Figure 2.

Responsiveness and peripheral frequency of beta cell–specific CD8⁺ T cells in 8.3-NOD mice. (A) Proliferation of splenic CD8⁺ T cells from 8.3-NOD and 8.3–TCR-β–transgenic NOD mice to islet cells. 2 × 10⁴ splenic CD8⁺ T cells were incubated with γ-irradiated islet cells for 3 d, pulsed with [³H]thymidine, harvested, and counted. Bars show the standard error of the means. (B) Peripheral frequency of beta cell–reactive CD8⁺ T cells in 8.3-NOD and 8.3–TCR-β–transgenic NOD mice. 12 replicate cultures of serial dilutions of splenocytes (10¹– 10⁵ cells/well) were stimulated with irradiated NOD islets (8/well) for 4 d, expanded in rIL-2 (0.5 U/ml) for 10 d and restimulated once with islets and rIL-2. The cultures were then challenged with 10⁴ NIT-1 or L929-Kd cells for 24 h, and the supernatants collected to measure their TNF-α content. Cultures that secreted TNF-α in response to NIT-1, but not L929-Kd, cells were considered to contain beta cell–reactive CD8⁺ T cells. (C) General proliferative activity of splenic CD8⁺ T cells of 8.3-NOD and 8.3–TCR-β–transgenic NOD mice. 2 × 10⁴ splenic CD8⁺ T cells were incubated with 10-fold serial dilutions of plate-bound KJ16 in rIL-2–containing CM for 3 d, pulsed with [³H]thymidine, harvested, and counted.