The Role of the Peritoneum in the Pathogenesis of Endometriosis

项目主任/主要研究者（最后，第一，中间）：Nowak，Romana A. 子宫内膜异位症是一种炎症性疾病，影响10%的妇女，但目前在高达70%的妇女 盆腔疼痛和30%的不孕症患者子宫内膜异位症的估计成本超过220亿美元， 仅US（1），不计算相关慢性疾病（偏头痛、肠易激综合征和 间质性膀胱炎（2，3）。因此，可以肯定地说，子宫内膜异位症是最昂贵的生殖疾病之一， 妇女不仅在治疗方面，而且在受影响患者中丧失了生产力和生活质量。最 子宫内膜异位症的发展被广泛接受的假设是逆行月经，导致 子宫内膜碎片植入内脏或腹膜壁。植入这些碎片 被认为受免疫细胞和子宫内膜细胞局部分泌的因子调节。但 子宫内膜细胞能够穿过腹膜间皮形成病变的机制 以及这些病变如何导致炎症增加和盆腔疼痛尚不清楚。我们的中央 假设腹膜间皮细胞经历上皮-间充质转化（EMT）， 与子宫内膜细胞及其分泌因子或内分泌干扰化学品接触， 邻苯二甲酸盐这导致腹膜完整性的丧失，允许子宫内膜细胞进入子宫内膜。 并诱导促炎细胞因子、趋化因子和其它因子的表达， 外泌体/微泡中的间皮细胞分泌。本研究提案的目标是开始测试 该假设使用子宫内膜异位症的体内小鼠模型和体外人间皮细胞培养物 系统，以研究腹膜间皮在促进肿瘤病变发生中的作用 以及内分泌干扰物邻苯二甲酸酯如何影响腹膜间皮瘤并促进 局部炎症环境。我们将在以下两个具体目标中检验这些假设： 1)表征经历EMT并丧失屏障的人间皮细胞的分泌组 在体外模型系统中的完整性。我们将进行小RNA测序和蛋白质组学分析， 在体外培养中经历EMT的原代人间皮细胞脱落的外来体/微泡 系统我们将测量跨上皮电阻的变化，以评估EMT期间屏障完整性的丧失。 2)利用体内小鼠模型，研究EMT在间皮中的过程， 邻苯二甲酸酯对间皮细胞EMT的影响。我们将评估 已知的EMT和炎性标志物在早期和建立的增生性病变中的作用 不同剂量的邻苯二甲酸盐这些研究的成功完成有望开辟一个全新的 子宫内膜异位症研究和内分泌干扰物领域的方向，为未来的研究奠定基础。 研究重点是炎症和EMT的基本机制，以及 内分泌干扰物可能导致慢性炎症。 PHS 398/2590（Rev.06/09）

Program Director/Principal Investigator (Last, First, Middle): Nowak, Romana A. Endometriosis is an inflammatory disease that affects 10% of women but is present in up to 70% of women with pelvic pain and 30% of women with infertility. The estimated cost of endometriosis is over $22 billion in the US alone (1), not counting associated chronic illnesses (migraine headache, irritable bowel syndrome and interstitial cystitis (2, 3). Thus it is safe to say that endometriosis is one of the costliest reproductive diseases in women not only in terms of treatment but also lost productivity and quality of life in affected patients. The most widely accepted hypothesis for development of endometriosis is retrograde menstruation that results in implantation of endometrial fragments on the visceral or peritoneal walls. Implantation of these fragments is thought to be regulated by factors secreted locally by immune cells and endometrial cells. However, the mechanisms by which endometrial cells are able to traverse the peritoneal mesothelium to establish lesions and how these lesions lead to increased inflammation and pelvic pain are not well understood. Our central hypothesis is that peritoneal mesothelial cells undergo epithelial-mesenchymal transition (EMT) in response to contact with endometrial cells and their secreted factors or with endocrine disrupting chemicals such as phthalates. This leads to a loss of peritoneal integrity allowing passage of endometrial cells into the peritoneum and induces expression of proinflammatory cytokines, chemokines and other factors that are secreted by mesothelial cells in exosomes/ microvesicles. The goal of this research proposal is to begin testing this hypothesis using an in vivo mouse model of endometriosis and an in vitro human mesothelial cell culture system to investigate the role of the peritoneal mesothelium in promoting the initiation of endometriotic lesions and how endocrine disrupters, phthalate, may affect peritoneal mesothelium and promote the development of a local inflammatory environment. We will test these hypotheses in the following two specific aims: 1) Characterize the secretome of human mesothelial cells undergoing EMT with loss of barrier integrity in an in vitro model system. We will perform small RNA sequencing and proteomic analysis of exosomes/microvesicles shed by primary human mesothelial cells undergoing EMT in an in vitro culture system. We will measure changes in trans-epithelial resistance to assess loss of barrier integrity during EMT. 2) Utilize an in vivo mouse model to investigate the process of EMT in mesothelium at sites of endometriotic lesions and the effects of phthalates on mesothelial cell EMT. We will assess changes in known EMT and inflammatory markers in early and established endometriotic lesions in mice treated with different doses of phthalates. Successful completion of these studies is expected to open up an entirely new direction in the field of endometriosis research and endocrine disrupters, providing a foundation for future studies focusing on basic mechanisms involved in inflammation and EMT and the mechanisms by which endocrine disrupters may contribute to chronic inflammation. PHS 398/2590 (Rev. 06/09) Page Continuation Format Page