP.(1)Effect of YYRD on ICCs of rats colon tissues: Compared with the control group,the positive expression of specific c-kit protein of ICCs of rats in model group was significantly reduced(P<0.01),indicating the number of ICCs in the colon significantly reduced.Compared with the model group,the positive expression of c-kit protein in all drug intervention groups increased,and the differences were statistically significant,indicating that the number of ICCs in the colon increased.(2)The effect of YYRD on the plasma 5-HT and VIP concentrations in rats: Compared with the control group,the 5-HT concentration in the model group decreased significantly(P<0.01).Compared with the model group,the plasma 5-HT concentration in all TCM and LCBL groups increased,and the differences were statistically significant.Compared with the control group,the plasma VIP concentration of the model group increased significantly(P<0.05).Compared with the model group,the plasma VIP concentration of all TCM groups decreased,and the differences were statistically significant.3.YYRD improved the composition of intestinal flora and its metabolite-SCFAs concentrations.(1)Effects of YYRD on the composition and abundance of intestinal flora in rats: At the phylum level,the relative abundance of 7 communities represented by Firmicutes in the model group increased,and The relative abundance of communities represented by Bacteroidetes and Proteobacteria was reduced.The relative abundance of the three communities represented by Firmicutes and Proteobacteria was increased in the YYRD medium-dose group,and the relative abundance of the 8 communities represented by Bacteroidetes was reduced.At the genus level,the relative abundance of the 12 genera represented by Bacteroides,Prevotella,and Blautia in the model group increased.The relative abundance of the 8 genera represented by Oscillospira,Lactobacillus and Coprococcus was reduced.The relative abundance of 11 genera represented by Phascolarctobacterium and Oscilillospira was increased and the 8 genera represented by Bacteroidetes reduced in the YYRD medium-dose group.Based on the relative abundance of genus-level species,YYRD's differential flora was selected,including Clostridium,Alisipes,Treponema,Staphylococcus,and Turicibacter.The results suggested that YYRD could increase the relative abundance of bacteria,such as Clostridium and Alisipes,that have potential to promote intestinal peristalsis and reduce the relative abundance of conditioned pathogenic bacteria such as Treponema,Staphylococcus and Turicibacter.(2)The effect of YYRD on the concentrations of intestinal flora metabolites-SCFAs in rat intestinal contents: Compared with the control group,the concentration of SCFAs(acetic acid,propionic acid and butyric acid)in the model group increased significantly(P<0.05);Compared with the model group,the concentration of SCFAs in all TCM and LCBL groups showed a downward trend(P>0.05).Conclusion: 1.YYRD can promote the expression of interstitial cells of Cajal in the colon,increase the concentration of serotonin and reduce the concentration of vasoactive intestinal peptide,thereby promoting intestinal transmission,improving intestinal motility,and alleviating STC.2.YYRD can improve the slow transit state of the intestine by adjusting the balance of intestinal flora and the concentration of short-chain fatty acids in rats with STC.3.YYRD's improvement of intestinal flora imbalance and intestinal motility disorder is one of its mechanisms for prevention and treatment of STC. |
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