Figure 5

Parenteral BCG-prime and nApa-subunit-boost by a homologous route impart a stronger specific cellular booster effect in the pulmonary and systemic compartments.

(a) Schematic representation of the prime–boost vaccination regimens employing the homologous parenteral or mucosal routes. Mice were primed once with BCG vaccine by the i.n. or s.c. route. Sixteen months later, mice received two booster doses of nApa (1 μg) in a DDA/MPL adjuvant at a 3-week interval by a homologous route. The BCG-primed mice which were left untreated or boosted (×2) with a saline served as controls. Five weeks after the last subunit-boost, mice (n = 4–5/group) were euthanized and the magnitude and quality of the cellular and humoral immune response was investigated. (b) The nApa- and WCL-specific IFN-γ, IL-17 and IL-4 response in the lung, CLN, spleen or ILN (pooled) using a cultured ELISPOT assay. Data are SFU/10⁶ organ cells and error bars represent s.d. of triplicate cultures. Significant using a one-way ANOVA followed by Bonferroni correction comparing cytokine responses of nApa vaccine-boosted groups with those of respective saline-boosted controls. A number in the parenthesis indicates a fold change in the total cytokine SFU in the nApa-boosted group with a higher response over the nApa-boosted group with a lower response (i.e., comparing the nApa vaccine-boosted two regimens employing different routes). (c,d) The frequency (%) of nApa-specific (c) and WCL-specific (d) IFN-γ or IL-2-producing cells among CD4⁺ and CD8⁺ T lymphocytes of the lung and spleen using a flow cytometry. Data are means + s.e.m. (n = 4–5 individually analyzed mice/group). *P < 0.05; **P < 0.01 and ***P < 0.001 using the ANOVA and Bonferroni correction (in b–d). s.c., subcutaneous; i.n., intranasal; vac, vaccine; WCL, whole cell lysate of Mtb; SFU, spot forming units.