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. 2020 Jul 27;130(9):4546–4560. doi: 10.1172/JCI136068

Figure 8. S1P/S1PR3 signaling is required for phagocytosis, ROS production, and intracellular killing of C. neoformans Δgcs1.

Figure 8

(A) Primary alveolar macrophages isolated from WT (n = 8) and S1PR3 deficient (n = 6) mice were coincubated with opsonized C. neoformans Δgcs1 and phagocytic index was calculated by microscopic observation. Experiment was conducted 3 times. (B) Phagocytic index for primary alveolar macrophages isolated from mice of indicated genotype (n = 5 each) with and without S1P supplementation were calculated as in A. Experiment was conducted 3 times. (C) Primary alveolar macrophages isolated from mice of indicated genotype (n = 3) with and without S1P supplementation were coincubated with opsonized C. neoformans Δgcs1. After incubation, culture media were plated onto YPD agar. Percentage of killing was calculated as the difference in CFUs of Δgcs1 incubated with or without macrophages. Experiment was conducted 5 times. (D) Primary alveolar macrophages of the indicated genotype (n = 3) were analyzed for ROS production after coincubation with opsonized C. neoformans Δgcs1. Experiment was conducted 5 times. All error bars represent SEM and statistical comparisons were done using 2-sided Student’s t test (*P = 0.0378, **P = 0.0222) or 1-way ANOVA with Bonferroni’s multiple comparisons post hoc test. P values were corrected for multiplicity using the Bonferroni’s adjustment (***P < 0.001, ****P < 0.0001, compared with control).