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Characterization and use of the ECV304 autoantigenic citrullinome to understand anti-citrullinated protein/peptide autoantibodies in rheumatoid arthritis.
de França, Natalia Regine; Ménard, Henri André; Lora, Maximilien; Zhou, Zhijie; Rauch, Joyce; Hitchon, Carol; Andrade, Luís Eduardo Coelho; Colmegna, Inés.
Afiliação
  • de França NR; Division of Rheumatology, Department of Medicine, McGill University, The Research Institute of the McGill University Health Centre, 1001 Décarie Boulevard, Montréal, QC, H4A 3J1, Canada.
  • Ménard HA; Division of Rheumatology, Paulista School of Medicine, Federal University of Sao Paulo, Sao Paulo, Brazil.
  • Lora M; Division of Rheumatology, Department of Medicine, McGill University, The Research Institute of the McGill University Health Centre, 1001 Décarie Boulevard, Montréal, QC, H4A 3J1, Canada.
  • Zhou Z; Division of Rheumatology, Department of Medicine, McGill University, The Research Institute of the McGill University Health Centre, 1001 Décarie Boulevard, Montréal, QC, H4A 3J1, Canada.
  • Rauch J; Division of Rheumatology, Department of Medicine, McGill University, The Research Institute of the McGill University Health Centre, 1001 Décarie Boulevard, Montréal, QC, H4A 3J1, Canada.
  • Hitchon C; Division of Rheumatology, Department of Medicine, McGill University, The Research Institute of the McGill University Health Centre, 1001 Décarie Boulevard, Montréal, QC, H4A 3J1, Canada.
  • Andrade LEC; Section of Rheumatology, Department of Medicine, University of Manitoba, Winnipeg, MB, Canada.
  • Colmegna I; Division of Rheumatology, Paulista School of Medicine, Federal University of Sao Paulo, Sao Paulo, Brazil.
Arthritis Res Ther ; 24(1): 23, 2022 01 13.
Article em En | MEDLINE | ID: mdl-35027076
BACKGROUND: Anti-citrullinated protein antibodies (ACPAs) are highly specific for rheumatoid arthritis (RA). In vivo, ACPAs target peptidyl-citrulline epitopes (cit-) in a variety of proteins (cit-prot-ACPAs) and derived peptides (cit-pept-ACPAs) generated via the peptidylarginine deiminase (PAD) isoenzymes. We aimed to identify a cell line with self-citrullination capacity, to describe its autoantigenic citrullinome, and to test it as a source of autocitrullinated proteins and peptides. METHODS: Human cell lines were screened for cit-proteins by Western blot. PAD isoenzymes were identified by RT-PCR. Autocitrullination of ECV304 was optimized, and the ECV304 autocitrullinomes immunoprecipitated by sera from three RA patients were characterized by mass spectrometry. Cit-pept-ACPAs were detected using anti-CCP2 ELISA and cit-prot-ACPAs, by an auto-cit-prot-ECV304 ELISA. Sera from 177 RA patients, 59 non-RA rheumatic disease patients and 25 non-disease controls were tested. RESULTS: Of the seven cell lines studied, only ECV304 simultaneously overexpressed PAD2 and PAD3 and its extracts reproducibly autocitrullinated self and non-self-proteins. Proteomic analysis of the cit-ECV304 products immunoprecipitated by RA sera, identified novel cit-targets: calreticulin, profilin 1, vinculin, new 14-3-3 protein family members, chaperones, and mitochondrial enzymes. The auto-cit-prot-ECV304 ELISA had a sensitivity of 50% and a specificity of 95% for RA diagnosis. CONCLUSIONS: ECV304 cells overexpress two of the PAD isoenzymes capable of citrullinating self-proteins. These autocitrullinated cells constitute a basic and clinical research tool that enable the detection of cit-prot-ACPAs with high diagnostic specificity and allow the identification of the specific cit-proteins targeted by individual RA sera.
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Texto completo: 1 Base de dados: MEDLINE Assunto principal: Artrite Reumatoide / Autoanticorpos Idioma: En Ano de publicação: 2022 Tipo de documento: Article

Texto completo: 1 Base de dados: MEDLINE Assunto principal: Artrite Reumatoide / Autoanticorpos Idioma: En Ano de publicação: 2022 Tipo de documento: Article