Immunogenic HLA-DR-Presented Self-Peptides Identified Directly from Clinical Samples of Synovial Tissue, Synovial Fluid, or Peripheral Blood in Patients with Rheumatoid Arthritis or Lyme Arthritis

Abstract

HLA-DR-molecules are highly expressed in synovial tissue (ST), the target of the immune response in chronic inflammatory forms of arthritis. Here, we used LC-MS/MS to identify HLA-DR-presented self-peptides in cells taken directly from clinical samples: ST, synovial fluid mononuclear cells (SFMC), or peripheral blood mononuclear cells (PBMC) from five patients with rheumatoid arthritis (RA) and eight with Lyme arthritis (LA). We identified 1,593 non-redundant HLA-DR-presented peptides, derived from 870 source proteins. Sixty-seven percent of the peptides identified in SFMC and 55% of those found in PBMC were found in ST, but analysis of SFMC/PBMC also revealed new antigen-presented peptides. Peptides were synthesized and examined for reactivity with the patients’ PBMC. To date, three autoantigens in RA and four novel autoantigens in LA, presented in ST and/or PBMC, were shown to be targets of T- and B-cell responses in these diseases; ongoing analyses may add to this list. Thus, immunoprecipitation and LC-MS/MS can now identify hundreds of HLA-DR-presented self-peptides from individual patients’ tissues or fluids with mixed cell populations. Importantly, identification of HLA-DR-presented peptides from SFMC or PBMC allows testing of more patients, including those early in the disease. Direct analysis of clinical samples facilitates identification of novel immunogenic T-cell epitopes. Data are available via ProteomeXchange (identifier PXD003051).

Interferon‐gamma production in Lyme arthritis synovial tissue promotes differentiation of fibroblast‐like synoviocytes into immune effector cells

“…Based on an analysis of HLA‐DR‐presented peptides in LA synovial tissue (Wang et al, ), we previously identified four LA‐associated autoantigens: EC growth factor, matrix metalloproteinase‐10, apolipoprotein B‐100, and annexin A2 (Crowley et al, ; Crowley et al, ; Drouin et al, ; Pianta et al, ). These four self‐proteins are abundant in postinfectious LA synovial tissue and are involved in tissue damage and/or wound repair.…”

Two rheumatoid arthritis–specific autoantigens correlate microbial immunity with autoimmune responses in joints

“…A detailed description of the methods for the isolation and identification of HLA-DR-presented peptides is given in our previous publications (19,20). In the current study, all nonre-jci.org…”

“…In a recent study (20), we identified HLA-DR-presented peptides in synovial tissue, SFMCs, and PBMCs from 5 patients with RA using LC-MS/MS, and the immunogenicity of the peptides was determined using patients' samples in enzyme-linked immunospot (ELISpot) assays. The findings from 1 patient (referred to here as RA1) were of particular interest.…”

“…In patient RA1, 2 immunogenic HLA-DR-presented human self-peptides derived from GNS and FLNA were also identified from her synovial tissue, and the same FLNA peptide was also found in her PBMCs (20). The HLA-DR-presented peptide derived from GNS was predicted to be promiscuous, binding to 24 of the 25 HLA-DR molecules modeled in the program TEPITOPE (26), (27).…”

“…We have developed an approach for antigen detection in chronic inflammatory arthritides, in which HLA-DRpresented peptides (T cell epitopes) are identified directly from patients' inflamed synovial tissue, synovial fluid mononuclear cells (SFMCs), or peripheral blood mononuclear cells (PBMCs) by liquid chromatography-tandem mass spectrometry (LC-MS/MS) and tested for immunogenicity using patients' samples (19)(20)(21)(22)(23)(24).…”