Extracellular matrix molecules regulate endothelial cell migration stimulated by lysophosphatidic acid

Background: Lysophosphatidic acid (LPA) and sphingosine I-phosphate (SIP) are lipids that bind G-protein coupled receptors and differentially promote transmigration of endothelial cells. Objective: To determine if endothelial cell transmigration stimulated by LPA, not S I P, is dependent on the extracellular matrix. Methods: Bovine pulmonary artery (BPAE) endothelial cell transmigration and locomotion were measured using a modified-Boyden chamber and video microscopy, respectively. Results were related to strength of adhesion and characteristics of cell adhesive contacts. Results and Conclusions: BPAEs responded to LPA by transmigration through gelatin- or collagen-coated filters, but not through fibronectin-, vitronectin-, or fibrinogen-coated filters. Fewer cells adhered to collagen or gelatin than to fibronectin in a static cell adhesion assay or after application of a g-force to detach cells. Video microscopy revealed that SIP stimulates large lamellipodia on two-dimensional fibronectin substrate. LPA stimulated lamellipodia on fibronectin, but the trailing edge remained attached, resulting in sting ray-shaped cells in video microscopy. LPA-treated cells on gelatin released the trailing edge. To understand how the extracellular matrix may regulate endothelial cell shape during movement, we surveyed changes in focal adhesion proteins. More Hic-5, a paxillin homolog, was detected in the detergent insoluble fraction of BPAEs attached to gelatin than fibronectin. No such difference was found in paxillin. In BPAEs, Hic-5 was localized to smaller punctate structures on fibronectin and longer, thinner focal adhesions on gelatin. These results indicated that localization of Hic-5 and strength of adhesion correlate with endothelial cell transmigration stimulated by LPA, but not with transmigration stimulated by SIP.