Anti-cytokine autoantibodies promote inflammation and fibrosis in a rare and fatal autoimmune disease
Abstract/Contents
- Abstract
- IgG4-Related Disease (IgG4-RD) is a fibroinflammatory condition characterized by multi-organ inflammation, tissue infiltration by IgG4-expressing plasma cells, and storiform fibrosis.1,2 Autoantibodies against various self-antigens have been previously described in IgG4-RD.3-5 However, these autoantibodies are predominantly tissue-specific and fail to account for the inflammation, fibrosis, and diverse organ manifestations observed in this disease.1 The contribution of B cells and autoantibodies to IgG4-RD remains unclear. Here, we developed and utilized next-generation sequencing, high-throughput proteomics, and in vitro functional assays to investigate the plasmablast and autoantibody responses in patients with IgG4-RD. In Chapter 2, we performed deep sequencing of the plasmablast antibody repertoires of patients with IgG4-RD, multiple sclerosis (MS), and systemic lupus erythematosus (SLE), and compared the molecular characteristics of the plasmablast antibodies between these autoimmune diseases. We found that the percentage of plasmablasts expressing IgG antibodies was higher in patients with these autoimmune conditions compared to healthy subjects. Identification of the clonal family plasmablasts in the patients' antibody repertoires revealed that the percentage of plasmablasts representing clonal-lineages in IgG4-RD and MS were higher than in SLE. In addition, we discovered a preferential usage of the heavy chain variable genes in the clonal IgG plasmablasts unique to each autoimmune condition and shared across these diseases. Comparison of the IgG subclass distributions of the IgG plasmablasts in the autoimmune conditions revealed a higher percentage of IgG2- and IgG4-expressing plasmablasts in IgG4-RD. In Chapter 3, we performed human cytokine microarray analysis on the plasma samples of patients with IgG4-RD and identified cytokine antigens targeted by IgG autoantibodies. We identified two IgG4-RD patient-derived clonal plasmablast antibodies that bound to IL-1 receptor antagonist (IL-1RA) and macrophage inflammatory protein-3 (MIP-3/CCL23). Plasma levels of anti-IL-1RA and -MIP-3 autoantibodies were significantly higher in patients with IgG4-RD compared to healthy subjects. In an independent validation cohort, we detected significantly higher levels of anti-IL-1RA autoantibodies in patients with IgG4-RD compared to healthy subjects with 16% of IgG4-RD patients possessing autoantibodies to IL-1RA. Elevated levels of anti-IL-1RA were correlated with renal involvement. However, high titers of anti-IL-1RA autoantibodies were also detect in IgG4-RD patients with other organ manifestations who did not have overt renal disease. We also observed significantly higher levels of anti-IL-1RA autoantibodies in SLE patients compared to healthy controls, with 28% of the SLE patients possessing autoantibodies to IL-1RA. In Chapter 4, we developed and utilized three in vitro cellular assays to characterize the ability of anti-IL-1RA autoantibodies to antagonize IL-1RA. We demonstrated significantly greater antagonism of IL-1RA in IL-1β reporter cells incubated with plasma from patients with IgG4-RD compared to healthy subjects, with 54% of IgG4-RD patients possessing neutralizing autoantibodies to IL-1RA. We selected plasma samples from three IgG4-RD patients with high titers of anti-IL-1RA and/or neutralizing capacity of IL-1RA in the IL-1β reporter assay to evaluate their ability to inhibit IL-1RA in A549 epithelial cells and MRC-5 fibroblasts. Plasma samples from all three patients inhibited IL-1RA in both cells lines, restoring the mRNA and protein levels of several inflammatory mediators. Treatment of A549 cells with anakinra, an FDA-approved IL-1RA protein therapeutic, in the presence of IL-1α, IL-1RA, and IgG4-RD patient plasma, significantly reduced the protein supernatant levels of inflammation- and fibrosis-associated cytokines. We also identified an anti-murine/human IL-1RA mAb that potently neutralized IL-1RA in vitro, indicating that targeting of a single IL-1RA peptide epitope is able to directly inhibit IL-1RA activity. Moreover, treatment of IL-1α-stimulated A549 cells with IL-1RA and a commercial anti-IL-1RA pAb restored the mRNA and protein levels of immune mediators implicated in pathogenic inflammation and fibrosis. Lastly, we demonstrated high expression levels of IL-1RA at the sites of pathology in IgG4-RD. Taken together, these studies represent the first report of anti-cytokine and anti-IL-1RA autoantibodies in IgG4-RD and autoimmunity, respectively. We demonstrate that anti-IL-1RA autoantibodies from IgG4-RD patients directly inhibit IL-1RA binding to promote the expression and production of pro-inflammatory and -fibrotic mediators implicated in the pathogenesis of this disease. We discovered neutralizing antibodies to IL-1RA in a subset of IgG4-RD patients and demonstrated expression of IL-1RA within inflamed IgG4-RD lesions of multiple organs. These findings provide a potential molecular basis for the diverse organ involvement observed in IgG4-RD. In addition, our discovery of anti-IL-1RA autoantibodies in a subset of patients with SLE suggests that these antibodies may also influence the pathophysiology of other autoimmune diseases with similar clinical features. Ultimately, our investigation of anti-cytokine and anti-IL-1RA autoantibodies provides new mechanistic insights to the pathology of IgG4-RD and autoimmunity towards the development of novel treatments for patients afflicted with these life-altering conditions.
Description
| Type of resource | text |
|---|---|
| Form | electronic resource; remote; computer; online resource |
| Extent | 1 online resource. |
| Place | California |
| Place | [Stanford, California] |
| Publisher | [Stanford University] |
| Copyright date | 2018; ©2018 |
| Publication date | 2018; 2018 |
| Issuance | monographic |
| Language | English |
Creators/Contributors
| Author | Jarrell, Justin Ansel |
|---|---|
| Degree supervisor | Robinson, William (William Hewitt) |
| Thesis advisor | Robinson, William (William Hewitt) |
| Thesis advisor | Bendall, Sean, 1979- |
| Thesis advisor | Davis, Mark M |
| Thesis advisor | Meyer, Everett |
| Degree committee member | Bendall, Sean, 1979- |
| Degree committee member | Davis, Mark M |
| Degree committee member | Meyer, Everett |
| Associated with | Stanford University, Department of Immunology. |
Subjects
| Genre | Theses |
|---|---|
| Genre | Text |
Bibliographic information
| Statement of responsibility | Justin Ansel Jarrell. |
|---|---|
| Note | Submitted to the Department of Immunology. |
| Thesis | Thesis Ph.D. Stanford University 2018. |
| Location | electronic resource |
Access conditions
- Copyright
- © 2018 by Justin Ansel Jarrell
- License
- This work is licensed under a Creative Commons Attribution 3.0 Unported license (CC BY).
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