Semaphorin 4C/Plexin-B2信号途径调控高内皮微静脉新生在肾间质纤维化启动阶段的作用及其干预研究

Tertiary lymphoid organs (TLO) neogenesis is one of the most important histological events in the priming phase of renal interstitial fibrosis, which is a relentless driver of the vicious cycle of inflammation, tissue damage and fibrosis. Our previous data have showed that high endothelial venules (HEV) development in renal interstitial area is closely related with lymphoid neogenesis. And the phenotypic characteristics of HEV, which is regulated by dendritic cells (DC), are also involved in HEV-mediated lymphocyte homing. Our past studies have found that semaphorin 4C (Sema4C) expression is upregulated in injured renal tubular epithelial cells and plays an important role in progressive renal interstitial fibrosis. Recently other researchers have detected that Sema4C is expressed in HEV endothelial cells, and its ligand, Plexin-B2, is expressed in both HEV endothelial cells and DC, which is involved in the regulation of angiogenesis and DC activity. Therefore Sema4C/Plexin-B2 signaling pathway is thought to have the potential to regulate HEV neogenesis and phenotypic conversion. In this project, we will plan to clarify the mechanisms of HEV-mediated TLO neogenesis via Sema4C/Plexin-B2 signaling pathway and explore the new preventive and therapeutic strategy in the early stage of renal fibrosis by using CD11c-DTR mice for DC deletion and transcription activator-like effectors nucleases (TALENs) technique for targeted gene editing.

肾内类淋巴组织样结构(TLO)形成是肾间质纤维化启动阶段重要的组织学事件,是炎症-组织损伤-肾脏纤维化恶性循环的驱动者。申请者发现:肾间质内高内皮微静脉(HEV)新生与TLO形成密切相关。而HEV功能又与HEV表型特点有关,受树突状细胞(DC)调控。申请者既往还发现:Semaphorin 4C(Sema4C)在损伤后的肾小管上皮高表达,是肾间质纤维化重要的调控分子。近期其他学者发现:Sema4C表达于HEV内皮细胞;其配体Plexin-B2表达于HEV内皮细胞和DC,参与调控血管生成和DC活化。可见:Sema4C/Plexin-B2具备调控HEV新生和表型特点的潜能。本项目拟从临床、动物、细胞三个层面,结合CD11c-DTR转基因小鼠的应用和TELANs基因靶向修饰技术,阐明Sema4C/Plexin-B2通路调控HEV介导的TLO形成的机制,为探索肾间质纤维化防治新策略提供实验依据。

本课题旨在探究肾脏纤维化启动阶段，局部炎症微环境形成的新机制——高内皮微静脉介导的类淋巴组织样结果形成以及树突状细胞的调控作用。我们选取原发、继发性肾脏病以及肾移植术后肾活检标本，通过免疫组化等方法检测肾间质DC、HEV新生现象以及HEV表型特点与肾脏局部类淋巴组织样结构新生之间的关系。结果表明，慢性肾移植组织存在HEV新生，可能参与肾脏炎症微环境的形成；DC可能与HEV的形成有关；而在原发及继发性肾脏疾病中，表达无统计学差异。.基于前期研究结果及综合分析TLO的最新研究进展，证实IL-7在TLO的形成及慢性炎症中的重要作用。因此，我们通过免疫组化、masson染色等方法，发现在IgA肾病以及动物UUO模型中，IL-7主要表达于肾小管上皮细胞，并且随着肾脏纤维化程度加重表达呈上升趋势。为明确IL-7的作用及其作用机制，我们通过免疫印迹等方法在体外研究中证实IL-7主要表达于肾小管上皮细胞，其表达量与肾脏纤维化呈正相关。外源性添加IL-7可通过活化mTOR信号通路，抑制肾小管上皮细胞自噬，减少细胞外基质的表达从而抑制肾脏纤维化。我们通过检测IgA肾病患者血清IL-7表达水平，发现血清IL-7水平与肾脏疾病进展以及预后密切相关。本研究证实肾小管上皮细胞通过IL-7-mTOR信号通路抑制肾小管上皮细胞自噬，从而抑制肾脏纤维化，进一步丰富和发展了肾脏纤维化的理论基础；通过检测血清IL-7水平，说明血清IL-7水平与IgA肾病预后存在相关性，为临床判断及干预肾脏疾病的预后提供新的思路，有望为肾脏纤维化提供新的诊疗策略。