Fig. 7. Lymphatic vessel defects in the heart and diaphragm of Nrp2 mutants. (A,B) Whole-mount X-gal staining of P0 hearts showing the reduction of capillaries in the Nrp2^–/– heart (B). (C) Double-staining of the X-gal-positive vessels present at the epicardial surface of Nrp2^+/– embryos (arrowhead) with anti-VEGFR3 (brown staining). (D,E) Transverse sections of E15 hearts stained with anti-VEGF-C antibodies. Note staining of the superficial epicardium in both Nrp2^+/– and Nrp2^–/– (D,E, arrows). (F,G) Whole-mount X-gal staining of P0 diaphragms (thoracic view). Note the reduction of peripheral X-gal-positive lymphatic capillaries (arrowheads) in the Nrp2^–/– diaphragm. Brackets outline the central portion of the diaphragm, which is unaffected. Weaker NRP2 expression in veins is also indicated (arrows). H. Double-staining of the X-gal-positive vessels present in the diaphragm of Nrp2^+/– embryos with anti-VEGFR3 (brown staining) confirms that they are lymphatics. (I) Higher magnification of the boxed region in H. Scale bars: in A, 580 µm for A,B; in C, 50 µm; in D, 360 µm for D,E; in F, 1400 µm for F,G; in H, 270 µm; in I, 40 µm.

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