Michael Paley and Lynn HassmanAnalysis of HLA-B27+ anterior uveitis Uveitis is associated with multiple rheumatic diseases, from inflammatory arthritis to vasculitis. In particular, approximately half of patients with HLA-B27+ spondyloarthritis develop anterior uveitis, however, the mediators of this eye disease are not well understood. In addition, incomplete control of ocular inflammation can lead to cataract, glaucoma, and permanent vision loss. As a result, there is a need to better understand the pathophysiology of ocular inflammation in order to better tailor uveitis therapy and provide critical insight into systemic rheumatologic disease. We hypothesize that human uveitis is driven by autoreactive T cells responding to ocular antigens. Our hypothesis is based on observations that CD4 and CD8 T cells infiltrate the eye in anterior uveitis and that the MHC class I molecule HLA-B27 is strongly associated with anterior uveitis. However, it is still unknown whether antigen presentation at the site of inflammation (e.g. eye) or at a distant organ (e.g. gut) is the initiating stimulus. Moreover, prior studies had technical limitations. To better elucidate the pathogenesis of this disease, we created a biorepository of blood and ocular samples from subjects with HLA-B27+ anterior uveitis. Preliminarily, we performed simultaneous single cell RNA sequencing (scRNAseq) and single cell T cell receptor (TCR) sequencing (scTCRseq) on 4 paired ocular and blood samples, allowing us to address our Specific Aims. |
Michelle ElvingtonEstablishing C3(H2O) as a biomarker in SLE and defining its role in B lymphocyte function Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by autoantibody production, immune complex formation, and complement activation and deposition in tissues. Complement activation by immune complexes drives type II & III hypersensitivity reactions, leading to inflammatory responses in target tissues. Failure to remove cellular debris, a process that is highly dependent on complement, is also an important tenet of SLE pathophysiology. The role of complement in the activation of immune cells is well-documented. Complement deposition in tissues serves as a diagnostic tool, especially in the kidney, and decreased serum complement components C4 and C3 are markers of active disease. C3 continuously activates at a low level in the fluid phase by spontaneous hydrolysis of the internal thioester bond. Within microseconds this form of C3 becomes covalently bound to a target (such as a microbe or cell) or interacts with water to form C3(H2O). Interest in C3(H2O) in disease processes has recently been renewed due to the discovery of the intracellular complement system (ICS) whose dysfunction is increasingly associated with debris clearance and autoimmunity. In a follow-up report to our identification of the ICS, I made an additional key observation of a process whereby C3(H2O) is loaded into the cell interior where it can be stored or cycled back out of the cell. Thus, C3(H2O) has two main functions: (1) serves as a surveillance system via being a trigger for alternative pathway (AP) activation on a target and (2) supplies this form of C3 for the ICS. In a process involving the uptake of C3(H2O), a variety of cells have been shown to cleave C3(H2O) intracellularly into C3a and C3b. Further, these cleavage products engage intracellular complement receptors and drive changes in immune cell cytokine production. Moreover, preliminary data indicates that serum C3(H2O) generation is substantially elevated in patients with SLE compared to healthy controls. While it is not known what drives increased C3(H2O) levels or what role it plays for immune cell function in patients with SLE, we suspect that elevated C3(H2O) may contribute to dysfunction of the ICS. While clinical assays exist for C3 and certain of its fragments, there was no quantitative and straight-forward assay to measure C3(H2O). My recent development of an ELISA specific for C3(H2O) which does not cross-react with native C3, C3b or C3b-derived fragments gives us the unique opportunity to investigate C3(H2O)’s role in human disease and to address how its presence may be participating in immunoinflammatory conditions. I hypothesize that increased C3(H2O) generation contributes to dysfunction in SLE and thus C3(H2O) serves as a marker of disease activity. |
Chieh-Yu Lin and Kory LavineCellular and Transcriptomic Landscape of Cardiac Sarcoidosis Sarcoidosis is a multisystem rheumatic disease without clear etiology. Cardiac involvement by sarcoidosis is identified in a subset of patients, with a prevalence ranging from 5-25%. Cardiac sarcoidosis has recently gained increasing attention, as it portends an unfavorable prognosis accounting for nearly 85% deaths in sarcoidosis patients. Today, it remains challenging to diagnose and effectively treat active cardiac sarcoidosis. Endomyocardial biopsy has limited sensitivity and diagnostic utility due to the patchy distribution and subepicardial predominance of disease. The pathognomonic findings of noncaseating granulomas are only identified in about 20% of the biopsy specimens. Given issues related to sampling bias, FDG PET imaging is the current standard of care, but the results are unsatisfactory. Importantly, treatment options for cardiac sarcoidosis are limited. Nonselective anti-inflammatory therapies such as corticosteroids and azathioprine represent the primary pharmacological treatment options. Unfortunately, these agents have limited efficacy and there is a paucity of clinical data to support their use. The rationale of using such modalities are largely extrapolated from non-cardiac sarcoidosis patients. TNF-alpha antagonists, such as infliximab, have shown promising results for severe/refractory sarcoidosis. However, the efficacy for cardiac sarcoidosis is unclear. These observations highlight a clinically unmet need to develop new approaches to diagnose and treat cardiac sarcoidosis. Surprisingly, little is understood regarding the pathological mechanisms that govern the initiation and progress of this devastating disease. We believe that a comprehensive understanding of the cellular composition and transcriptomic landscape of cardiac sarcoidosis in humans represents a critical first step towards obtaining the requisite information to begin to generate effective diagnostic tools and treatment options. |
| Iris Lee Complement regulation of the microbiome Secretory immunoglobulin A (sIgA) protects the host against pathogenic bacteria and maintains gut homeostasis by regulating host-microbiome interactions. Dysregulated sIgA is linked to dysbiosis and systemic inflammation, which are associated with autoimmunity. The proposed research will define the role of CD55, a complement regulator, in Peyer’s patch B cells, the sIgA repertoire, and microbiome composition. Preliminary data outlined in this proposal suggest that loss of CD55 increases IgA class-switching in Peyer’s patches. The central hypothesis is that CD55 regulates Peyer’s patch homeostasis by preventing the generation of complement fragments C3a/C5a. The specific aims of the proposal are to 1) Explore the mechanisms underlying the role of CD55 on IgA class-switching in Peyer’s patches and 2) Determine the role of CD55 on sIgA repertoire and microbiome development. The proposed studies will use flow cytometry, immunofluorescence staining, and high throughput sequencing to determine qualitative changes in sIgA production. |
| Michael Paley Mechanistic Role of ERAP1 in Axial Spondyloarthritis Pathogenesis Inheritance of the HLA-B*27 allele is the strongest genetic risk factor for developing axial spondyloarthritis (axSpA) and acute anterior uveitis (AAU).1 Specific alleles of the aminopeptidase ERAP1 potentiate this risk, but only in HLA-B*27+ individuals.1 The mechanism underlying this association, however, remains unclear. The “arthritogenic peptide” model proposes that HLA-B*27 contributes to disease pathogenesis by presentation of enteric bacterial antigens and ocular/articular self-antigens to cross-reactive, pathogenic CD8 T cells. This is supported by our recent work identifying shared (“public”) T cell receptors (TCRs) from CD8 T cells that were expanded in the eye and joint in patients with HLA-B*27+ AAU or axSpA.2 These TCRs recognized both a peptide from the bacterial YeiH protein and a self-peptide from GPER1. The YeiH protein is expressed by various pathogens (Shigella, Salmonella, Campylobacter, etc.) which are known to promote the development of axSpA after an intestinal infection. In parallel, public single cell RNA sequencing databases and our preliminary data suggest that GPER1 is specifically expressed in the iris pigmented epithelium, which are located within the inflamed ocular structures during AAU. Collectively, these data suggest that YeiH may be the initial antigenic trigger for pathogenic CD8 T cells that subsequently react to the self-antigen GPER1 in the eye and a yet-to-be-determined antigen in the joint. ERAP1 is an aminopeptidase that trims peptides to be loaded onto MHC class I molecules.1 Therefore, we hypothesize that specific ERAP1 alleles increase the risk of axSpA and AAU by facilitating the loading of the bacterial antigen YeiH onto HLA-B*27, which leads to the generation of YeiH-specific CD8 T cells. In this proposal, we aim to examine whether ERAP1 alleles facilitate cross-presentation of the YeiH antigen in vitro and whether ERAP1 alleles facilitate the generation of YeiH-specific CD8 T cells in vivo. Funds from the Pilot and Feasibility Grant will be used to support the following aims. |
| Tarin Bigley Identifying mechanisms of thymic disruption in MRV-induced autoimmunity Autoimmunity is a growing cause of human disease worldwide, but the etiology of many autoimmune diseases remains largely unknown. Both genetic and environmental factors contribute to development of autoimmune disease. Viral infections are considered to be a contributing factor to autoimmunity, but causality is often challenging to identify because the initial virus-induced immune dysregulation likely occurs before onset of overt autoimmune disease. This is especially true for chronic and latent infections such as herpesvirus infections. We have shown that neonatal infection with a herpesvirus, murine roseolovirus (MRV), induces autoimmunity that develops in adult mice, long after resolution of acute infection. These mice are prone to diverse autoantibodies as well as autoimmune gastritis, but are also predisposed to systemic autoimmunity with features of systemic lupus erythematosus after toll-like receptor 7 (TLR7) stimulation. This phenotype seems to be due to loss of T cell tolerance in the thymus. We have therefore identified an in vivo system to perform mechanistic studies into how early life infection with a roseolovirus leads to immune dysregulation that manifests as limited autoimmunity with predisposition to systemic autoimmunity after additional immune perturbations. We have found that type I interferon (IFN) signaling during the acute infection is necessary for MRV-induced autoimmunity and propose to utilize transcriptomics and T cell receptor sequencing to identify the impact of type I IFN on molecular pathways and T cell development in the thymus during MRV infection. Moreover, we have found that induction of systemic autoimmunity in MRV infected mice is associated with CD8+ T cell activation in the thymus and thymic atrophy. We propose to use a well-established model of T cell recirculation to the thymus to evaluate if recirculated T cells are present and target the thymus in our lupus-like systemic autoimmunity model. The data generated from this proposal will provide a foundation on which we can build toward our overall goal of understanding the mechanistic role that early life roseolovirus infection plays in development of autoimmunity later in life. |