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T Cells Recognize Multiple GAD65 and Proinsulin Epitopes in Human Type 1 Diabetes, Suggesting Determinant Spreading

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Abstract

Human type 1 diabetes is thought to be mediated by autoreactive T cells specific for antigens expressed by pancreatic beta cells. However, it is unclear which autoantigens and determinants thereof are the targets of the autoimmune attack. Using comprehensive peptide libraries that cover the entire sequence of two major candidate autoantigens, GAD65 and proinsulin, we measured the in vivo frequencies of peptide-specific, IFN-γ-producing memory T cells in 27 diabetic patients, 14 high risk individuals, and 15 partially HLA-matched healthy controls. Compared to the controls, both a higher number of determinants on the islet cell antigens were recognized and the frequencies of peptide specific cells were increased in patients and high risk individuals. Inclusion of signal enhancing anti-CD28 antibody further accentuated this difference. Considerable heterogeneity in peptide recognition was seen even in DRB1*04, DQB1*0302 matched individuals. Unlike its peptides, the GAD protein antigen did not recall a T cell memory response. The highly heterogeneous recognition of a multitude of peptide determinants on both autoantigens, occurring in the absence of protein recognition, and the low functional avidity of the memory cells involved jointly suggest that the autoimmune T cell repertoire in human type 1 diabetes primarily targets cryptic determinants engaged by determinant spreading.

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REFERENCES

  1. Bonifacio E, Bingley PJ, Shattock M, Dean BM, Dunger D, Gale EA, Bottazzo GF: Quantification of islet-cell antibodies and prediction of insulin-dependent diabetes. Lancet 335:147–149, 1990

    Article  PubMed  Google Scholar 

  2. Verge CF, Gianani R, Kawasaki E, Yu L, Pietropaolo M, Jackson RA, Chase HP, Eisenbarth GS: Prediction of type I diabetes in first-degree relatives using a combination of insulin, GAD, and ICA512bdc/IA-2 autoantibodies. Diabetes 45:926–933, 1996

    PubMed  Google Scholar 

  3. Seissler J, Hatziagelaki E, Scherbaum WA: Modern concepts for the prediction of type 1 diabetes. Exp Clin Endocrinol Diabetes 109:S304–S316, 2001

    PubMed  Google Scholar 

  4. Leslie RD, Atkinson MA, Notkins AL: Autoantigens IA-2 and GAD in type I (insulin-dependent) diabetes. Diabetologia 42:3–14, 1999

    PubMed  Google Scholar 

  5. Greeley SA, Katsumata M, Yu L, Eisenbarth GS, Moore DJ, Goodarzi H, Barker CF, N aji A, Noorchashm H: Elimination of ma-ternally transmitted autoantibodies prevents diabetes in nonobese diabetic mice. Nat Med 8:399–402, 2002

    PubMed  Google Scholar 

  6. Bendelac A, Carnaud C, Boitard C, Bach JF: Syngeneic trans-fer of autoimmune diabetes from diabetic NOD mice to healthy neonates. Requirement for both L3T4+ and Lyt-2+ T cells. J Exp Med 166:823–832, 1987

    PubMed  Google Scholar 

  7. Baekkeskov S, Kanaani J, Jaume JC, Kash S: Does GAD have a unique role in triggering IDDM? J Autoimmun 15:279–286, 2000 8. Wegmann DR, Eisenbarth GS: It's insulin. J Autoimmun 15:286–291, 2000

    PubMed  Google Scholar 

  8. Kaufman DL, Clare-Salzler M, Tian J, Forsthuber T, Ting GS, Robinson P, Atkinson MA, Sercarz EE, Tobin AJ, Lehmann PV: Spontaneous loss of T-cell tolerance to glutamic acid decar-boxylase in murine insulin-dependent diabetes. Nature 366:69–72, 1993

    PubMed  Google Scholar 

  9. Tisch R, Yang XD, Singer SM, Liblau RS, Fugger L, McDevitt HO: Immune response to glutamic acid decarboxylase correlates with insulitis in non-obese diabetic mice. Nature 366:72–75, 1993

    PubMed  Google Scholar 

  10. Tian J, Atkinson MA, Clare-Salzler M, Herschenfeld A, Forsthuber T, Lehmann PV, Kaufman DL: Nasal administration of glutamate decarboxylase (GAD65) peptides induces Th2 responses and pre-vents murine insulin-dependent diabetes. J Exp Med 183:1561–1567, 1996

    PubMed  Google Scholar 

  11. Elliott JF, Qin HY, Bhatti S, Smith DK, Singh RK, Dillon T, Lauzon J, Singh B: Immunization with the larger isoform of mouse glutamic acid decarboxylase (GAD67) prevents autoimmune diabetes in NOD mice. Diabetes 43:1494–1499, 1994

    PubMed  Google Scholar 

  12. Yoon JW, Yoon CS, Lim HW, Huang QQ, Kang Y, Pyun KH, Hirasawa K, Sherwin RS, Jun HS: Control of autoimmune diabetes in NOD mice by GAD expression or suppression in beta cells. Science 284:1183–1187, 1999

    PubMed  Google Scholar 

  13. Zekzer D, Wong FS, Ayalon O, Millet I, Altieri M, Shintani S, Solimena M, Sherwin RS: GAD-reactive CD4 C Th1 cells induce diabetes in NOD/SCID mice. J Clin Invest 101:68–73, 1998

    PubMed  Google Scholar 

  14. Wong FS, Karttunen J, Dumont C, Wen L, Visintin I, Pilip IM, Shastri N, Pamer EG, Janeway CA, Jr: Identification of an MHCclass I-restricted autoantigen in type 1 diabetes by screening an organ-specific cDNA library. Nat Med 5:1026–1031, 1999

    PubMed  Google Scholar 

  15. Daniel D, Wegmann DR: Protection of nonobese diabetic mice from diabetes by intranasal or subcutaneous administration of in-sulin peptide B-(9-23). Proc Natl Acad Sci U.S.A 93:956–960, 1996

    PubMed  Google Scholar 

  16. Muir A, Peck A, Clare-Salzler M, Song YH, Cornelius J, Luchetta R, Krischer J, Maclaren N: Insulin immunization of nonobese dia-betic mice induces a protective insulitis characterized by diminished intraislet interferon-gamma transcription. J Clin Invest 95:628–634, 1995

    PubMed  Google Scholar 

  17. Haskins K, McDuffie M: Acceleration of diabetes in young NOD mice with a CD4 C islet-specific T cell clone. Science 249:1433–1436, 1990

    PubMed  Google Scholar 

  18. Nepom GT, Lippolis JD, White FM, Masewicz S, Marto JA, Herman A, Luckey CJ, Falk B, Shabanowitz J, Hunt DF, Engelhard VH, Nepom BS: Identification and modulation of a naturally processed T cell epitope from the diabetes-associated autoantigen human glu-tamic acid decarboxylase 65 (Hgad65). Proc Natl Acad Sci U.S.A 98:1763–1768, 2001

    PubMed  Google Scholar 

  19. Reijonen H, Elliott JF, van Endert P, Nepom G: Differential pre-sentation of glutamic acid decarboxylase 65 (GAD65) T cell epi-topes among HLA-DRB1*0401-positive individuals. J Immunol 163:1674–1681, 1999

    PubMed  Google Scholar 

  20. Alleva DG, Crowe PD, Jin L, Kwok WW, Ling N, Gottschalk M, Conlon PJ, Gottlieb PA, Putnam AL, Gaur A: A disease-associated cellular immune response in type 1 diabetics to an immunodominant epitope of insulin. J Clin Invest 107:173–180, 2001

    PubMed  Google Scholar 

  21. Schlosser M, Strebelow M, Wassmuth R, Arnold ML, Breunig I, Rjasanowski I, Ziegler B, Ziegler M: The Karlsburg type 1 diabetes risk study of a normal schoolchild population: Association of beta-cell autoantibodies and human leukocyte antigen-DQB1 alleles in antibody-positive individuals. J Clin Endocrinol Metab 87:2254–2261, 2002

    PubMed  Google Scholar 

  22. Erlich H, Bugawan T, Begovich AB, Scharf S, Griffith R, Saiki R, Higuchi R, Walsh PS: HLA-DR, DQ and DP typing using PCR amplification and immobilized probes. Eur J Immunogenet. 18:33–55, 1991

    PubMed  Google Scholar 

  23. Geysen HM, Rodda SJ, Mason TJ, Tribbick G, Schoofs PG: Strate-gies for epitope analysis using peptide synthesis. J Immunol Methods 102:259–274, 1987

    PubMed  Google Scholar 

  24. Gammon G, Geysen HM, Apple RJ, Pickett E, Palmer M, Ametani A, Sercarz EE: T cell determinant structure: Cores and determinant envelopes in three mouse major histocompatibility complex haplo-types. J Exp Med 173:609–617, 1991

    PubMed  Google Scholar 

  25. Helms T, Boehm BO, Asaad RJ, Trezza RP, Lehmann PV, Tary-Lehmann M: Direct visualization of cytokine-producing recall antigen-specific CD4 memory T cells in healthy individuals and HIV patients. J Immunol 164:3723–3732, 2000

    PubMed  Google Scholar 

  26. Semana G, Gausling R, Jackson RA, Hafler DA: T cell autoreactivity to proinsulin epitopes in diabetic patients and healthy subjects. J Autoimmun 12:259–267, 1999

    PubMed  Google Scholar 

  27. Lohmann T, Leslie RD, Hawa M, Geysen M, Rodda S, Londei M: Immunodominant epitopes of glutamic acid decarboxylase 65 and 67 in insulin-dependent diabetes mellitus. Lancet 343:1607–1608, 1994

    PubMed  Google Scholar 

  28. Schloot NC, Roep BO, Wegmann DR, Yu L, Wang TB, Eisenbarth GS: T-cell reactivity to GAD65 peptide sequences shared with coxsackie virus protein in recent-onset IDDM, post-onset IDDM patients and control subjects. Diabetologia 40:332–338, 1997

    PubMed  Google Scholar 

  29. Manca F, Habeshaw J, Dalgleish A: The naive repertoire of human T helper cells specific for gp120, the envelope glycoprotein of HIV. J Immunol 146:1964–1971, 1991

    PubMed  Google Scholar 

  30. Roep BO, Atkinson MA, van Endert PM, Gottlieb PA, Wilson SB, Sachs JA: Autoreactive T cell responses in insulin-dependent (Type 1) diabetes mellitus. Report of the first international workshop for standardization of T cell assays. J Autoimmun 13:267–282, 1999

    PubMed  Google Scholar 

  31. Panina-Bordignon P, Lang R, van Endert PM, Benazzi E, Felix AM, Pastore RM, Spinas GA, Sinigaglia F: Cytotoxic T cells specific for glutamic acid decarboxylase in autoimmune diabetes. J Exp Med 181:1923–1927, 1995

    PubMed  Google Scholar 

  32. Reijonen H, Novak EJ, Kochik S, Heninger A, Liu AW, Kwok WW, Nepom GT: Detection of GAD65-specific T-cells by major histo-compatibility complex class II tetramers in type 1 diabetic patients and at-risk subjects. Diabetes 51:1375–1382, 2002

    PubMed  Google Scholar 

  33. Viglietta V, Kent SC, Orban T, Hafler DA: GAD65-reactive T cells are activated in patients with autoimmune type 1a diabetes. J Clin Invest 109:895–903, 2002

    PubMed  Google Scholar 

  34. Eisenbarth GS, Kotzin BL: Enumerating autoreactive T cells in peripheral blood: A big step in diabetes prediction. J Clin Invest 111:179–181, 2003

    PubMed  Google Scholar 

  35. Wicker LS, Chen SL, Nepom GT, Elliott JF, Freed DC, Bansal A, Zheng S, Herman A, Lernmark A, Zaller DM, Peterson LB, Roth-bard JB, Cummings R, Whiteley PJ: Naturally processed T cell epi-topes from human glutamic acid decarboxylase identified using mice transgenic for the type 1 diabetes-associated human MHC class II allele, DRB1*0401. J Clin Invest 98:2597–2603, 1996

    PubMed  Google Scholar 

  36. Endl J, Otto H, Jung G, Dreisbusch B, Donie F, Stahl P, Elbracht R, Schmitz G, Meinl E, Hummel M, Ziegler AG, Wank R, Schendel DJ: Identification of naturally processed T cell epitopes from glutamic acid decarboxylase presented in the context of HLA-DR alleles by T lymphocytes of recent onset IDDM patients. J Clin Invest 99:2405–2415, 1997

    PubMed  Google Scholar 

  37. Peakman M, Tree TI, Endl J, van Endert P, Atkinson MA, Roep BO: Characterization of preparations of GAD65, proinsulin, and the islet tyrosine phosphatase IA-2 for use in detection of autoreactive T-cells in type 1 diabetes: Report of phase II of the second international immunology of diabetes society workshop for standardization of T-cell assays in type 1 diabetes. Diabetes 50:1749–1754, 2001

    PubMed  Google Scholar 

  38. Gebauer BS, Hricik DE, Atallah A, Bryan K, Riley J, Tary-Lehmann M, Greenspan NS, Dejelo C, Boehm BO, Hering BJ, Heeger PS: Evolution of the enzyme-linked immunosorbent spot assay for post-transplant alloreactivity as a potentially useful immune moni-toring tool. Am J Transplant </del>2:857–866, 2002

    PubMed  Google Scholar 

  39. Targoni OS, Lehmann PV: Endogenous myelin basic protein inacti-vates the high avidity T cell repertoire. J Exp Med 187:2055–2063, 1998

    PubMed  Google Scholar 

  40. Lehmann PV, Targoni OS, Forsthuber TG: Shifting T-cell activation thresholds in autoimmunity and determinant spreading. Immunol Rev 164:53–61, 1998

    PubMed  Google Scholar 

  41. Harrington CJ, Paez A, Hunkapiller T, Mannikko V, Brabb T, Ahearn M, Beeson C, Goverman J: Differential tolerance is induced in T cells recognizing distinct epitopes of myelin basic protein. Immunity 8:571–580, 1998

    PubMed  Google Scholar 

  42. Sercarz EE, Lehmann PV, Ametani A, Benichou G, Miller A, Moudgil K: Dominance and crypticity of T cell antigenic deter-minants. Annu Rev Immunol 11:729–766, 1993

    Google Scholar 

  43. Mor F, Cohen IR: Pathogenicity of T cells responsive to diverse cryptic epitopes of myelin basic protein in the Lewis rat. J Immunol 155:3693–3699, 1995

    PubMed  Google Scholar 

  44. Lehmann PV, Forsthuber T, Miller A, Sercarz EE: Spreading of T-cell autoimmunity to cryptic determinants of an autoantigen. Nature 358:155–157, 1992

    Google Scholar 

  45. Miller SD, Mcrae BL, Vanderlugt CL, Nikcevich KM, Pope JG, Pope L, Karpus WJ: Evolution of the T-cell repertoire during the course of experimental immune-mediated demyelinating diseases. Immunol Rev 144:225–244, 1995

    PubMed  Google Scholar 

  46. Yu L, Robles DT, Abiru N, Kaur P, Rewers M, Kelemen K, Eisenbarth GS: Early expression of antiinsulin autoantibodies of humans and the NOD mouse: Evidence for early determination of subsequent diabetes. Proc Natl Acad Sci U.S.A 97:1701–1706, 2000

    PubMed  Google Scholar 

  47. Bonifacio E, Lampasona V, Bernasconi L, Ziegler AG: Matura-tion of the humoral autoimmune response to epitopes of GAD in preclinical childhood type 1 diabetes. Diabetes 49:202–208, 2000

    PubMed  Google Scholar 

  48. Naserke HE, Ziegler AG, Lampasona V, Bonifacio E: Early devel-opment and spreading of autoantibodies to epitopes of IA-2 and their association with progression to type 1 diabetes. J Immunol 161:6963–6969, 1998

    PubMed  Google Scholar 

  49. Brooks-Worrell B, Gersuk VH, Greenbaum C, Palmer JP: Inter-molecular antigen spreading occurs during the preclinical period of human type 1 diabetes. J Immunol 166:5265–5270, 2001

    PubMed  Google Scholar 

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Authors and Affiliations

  1. Department of Pathology, Case Western Reserve University, Cleveland, Ohio

    Patrick A. Ott, Marcus T. Dittrich, Bernhard A. Herzog, Robert Guerkov, Magdalena Tary-Lehmann & Paul V. Lehmann

  2. Section of Endocrinology, University Hospital of Ulm, Ulm, Germany

    Marcus T. Dittrich, Bernhard A. Herzog, Ivana Durinovic-Bello & Bernhard O. Boehm

  3. Barbara Davis Center of Childhood Diabetes, University of Colorado Health Sciences Center, Denver, Colarado

    Peter A. Gottlieb & Amy L. Putnam

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Ott, P.A., Dittrich, M.T., Herzog, B.A. et al. T Cells Recognize Multiple GAD65 and Proinsulin Epitopes in Human Type 1 Diabetes, Suggesting Determinant Spreading. J Clin Immunol 24, 327–339 (2004). https://doi.org/10.1023/B:JOCI.0000029120.77824.41

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