Pemphigus IgG Internalization and Desmosome Disassembly

天疱疮 IgG 内化和桥粒拆卸

• 批准号: 6805626
• 负责人: ANDREW P. KOWALCZYK
• 金额: $ 15.3万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-09-30 至 2006-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6805626
• 关键词: antibody formation; autoantibody; biological signal transduction; cadherins; cell adhesion molecules; cell membrane; clinical research; endocytosis; enzyme linked immunosorbent assay; fluorescence microscopy; human tissue; immunoglobulin G; immunoprecipitation; intercellular connection; intermolecular interaction; keratinocyte; molecular pathology; pemphigus; protein binding; scintillation counter; tissue /cell culture; western blottings

DESCRIPTION (provided by applicant): Pemphigus vulgaris (PV) is a severe autoimmune blistering disease characterized by the disruption of adhesion between epithelial cells in the oral mucosa and in the lower layers of the epidermis. The antigen targeted by autoantibodies in this disease is the desmosomal cadherin desmoglein-3 (Dsg3). Desmosomes are cell-cell adhesion structures that play critical roles in the establishment and maintenance of epidermal integrity. A major consequence of Dsg3 autoantibody production is the disruption of desmosomal cell-cell adhesion, and it is likely that PV antibodies directly interfere with cell-cell adhesion mediated by Dsg3. However, numerous studies have also demonstrated that desmosomal components are internalized by keratinocytes exposed to PV sera, both in vivo and in vitro. In preliminary studies, we have found that PV sera as well as pathogenic monoclonal antibodies directed against Dsg3 are internalized by keratinocytes and enter the endocytic pathway. These observations raise the possibility that endocytosis of Dsg3 leads to a significant reduction of cell surface levels of Dsg3, and that Dsg3 internalization in response to PV antibodies may be causally related to pemphigus disease pathogenesis. Therefore, by inhibiting Dsg3 endocytosis in response to PV IgG ligation, it might be possible to blunt the impact ofPV autoantibodies on epithelial cellcell adhesion. In order to address this possibility, a sophisticated understanding of how membrane trafficking systems interface with desmosomal adhesion molecules is needed. This R21 proposal will test the hypothesis that pathogenic Dsg3 antibodies cause the internalization of this cell surface adhesion molecule. Two aims are proposed. First, we will establish as series of quantitative assays to monitor PV IgG internalization, and use these assays in combination with pharmacological and dominant negative approaches to determine the basic mechanisms by which Dsg3 is internalized. Secondly, we will determine if PV IgG and Dsg3 are internalized alone, or as a complex with other desmosomal molecules. We will then test the possibility that components of the desmosome regulate Dsg3 internalization. These experiments will form the foundation for a subsequent R01 application. The long-term goal of these studies is to expose new cellular pathways that might be targeted therapeutically to make keratinocytes more resistant to PV IgG in patients suffering from this devastating disease.

描述(由申请人提供)： 寻常型天疱疮(PV)是一种严重的自身免疫性水疱病，其特征是口腔粘膜和表皮下层上皮细胞之间的黏附断裂。在这种疾病中，自身抗体靶向的抗原是桥粒钙粘蛋白桥粒桥粒蛋白-3(Dsg3)。桥粒是细胞间的黏附结构，在建立和维持表皮完整性方面起着关键作用。Dsg3自身抗体产生的一个主要后果是破坏桥粒细胞与细胞的黏附，可能是PV抗体直接干扰Dsg3介导的细胞与细胞的黏附。然而，大量的研究也表明，无论是在体内还是在体外，接触PV血清的角质形成细胞都能内化桥粒成分。在初步研究中，我们发现PV血清以及针对Dsg3的致病单抗被角质形成细胞内化并进入内吞途径。这些观察结果表明，Dsg3的内吞作用导致细胞表面Dsg3水平显著降低，并且Dsg3对PV抗体的内化可能与天疱疮的发病有关。因此，通过抑制Dsg3的内吞反应来回应PV的免疫球蛋白G，有可能减弱PV自身抗体对上皮细胞黏附的影响。为了解决这种可能性，需要对膜运输系统如何与桥粒黏附分子相互作用进行深入的了解。这项R21提案将检验致病Dsg3抗体导致这种细胞表面黏附分子内化的假设。提出了两个目标。首先，我们将建立一系列的定量检测方法来监测PV-Ig G内化，并结合药理学和显性阴性方法来确定Dsg3内化的基本机制。其次，我们将确定PV Ig G和Dsg3是单独内化，还是与其他桥粒分子形成复合体。然后我们将测试桥粒的组成部分调节Dsg3内化的可能性。这些实验将为后续的R01应用奠定基础。这些研究的长期目标是揭示新的细胞通路，这些通路可能是治疗上的靶向，以使角质形成细胞对这种毁灭性疾病患者的PV IgG具有更强的抵抗力。