Interaction Between The Gegen Qinlian Decoction And The Gut Microbiota Of Infectious Diarrheal Mini-pigs

Objective:The aim of this study was to investigate the mechanism for metabolism differences of the bioactive components of GQD between the diarrheal and healthy gut microbiota(GM).GQD regulated GM strucute,Short-chain fatty acids(SCFAs),and mucosal inflammation in Escherichia coli(E.coli)-induced diarrheal mini-pigs was studied.It is suggested that the interaction between the GQD and GM of mini-pigs could further understand the mechanism of GQD for the treatment of diarrhea.Methods:(1)A HPLC-MS/MS method for simultaneous analysis of 16 major ingredients from GQD in bacterial solution was developed and applied to a comparative study on microbial metabolism between diarrheal and normal state.(2)Mechanism of the metabolism differences of GQD components between diarrheal and healthy state was studied by the type,activities,and sources of microbial enzymes.(3)Effect of GQD on the modulation of GM was detected by 16S rRNA high throughout sequencing analysis.(4)Effect of GQD on the levels of fecal SCFAs was analyzed by GC-MS;Effect of GQD on the mucosal inflammation was assessed by immunohistochemical and western blotting analysis.(5)During the administration,pharmacokinetics of 11 components of GQD and levels of three SCFAs in the mini-pig blood were analyzed to study their changes in vivo.Results:After co-incubation,hydrolysis of glycosides and production of glycones was accelerated by E.coli-induced diarrheal GM,and degradation of alkaloids was facilitated.(2)?-Glucosidase,?-glucuronidase,and nitroreductase were the major microbial enzymes involved in the metabolism of ingredients from GQD.The activities of these enzymes were higher in diarrheal state,which is mainly due to the proliferation of Escherichia spp.(3)GQD alleviated diarrheal symptoms of mini-pigs and modulated structure of GM.GQD significantly increased and reduced the abundances of Bacteroideles and Proteobacteria,respectively,which facilitates the growth of symbiotic bacteria.(4)GQD remarkably improved levels of fecal SCFAs,alleviated mucosal TNF-? and IL-6 levels,HDAC activity,and NF-?B p65 expression,which suggests that anti-inflammatory efficacy of GQD may be due to an inhibitory effect of SCFAs on HDAC and NF-?B pathways.(5)During the experiments,plasma levels of ingredients in GQD were reduced to the normal gradually,and levels of serum SCFAs were increased to the healthy condition,indicating the modelation of GM.Conclusions:Diarrheal GM could accelerate the transformation of the glycosides of GQD into the glycones.These results may be due to the proliferation of Escherichia spp.,which facilitates the secretion of metabolized enzymes and increases its activity.The anti-diarrheal and anti-inflammatory activities of glycones are better than those of their prototype,indicating the better efficacy of GQD after microbial transformation.GQD could inhibit growth of pathogenic bacteria,promote growth and colonization of symbiotic bacteria,which modulates structure of GM in diarrheal mini-pigs.GQD could facilitate production of SCFAs,alleviate mucosal inflammation by inhibiting HDAC and NF-?B pathways,which restores intestinal micro-ecology.Our findings may elucidate the mechanism of GQD on the treatment of infectious diarrhea.