胆绿素还原酶调节巨噬细胞活化状态对肾间质纤维化的影响和干预研究

As increasing doubts on renal tubular epithelial-mesenchymal transition (EMT), macrophages infiltration, the common feature of renal injury, has been recently proved that they play an important role in kidney fibrosis. The role of macrophages in mediating renal injury is related to their activation status or subsets. However, there needs to be a much greater understanding of the therapeutic targets of those macrophages. Biliverdin reductase (BVR), the key regulator of oxidative stress, has been recently characterized in activity profile of signal transduction pathways and inflammatory response. It neutralizes ROS, increases IL-10 expression in macrophages, and inhibits the TNFα pathway. This suggests it has the potential to skew macrophages toward a M2 "healer" phenotype. We hypothesize that the correction of the aberrant macrophages phenotypes by BVR could inhibit the kidney destructive inflammation and early-onset renal fibrosis. To test this hypothesis, using UUO mice in comparison to normal mice or human renal biopsy samples we will determine whether: (A) macrophage phenotypes are aberrant and skewed towards M1 "destroyers" and defective in shifting towards M2 "healers", thereby driving the onset of renal fibrosis; (B) Aberrant macrophage are a result of renal injury and/or inflammation (environment); (C) BVR could regulate macrophages activation status or subsets to a M2 "healer" phenotype; (D) By deliver BVR gene, macrophages can be genetically engineered to inhibit inflammatory or signaling pathways, providing long-lasting expression and site specific migration to inhibit kidney inflammation and early-onset renal fibrosis, which will offer a unique opportunity to identify novel therapeutic strategies for human renal fibrosis.

随着"肾小管上皮细胞-间充质转分化"理论的被质疑和边缘化，浸润肾间质的主要细胞- - 巨噬细胞在肾脏纤维化中的作用逐渐受到关注。巨噬细胞调节炎症和纤维化的机制与巨噬细胞活化状态有关。然而，目前尚缺乏调节肾脏巨噬细胞活化状态的基因或生物治疗。抗氧化家族的重要成员胆绿素还原酶（BVR），同时是一种细胞内信号分子，能够清除巨噬细胞内ROS，阻断TNF-α信号，促进巨噬细胞IL-10分泌，因而具备调节巨噬细胞表型向M2（保护性）转化的潜能。本项目拟证实BVR对巨噬细胞M2（保护性）的激活和M1（损伤性）的抑制，明确BVR对巨噬细胞活化状态的调节效应和信号机制，并通过将BVR基因靶向转染小鼠自体骨髓来源巨噬细胞，导入肾脏局部，持续调节肾间质巨噬细胞的活化状态，从而阻断或逆转肾脏纤维化。本项目将明确一个调节巨噬细胞活化状态干预肾纤维化的新靶点，将为肾纤维化发病机理提供新理论基础，为防治肾纤维化探索新途径。

巨噬细胞调节炎症和纤维化的机制与巨噬细胞活化状态有关。抗氧化家族的重要成员胆绿素还原酶（BVR），同时是一种细胞内信号分子，能够清除巨噬细胞内ROS，阻断TNF-α信号，促进巨噬细胞IL-10分泌，因而具备调节巨噬细胞表型向M2转化的潜能。.通过建立小鼠UUO模型，观察肾组织浸润巨噬细胞的表型、活化状态和BVR阳性巨噬细胞的比例，完成不同亚群BVR阳性巨噬细胞浸润程度与尿蛋白、炎性细胞浸润度和肾脏病变活动度的相关性分析。体外培养高表达BVR的小鼠自体骨髓来源巨噬细胞系，完成分离和功能鉴定，并检测其对炎性细胞活化状态的影响。检测体外不同活化亚型巨噬细胞和高/低表达BVR巨噬细胞对肌成纤维细胞活化和增殖的影响。观察巨噬细胞对肾脏纤维化和修复过程中的细胞因子的影响，发现IL-34及CSF-1在肾脏纤维化中与巨噬细胞密切相关。为了明确肾脏纤维化过程中肌成纤维细胞主要来源，构建异体共生模型，发现肾脏纤维化过程中肌成纤维细胞主要来源于肾脏原位细胞。.本项目明确BVR对巨噬细胞活化状态的调节效应和信号机制，可作为一个调节巨噬细胞活化状态干预肾纤维化的新靶点，为肾纤维化发病机理提供新理论基础，为防治肾纤维化探索出新途径。