Figure 6.

Intersectional lineage tracing reveals the contribution of endocardial-derived MCs to fibroblasts and pericytes in adult hearts. A, the strategy for tracing endocardial-derived cushion MCs in adult hearts. Tamoxifen (Tam) was administrated at E9.5, and the hearts were harvested for analysis at the adult stage. B, immunostaining for tdTomato on adult heart sections with indicated genotypes. Scale bars, 2 mm. C, immunostaining for tdTomato and PDGFRa on heart sections from adult Nfatc1–Dre;Sox9–CreER;Ai66. The arrows indicate tdTomato⁺PDGFRa⁺ cells. The boxed regions in the left panel are magnified and split channels in the right panels. Scale bars, 100 μm. D, the cell counting result showing the percentage of tdTomato⁺ fibroblasts in ventricular free walls and septum (n = 4 mice). E, immunostaining for tdTomato, CDH5, and PDGFRb on heart sections from adult Nfatc1–Dre;Sox9–CreER;Ai66. The arrows indicate tdTomato⁺PDGFRb⁺ pericytes. The boxed regions in the left panel are magnified and split channels in the right panels. Scale bars, 100 μm. F, the cell counting result showing the percentage of tdTomato⁺ pericytes in ventricular free walls and septum (n = 4 mice). G and H, the percentage of tdTomato⁺ fibroblasts and pericytes in ventricles from adult Nfatc1–Dre;Sox9–CreER;Ai66 were assessed by flow cytometry. IgG were controls for PDGFRa or PDGFRb antibodies (n = 3 mice for each experiment). I, immunostaining for tdTomato, NG2, and CDH5 on heart sections from adult Nfatc1–Dre;Sox9–CreER;Ai66. The arrows indicate tdTomato⁺NG2⁺ pericytes. DAPI, 4′,6′-diamino-2-phenylindole; FSC, forward scatter; LV, left ventricle. Scale bars, 100 μm.