Podocyte-specific human PLA2R transgenic mouse model of membranous nephropathy

足细胞特异性人PLA2R转基因小鼠膜性肾病模型

• 批准号: 8103239
• 负责人: DAVID J SALANT
• 金额: $ 20.91万
• 依托单位: BOSTON MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-07-02 至 2012-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8103239
• 关键词: Animal Testing; Antibodies; Antigen Targeting; Antigens; Autoantibodies; Binding; Biopsy; Cavia; Cell Aging; Chinchilla (genus); Clinical; Complement; Deposition; Development; Disease; Eicosanoid Production; Epitopes; Future; Goals; Human; Idiopathic Membranous Nephropathy; IgG1; IgG4; Immune; Immunoglobulin G; Injury; Kidney; Laboratory Animals; Ligand Binding; Location; Mediating; Membranous Glomerulonephritis; Modeling; Mus; Oryctolagus cuniculus; Pathogenicity; Patients; Phospholipase; Play; Positioning Attribute; Proteinuria; Rattus; Role; Serum; Specificity; System; Techniques; Testing; Transgenic Mice; Transgenic Organisms; complement pathway; in vivo; model development; mouse model; nephrin; podocyte; promoter; public health relevance; receptor; tool

DESCRIPTION (provided by applicant): The primary goal of this exploratory proposal is to develop a tool to determine if circulating autoantibodies directed at the M-type phospholipase receptor (PLA2R) from patients with idiopathic membranous nephropathy (MN) are pathogenic. We have established that about 75% of patients with idiopathic MN have circulating anti-PLA2R autoantibodies whereas none of the disease or normal controls has tested positive, thus indicating a high degree of specificity. The anti-PLA2R antibodies are IgG4, the subclass that predominates in idiopathic but not secondary MN, the antigen PLA2R is expressed on podocytes and colocalizes with IgG4 in the glomerular immune deposits of patients with MN, and the antibodies can be eluted from the glomerular immune deposits of MN renal biopsies. Moreover, the presence of circulating anti-PLA2R antibodies correlates with disease activity in MN. Although these findings strongly suggest that anti-PLA2R antibodies are responsible for, or at least contribute to disease development, proof of pathogenicity has yet to be established. Whereas simple transfer of the disease to laboratory animals with human anti-PLA2R would readily answer this question, human anti-PLA2R does not recognize PLA2R in the glomeruli of any small laboratory animal tested to date. Therefore, we plan to take advantage of both the lack of immunoreactive PLA2R in mouse glomeruli and the refinement of podocyte- specific transgenic techniques to develop a mouse model in which the human PLA2R is expressed in podocytes. To that end we propose the following specific aims: Specific Aim 1: Produce a transgenic mouse utilizing the NPHS1 (nephrin) promoter to drive expression of human PLA2R exclusively in mouse podocytes. Specific Aim 2: Verify that human PLA2R is expressed on mouse podocytes in vivo. Specific Aim 3: a) Determine if injected human anti-PLA2R autoantibodies bind to human PLA2R and form immune deposits typical of MN in the transgenic mouse glomeruli. b) Establish if injected human anti-PLA2R antibodies are able to cause proteinuria in the human PLA2R transgenic mouse. Specific Aim 4: Determine if the human PLA2R transgenic mice develop anti-PLA2R antibodies and MN when actively immunized with human PLA2R. Successful development of this model will pave the way for studies to both establish the pathogenic role of human anti-PLA2R autoantibodies and define the mechanisms of injury. PUBLIC HEALTH RELEVANCE: Having demonstrated that the M-type phospholipase receptor (PLA2R) is a specific target antigen of circulating autoantibodies in a high proportion of patients with idiopathic membranous nephropathy, we propose to develop a mouse model to determine if the antibodies are responsible for the clinical (proteinuria) and pathological manifestations of the disease. To that end, we will produce transgenic mice in which the human PLA2R is expressed on mouse podocytes, the location of the antigen in human kidneys.

描述（由申请人提供）：该探索性提案的主要目标是开发一种工具来确定特发性膜性肾病（MN）患者针对 M 型磷脂酶受体（PLA2R）的循环自身抗体是否具有致病性。我们已经确定，约 75% 的特发性 MN 患者具有循环抗 PLA2R 自身抗体，而疾病或正常对照均未检测出阳性，因此表明具有高度特异性。抗 PLA2R 抗体是 IgG4，该亚类在特发性而非继发性 MN 中占主导地位，抗原 PLA2R 在足细胞上表达，并与 MN 患者肾小球免疫沉积物中的 IgG4 共定位，并且可以从 MN 肾活检的肾小球免疫沉积物中洗脱抗体。此外，循环抗 PLA2R 抗体的存在与 MN 的疾病活动相关。尽管这些发现强烈表明抗 PLA2R 抗体是疾病发展的原因，或至少有助于疾病的发展，但致病性的证据尚未确定。虽然简单地将疾病转移到具有人抗 PLA2R 的实验动物身上就可以很容易地回答这个问题，但人抗 PLA2R 不能识别迄今为止测试的任何小型实验动物的肾小球中的 PLA2R。因此，我们计划利用小鼠肾小球中缺乏免疫反应性PLA2R和足细胞特异性转基因技术的改进来开发一种在足细胞中表达人PLA2R的小鼠模型。为此，我们提出以下具体目标： 具体目标 1：利用 NPHS1（去氧肾上腺素）启动子产生转基因小鼠，仅在小鼠足细胞中驱动人 PLA2R 的表达。 具体目标 2：验证人 PLA2R 在体内小鼠足细胞上表达。 具体目标 3：a) 确定注射的人抗 PLA2R 自身抗体是否与人 PLA2R 结合并在转基因小鼠肾小球中形成 MN 典型的免疫沉积物。 b) 确定注射的人抗 PLA2R 抗体是否能够在人 PLA2R 转基因小鼠中引起蛋白尿。 具体目标 4：确定人 PLA2R 转基因小鼠在用人 PLA2R 主动免疫时是否会产生抗 PLA2R 抗体和 MN。该模型的成功开发将为确定人类抗 PLA2R 自身抗体的致病作用和确定损伤机制的研究铺平道路。 公共健康相关性：在证明 M 型磷脂酶受体 (PLA2R) 是大部分特发性膜性肾病患者循环自身抗体的特异性靶抗原后，我们建议开发一种小鼠模型，以确定这些抗体是否与该疾病的临床（蛋白尿）和病理表现有关。为此，我们将生产转基因小鼠，其中人类 PLA2R 在小鼠足细胞（人类肾脏中抗原的位置）上表达。