巨噬细胞的极化异质性对卵巢癌腹水的分子调控机理

Change of vascular permeability is a key factor in ascites production in ovarian cancer, in which tumor-associated macrophages play a crucial role. Targeting macrophages, on one hand, abrogated the formation of ascites, indicating a rescue of vascular leakage; on the other hand, resulted in an increase of permeability, shown as edema, suggesting the heterogeneity of macrophage-mediated vascular permeability. Our previous findings revealed that differentially polarized macrophages have disparate regulatory impact on vascular permeability, i.e., M1-macrophages induce permeability, whereas M2-macrophages protect the vascular barrier. To identify polarization-specific molecules that regulate vascular permeability is the foundation and prerequisite for the study of ascites regulation by the heterogeneity of macrophage polarization. RNA-seq studies in the past using bulk samples were unable to dissect the heterogeneity on the single-cell level. Thus, using recently developed single-cell RNA-seq technique we have demonstrated the heterogeneity in macrophage polarization in murine ovarian cancer model and identified certain cell adhesion molecules (CAMs) being differentially expressed in a polarization-specific manner. This project aims to determine the function and molecular mechanism of the CAMs in regulating vascular permeability using cellular and animal models. Also, we aim to study the heterogeneity of macrophages in ovarian cancer patient samples and its regulating mechanism on vascular permeability, as well as its relevance to ascites production. This project will provide new therapeutic strategies to treat ascites and macrophage-targeting immunotherapies.

血管通透性改变是影响卵巢癌腹水形成的重要因素，肿瘤相关巨噬细胞参与并调节该病理过程。抑制卵巢癌巨噬细胞可减少腹水，却并发严重的颜面部和肢端水肿。该现象提示巨噬细胞对血管通透性的调节存在异质性。我们前期研究支持该观点，并发现不同极化的巨噬细胞对血管通透性的作用迥异：M1亚型促进血管通透，而M2亚型保护血管屏障。明确调节血管通透性的极化特异性分子是研究巨噬细胞极化异质性对腹水调控的基础与前提。过去对大量混合细胞的转录组测序无法在单细胞层面研究异质性，我们采用单细胞测序在小鼠中成功证明卵巢癌巨噬细胞的极化异质性，且发现一些细胞粘附分子在不同极化巨噬细胞中的表达差异。本项目拟在细胞和动物模型中研究这些极化特异性细胞粘附分子对血管通透性的调节功能及机理，并在临床样本中探讨卵巢癌巨噬细胞的极化异质性、调控机制及其与腹水生成的关系。本研究将为卵巢癌腹水的临床治疗和以巨噬细胞为靶标的免疫治疗提供新策略。

血管通透性改变是影响卵巢癌腹水形成的重要因素，本团队前期研究发现巨噬细胞参与并调节该病理过程。然而，腹水微环境中巨噬细胞异质性大，调节血管通透性的巨噬细胞类型及作用机制尚未可知。本项目研究发现：1）M2型巨噬细胞与内皮细胞接触后下调整合素VLA4分子表达，并通过抑制内皮细胞Rac1/ROS/p-Pyk2/p-VE-cad通路保护血管屏障；2）进一步证明该通路可与VEGF通路协同调控血管通透性，同时阻断VLA4和VEGF可协同抑制卵巢癌腹水生成；3）通过对卵巢癌不同病灶的免疫细胞单细胞转录组测序，发现一类富集于腹水中且高表达脂质分化相关蛋白PLIN2的巨噬细胞亚型，将其命名为“腹水相关巨噬细胞”（AAM）。随后通过体内外实验证明，巨噬细胞PLIN2可促进血管通透性，且通过整合素与CD4+ T细胞相互作用起到抑制免疫的功能。此外，我们还探索了血管功能调控新分子DDX24及其上下游调控机制，并揭示了血管通透性介导的病毒相关肠道炎症的致病机理。本研究以卵巢癌腹水为例，深入阐明了血管通透性的调控新机制，为恶性腹水的临床治疗和以巨噬细胞为靶标的免疫治疗提供新策略。