化瘀解毒方重建肠道菌群改善子宫内膜异位症炎性微环境机制研究

Endometriosis is an estrogen-dependent inflammatory disorder characterized by the.presence of endometrial tissue outside the uterine cavity. Inflammation is an.important characteristic of endometriosis. The study found that Lipopolysaccharide levels in the peritoneal fluid of patients with endometriosis increased significantly, and it induced the inflammatory cascade reaction through the TLR4 / NF-kappa B signal path. We hypothesized that imbalance of intestinal flora of patientswith endometriosis leaded to Lipopolysaccharide translocation to the peritonealfluid, which resulted in inflammation. Blood stasis is the key link in the process of EM development, and Huayu Jiedu decoction is an effective prescription for treating EM. Our previous study showed that the intestinal flora of EM model mice treated by Huayu Jiedu decoction was more similar to that of normal mice. This project intends to explore the characteristics of intestinal flora, metabolite, and the inflammatory microenvironment in peritoneal fluid of EM model mice adopting the 16S high throughput sequencing, nuclear magnetic resonance technology, inflammatory cytokines chip etc. This project are still to clarify the effect of Huayu Jiedu decoction on the intestinal flora, metabolic, and the inflammatory microenvironment. We would illuminate biological basis of Huayu Jiedu decoction for the treatment of EM from the perspective of intestinal flora reconstruction, and lay the solid foundation for its wide application in clinic.

子宫内膜异位症（EM）是子宫内膜腺体和间质种植于子宫以外的疾病，炎症是EM的重要特征。研究发现EM患者腹腔液中内毒素脂多糖（LPS）水平显著升高，且通过TLR4/NF-κB产生炎症级联反应。我们提出假说：EM患者可能存在肠道菌群稳态失衡，出现LPS移位至腹腔液，诱发炎症的产生。血瘀是贯穿EM发生发展过程中的中心环节，化瘀解毒方是临床治疗EM的有效方剂。前期实验研究提示经过化瘀解毒方灌胃处理的EM模型小鼠肠道菌群发生改变，与正常小鼠更为相近。因此，本项目拟建立动物模型，采用16s高通量测序、核磁共振技术、炎性因子芯片等方法探讨EM肠道菌群、代谢组特征及腹腔炎性微环境特点；明确化瘀解毒方对EM肠道菌群、代谢组，以及外周血、腹腔液炎症的干预作用。从肠道菌群重建角度进一步阐明EM炎性微环境形成的机制，揭示化瘀解毒方治疗EM的生物学基础，为其在临床上的广泛应用奠定坚实的基础。

子宫内膜异位症（EM）是雌激素依赖的炎性疾病。炎症是EM的重要特征。肠道菌群失调可致肠道黏膜屏障破坏，诱发炎症反应。研究发现EM患者腹腔液中内毒素脂多糖（LPS）水平显著升高。血瘀是贯穿EM发生发展过程中的中心环节，化瘀解毒方（HYJDF）是临床治疗EM的有效方剂。本研究通过建立动物模型，研究分析EM小鼠模型肠道菌群、粪便代谢组结构特征；收集临床样本，检测子宫内膜异位症患者肠道菌群、外周血炎性因子。进一步对EM模型小鼠进行化瘀解毒方及差异代谢产物α-亚麻酸(ALA)干预，明确对EM模型小鼠肠道菌群、粪便代谢组及炎性表征的作用及对腹腔液巨噬细胞、肠壁屏障等的影响。结果表明：（1）EM模型小鼠肠道菌群的多样性和丰度降低。本研究筛选出156个有命名的差异代谢物，发现次生胆汁酸生物合成、α-亚麻酸代谢通路在内异症小鼠肠道菌群和代谢物交互过程中扮演着重要作用。这2条通路涉及4种重要差异代谢物，包括ALA。（2）EM患者与健康女性血清IL-17A与IL-8浓度水平存在显著性差异，两组受试者肠道菌群丰度在门和属水平存在差异，且内异症组患者肠道属水平菌群丰度与血清激素及炎性因子之间存在相关性。（3）经HYJDF治疗后，内异症小鼠肠道内多种重要差异代谢物和菌属水平均向正常对照组水平改善；腹腔LPS含量显著降低，巨噬细胞表型和吞噬功能改善；内异灶纤维化水平、TLR4及下游相关蛋白表达降低。（4）ALA能够改善内异症小鼠肠道菌群及肠道菌群功能富集通路，改善内异症小鼠肠壁屏障和腹腔免疫微环境。本研究从肠道菌群角度阐明了EM腹腔炎性微环境形成的机制，说明肠道微生物的改变有可能影响子宫内膜异位症的发生发展。明确了化瘀解毒方治疗EM在重建肠道菌群、改善炎性微环境的作用，从肠道菌群重建角度阐明化瘀解毒方治疗EM的生物学基础，为中医药防治EM奠定坚实理论基础。