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 [methyl3H]thymidine uptake, intracellular [Ca2+] ([Ca2+]i) was measured with the Ca(2+)-sensitive indicator fura-2, and tyrosine phosphorylation was measured by Western blotting techniques. All three isoforms of PDGF stimulated [methyl3H]thymidine uptake concentration dependently, with similar potency. PDGF-AB induced significantly greater [methyl3H]-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.
Hughes
A D
AD
Department of Clinical Pharmacology, St. Mary's Hospital Medical School, Imperial College of Science, Technology and Medicine, London, England.
Patel
M
M
Wijetunge
S
S
Clunn
G
G
Schachter
M
M
D015153:Blotting, Western; D002118:Calcium / Q000502:physiology; D002478:Cells, Cultured; D004247:DNA / Q000096:biosynthesis*; D006801:Humans; D007536:Isomerism; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D010766:Phosphorylation; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology*; D012501:Saphenous Vein / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*; D013936:Thymidine / Q000378:metabolism; D014443:Tyrosine / Q000378:metabolism
D010982:Platelet-Derived Growth Factor; D014443:Tyrosine; D004247:DNA; D002118:Calcium; D013936:Thymidine