寻常痤疮皮损的微生物群研究

Acne vulgaris (acne) is amongst the most typical skin diseases in the world and primarily impacts younger people face, with huge disease burden. Propionibacterium acnes is considered to be the pathogen and treatment target of acne. Our prior research discovered that there initially were various microorganisms, for instance bacteria and fungi, in the lesions of acne, indicating that microbial imbalance of skin is closely associated with acne. This project intends to describe the relationship between acne and microorganism scientifically and comprehensively by both qualitative and quantitative methods. With polarized light dermoscopy, ultraviolet light dermoscopy, Garm's staining, and fluorescence staining, the lesions of acne and morphology of microorganisms are observed. With Real-time fluorescence quantitative polymerase chain reaction, the composition of bacteria/fungi is quantitatively analyzed. Moreover, the high-throughput sequencing is utilized to create the metagenomic library, and also the microbial genetic composition and community function are analyzed through the methods of OTU cluster analysis, the community structure statistics, and UniFrac analysis. The project is aimed to solve the following scientific problems: 1) Is acne only the infection of Propionibacterium acnes or the presence of microbial imbalance? 2) Are Staphylococcus spp. and Malassezia spp. or other microorganisms mixed up in the development of acne? 3) Exactly what is the role of Malassezia spp. in various stages of acne? Eventually, we are going to explore new theoretical guidance to the treatment and prevention of acne vulgaris.

寻常痤疮（痤疮）是青年人面部最常见皮肤病，疾病负担重，痤疮丙酸杆菌被认为是致病菌和治疗靶标。我们观察到痤疮皮损内有细菌和真菌等多种微生物，提示皮肤微生态失衡与痤疮发病相关。本课题拟通过形态学定性和分子生物学定量方法全面分析痤疮皮损微生态。用偏振光和紫外光皮肤镜、革兰和荧光染色观察皮损及细菌或真菌形态及数量，组织病理、扫描电镜及透射电镜观察病理改变及各微生物超微结构。提取皮损DNA，用细菌和真菌通用和特异性引物实时荧光定量PCR分析细菌/真菌的总量和各菌种构成；高通量测序构建微生物宏基因组文库，经OTU聚类、群落结构统计、UniFrac等分析皮损所含微生物遗传组成及其群落功能。动态分析治疗后微生态变化。拟解决科学问题：1）痤疮是痤疮丙酸杆菌感染还是存在微生态失衡？2）痤疮皮损中的葡萄球菌和马拉色菌等是否参与痤疮发病？3）马拉色菌在不同阶段痤疮皮损中起何作用？为痤疮防治提供新理论。

寻常痤疮，简称痤疮，是一种常见的慢性炎症性皮肤病，主要累及青少年。痤疮发病机制尚不明确。随着微生物组学的兴起，研究者发现皮肤天然微生物成分的改变引起“生态失调”，可能导致痤疮的发生。目前对痤疮徼生态的研究主要集中在“痤疮患者”与“健康人”的皮肤和肠道菌群组成和差异上。本课题旨在探寻寻常痤疮患者本身皮肤表面非炎症性皮损和炎症性皮损之间的微生物菌群组成及差异，以及主要菌群间相互作用机制，为寻常痤疮以及其他炎症性皮肤疾病的病因及治疗提供新的思路和线索。研究内容包括：寻常痤疮患者非炎症性皮损和炎症性皮损的形态学观察、菌种收集和分离；寻常痤疮患者非炎症性与炎症性皮损内微生物菌群的定性及定量研究；限制性马拉色菌（Malassezia restricta）对表皮葡萄球菌 (Staphylococcus epidermidis)，金黄色葡萄球菌（Staphylococcus aureus）和痤疮丙酸杆菌（Cutibacterium acnes）的生物学特征的影响及作用机制研究。本课题收集临床痤疮患者炎症性和非炎症性两类皮损，发现C. acnes在炎症性皮损中数量低于非炎症性皮损，但在细菌中相对丰度升高；M. restricta在数量上是M. globosa的9.42倍，是痤疮皮损中最多的真菌。两组间S. epidermidis、C. acnes和M. restricta细胞数量的相关性分析显示，C. acnes分别与S. epidermidis和M. restricta呈正相关（p<0.05），说明这三种菌在从非炎症性皮损到炎症性皮损的转化过程中，具有类似的增殖趋势，由于C. acnes目前被认为是导致痤疮发病的重要细菌，提示S. epidermidis和M. restricta在痤疮皮损的微生态平衡中以及痤疮发病机制中可能起到重要作用。M. restricta在体外可能通过上调C. acnes的精氨酸合成通路，抑制生物膜形成，精氨酸也被证明在体外可抑制C. acnes生物膜形成，但具体的作用机制不清。M. restricta在体外可能通过下调S. aureus的群集感应通路，抑制毒素（δ溶血素、肠毒素）产生，抑制生物膜形成。