FIG. 4.
Increased protective immunity to L. major in Bim−/− mice. (A) Experimental design of infection, challenge, and analysis. Briefly, groups of either Bim+/+ or Bim−/− mice (three mice/group) were infected intradermally in the left ear with 3 × 103 L. major promastigotes. Four months after this primary infection, mice from each group were challenged in the right ear with 3 × 103 L. major promastigotes. As a control, naïve Bim+/+ and Bim−/− mice (no primary infection) were challenged at the same time. Three weeks after this challenge, mice were sacrificed and the numbers of parasites at the chronic site (left ear) and the challenge site (right ear) and in the LNs draining either site were quantified. (B and C) Numbers of parasites (expressed as 1/dilution with detectable parasites) in either (B) the ear or (C) the draining LNs. The symbols indicate values for individual mice as follows: naïve mice, Bim+/+ (squares) and Bim−/− (circles); chronic and challenged mice, Bim+/+ (open squares and circles) and Bim−/− (filled squares and circles). The dotted lines indicate the limit of detection. The Bim−/− mice exhibited a significant reduction in the parasite burden at the challenge site (indicated by an asterisk; P < 0.037 for ears and P < 0.025 for LNs, Student's t test) compared to the naïve Bim−/− controls. In addition, in Bim−/− mice there were significant reductions in the numbers of parasites at the chronic site (P < 0.005 for ears and P < 0.033 for LNs) and at the challenge site (P < 0.032 for ears and P < 0.043 for LNs) compared to Bim+/+ controls. The numbers of parasites in challenged Bim+/+ mice were not significantly different from the numbers in naïve Bim+/+ controls (P < 0.160 for ears and P < 0.29 for LNs). n.s., not significant. (D) Representative dot plots showing IFN-γ production by CD4+ T cells of naïve mice (3 weeks after infection) or at the chronic site (19 weeks after infection) and challenge site (3 weeks after challenge) of either Bim+/+ (WT) or Bim−/− mice. (E) Total numbers of IFN-γ+ CD4+ T cells in Bim+/+ mice (open bars) and Bim−/− mice (filled bars) at either the chronic site or the challenged site. The error bars indicate the standard errors of the means. A significant increase in the number of IFN-γ+ cells at the challenge site was observed in Bim−/− mice compared to Bim+/+ mice (P < 0.03, Student's t test). The data are representative of two independent experiments in which similar results were obtained.