| Purpose:To evaluate the clinical efficacy of optimized kuijie prescription in the treatment of ulcerative colitis(UC)with dampness-heat syndrome of large intestine.Metabolomics and immunohistochemistry techniques were used to study the blood components and anti-inflammatory targets of optimized kuijie prescription for the treatment of UC patients with dampness-heat syndrome of large intestine,so as to reveal the pharmacodynamic and anti-inflammatory mechanisms of optimized kuijie prescription for the treatment of UC patients with dampness-heat syndrome of large intestine.Material and method:1.From March 2021 to September 2021,UC patients with dampness and heat of large intestine who were admitted to the outpatient department and inpatient department of Anorectal Department of Hulun Buir Zhong Meng Hospital were collected.Sixty patients were randomly divided into treatment group(optimized kuijie prescription)and control group(sulfasalazine enteric-coated tablets).Respectively the baseline data of two groups before and after drug intervention(age,gender,BMI,course of diseases,disease severity,disease classification),clinical curative effect and syndrome curative effect of traditional Chinese medicine,Chinese medicine syndrome integral,improved the Mayo score,under endoscopic score,colonoscopy mucosal healing score,quality of life index(IBDQ score),Laboratory physical and chemical index(Serum CRP,Esr),safety and adverse reactions were observed,compared and analyzed,to evaluate the clinical effect of optimized Kuijie prescription on UC of damp-heat Syndrome of large intestine.2.In the UHK group,10 UC patients were randomly selected,and in the UHK group,6patients were randomly selected to be treated with the optimized Kuijie prescription and the intervention of MY group(dosing after 6 hours)and M66 group(dosing after 12 hours),at the same time 10 randomly selected from the same period of clinic health check-up NK group(Natural Control Group)and so on four groups of serum sample under test,the metabolites in the samples were detected,screened and identified by non-targeted metabonomics technology,and matched with the material information in the database,in order to understand the differential metabolites of optimized Kuijie prescription in the treatment of UC with damp-heat Syndrome of large intestine,the possible metabolic pathway of optimized Kuijie prescription in the treatment of UC with damp-heat syndrome of large intestine was explored through analysis,identification and screening.3.Four groups of samples(UHK group,MY group,M66 group and NK group)were detected by target metabonomics,the metabolic products of 347 common intestinal flora(including amino acids,fatty acids,bile acids,sugars,benzenoids and nucleotides)were identified and Quantitative analysis.By evaluating the quality of experimental data,multidimensional statistical analysis,differential metabolite screening,KEGG pathway analysis and other data analysis,the selective and specific detection and verification of the possible target metabolites of blood components found in the non-targeted metabolomics were carried out.4.The expression of Il-17 a,NF-KB,Ikba,CCL2 and CXCL8 in intestinal mucosa biopsies of UC patients with damp-heat syndrome of large intestine before and after intervention were observed by immunohistochemical technique,therefore,the anti-inflammatory mechanism of optimized Kuijie prescription in treating UC of damp-heat syndrome of large intestine is expounded.Results:1.A total of 58 patients finally completed the study(treatment group 30 cases,28 cases in the control group,2 cases,because during treatment medication treatment,not according to stipulations poor adherence to withdraw from studies),the two groups before treatment baseline data(age,gender,BMI,course of the disease,the disease severity,disease classification)was no significant difference(P>0.05).After drug intervention,the total effective rate of the treatment group(90.0%)was better than that of the control group(78.6%),and there was no significant difference between the two groups(P>0.05).After treatment,the total effective rate of the treatment group(90.0%)was better than that of the control group(67.9%),the difference was statistically significant(P<0.05).TCM syndrome integral,the two groups before treatment improved the Mayo grade,under endoscopic score,colonoscopy mucosal healing and IBDQ score no statistical difference(P>0.05),treatment group after treatment in improving the TCM syndrome integral,improved the Mayo grade,under endoscopic score,colonoscopy mucosal healing and IBDQ score are better than that in control group,There were statistically significant differences(P<0.05),and there were statistically significant differences(P<0.05)between the two groups in improved TCM syndrome score,improved Mayo Mayo score,endoscopic score,mucosal healing score under colonoscopy and IBDQ score before and after treatment,which were all improved compared with before treatment.Before treatment,there was no significant difference in the laboratory physical and chemical indexes(serum CRP and ESR)between the two groups(P>0.05).After treatment,the treatment group was better than the control group in improving the expression changes of CRP and ESR(P<0.05).There was one mild rash in the control group,and no local and systemic adverse reactions occurred in the treatment group.2.A total of 891 metabolites were identified by non-targeted metabonomics techniques in both positive and negative ion modes,all metabolites in M66 vs MY group,M66 vs NK group,MY group vs NK group,UHK group vs M66 group were analyzed by univariate and multivariate statistical analysis.In order to explore and identify the potential biomarkers of optimized Kuijie prescription for the treatment of UC with damp-heat syndrome of large intestine,14 potential biomarkers including ARA,DHA,OA,LA and PA related to the therapeutic effect of optimized Kuijie prescription were determined by comparing the M66 group and the NK group.Twenty-five metabolic pathways were screened by KEGG pathway enrichment analysis,mainly affected the lipid synthesis and metabolism of relevant unsaturated fatty acid biosynthesis pathway,linoleic acid metabolic pathways,and fatty acid biosynthesis pathway,shows that optimized Kuijie prescription can regulate the metabolic disorder of UC patients,especially by regulating lipid synthesis and metabolism in the treatment of UC.3.Using targeted metabolomics detection technology,a total of 216 metabolites were identified in positive and negative ion switching detection mode by targeting 347 metabolites.All metabolites detected in the M66 group and the NK group were analyzed for inter-group differences.A total of 21 metabolites with significant differences were screened,including Aminoadipic acid,L-glutamic acid,L-isoleucine,Pyroglutamic acid,Palmitoleic acid and Glycodeoxycholic acid.Thirty-four metabolic pathways were screened by KEGG pathway enrichment analysis,which mainly affected 2-oxocarboxylic acid metabolism and Biosynthesis of Amino acids,etc.These results indicate that optimized Kuijie prescription can restore the intestinal mucosal barrier function and improve the intestinal inflammatory state of UC patients by affecting amino acid metabolism and synthesis.4.After drug intervention in the treatment group and the control group,immunohistochemical detection of intestinal mucosal biopsy tissues of patients in the two groups showed a statistically significant difference in the staining number of IL-17 A,NF-κB,Ik Ba,CCL2 and CXCL8 positive cells(P<0.05),and the treatment group was superior to the control group.Conclusion:1.This research through the untargeted metabolomics and targeted metabolomic analysis,found that the optimized Kuijie prescription through regulating lipid synthesis and metabolic pathways and amino acid metabolism synthesis pathways such as 2-oxocarboxylic acid metabolism,biosynthesis of amino acids,impact on metabolic product,thus promote the intestinal mucosa repair,and improve the intestinal inflammation.2.The expression of pro-inflammatory cytokines IL-17 A,transcription factor NF-κB and inflammatory chemokine CCL2 and CXCL8 were inhibited,and the expression of inhibitory protein Ik Ba was increased by optimized Kuijie prescription in treating UC with damp-heat syndrome of large intestine,suggesting that the anti-inflammatory mechanism might be related to the inhibition of IL-17 signaling pathway.3.Through clinical research,we found that optimized Kuijie prescription can relieve clinical symptoms,promote mucosal healing,improve patients’ quality of life and reduce the occurrence of adverse reactions.4.The study further found that optimized Kuijie prescription has a bidirectional anti-inflammatory effect by regulating the overexpression of related metabolites generated by arachidonic acid metabolism pathway related to linoleic acid metabolism and inhibiting IL-17 signaling pathway,thus producing therapeutic effects for UC patients with large intestine dampness and heat syndrome. |
