Fig. 3: Higher levels of CD4⁺ regulatory T (Treg) cells during second pregnancies.

a Experimental setup: age-matched Fir/Tiger mice were allogenically mated to Balb/c males once or twice, respectively, and evaluated on various time points during pregnancy (gd 3.5: n = 10, gd 7.5: n = 6 (10), gd 15.5: n = 12). b–g Flow cytometry analysis included frequencies of CD4⁺ Treg cells and associated ex-vivo IL-10 expression in (b–d) lymph node (b gd 3.5: p = 0.0028, gd 7.5: p = 0.002, gd 15.5: p = 0.0042; c gd 3.5: p = 0.042) and (e–g) uterus (e gd 3.5: p = 0.000024, gd 7.5: p = 0.0086, gd 15.5: p = 0.011; f gd 3.5: p = 0.078), respectively. Pseudocolour plots and respective numbers presented correspond to CD4 as parent population. Additional plots are shown in Supplementary Fig. 3.h Serum progesterone levels were assessed throughout pregnancy. i–k Flow cytometry analysis assessed (i, j) ex-vivo IL-10 expression (gd 3.5: p = 0.029, gd 7.5: p = 0.048, gd 15.5: p = 0.010) and (k) CD80⁺/CD86⁺ expression (gd 3.5: p = 0.02, gd 7.5: p = 0.03, gd 15.5: p = 0.04) on CD11c⁺ dendritic cells (DC). Data are presented as violin plots with individual point and median and quartiles. The statistical significance was calculated using Multiple unpaired t-tests (* p < 0.05, ** p < 0.01, *** p < 0.001). Source data are provided as a Source Data file.