RPMs require PPARγ intrinsically for their development. (A) Total RPM counts from Pparg^fl/fl and Vav1-Cre/Pparg^fl/fl mice (gated as shown in Fig. 2 A) at indicated days after birth (postnatal day [PND]). (B) PPARγ expression by RPMs from representative WT (Pparg^fl/fl) mice at indicated PNDs and in adults shown as histograms (left) and as averages of MFI (normalized to FMO controls) from groups of WT and KO mice (right). (C and D) Bar graphs showing percentages of Ki67⁺ (C) and EdU⁺ (D) RPMs at indicated time points. (E) VCAM-1 expression gated on F4/80⁺ RPMs shown as dot plots from representative individuals at PND 7 (left) and bar graphs showing MFI (mean and SD) of groups of Pparg^fl/fl and Vav1-Cre/Pparg^fl/fl at PND 3, 5, and 7 (different color symbols indicate separate experiments; right). (F) Graphical scheme of experiments aiming at blocking (G) and inducing (H) PPARγ expression. (G) Total RPM counts from adult C57BL/6 mice treated i.p. with 1 mg/kg of PPARγ inhibitor GW9662 (left) or with 10 mg/kg of PPARγ ligand rosiglitazone (right) for 7 consecutive days prior to analysis at days indicated in the scheme. (H) Total RPM counts, frequency of Ki67⁺, and normalized MFI of PPARγ in RPMs from adult C57BL/6 mice at day 4 after hydrodynamic IL-4 gene delivery (plasmid pTT5-Il4). (I) Total RPM counts from mice of indicated genotypes. All mice were 8–12 wk old; RPMs gated as in Fig. 2 A. (J) Graphical presentation of experiments designed to test the fetal monocyte intrinsic requirement of PPARγ for RPM development. CD45⁺ cells were sorted from fetal livers of E14.5 embryos (CD45.1) and subsequently transferred to Pparg^fl/fl and Vav1-Cre/Pparg^fl/fl newborns. Analysis of the RPM reconstitution was performed 8 wk after transfer. (K) Dot plots showing RPM origin in recipient mice 8 wk after transfer. RPMs are gated as live CD11b^intF4/80⁺VCAM-1⁺CD11c^lo; red gates indicate CD45.1⁺ RPMs (derived from transferred fetal progenitors). (L) RPM counts of mice supplemented with fetal monocyte precursors or not (left), and frequencies of donor-derived and intrinsic RPMs in mice adoptively transferred with fetal precursors (right). The presented data are representative of at least two independent experiments, except C and H, which come from a single experiment. Symbols in column graphs represent values of individuals and the column size mean (± SD). *, P < 0.05 **, P < 0.01; ***, P < 0.001; ****, P < 0.0001 (unpaired two-tailed Student’s t test).