Pharmacological Effects And Pharmacological Basis Of Shengjiangxiexiin Tang On Mice With Clostridium Difficile-associated Diarrhea

Rationale: In recent years,the incidence of Clostridium Difficile Associated Diarrhea(CDAD)has been increasing,with more patients experiencing recurrence and severity,posing a serious threat to public health.Currently,available clinical treatment programs have their limitations,highlighting the urgent need for new treatment plans and alternative medications.Shengjiangxiexin Tang(SJT),a traditional Chinese medicine recorded in "Treatise on Cold-Damage Diseases," is composed of eight medicinal herbs including fresh rioma of Zingiber officinale Rosc.,dried rioma of Zingiber officinale Rosc.,Panax ginseng C.A.Mey.,Pinellia ternata Breit.,Glycyrrhiza uralensis Fisch.,Scutellaria baicalensis Georgi.,Coptis chinensis Franch.,Ziziphus jujuba Mill.,and is a classic prescription for treating diarrhea which has been widely used in practical treatments.In our previous studies,SJT was used as an adjuvant therapy drug for CDAD,and significant curative effects were observed.However,its pharmacological substance basis and therapeutic mechanism are still unclear,which has limited its clinical application.Therefore,this study replicated the CDAD model and explored the mechanism and pharmacological substance basis of SJT in treating CDAD at the pharmacodynamic,intestinal microbiological,metabolomic,mass spectrometry imaging,and other levels.Objective: This study aimed to systematically analyze the metabolic mechanism and key pharmacodynamic components of SJT in CDAD mice by combining pharmacodynamics,intestinal microbial levels,metabolomics,and mass spectrometry imaging.The results of this study enrich the scientific evidence of the active substance components of Shengjiangxiexin Tang.Materials and methods:1)This study aimed to observe the therapeutic effects of SJT on CDAD by establishing a CDAD mouse model and assessing parameterssuch as body weight,toxin levels,pathology,and immunity.2)We analyzed the changes in intestinal flora of CDAD mice after SJT treatment using 16 S r DNA high-throughput sequencing technology.3)We applied non-targeted serum metabolomics and serum pharmacochemistry to study the metabolic mechanism and active substance composition of SJT in CDAD.4)We used the air-flow-assisted desorption electrospray ionization mass spectrometry imaging(AFADESI-MSI)technique to obtain the distribution of drug components in the tissues of SJT-treated CDAD mice and screened the important tissue prototype components.5)Finally,we constructed a multi-scale and multi-factor network based on Spearman correlation analysis for comprehensive visualization and analysis.Results:1)The administration of SJT treatment significantly increased the body weight of CDAD mice and reduced fecal toxin levels.HE staining showed that SJT significantly reduced colonic damage.At the immune cell level,the CD25 percentage increased(P < 0.05),and LY6 G and CD11 B percentages decreased(P < 0.05)after SJT treatment.At the molecular level,CXCL,IL1-β,and TNF-αexpression were reduced(P < 0.001),and IL-22 expression was increased(P < 0.001)in the SJT group.2)The SJT group restored the C.difficile-induced disorder of intestinal microbiota and partially restored the composition of intestinal flora in three phyla of Firmicutes,Proteobacteria,and Bacteroidetes horizontal flora,and Muribaculaceae,Blautia,Parasutterella,Lachnoclostridium,Akkermansia,and Anaerotruncus abundance of six genera.3)Non-targeted serum metabolomics studies showed that SJTimproved the metabolic profile of CDAD mice by significantly back-regulating 24 differential metabolites involving eight metabolic pathways,including taurine and hypotaurine metabolism,ascorbate and aldehyde metabolism,glycerolipid metabolism,and pentose and glucuronide interconversion.Seventeen SJT entry components were screened by serum medicinal chemistry strategy,and biomarkers were correlated with the entry components to screen for kaempferol-3-arabofuranoside,Suffruticoside A,methyl rutin(methoxylutcolin),6 gingerol(6-Shogaol),6-gingerol,feruloyltyramine,3-Hydroxyglabrol(II),wogonin,berberrubine,and nine other potent components potentially effective for CDAD treatment.4)Based on the comparison of mass spectrometry imaging maps of SJT and MOD by AFADESI-MSI technique,16 tissue prototype components of SJT for CDAD treatment were screened,including baicalin(wogonin),(E)-8β,17-EPoxylabd-12-ene15,16-dial,gingerol((e)-[4]-shogaol),glycyrrhizic acid,and feruloyltyramine(N-trans-Feruloyltyramine).5)The comprehensive correlation network of "pharmacodynamic index-intestinal flora-blood component-tissue component" showed that the key parameters of SJT for CDAD included baicalin,mycophenol,ginsenoside Rf,ginseng diol,lithophan,hormothamnione,glycyrrhizic acid,CD11 b,LY6g,and LY6 g.CONCLUSION: In this study,we utilized various tools to investigate the therapeutic effects and potential active substance components of SJT in treating CDAD mice.Our comprehensive analysis of phenotypic information,intestinal flora,serum metabolomics,mass spectrometry imaging,and related networks revealed that the SJT histone formula was significantly effective in treating CDAD mice.We successfully identified the pharmacodynamic substance components contained in the histone formula,including baicalein(wogonin),hormothamnione,gingerenol(Shogaol,gingerol,sodium sodium,(e)-7-hydroxy-1,7-bis(4-hydroxy Phenyl)he Pt-5-ene-3s-sulfonate,feruloyltyramine,glycyrrhizic acid,Gmelofuran,and Ginsenoside Rf(Ginsenoside-Rf).These components,attributed to Zingiber officinale Rosc.,Glycyrrhiza uralensis Fisch.,Ziziphus jujuba Mill.,Panax ginseng C.A.Mey.,and Scutellaria baicalensis Georgi.,play an important role in the treatment of CDAD by SJT and may serve as the pharmacological substance basis of SJT.These findings not only provide a reference basis for SJT quality control studies but also offer scientific evidence for the clinical application of SJT.