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.
Department of Pediatrics, Osaka City University Graduate School of Medicien, Osaka, Japan.
Takashi
T
Higashide
Tomoyuki
T
Kawamura
Masao
M
Nagata
Reiko
R
Kotani
Kayo
K
Kimura
Masakazu
M
Hirose
Hiroshi
H
Inada
Shizuhiro
S
Niihira
Tsunekazu
T
Yamano
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