PADs, through a hapten/carrier model. Here, we test this model in mice. Methods HLA-DRB1*04:01 transgenic mice were immunised subcutaneously with PADs or phosphate buffered saline (PBS) in Freund's complete adjuvant (CFA). Three booster injections of PAD or PBS in Freund' incomplete adjuvant (IFA) were given subcutaneously 15, 35 and 55 days later. Mice were: 1. tested for anti-PAD antibodies by ELISA. 2. tested for T cell responses to PADs, native or citrullinated fibrinogen 65 days after PAD immunisation. 3. tested for anti-citrullinated fibrinogen antibodies by ELISA using fibrinogen peptides under citrullinated and native form.Results HLA-DRB1*04:01 transgenic mice immunised with PADs developed antibodies and T cells to PADs and IgG antibodies to citrullinated peptides from fibrinogen, in the absence of T cell response to native or citrullinated fibrinogen. Conclusions T cell immunisation to PAD proteins triggers ACPAs through a hapten carrier mechanism in which the carrier is PAD which performs citrullination and the hapten any protein being citrullinated by PAD. Introduction Differences in enzymatic activity and pathogenic impact of Porphyromonas gingivalis (P.g.) peptidylarginine deiminase (PPAD) for development of RA have been published, confounded by different PPAD variations and methods used. Objectives Enzymatic active PPAD isolated from an RA patient (RA-PPAD) was first time linked to citrullination of RA autoantigens, diagnosis, therapy response and RA-onset. Methods Recombinant RA-PPAD cloned, verified by DNA sequencing and expressed in Escherichia coli and purified. RA-PPAD and its enzymatic activity was analysed using 2D-Elektrophoresis, mass spectrometry (MS), immunoblot and ELISA. Results RA-PPAD autocitrullinates amino acid position 63 (aa 63 ) and exhibits so far two new amino acid mutations aa 73 F to L and aa 447 E to V. Anti-citrullinated RA-PPAD antibodies were detected in 38% (n=36) of patients with RA, but were absent in Systemic lupus erythematosus (n=30), Osteoarthritis (n=36) and control sera (n=23). Twenty percent of RA patients (n=30) showed an increase in antibody-titre against citrullinated RA-PPAD after RA onset. High antibody titre against the cit-PPAD-peptide of 15aa (CPP) derived from the autocitrullination site (R 63 ) correlates with TNFa-inhibitor (TBA) non-response (n=61). Anti-CPP levels correlate with DAS28,rheumatoid factor, a-CCP 2 levels and increase with age. RA-PPAD is able to citrullinate internal arginines in fibrinogen, vimentin, hnRNP-A2/B1 and histone H1. This internal citrullination-sites are recognised by RA sera and able to bind HLA401. Conclusions Failure of P.g. clearance in RA patients may lead to excessive exposure of citrullinated self-antigens and bacterial antigens inducing immune-mimicry. P.g. infection can be linked to RA and its correlation to TBA non-response leads to the suggestion to clear P.g infection before a-TNF treatment. Introduction Genome-wide association studies of multiple autoimmune diseases, including Sjögren's syndrome, systemic lu...
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