The glomerular basement membrane (GBM) is an essential part of the blood filtration barrier and the primary site of injury in aggressive forms of glomerulonephritis mediated by anti-GBM antibodies. Pathogenic antibodies are elicited by autoimmune or alloimmune mechanisms and target specific sites on a3a4a5(IV) collagen, the major GBM component. These sites are located within the non- collagenous (NC1) domains, which mediate the specific assembly of collagen IV chains into quaternary structures, forming NC1 hexamers in the native tissues. Incompletely understood are the molecular mechanisms by which immune tolerance to collagen IV chains is normally established in health, broken in autoimmune anti-GBM disease, or selectively deficient in alloimmune Alport post-transplant nephritis. The objective of this project is to determine at molecular level how the adaptive immune system is abnormally engaged, initiating anti-GBM nephritis. The premise is that assembly of collagen IV chains into quaternary structures profoundly affects adaptive immune responses to this antigen in health and disease, because the formation of NC1 hexamers: a) hides cryptic B cell epitopes while creating novel quaternary epitopes;b) affects the processing and presentation of T cell epitopes;and c) shapes immune tolerance by determining which collagen IV chains associate and their native structure in tissues. Anti-GBM antibodies are the major focus of this proposal because they couple the activation of antigen-specific B and T cells to effector pathways causing tissue damage. We hypothesize that the formation of pathogenic antibodies is determined by both molecular structure of the antigen and the host genetic background (antigen-deficient or -sufficient). The recurring theme is that the nephritogenicity of anti-GBM antibodies is contingent upon their ability to bind to accessible sites in the GBM and engage inflammatory effectors. The objective set forth in this proposal will be achieved by pursuing four specific aims.
Aim 1 will characterize novel subsets of anti-GBM autoantibodies found only in the patient kidneys and determine their pathogenic significance by passive transfer studies.
Aim 2 will identify the pathogenic determinants of the inciting alloantigen and of the anti-GBM alloantibodies causing post-transplant nephritis in X-linked Alport patients.
Aim 3 will characterize the molecular features of the antigen and antibodies determining the development of anti-GBM nephritis in mouse models, and their effector mechanisms.
Aim 4 will establish how genetic deficiency of collagen IV chains in Alport syndrome is permissive for pathogenic alloimmune responses by identifying abnormal B cell and T cell responses to collagen IV in antigen-knockout mice. The characterization of novel autoantigenic and alloantigenic sites on a3a4a5(IV) collagen and the identification of pathogenic determinants and effector mechanisms of anti-GBM antibodies will facilitate the development of targeted strategies for prevention, detection, and treatment of human anti-GBM nephritis.

Public Health Relevance

A very aggressive, life-threatening form of kidney disease is caused by abnormally produced antibodies that target the glomerular basement membrane (GBM), an important part of the blood filtration barrier in the kidneys. Detailed knowledge of the specific GBM components attacked by disease-producing antibodies and studies of a similar illness produced in laboratory mice or rats will helps us understand individual steps in the sequence of events leading to severe glomerular disease. These studies may identify factors that increase the risk of disease, suggest how to avoid disease in people at risk, and yield better treatments of established disease.

National Institute of Health (NIH)
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Research Project (R01)
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Pathobiology of Kidney Disease Study Section (PBKD)
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Flessner, Michael Francis
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Vanderbilt University Medical Center
Internal Medicine/Medicine
Schools of Medicine
United States
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Borza, Dorin-Bogdan (2014) IgG4-restricted anti-glomerular basement membrane autoantibodies targeting quaternary epitopes of native ?345(IV) collagen. Am J Kidney Dis 64:156-7
Olaru, Florina; Luo, Wentian; Suleiman, Hani et al. (2014) Neonatal Fc receptor promotes immune complex-mediated glomerular disease. J Am Soc Nephrol 25:918-25
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Olaru, Florina; Wang, Xu-Ping; Luo, Wentian et al. (2013) Proteolysis breaks tolerance toward intact ýý345(IV) collagen, eliciting novel anti-glomerular basement membrane autoantibodies specific for ýý345NC1 hexamers. J Immunol 190:1424-32
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Luo, Wentian; Wang, Xu-Ping; Kashtan, Clifford E et al. (2010) Alport alloantibodies but not Goodpasture autoantibodies induce murine glomerulonephritis: protection by quinary crosslinks locking cryptic ýý3(IV) collagen autoepitopes in vivo. J Immunol 185:3520-8
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