Interleukin-1β Expression Is Required for Lysophosphatidic Acid-Induced Lymphangiogenesis in Human Umbilical Vein Endothelial Cells

Figure 4

Lysophosphatidic acid- (LPA-) stimulated endothelial cell tube formation in vitro is mediated through endothelial growth factor receptor (EGFR) transactivation-, matrix metalloproteinase- (MMP-), LPA1/3-, and interleukin- (IL-) 1R-dependent mechanisms. Starved HUVECs were pretreated with AG1478 (100 nM), AF12198 (10 nM), GM6001 (10 𝜇M), Ki16425 (10 𝜇M), and toxin B (10 nM) for 1 h, followed by LPA (5 𝜇M) treatment for an additional 24 h, and then cells were seeded onto Matrigel-coated plates. After 6 h of plating, Matrigel of each experiment was fixed and subjected to an immunocytochemical assay. The Matrigel was stained with PECAM-1 or a Prox-1 primary antibody followed by an FITC-conjugated secondary antibody. Images were taken after plating and visualized by immunofluorescence microscopy.