CD40 induces macrophage anti–Toxoplasma gondii activity by triggering autophagy-dependent fusion of pathogen-containing vacuoles and lysosomes
J. Clin. Invest. Rosa M. Andrade, et al. 116:2366 doi:10.1172/JCI28796 [Go to this article.]

Figure 1
CD40 stimulation induces vacuole/lysosome fusion in human and mouse macrophages infected withT. gondii . (A) Control and CD154-stimulated human macrophages were incubated with LysoTracker Red, then infected with T. gondii–YFP. Macrophages incubated with opsonized T. gondii were used as controls. Cells were examined by confocal microscopy 6 hours after infection or 2 hours after addition of opsonized parasites. Macrophages shown contain 1 tachyzoite of T. gondii. (B–D) Control and CD40-activated human macrophages were infected with T. gondii–YFP. Macrophages incubated with opsonized T. gondii were used as controls. Macrophages were incubated with anti–LAMP-1 (B), anti-CD63 (C), or anti-cathepsin D (D) Abs; this was followed by addition of secondary antibodies. Cells were examined by confocal microscopy at 6 hours after infection or 1 hour after addition of opsonized T. gondii. CD40-activated macrophages show colocalization of LAMP-1, CD63, and cathepsin D (rings) around T. gondii–containing vacuoles (arrowheads). Cath, cathepsin. Scale bars: 5 μm. (E and F) Quantification of colocalization of late endosomal/lysosomal markers around vacuoles containing T. gondii within human (E) or mouse (F) primary macrophages. Monolayers were examined at 6 and 8 hours after challenge in the case of human and mouse macrophages, respectively. In the case of macrophages incubated with opsonized T. gondii, M6PR was assessed at 15 minutes while LAMP-1, LAMP-2, CD63, and cathepsin D expression were assessed at 1 hour. Percentages indicate the mean ± SD. Results shown are representative of 3–4 independent experiments. Cath, cathepsin; ctr, control; DIC, differential interface contrast; ops, opsonized.