Figure 2. TATD blocks DQ8-restricted T cell responses and prevents diabetes in NOD mice.
(A) Chemical structure of TATD. (B) TATD blocked an in vitro DQ8–restricted T cell response to insulin B:13-23 (Clone 5) and deamidated α-gliadin228-240 (489 TCR). Data represent the mean ± SEM and are representative of 3 independent experiments. No antigen addition to the culture resulted in IL-2 levels below 3 pg/ml. (C) TATD was cultured with recombinant α-gliadin/DQ8 protein and free peptide in conditions to allow peptide exchange. After washing, the recombinant protein was used to stimulate 489, and IL-2 secretion was measured. Data are from triplicate wells and are representative of 3 independent experiments. 489 in culture without protein resulted in IL-2 levels below 3 pg/ml. (D) Female NOD mice were treated from 4 to 12 weeks of age with 20 mg/kg TATD (n = 10) or PBS (n = 13) by intraperitoneal injection daily for 5 days each week. P = 0.006, by log-rank test. (E) Peak serum insulin autoantibody (IAA) levels were measured using a fluid-phase RIA during the 40-week prevention study. **P = 0.003, by Mann-Whitney U test. The dotted line at 0.01 indicates a positive value. (F) Blood glucose levels during the late prevention study, in which female NOD mice were treated with 30 mg/kg TATD orally each day (n = 9), 50 μg anti-CD3 monoclonal antibody intraperitoneally for 5 consecutive days (n = 10), or PBS (n = 10) beginning at 12 weeks of age and ending at 25 weeks. Data represent the mean ± SEM. *P < 0.05, by ANOVA for comparison of TATD versus PBS. (G) Intraperitoneal GTT following cessation of the study treatments; each dot represents an individual mouse. ***P < 0.01, by 2-tailed, unpaired t test. (H) Representative H&E-stained pancreatic sections from PBS- and TATD-treated mice; insulin staining is shown in brown. Original magnification, ×15. (I) Insulitis scoring from at least 100 separate islets from TATD-treated (n = 3) and PBS-treated (n = 5) mice.