Pathobiology of Complement C3 effects in ADPKD

描述(由申请人提供)： 由于常染色体显性遗传性多囊肾病(ADPKD)的诊断要到40岁才能排除，因此许多ADPKD患者都是退伍军人。本研究的目的是明确靶向补体C3(补体途径的轴向成分)对ADPKD的治疗潜力。随着局部C3产生的新的基本作用的发现而导致的范式转变，我们的研究指出，肾内C3的产生是决定肾脏囊变速度的途径的调节因素。在PKD模型中，我们基于对肾脏的全基因组表达分析形成了这一假说。我们证明了这一假说：(I)排列在ADPKD和常染色体隐性(AR)PKD囊内的肾小管细胞中C3mRNA和蛋白的存在以及这些细胞激活C3的能力；(Ii)ARPKD和ADPKD及其模型中具有生物活性的C3裂解产物的肾脏含量增加；(Iii)在PKD模型中，肾脏C3的表达与肾脏囊变的速度和C3低活性基因变异之间存在显著的相关性；以及(Iv)在遗传性PKD模型中，C3缺乏对肾囊变具有显著的保护作用。虽然C3可能通过不同的途径发挥作用，但我们发现加速囊变与补体受体CR3调节的途径有一致的关联。CR3(或Mac-1)是C3片段iC3b的主要受体，在囊性肾脏中含量丰富，在单核/巨噬细胞的分化、附着和存活中起着核心作用。虽然我们已经确定C3、巨噬细胞标记物CD14和C3诱导因子MCP-1是PKD预后的候选预测因子，但其他研究表明，巨噬细胞耗竭通过减少囊性小管的增殖来减少ADPKD直系模型的囊变。CR3还可诱导促囊性肿瘤坏死因子的释放，并直接激活肾小管上皮细胞的c-Src。由于肾小管细胞在无液体流动时产生并激活C3，我们认为在PKD中C3效应随着囊性小管扩张而增加，形成一个恶性的生囊循环。具体地说，我们假设C3途径的激活通过CR3依赖的过程加速了ADPKD的囊性形成。我们提出这一假说有三个相互关联的目的：1)剖析C3产生对PKD1途径的囊变作用的机制；2)确定肾小管来源的C3在ADPKD发病机制中的作用；3)确定补体成分受体CR3(或MAC1)在PKD1诱导的囊变中的作用。这些目标的目标将通过整合：(I)询问肾脏囊变的新调控机制的高度创新的研究设计与(Ii)新技术试剂的产生(例如，用于条件C3靶向)来实现。通过将已建立的和新的致囊途径与C3-CR3联系联系起来，实现拟议的目标将允许整合现有的知识。拟议的研究代表着实现我们的长期目标的下一步，即开发一种安全的基于C3的ADPKD和其他肾脏疾病的治疗方法。

DESCRIPTION (provided by applicant): Because the diagnosis of autosomal dominant polycystic kidney disease (ADPKD) cannot be excluded until age 40, many ADPKD patients are veterans. The objective of this proposal is to define therapeutic potential of targeting C3 (the axial component of complement pathway) in ADPKD. In line with paradigm shifts resulting from discoveries of novel essential roles of local C3 production, our studies point to intra-renal production of C3 as a regulator of a pathway that dictates the pace of renal cystogenesis. We formulated this hypothesis based on our genome-wide expression analyses of kidneys with rapid vs slow pace of cystogenesis in a PKD model. We supported this hypothesis by demonstrating: (i) presence of both C3 mRNA and protein in renal tubular cells that line ADPKD and autosomal recessive (AR) PKD cysts and the ability of these cells to activate C3; (ii) increased renal content of biologically active C3 split products i ARPKD and ADPKD and their models; (iii) strong correlation between renal C3 expression and pace of renal cystogenesis in a PKD model and C3 hypoactive gene variant with the disease progression among ADPKD patients; and (iv) significant protective effect of C3 deficiency on renal cystogenesis in genetic PKD models. While C3 may act through different pathways, we have found consistent association of accelerated cystogenesis specifically with the pathway regulated by complement receptor CR3. CR3 (or Mac- 1), a major receptor for C3 fragment iC3b, which is highly abundant in cystic kidneys, plays a central role in differentiation, attachment and survival of monocytes/macrophages. While we have identified C3, macrophage marker CD14 and C3-inducible factor MCP-1 as candidate predictors of PKD outcomes, others have demonstrated that macrophage depletion attenuates cystogenesis in orthologous models of ADPKD by reducing the proliferation of cystic tubules. CR3 may also induce pro-cystogenic TNF release and directly activate c-Src in renal tubule cells. Since renal tubule cells produce and activate C3 when fluid flow is absent we suggest that the C3 effects in PKD increase as cystic tubules dilate, forming a vicious cystogenic cycle. Specifically, we hypothesize that C3 pathway activation accelerates cyst formation in ADPKD through a CR3 dependent process. We address this hypothesis in three inter-related aims: 1) Dissect mechanisms underlying cystogenic effects of C3 production on Pkd1 pathway; 2) Determine effects of renal tubule-derived C3 in the pathogenesis of ADPKD.; and 3) Determine the role of complement component receptor CR3 (or Mac1) in Pkd1-induced cystogenesis. Objectives of these Aims will be accomplished by integrating: (i) highly innovative study design of interrogating novel regulatory mechanisms of renal cystogenesis with (ii) generation of novel state of the art reagents (e.g., for conditional C3 targeting). Achieving the proposed aims will allow integration of existing knowledge by linking established and novel cystogenic pathways to the C3-CR3 nexus. The proposed studies represent the next step for attaining our long-term goal of developing a safe C3-based therapy for ADPKD and other renal disorders.