1. Autoantibody and T cell responses to oxidative post-translationally modified insulin neoantigenic peptides in type 1 diabetes
- Author
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Rocky Strollo, Chiara Vinci, Y. K. Stella Man, Sara Bruzzaniti, Erica Piemonte, Ghadeer Alhamar, Silvia Irina Briganti, Ilaria Malandrucco, Flavia Tramontana, Chiara Fanali, James Garnett, Roberto Buccafusca, Perrin Guyer, Mark Mamula, Eddie A. James, Paolo Pozzilli, Johnny Ludvigsson, Paul G. Winyard, Mario Galgani, Ahuva Nissim, Strollo, R., Vinci, C., Man, Y. K. S., Bruzzaniti, S., Piemonte, E., Alhamar, G., Briganti, S. I., Malandrucco, I., Tramontana, F., Fanali, C., Garnett, J., Buccafusca, R., Guyer, P., Mamula, M., James, E. A., Pozzilli, P., Ludvigsson, J., Winyard, P. G., Galgani, M., and Nissim, A.
- Subjects
Insulin neoepitope peptide ,Endocrinology, Diabetes and Metabolism ,Oxidative post-translational modification ,Autoimmunity ,Endocrinology and Diabetes ,Immune response ,Insulin ,Insulin autoantibodies ,Neoantigen ,Neoepitope ,Oxidative post-translational modifications ,Post-translational modifications ,Endokrinologi och diabetes ,Internal Medicine ,Insulin autoantibodie - Abstract
Aims/hypothesis Antibodies specific to oxidative post-translational modifications (oxPTM) of insulin (oxPTM-INS) are present in most individuals with type 1 diabetes, even before the clinical onset. However, the antigenic determinants of such response are still unknown. In this study, we investigated the antibody response to oxPTM-INS neoepitope peptides (oxPTM-INSPs) and evaluated their ability to stimulate humoral and T cell responses in type 1 diabetes. We also assessed the concordance between antibody and T cell responses to the oxPTM-INS neoantigenic peptides. Methods oxPTM-INS was generated by exposing insulin to various reactive oxidants. The insulin fragments resulting from oxPTM were fractionated by size-exclusion chromatography further to ELISA and LC-MS/MS analysis to identify the oxidised peptide neoepitopes. Immunogenic peptide candidates were produced and then modified in house or designed to incorporate in silico-oxidised amino acids during synthesis. Autoantibodies to the oxPTM-INSPs were tested by ELISA using sera from 63 participants with new-onset type 1 diabetes and 30 control participants. An additional 18 fresh blood samples from participants with recently diagnosed type 1 diabetes, five with established disease, and from 11 control participants were used to evaluate, in parallel, CD4+ and CD8+ T cell activation by oxPTM-INSPs. Results We observed antibody and T cell responses to three out of six LC-MS/MS-identified insulin peptide candidates: A:12–21 (SLYQLENYCN, native insulin peptide 3 [Nt-INSP-3]), B:11–30 (LVEALYLVCGERGFFYTPKT, Nt-INSP-4) and B:21–30 (ERGFFYTPKT, Nt-INSP-6). For Nt-INSP-4 and Nt-INSP-6, serum antibody binding was stronger in type 1 diabetes compared with healthy control participants (p≤0.02), with oxidised forms of ERGFFYTPKT, oxPTM-INSP-6 conferring the highest antibody binding (83% binders to peptide modified in house by hydroxyl radical [●OH] and >88% to in silico-oxidised peptide; p≤0.001 vs control participants). Nt-INSP-4 induced the strongest T cell stimulation in type 1 diabetes compared with control participants for both CD4+ (p+ (p=0.049). CD4+ response to oxPTM-INSP-6 was also commoner in type 1 diabetes than in control participants (66.7% vs 27.3%; p=0.039). Among individuals with type 1 diabetes, the CD4+ response to oxPTM-INSP-6 was more frequent than to Nt-INSP-6 (66.7% vs 27.8%; p=0.045). Overall, 44.4% of patients showed a concordant autoimmune response to oxPTM-INSP involving simultaneously CD4+ and CD8+ T cells and autoantibodies. Conclusions/interpretation Our findings support the concept that oxidative stress, and neoantigenic epitopes of insulin, may be involved in the immunopathogenesis of type 1 diabetes. Graphical abstract
- Published
- 2023