基于16S rRNA测序探讨特应性皮炎湿证特征与皮肤菌群的相关性  

Research on the correlation between the characteristics of dampness syndrome and skin microflora in atopic dermatitis patients based on 16S rRNA sequencing method

作　　者：林颖 刘颖瑶[3] 张彩云 郑敏玲 罗晓晴 董雷 刘俊峰 屈平华 LIN Ying;LIU Yingyao;ZHANG Caiyun;ZHENG Minling;LUO Xiaoqing;DONG Lei;LIU Junfeng;QU Pinghua(Guangdong Provincial Key Laboratory of Chinese Medicine for Prevention and Treatment of Refractory Chronic Diseases Guangzhou 510120,China;The Second Affiliated Hospital of Guangzhou University of Chinese Medicine(Guangdong Provincial Hospital of Chinese Medicine),Guangzhou 510120,China;The Second Clinical Medical College of Guangzhou University of Chinese Medicine,Guangzhou 510120,China;Guangdong Women and Children Hospital,Guagnzhou 510010,China;School of Life Sciences,Sun Yat-Sen University,Guangzhou 510275,China;Guangdong-Hong Kong-Macao Joint Lab on Chinese Medicine and Immune Disease Research,Guangzhou 510120,China)

机构地区：[1]广东省中医药防治难治性慢病重点实验室,广州510120 [2]广州中医药大学第二附属医院(广东省中医院),广州510120 [3]广州中医药大学第二临床医学院,广州510120 [4]广东省妇幼保健院,广州510010 [5]中山大学生命科学学院,广州510275 [6]粤港澳中医药与免疫疾病研究联合实验室,广州510120

出　　处：《中华中医药杂志》2023年第11期5504-5509,共6页China Journal of Traditional Chinese Medicine and Pharmacy

基　　金：国家自然科学基金面上项目(No.31972856);广东省中医药防治难治性慢病重点实验室(No.MB2020KF05);广东省中医皮肤临床医学研究中心及广东省中医皮肤病研究所专项课题(No.PFK2021-02);广东省中医院拔尖人才项目(No.BJ2022YL08)。

摘　　要：目的:探讨特应性皮炎(AD)湿证特征与皮肤菌群的相关性,为提高中医辨证治疗的疗效和探索AD治疗的新策略、新方法提供理论依据。方法:纳入广东省中医院大学城医院皮肤科就诊的24例AD患者以及33名健康人进行肘部的皮肤样本采集。提取样本DNA,对细菌16S rRNA基因V4区进行扩增,Illumina Miseq2平台进行序列测定。Qiime 2(v2020.2.0)软件按照99%的相似水平对测序序列进行OTU聚类分析和比对注释、LefSe(LDA Effect Size)分析和Metastats差异分析,并采用统计软件Stata 13.1分析样本在属水平上的差异。按《中医湿证评估量表》对所有湿证条目进行评分,剔除2名湿证信息不全或模糊的健康人,分为湿证和非湿证,分析AD的湿证特征以及湿证与非湿证的皮肤微生态结构差异。结果:AD组湿证9例,非湿证15例,健康组湿证4名,非湿证27名,AD组湿证比例显著大于健康组(P=0.013)。AD患者出现湿证条目频率最高的是倦怠思睡和舌苔厚腻,其次为面色秽浊如有污垢、面部/头发油腻、口黏腻、渴不欲饮水。AD湿证组的显著物种是葡萄球菌属;AD非湿证组的显著物种是双歧杆菌属和甲基杆菌属;健康湿证组的显著物种是金黄杆菌属;健康非湿证组的显著物种是不动杆菌属、短波单胞菌属等。湿证和非湿证组间有显著差异的优势菌属为棒状杆菌属(P=0.038)、不动杆菌属(P=0.037)、金黄杆菌属(P=0.013)。AD湿证组和AD非湿证组差异较明显的优势菌属为不动杆菌属(P=0.095)、短波单胞菌属(P=0.094),但差异无统计学意义。AD非湿证组和健康非湿证组组间差异较明显的优势菌属为短波单胞菌属(P=0.00048)、莫拉菌属(P=0.022)、副球菌属(P=0.0013)、皮肤杆菌属(P=0.015)、葡萄球菌属(P=0.0000042)。结论:湿证是AD发病以及病情加重的主要证候,皮肤菌群的改变可能作为湿证症状的客观评价指标以及中药治疗的靶点,值得关注和进一步深�Objective:To explore the correlation between the characteristics of dampness syndrome and skin microflora in patients with atopic dermatitis(AD),so as to provide theoretical basis for improving the curative effect of traditional Chinese medicine syndrome differentiation treatment and exploring new strategies and methods for AD treatment.Methods:We collected skin samples from elbow fosses of 57 participants,including 33 healthy volunteers,as well as 24 AD patients,who were receiving treatment in the Department of Dermatology,Guangdong Provincial Hospital of Chinese Medicine,Guangzhou Higher Education Mega Center Hospital.We extracted total genomic DNAs of all samples,in which the V4 region of the bacterial 16S rRNA gene was PCR-amplified and sequenced on the Illumina MiSeq 2 platform.With the Qime 2(v2020.2.0)software,we performed multiple analyses on the sequence at the similarity level of 99%,such as OTU cluster analysis and comparison annotation,LefSe(LDA Effect Size)analysis and Metastats difference analysis.Besides,we performed a difference analysis on the samples at genus level,using the statistical software Stata 13.1.All dampness syndrome entries were scored according to TCM Dampness Syndrome Evaluation Scale.Two cases from the healthy control group were excluded for their syndrome information being incomplete or vague.The remaining samples in each group were then divided into two sub-groups,dampness syndrome group(D group)and non-dampness syndrome group(ND group),to analyze characteristics of dampness syndrome in adolescent and adult patients with AD,as well as the skin microecological structural differences between D group and ND group.Results:In patients with AD,there were 9 samples in D group and 15 in ND group.In healthy volunteers,the numbers were 4 and 27.The proportion of samples with dampness syndrome in AD group was obviously greater than that in healthy control group(P=0.013).The most frequent entries of dampness syndrome in AD patients were fatigue and sleepiness,thick and greasy tongue coating

关 键 词：特应性皮炎 湿证 皮肤菌群 16S rRNA测序 

分 类 号：R275.9[医药卫生—中西医结合]