COMPARISON OF EFFECTS OF PLATELET-DERIVED GROWTH-FACTOR ISOFORMS ON SIGNALING AND DNA-SYNTHESIS OF HUMAN CULTURED SAPHENOUS-VEIN CELLS

We examined the effect of platelet-derived growth factor (PDGF)-AA, PDGF-AB, and PDGF-BB isoforms on DNA synthesis, Ca2+ mobilization, and tyrosine phosphorylation in cultured human saphenous vein cells cultured by an explant technique; confluent cells derived from passage 3 were used for all studies. DNA synthesis was measured by [methyl(3)H]thymidine uptake, intracellular [Ca2+] ([Ca2+](i)) was measured with the Ca2+-sensitive indicator fura-2, and tyrosine phosphorylation was measured by Western blotting techniques. All three isoforms of PDGF stimulated [methyl(3)H]thymidine uptake concentration dependently, with similar potency. PDGF-AB induced significantly greater [methyl(3)H]thymidine uptake than PDGF-BB, and PDGF-AA was much less effective than either PDGF-AB or PDGF-BB. The effects of all three isoforms were inhibited by tyrphostin-23, a selective inhibitor of tyrosine kinases. PDGF-AB and PDGF-BB increased [Ca2+](i), although the maximum response to PDGF-AB was significantly less than that to PDGF-BB. Both isoforms increased [Ca2+](i) by stimulating influx and intracellular release of Ca2+. PDGF-AA had no measurable effect on [Ca2+](i). All three isoforms increased tyrosine phosphorylation of a 170-kDa protein detected by Western blotting. Quantitative densitometry indicated that PDGF-BB induced greater tyrosine phosphorylation than PDGF-AB and both PDGF-BB and PDGF-AB induced markedly more tyrosine phosphorylation than PDGF-AA. PDGF isoforms have differing efficacies in terms of DNA synthesis, Ca2+ mobilization, and tyrosine phosphorylation by human saphenous vein cells.