Prominent lymph node lymphangiogenesis in VEGFC transgenic mice. (A-B) Routine H&E stains of lymph nodes of non–tumor-bearing mice. (C-D) Double immunofluorescence staining of lymph nodes of non–tumor-bearing mice demonstrated a comparable pattern of CD31⁺/LYVE-1–negative blood vessels (green) and LYVE-1–positive sinusoids (red) in wild-type mice (C) and in VEGFC transgenic (TG; D) mice. Nonmetastatic sentinel lymph nodes of SCC-bearing VEGFC transgenic mice have increased numbers of enlarged LYVE-1–positive sinusoids (red; F), compared with control mice (E). An increased number of enlarged LYVE-1–positive lymphatic vessels (red) was also found in the metastatic sentinel lymph nodes of VEGFC transgenic mice (H), compared with control mice (G). The tumor-associated LYVE-1–positive vessels (red) in VEGFC transgenic mice showed high levels of BrdU staining in lymphatic endothelial cells (J; green), indicating active lymphatic proliferation (arrowheads) within sentinel lymph nodes. These cells also expressed Prox1 (I; green). Nuclei are stained blue (Hoechst stain). (A-F, I-J) Scale bars represent 100 μm; (G-H) scale bars represent 200 μm.