Figure 5.

Increased MOI from the non-virulent strain of M. tb inhibits RIPK3 gene expression, while the virulent strain favors RIPK1 and RIPK3 expression at the protein level. Human MDMs were obtained after seven days in culture. 2 × 10⁶ MDMs were infected at MOI 1 and MOI 10 with an avirulent (H37Ra) and at MOI 1 y MOI 5 with a virulent (H37Rv) strain of M. tb 24 h; 2 × 10⁶ MDMs were not infected as a control (Uninf). After postinfection, cells were recovered and prepared for qPCR or Western Blot. Relative expression of RIPK1 (a) and RIPK3 (b) genes was assessed by qPCR. Bar graphs show the mean ± SD from four independent experiments (n = 4 donors). Representative Western blot of p-RIPK1, p-RIPK3, and GAPDH (c). Band densities of p-RIPK1 (d) and p-RIPK3 (e) were normalized against GAPDH and quantified by densitometry analysis with the ImageJ software. Results are shown in relative units (RU) of concentration using ImageJ software. The bar graphs show the mean ± SD from two independent experiments (n = 2 to immunoblot, and n = 4 to qPCR were performed with two technical replicates). Statistical analysis was performed using Kruskal–Wallis analysis, followed by Dunn’s post hoc test. * p < 0.05.