Figure 5.

Lymphatic ring assay. (A): Visualization by optical microscopy of LEC spreading from mouse thoracic duct fragment embedded for 11 days in a 3D-type I collagen gel. (B): Outgrowing LEC display filopodia-like structures visible upon phalloidin staining under confocal microscopy. (C) LEC are labeled in green with an anti-LYVE-1 antibody and observed under a fluorescent microscope. Nuclei are counterstained in blue (Dapi). (D-H) Electron microscopy micrographs of 3D-cultures of lymphatic thoracic duct rings. (D): Long extended processes of LEC. Note the invagination process (arrow). (E, F): Establishment of an irregular lumen in tubular structures lined by long thin endothelial cell processes. (G, H): Sprouting endothelial cells display processes and intracellular vesicles. d = matrix degradation. d = matrix degradation; lu = lumen. Scale bars in (A): 500 μm, in (B): 40 μm, in (C): 100 μm, and in (D-H): 1 μm.