FIGURE 3.

Mucosal delivery of eDQ8d epitopes by L. lactis significantly decreases the DQ8d-induced DTH response and proliferative capacity of bulk spleen and inguinal lymph node cells and lamina propria cells. NOD AB° DQ8 transgenic mice were immunized by s.c. injection of 100 μg eDQ8d in CFA at day 1. Mice were orally treated with LL-eDQ8d or LL-pT1NX at days 1–10. Control mice received BM9. At day 10, mice were challenged with 10 μg eDQ8d in 10 μl saline in the auricle of the ear. DTH responses are expressed as the mean (±SEM) increase in ear thickness from baseline, 24 hours after injection (A). After the DTH measurements, spleens, inguinal lymph nodes, and lamina propria cells of the BM9 (control), LL-pT1NX and LL-eDQ8d groups were isolated and ex vivo stimulated with 50 μg/ml eDQ8d peptide or 50 μg/ml irrelevant peptide (irrel) (white bars: 3b–3d). eDQ8d-specific proliferative response of bulk splenocytes (P=0.048) (B) and inguinal lymph node cells (P=0.002) (C) and lamina propria cells (P=0.044) (D) were studied by thymidine incorporation, expressed as the mean (±SEM) cpm.

Cytokine measurements in the supernatant of splenocytes (3e-3f) and inguinal lymph node cells (3g) were performed 24 hours after ex vivo eDQ8d (black bars) or irrelevant peptide (irrel)(white bars) stimulation. Results represent the mean (±SEM) of cytokine secretion in pg/ml for at least two individual experiments including 6 mice in each group. ▼ indicates not detected (below detection threshold).