T cell epitope mapping study with insulin overlapping peptides using ELISPOT assay in Japanese children and adolescents with type 1 diabetes.
Type 1 diabetes (T1D) is a T cell-mediated autoimmune disease. Insulin seems to be a critical antigen recognized by autoreactive T cells. In this study, we performed T cell epitope mapping of insulin using serial overlapping peptides in Japanese patients with T1D. Serial overlapping insulin peptides comprising 23 peptides, which were each 15-amino acid long, were prepared based on insulin sequence. Cytokine secretion from peripheral T cells against these peptides was studied by enzyme-linked immunospot (ELISPOT) assay in 18 patients with recent-onset T1D and 12 patients with established T1D, and compared with 17 healthy control subjects. In ELISPOT assay, IFN-gamma-secreting T cells, but not IL-4, against several insulin peptides were observed in 77.8% of patients with recent-onset T1D, 50.0% of patients with established T1D, and 0% of healthy control subjects. All epitopes recognized by T cells were identified in the B-chain of insulin. The most frequent epitope existed at the B10-24 region (9/18), followed by B1-15 and B11-25 regions (6/18, each), with B4-18, B9-23, and B12-26 identified in some patients. These data did not correlate with insulin autoantibodies or HLA-DRB1 of the patients. This is the first report of T cell epitope mapping using one amino acid serial overlapping peptides of insulin in T1D. ELISPOT assay revealed the frequent existence of insulin peptide-specific T cells in patients with recent-onset and established T1D. The T cell epitopes of insulin were similar but not identical in our cohort, which probably explains the difficulty encountered in prevention of human T1D by using insulin.
Higashide
Takashi
T
Department of Pediatrics, Osaka City University Graduate School of Medicien, Osaka, Japan.
Kawamura
Tomoyuki
T
Nagata
Masao
M
Kotani
Reiko
R
Kimura
Kayo
K
Hirose
Masakazu
M
Inada
Hiroshi
H
Niihira
Shizuhiro
S
Yamano
Tsunekazu
T
D000293:Adolescent; D000818:Animals; D049109:Cell Proliferation; D002648:Child; D002675:Child, Preschool; D003470:Culture Media / Q000737:chemistry; D016207:Cytokines / Q000378:metabolism; D003922:Diabetes Mellitus, Type 1 / Q000235:genetics / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D018604:Epitope Mapping; D005260:Female; D006680:HLA Antigens; D006801:Humans; D007223:Infant; D007231:Infant, Newborn; D007328:Insulin / Q000235:genetics / Q000378:metabolism*; D007564:Japan; D008297:Male; D051379:Mice; D010455:Peptides / Q000235:genetics / Q000378:metabolism*; D013601:T-Lymphocytes / Q000166:cytology / Q000502:physiology*
D003470:Culture Media; D016207:Cytokines; D006680:HLA Antigens; D007328:Insulin; D010455:Peptides