Histological analyses
Embryos (E17) were fixed with 4% paraformaldehyhe for 14 hours, dehydrated and embedded in paraffin wax. Sections (7–8 um) were stained with Hematoxylin and Eosin (n≥3 for each genotype). For lineage tracing analyses, embryos were stained for β-galactosidase activity as described [43]. Briefly, the specimens (E11.0 – E11.5) were fixed in 4% paraformaldehyde for 30 minutes at room temperature, washed 3 times for 10 minutes in the detergent wash and developed for 2–12 hours in X-gal solution (n≥3 for each genotype). For immunohistochemistry, fixed sections from tissues harvested at E10.5 – E11.5 were stained with monoclonal α-smooth muscle actin (DAKO), cleaved caspase-3 (Cell Signaling) or phophohistone-H3 (Cell Signaling) antibodies. TUNEL assays were performed using the DeadEnd fluorometric TUNEL system (Promega). In each assay 3 or more embryos were analyzed for each genotype.