Figure captions
Figure. 1 Effect of recombinant TSA-1-C4 plus Tc24-C4 combination on NO and H2O2 production by peritoneal macrophages. Peritoneal cells isolated from naïve mice were cultured in supplemented DMEM for 48 h, non-adherent cells were removed through DMEM washing. Cells were stimulated with TSA-1-C4 and Tc24-C4 combination (100, 50, 25, 12.5 and 6.25 µg/ml) for 48 h. Supernatants were recovered to measure NO (indirectly by nitrite concentration)(A) and H2O2 (B)production. Data is represented by the mean ± S.D. of three independent experiments. Significance was calculated by one-way ANOVA and Tukey’s multiple-comparison test (* recombinant protein stimulated cells vs non stimulated cells; ^ Tc24-C4 stimulated cells vs TSA-1-C4 stimulated cells; & Tc24-C4 stimulated cells vs TSA-1-C4+Tc24-C4 stimulated cells). One, two, and three symbol characters were used to annotate the P values of <0.05, <0.01, and <0.001, respectively.
Figure. 2 Effect of recombinant TSA-1-C4 plus Tc24-C4 combination on cytokine production by peritoneal macrophages. Peritoneal cells isolated from naïve mice were cultured in supplemented DMEM for 48 h, adherent cells were stimulated with TSA-1-C4 and Tc24-C4 combination (100, 50, 25 and 12.5 and 6.25 µg/ml) for 48 h. Supernatants were recovery to measure TNF-α (A), IL-1β (B), IL-6(C) and IL-10 (D) production. The purple dotted line represents cytokine production by non-stimulated cells control. Data is represented by the mean ± S.D. of three independent experiments. Significance was calculated by one-way ANOVA and Tukey’s multiple-comparison test (% TSA-1-C4+Tc24-C4 stimulated cells vs TSA-1-C4 stimulated cells; & TSA-1-C4+Tc24-C4 stimulated cells vs Tc24-C4 stimulated; ^ Tc24-C4 stimulated cells vs TSA-1-C4 stimulated cells). One, two, and three symbol characters were used to annotate the P values of <0.05, <0.01, and <0.001, respectively.
Figure. 3 Effect of recombinant TSA-1-C4 plus Tc24-C4 combination on cytokine production by CD8+ T cells. Peritoneal cells isolated from naïve mice were stimulated with TSA-1-C4 and Tc24-C4 combination (100, 50, 25 and 12.5 µg/ml); after 48 h, supernatant was removed and cells were co-cultivated with CD8+ T cells from naïve mice. Supernatants were recovery to measure IFN-γ (A), TNF-α (B), IL-4(C) and IL-10 (D) production. The purple dotted line represents cytokine production by non-stimulated cells control. Data is represented by the mean ± S.D. of three independent experiments. Significance was calculated by one-way ANOVA and Tukey’s multiple-comparison test (% TSA-1-C4+Tc24-C4 stimulated cells vs TSA-1-C4 stimulated cells; & TSA-1-C4+Tc24-C4 stimulated cells vs Tc24-C4 stimulated). One, two, and three symbol characters were used to annotate the P values of <0.05, <0.01, and <0.001, respectively.
Figure. 4 (A ) Timeline for T. cruzi -experimental infection using BALB/c mice, prime-boost vaccination (red arrows), macrophages injection (purple arrow) and euthanize (blue arrow). (B ) Effect of the recombinant protein combination onin-vivo cytotoxicity assay. Representative histogram profile showed as percentage. Significance was calculated by one-way ANOVA and Tukey’s multiple-comparison test and is indicated as follows ***, P≤0.001 (comparing with the infected untreated control group).