基于肠道菌群调控宿主LPS/TLR4信号通路探讨归术益坤方治疗多囊卵巢综合征的机制

Polycystic ovary syndrome (PCOS) is one of the most commonly endocrine disorders in gynecology. It negatively impacts women’s health on both physical and psychological level. It is found that gut microbiota is closely related to PCOS and dysfunction of gut microbiota could disturb LPS/TLR4 signal pathway, which is known to associated with inflammation. Our previous PCOS research in rat model suggested that dysfunction of gut microbiota was triggered by letrozole. Meanwhile, Gui Zhu Yi Kun formula (GZYKF) showed reduced level of proinflammatory factors in PCOS rats. Therefore, we propose a hypothesis that the effects of GZYKF on PCOS pathological model are through improving gut microbiota function and inhibiting the LPS/TLR4 signaling. In this research PCOS rat model will be established and treated with GZYKF. We will explore the diversity and gene functions of gut microbiota using Metagenome Sequencing and 16S rRNA gene-sequencing; evaluate the key proteins and genes of LPS/TLR4 signal pathway using ELISA, Western-blot and Real-time PCR. Further reveal the effects of GZYKF on gut microbiota and LPS/TLR4 signal pathway and find potential targets of treating PCOS by GZYKF.

多囊卵巢综合征（PCOS）是最常见的生殖内分泌疾病，严重威胁女性身心健康。肠道菌群与PCOS密切相关。肠道菌群失调，可上调炎症相关LPS/TLR4通路。我们前期工作证实来曲唑诱导的PCOS大鼠模型存在肠道菌群失调的现象；且归术益坤方能降低PCOS炎症因子水平。由此我们提出假说：归术益坤方治疗PCOS是通过改善肠道菌群功能，下调LPS/TLR4信号通路实现的。本课题拟复制PCOS大鼠模型，以归术益坤方进行干预；采用宏基因组测序、16S rRNA测序技术，进行肠道菌群多样性分析和基因功能分析；采用ELISA、Western-blot、Real-time PCR法检测卵巢组织中LPS/TLR4通路关键蛋白及基因的表达；研究归术益坤方对肠道菌群及其介导的LPS/TLR4通路的影响；从而探讨归术益坤方治疗PCOS的作用靶点，揭示其治疗PCOS的潜在机制，为归术益坤方的进一步研发和临床推广奠定基础。

多囊卵巢综合征（PCOS）是严重影响女性身心健康的常见的生殖内分泌疾病。本研究通过来曲唑诱导PCOS大鼠模型，以归术益坤方、菌群移植、归术益坤方联合菌群移植及二甲双胍进行干预，检测大鼠体重、观察大鼠动情周期，检测血清中雄激素水平、空腹血糖、空腹胰岛素水平，观察大鼠卵巢形态改变; 通过ELISA检测大鼠血清中LPS、IL-6、IL-18、TNF-α水平，Western-blot、Real-time PCR法检测卵巢组织中LPS/TLR4通路关键蛋白、基因的表达。采用16s rRNA、宏基因测序技术，对大鼠肠道菌群多样性及和基因功能;结果表明归术益坤方可降低PCOS模型大鼠体重、血清睾酮、空腹血糖、调节大鼠动情周期，改善卵巢多囊性改变；降低血清LPS、TNF-α、IL-18水平，下调LPS、TLR4、NF-kB表达。增加PCOS大鼠肠道菌群的α多样性，降低厚壁菌门相对丰度，提高拟杆菌、乳杆菌和短链脂肪酸产生菌相对丰度；基因功能分析发现归术益坤方可能促进碳水化合物和蛋白质代谢。初步揭示了归术益坤方治疗PCOS的潜在机制可能是通过改善PCOS大鼠肠道菌群、下调肠道菌群介导的LPS/TLR4信号通路，降低相关促炎因子表达，改善PCOS大鼠慢性炎症状态，降低PCOS大鼠雄激素水平，进而改善PCOS卵巢多卵泡状态来实现的。为归术益坤方的进一步研发和临床推广奠定基础，同时也为其他代谢相关疾病的防治提供新的研究思路。