Figure 5.

ATG combinations with tacrolimus or rapamycin control humoral response induced by ATG treatment alone. (A) C57Bl/6 mice were subcutaneously immunized with NP-OVA + CFA and treated with 500μg of murine ATG or IgG control. A subgroup then received either 1 mg/Kg of tacrolimus or 0.5 mg/Kg of rapamycin, intraperitoneally, daily. Mice were euthanized at day 8 after immunization, and serum and lymph nodes were analyzed, as shown in Fig 2A. (B) Representative contour plots of Tfh (CD4+CXCR5+ PD-1+) cells in lymph nodes gated in CD4+ cells. (C) The frequency and absolute cell number per lymph node of Tfh cells. (D) Representative contour plots of GC B (B220+GL-7+ Fas+) cells in lymph nodes gated in B220+ cells. (E) The frequency and absolute cell number per lymph node of GC B cells. (F) Representative contour plots of B220+ IgG1+ cells in lymph nodes gated in B220+ cells. (G) The frequency and absolute cell number per lymph node of IgG1+ B cells. ELISA quantification of serum (H) NP-specific and (I) OVA-Specific IgG antibodies at day 8 after immunization. (A-I) Red dots represent the ATG-treated mice, black dots represent the IgG-treated mice, blue dots represent the ATG + tacrolimus-treated mice, and green dots represent the ATG + rapamycin-treated mice. Data as mean ± SD are shown (data from three independent experiments; with n = 4 per group; One-way ANOVA with Tukey multiple comparisons test).