Figure 5.

Progesterone reduces the polyfunctional cytokine profile of CD8⁺ and CD4⁺ maternal T cells and favors a more IL‐4 dominant profile. (A) A Boolean gating strategy was established to look at the percentage of CD8⁺ and CD4⁺ T cells expressing from zero to six cytokines from both the maternal (top) and control (bottom) donors (n = 10 of each group), where PBMCs had been treated with PHA and either DMSO (vehicle), or 1 or 10 μM progesterone. *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001, ****p ≤ 0.0001, one‐way ANOVA, repeated measures, and Bonferroni multiple comparison test. (B) The same strategy was then used to look at the percentage of CD8⁺ and CD4⁺ T cells expressing each individual cytokine and every possible combination up to six cytokines. The differential absolute expression between the vehicle and 10 μM progesterone was used to create a heat map indicating the change in cytokine combinations when maternal or male PBMCs were treated with PHA and 10 μM progesterone. The change in cytokine expression following progesterone treatment is represented by a range of colors of varying green (increase) and red (decrease) intensity, denoting the percentage change in cytokine production with progesterone treatment. The map has been laid out in three sections; the left section is combination of cytokines including IFN‐γ but excluding IL‐4, the middle section is combination including both IFN‐γ and IL‐4, and the right section is inclusive of IL‐4 but excludes IFN‐γ.