Tolerant anti-CD3–treated BALB/c and C57BL/6 mice are polarized toward a Th1 cytokine profile. Splenocytes were pooled from anti-CD3–treated, HBSS-treated, or untreated animals (n = 2-3/treatment group) 1 week after the final of 4 FVIII immunizations (1 of 3 and 1 of 2 representative experiments for BALB/c and C57BL/6 mice, respectively). Cells were restimulated in vitro with a combination of anti-CD3 and anti-CD28 mAbs, FVIII, or were not stimulated (unstimulated). Both the BALB/c and C57BL/6 mice treated with anti-CD3 (in vivo) were able to produce Th1 cytokines in vitro (IL-2 and IFN-γ; A) but were limited in their ability to secrete Th2 cytokines (IL-4, IL-5, and IL-10) compared with HBSS-treated and untreated animals after antigen-specific restimulation (B). “Anti-CD3 (tol.)” and “anti-CD3 (nt)” refer to anti-CD3–treated C57BL/6 mice inhibitor negative or positive, respectively, 1 week after FVIII immunizations. *P < .01, **P < 1.0 × 10−5 for anti-CD3 compared with FVIII control; +P < .02, ++P < .002, +++P < 1.0 × 10−4 for anti-CD3 (tol.) compared with anti-CD3 (nt).
