Cell Interactions with Collagen MatricesIn VivoandIn VitroDepend on Phosphatidylinositol 3-Kinase and Free Cytoplasmic Calcium

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Abstract
We have investigated the role of phosphatidylinositol 3-kinase (PI3-kinase) in cellular interactions with collagenous matrices. Platelet-derived growth factor-BB (PDGF-BB) elicited a mobilization of intracellular Ca2+ in pig aortic endothelial (PAE) cells transfected with wild type PDGF β-receptor. This response was greatly reduced in PAE cells transfected with PDGF β-receptors mutated at positions Y740 and Y751 to prevent PI3-kinase binding. The experimental drug 1D-myo-inositol 1,2,6-trisphosphate (α-trinositol) induced a rapid increase and subsequent oscillations of the cytoplasmic Ca2+ concentration in cultured fibroblasts. This response was not due to an effect of α-trinositol on inositol 1,4,5-trisphosphate (IP3) receptors. α-Trinositol did not influence PDGF-BB elicited chemotaxis through collagen-coated membranes of PAE cells transfected with the wild-type PDGF β-receptor, but restored PDGF-BB elicited chemotaxis of PAE cells transfected with the PI3-kinase binding-site mutated PDGF β-receptor. Collagen gel contraction has been suggested to serve as a model for cellular control of interstitial fluid pressure (PIF) in dermis. The PI3-kinase inhibitors wortmannin (50 nM) and LY294002 (5μmlM) inhibited the stimulation of fibroblast-mediated collagen gel contraction by 0.4 nM PDGF-BB. Injection of wortmannin in rat paw skin induced a lowering of PIF, and this effect was abolished in animals pre-treated with α-trinositol. Pretreatment of rats with α-trinositol abolished the decrease in PIF induced by injecting monoclonal anti-rat α2β1 integrin IgG in rat paw skin. Taken together our data indicate that cell-collagen interactions in vivo and in vitro depend on PI3-kinase, and that this dependence can be bypassed by a drug eliciting intracellular Ca2+ mobilization.

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