特络细胞调控巨噬细胞免疫活性在子宫内膜异位症发生机制中的作用研究

Telocytes (TCs), a newly discovered peculiar type of stromal cells, have been identified in many organs, including uterus and oviduct tissues, with proposed multiple potential bio-functions, including intercellular signalling, immunoregulation/ immunosurveillance, and etc,. It is necessary to clarify the pathophysiological roles of TCs, either in normal or disease-affected tissues. Previously, we have demonstrated that in rat model of endometriosis (EMs), TCs connected to multiple activated immunocytes and developed direct intercellular contacts, just mimic the structure of immunological synapse (IS). Furthermore, in exposure to TCs conditional media (TCM) for 48 h in vitro, SD rat peritoneal macrophages demonstrated with activation and stable high expression of IL-6, with unknown mechanism and biological functions. However, we speculated that, TCs regulate macrophage function probably through direct IS or through paracrine effects indirectly, and subsequently might play an important role in development of EMs. To verify this hypothesis. We design to investigate modulation effects of TCs on immunological functions of macrophage, and subsequent effects on biological behavior of endometrial stromal cell (ESC), and intend to seek potential pathway and key proteins/ molecules under this process, by means of in vitro cells co-culture. And then followed with in vivo experiments in EMs rat model to examine the above results. Aim to elucidate regulation effects of TCs on macrophage and ESC, unveil the underlying pathway and corresponding key proteins. This application will give an excellent insight into the function of TCs in immunoregulation/ immunosurveillance and roles in development of EMs.

特络细胞（telocytes，TCs）是一种新型的间质细胞，其功能是目前的研究难点，推测与信号传导、免疫调节等有关。我们前期首次发现，内异症（EMs）病灶内TCs与活化的诸多免疫细胞间形成免疫突触（IS）样结构；且离体TCs活化巨噬细胞（MΦ）后稳定高表达IL-6，但机制不清。故推测TCs对MΦ的直接／间接免疫调节效应参与EMs的免疫发病机制。课题拟在分子、细胞和SD大鼠EMs模型层面，以TCs对MΦ的免疫调节为切入点，首先建立体外细胞共培养平台，深入研究TCs与MΦ建立的IS样结构，和TCs的旁分泌效应，对MΦ免疫活性的直接／间接调节效应；以及TCs处理后的MΦ，对离体内膜间质细胞（ESC）增殖等行为的调节效应；最后在模型层面予以验证。旨在解析TCs调节MΦ免疫活性、影响ESC增殖／凋亡的关键通路与分子。本申请将为EMs的研究引入TCs与免疫调节相结合的新内容，亦为EMs防治提供新靶点。

背景：子宫内膜异位症（EMs），是一种以子宫内膜出现在宫腔外为特征的妇科疾病，现有经典理论如月经逆行等均不足以完全解释EMs的发生和发展。一直以来，对EMs发病机制的研究，主要集中在：子宫内膜腺上皮、子宫内膜间质细胞（ESCs），以及腹腔液中以巨噬细胞（pMACs）为主的诸多免疫细胞，并认为靶向或选择性的调节pMACs活化与功能，是防治EMs的一个方向。特络细胞（telocytes，TCs）是一种新型的间质细胞，其功能是目前的研究难点，推测与信号传导、免疫调节等有关。目前EMs中TCs对pMACs的直接与间接调控作用，关键信号通路与分子，以及对下游ESCs的影响均是未知领域。本研究则是基于这样的构想，深入探索TCs在EMs中的机制。..主要研究内容：通过小鼠原代子宫TCs与腹腔pMACs的直接与间接共培养，观察到通过线粒体途径，对pMACs产生了明显的活化作用，包括pMACs的形态学改变，细胞活力与侵袭能力的显著增强，以及免疫相关细胞因子的分泌，pMACs的凋亡抑制；TCs处理离体巨噬细胞系（RAW264.7），以及EMs模型中的在体pMACs，观察到通过抑制线粒体介导的BAX/bcl—caspase9—caspase-3通路途径，抑制离体、在体腹腔液、以及EMs病灶中，巨噬细胞的凋亡，诱导M1分化，增强其活性，发挥抗炎、吞噬功能，有利于清除EMs病灶，抑制EMs发生、发展。此外，子宫TCs与ESCs共培养，可通过ERK途径，介导增强ESCs的增殖，粘附和运动功能。子宫TCs还通过其外分泌（WNTs生成）效应，激活ESCs中WNT/β-catenin途径，介导增强ESCs的蜕膜化与间质-上皮转化（MET）效应。..科学意义：通过直接的细胞间连接，或间接的旁分泌效应，TCs可对盆腔局部pMACs与ESCs发挥免疫调节效应，调控pMACs与ESCs的活性来发挥对EMs 病理过程的抑制效应。