Figure 6.

BCG vaccination promotes predominantly Th1 responses. (A) Wildtype mice were adoptively transferred with 4 x 10⁴ T cells purified from P25 TCR-Tg GFP⁺ mice 16 hours prior to vaccination with BCG (B), or peptide-25 adjuvanted with either alum (A) or LASTS-C (LC). The mice were sacrificed 6 days after vaccination, and splenocytes (n=5) were analyzed by FACS for master transcription regulators and key markers for Th1 (Tbet) and Tfh (CXCR-5 and Bcl-6). Multiple comparisons were analyzed by Kruskal-Wallis one-way ANOVA (**p < 0.01, ***p < 0.001). (B) Wildtype mice were adoptively transferred with 4 x 10⁴ T cells purified from P25 TCR-Tg GFP⁺ mice 16 hours prior to vaccination with BCG or with the P25 peptide adjuvanted with LASTS-C. Formalin-fixed and paraffin-embedded spleens were cut into thin sections for immunocytochemistry with anti-GFP followed by hematoxylin and eosin counterstaining. (C) High magnification of boxed areas in (B) to visualize the P25 specific T cells as dark colored spots. Follicles (F), germinal centers (GC), and P25 Th cells were quantified manually by a blinded observer as the number of GC per follicle in (B), and the number of P25 Th cells per GC in (C). Medians with interquartile ranges for 6-7 sections derived from 3 mice from each group are plotted. Mann-Whitney test was used for pairwise comparison (*p < 0.05, **p < 0.01, ***p < 0.001).