The Imbalance Of Gut Microbiota And Its Correlation With Plasma Inflammatory Cytokines In Pemphigus Vulgaris Patients

Objective:The purpose of this study was to analyze the gut microbiota of pemphigus vulgaris patients and healthy control group,assessing potential correlation with circulating inflammatory cytokines,and preliminarily explore the possible mechanism of pemphigus.Methods:Peripheral blood and fecal samples were obtained from 18 pemphigus vulgaris patients and 14 age-and gender-matched healthy controls.The Thl7/Th1/Th2 cells related cytokines in serum which extracted from peripheral blood were detected by Luminex 200TM System,then analyzed the different levels of serum cytokines between the two groups.Microbial DNA was extracted and purified from fecal sample by the QIAamp DNA stool minikit,the total DNA was tested.The sequencing of V3-V4 16S rDNA gene region were performed by the Kit Ion 16sTM Metagenomics Kit and SimpliAmp thermal cycler platform.DNA Library was builded by Ion Plus fragment Library kit.The purified and amplified 16SrDNA was sequenced by Illumina HiSeq PE250 platform.The diversity of species inpemphigus vulgaris(PV)patients and healthy controls was analyzed by Alpha(a)diversity.The differences of gut microbiota between pemphigus vulgaris and healthey groups were analyzed by the analysis of beta(?)-diversity,Principle coordinates analysis(PCoA),MRPP analysis and the analysis of the Heatmap clustering.The different microbes between pemphigus vulgaris and healthey samples was identifyied by LefSe analysis,and Statistics analysis of gut microbial differences between pemphigus and control samples was performed by Wilcox.test and Kruskal.test.The functional annotation of the differential gene was analysis by the Kyoto Encyclope-d ia of Genes and Gemos(KEGG)bioinformatics database,and the genetic function was predicted by the PICRUSt and STAMP software platform.Correlation between serum cytokines and different microbial taxa was assessed by Person correlation test.Results:The study showed that there was no difference between patients and healthy controls with respect to gut microbiota diversity,as assessed by a-diversity.The analysis of?-diversity,PCoA,MRPP analysis and the analysis of the Heatmap clustering showed that There were exsited differences of gut microbiota between pemphigus vulgaris and healthey groups.LefSe analysis found that there were 19 different bacteria between the two groups.The non-parametric test verified the above 19 difference bacteria,and 14 taxa were found to be significantly different between the two groups.At the genera level,Lachnospiracea incertae sedis and Coprococcus were decreased,while Granulicatella,Flavonifractor was increased in PV.Based on the function analysis of KEGG,we found 10 and 35 differently abundant pathways at 2 and 3 levels.Serum levels of C5a,interleukin(IL)-2R,IL-6,IL-8,IL-7,IL-1?,IL17A,IL-5and IL-21 were significantly increased in PV.Flavonifractor exhibited a positive correlation with C5a,IL-6,IL-8,IL-7,IL-1?,IL17AandIL-21.Lachnospiracea_incertae sedis and Coprococcus showed a negative correlation with IL-17A.Conclusion:PV patients have gut microbial dysbiosis which was significantly associated with Thl/Th2/Th17 related cytokines.It indicated that gut microbiota might contribute to the immune disorder and the development of PV.