生命早期三氯生暴露对婴幼儿肠道菌群及肥胖的影响

Obesity epidemic has become an important public health problem. Recent studies suggest that gut microbiota is closely related to the development of obesity. Triclosan (TCS) is a broad-spectrum antimicrobial chemical widely used in personal care products including toothpaste, thus the exposure to TCS is common at population level. Animal experiment shows that TCS exposure changes intestinal microbial composition. Early life is a critical period for gut microbial colonization and development. It is unknown whether exposure to TCS during early life affects gut microbiota and the risk of obesity. Our pilot study suggested that prenatal TCS exposure was associated with increased abdominal adiposity in children. This is the first study to evaluate whether prenatal exposure to TCS alters maternal vaginal microbial composition, interferes with neonatal gut microbial colonization and development, and leads to childhood obesity. This proposal is built on our prospective birth cohort study in Shanghai. In this project, a nested case-control study will be conducted among 150 obese and 150 normal-weight children born by vaginal delivery to test our hypothesis. We will examine the associations between maternal urinary TCS concentration and composition of vaginal microbiota, and analyze the sequencing of 16S rDNA of microbiota in meconium and children’s fecal samples at 6 weeks, 6 months, 12 months, and 3 years after births. The exposure markers of certain gut microbiome will also be measured. We will analyze the development of the intestinal microbiota in children, and its association with childhood obesity. Our study will provide insight on early prevention of obesity, and new clues to conquering epidemic of obesity.

肥胖流行是重要的公共卫生问题。近年研究提示，肥胖的发生可能与肠道菌群相关。三氯生（TCS）是广谱抗菌剂，添加于个人护理产品(牙膏等)，人群暴露广泛。动物实验显示，TCS可影响肠道菌群。生命早期是肠道菌群定植和形成的关键时期，但生命早期TCS暴露是否干扰肠道菌群形成、增加肥胖的风险，国内外尚无研究。我们的预实验结果提示，孕期TCS暴露与幼儿脂肪增多有关。本项目将首次研究围产期TCS暴露是否影响阴道菌群，干扰婴幼儿肠道菌群定植和形成，增加儿童肥胖发病风险。本项目依托上海出生队列平台，采用巢式病例对照研究（150例肥胖和150例正常体重3岁儿童，均为自然分娩），研究孕期尿TCS水平与阴道菌群组成和丰度的关系；对子代不同时间（胎粪、婴幼儿6周，6、12个月及3岁）粪便肠道菌群细菌 16S rDNA测序分析、检测相关菌群标记，分析与儿童肥胖的关系。研究结果可为生命早期预防肥胖、遏制其流行提供新线索。

三氯生（TCS）是人工合成的广谱抗菌剂。TCS因其广谱抗菌作用而被广泛添加于超过两千多种的日常生活用品，使用后，TCS随之经污水被排放进自然界水生态系统，可造成环境的污染。我们分析了620例孕妇尿样三氯生浓度，TCS可在97.9%的孕妇尿样中检出。生命早期是肠道微生物发育的关键时期，甚至在宫内就可能早期暴露于胎盘的少量菌群。对足月妊娠、无临床感染指征的剖宫产产妇胎盘和脐带血样本，我们应用Illumina MiSeq测序系统通过微生物16S rRNA基因V3-V4高变区测序分析，发现微生物可在所有胎盘标本中检出。在门水平，胎盘的微生物群主要由4个门（拟杆菌门、厚壁菌门、放线杆菌门和变形杆菌门）和10个优势菌属组成。妊娠期糖尿病孕妇的胎盘微生物菌群多样性更高。在检出菌群的部分脐带血样中，菌群构成在门和属水平上与其对应的胎盘菌群相似，提示胎盘菌群可能是其潜在的来源。我们采集了300例孕妇孕晚期尿样和阴道分娩的新生儿胎皮脂，对孕妇的尿样检测包括TCS在内的酚类环境污染物。采用16S rRNA基因测序对新生儿胎皮脂标本进行菌群的检测和分析，探讨孕妇尿TCS浓度与阴道分泌物菌群及新生儿皮肤上胎皮脂菌群的关系。采用宏基因组测序对胎粪、42天、3个月、6个月婴儿粪便进行菌群分析。分析母亲孕期TCS暴露对子代早期菌群及儿童肥胖和代谢健康的影响。我们通过斑马鱼染毒试验，探究TCS暴露对消化道菌群的影响。TCS暴露浓度分别设定为0.03、0.3、3、30、100、300 ng/mL，以0.01% 的二甲基亚砜(DMSO)作为溶剂对照组。TCS长期暴露实验，从斑马鱼受精卵孵化为幼鱼开始暴露至斑马鱼120天；短期暴露实验为4月龄斑马鱼TCS暴露7天。通过微生物16S rRNA基因V3-V4高变区测序分析检测斑马鱼消化道微生物。结果发现：斑马鱼从幼鱼到成年期，长期暴露于高TCS浓度，可能会减少其消化道微生物的多样性，改变消化道微生物群的组成。TCS短期暴露对斑马鱼消化道菌群的多样性未见影响，但在TCS暴露组，微生物组成存在轻微改变。此外，我们也在研究中发现，孕妇尿TCS浓度高与GDM患病风险增高相关，并与女婴出生体重正相关；孕期TCS暴露也可影响孕妇和新生儿甲状腺激素水平，内分泌干扰作用有待进一步研究。