巨噬细胞与促纤维化Niche形成的反馈调控加速肾纤维化进程

Macrophage infiltration and excessive extracellular matrix (ECM) deposition are the characteristic pathological features during progression of renal fibrosis in chronic kidney disease (CKD). However, whether macrophage activation and aberrant ECM deposition are mutually regulated in CKD remains elusive. Here, we have demonstrated that activation of the fibrinolytic system in macrophages is mediated by HGF/c-met signaling, which physically regulates ECM degradation to maintain kidney homeostasis. In CKD, the capacity of macrophages to mediate ECM proteolytic degradation was largely impaired due to significant downregulation of HGF signaling in vivo, resulting in excessive ECM synthesis and deposition and a sequentially formed pro-fibrotic microenvironment, the Fibrogenic Niche. The fibrogenic niche could reciprocally anchor and regulate biologic activities of the kidney resident or infiltrated cells and eventually leading to renal fibrosis. Based on our findings, we hypothesize that the fibrogenic niche could mediate macrophages polarization and successively inhibit ECM proteolysis by which to “protect” itself from degradation through an established feedback loop. In other words, the imbalance of the macrophage-fibrogenic niche feedback loop could accelerate renal fibrosis in CKD. In the current application, we aim first to validate the presence and molecular mechanisms in regulating this feedback loop both in vitro and in vivo. Then, we will further explore the possibility of effectively retard renal fibrosis through disrupting the macrophages-fibrogenic niche feedback loop in multiple CKD models via ①recovering and enhancing the proteolytic ability of macrophages or ② repressing formation of the fibrogenic niche, respectively. To our knowledge, the model proposed in this application will serve as a novel experimental tool in delineating the molecular targets holds promise for future therapies of CKD.

巨噬细胞浸润和细胞外基质（ECM）沉积是肾纤维化发生发展的重要病理学事件，两者之间是否相互调控尚未明确。申请人发现，巨噬细胞纤溶系统受肝细胞生长因子（HGF）信号调控，促进ECM降解。CKD时，体内HGF信号显著减弱，导致巨噬细胞水解ECM能力降低；异常沉积的ECM形成促纤维化巢（Niche），反向调控肾组织细胞，加重肾纤维化。申请人认为，促纤维化Niche也可反馈作用于巨噬细胞，调节炎症表型并抑制其水解ECM，以“保护”自身不被降解；巨噬细胞-促纤维化Niche间形成的反馈调控加速肾纤维化进程。本项目拟首先通过体内、体外实验明确该反馈调控是否存在；在此基础上，深入探讨其分子机制；并分别从①恢复巨噬细胞水解ECM功能②抑制促纤维化Niche形成两方面破坏巨噬细胞-促纤维化Niche反馈环，在多个CKD模型中探讨该方法是否能有效抑制肾纤维化进程；以期为后续转化应用提供新的理论模型和干预靶点。

肾纤维化是各种慢性肾病（CKD）进展至终末期肾病的病理基础和共同结局，是导致肾功能丧失的决定性因素。肾脏组织微环境在该病理过程中具有重要作用，然而，其分子组成和动态调控仍不清楚。在该项目的资助下，我们发现，肝细胞生长因子（HGF）/c-met信号对调控肾组织微环境稳态，维持肾功能具有重要作用。课题组通过深度挖掘转录组学和单细胞测序结果，联合数学建模，采用生物学实验验证，发现并证实，（HGF）/c-met信号主要通过调控肾小管上皮细胞和巨噬细胞发挥肾保护作用。病理条件下，体内HGF水平低下，导致HGF/c-met信号减弱，肾小管上皮细胞表型发生转化，合成大量细胞外基质；巨噬细胞发生极性转化，纤溶酶原激活物uPA、tPA分泌减少，水解细胞外基质能力减弱。由于上述两类细胞功能异常引起的细胞外基质合成与降解失衡，导致细胞外基质异常堆积，诱发并推动肾纤维化的发生发展。在细胞外基质沉积的同时，微环境的重要组建者，细胞外基质糖蛋白Tenascin C（TNC）沉积越来越多，逐渐形成促纤维化样病灶，激活肾小管上皮细胞整合素受体αvβ6，活化下游FAK信号通路，从而破坏肾小管上皮细胞完整性并诱发上皮-间充质转分化，加重肾纤维化进展。体内采用中和抗体封闭αvβ6或小分子抑制剂干预下游FAK信号，可有效抑制肾纤维化进程。受损的肾小管上皮细胞还分泌细胞基质Fibrillin-1（FBN），活化内皮细胞αvβ6/TGF-β信号，抑制细胞增殖并诱导凋亡。体内阻断该通路可显著减轻内皮细胞损伤并恢复肾组织血管网密度。此外，上述微环境分子还具有重要的诊断价值，临床检测发现，CKD患者尿液TNC和血清FBN水平显著升高，升高程度与病情进展和纤维化程度呈正相关。通过该项目研究，我们从微环境角度深入认识了肾纤维化发生发展机制，发现潜在的药物靶点并提出新的临床诊治策略；同时，我们还创建了基于组织微环境的体外研究策略和全新药物筛选平台，为后续转化研究奠定基础。