Quantitation of platelet-derived growth factor receptors in human arterial smooth muscle cells in vitro.
Platelet-derived growth factor (PDGF) is suggested to play an important role in the development of atherosclerosis as a migratory and mitogenic stimulus to arterial smooth muscle cells (ASMCs). Stimulated and unstimulated ASMCs were studied with respect to PDGF receptor (PDGF-R) mRNA and protein expression. Quantitative RT-PCR was developed for simultaneous evaluation of both PDGF-R alpha and -R beta mRNA expression and a quantitative ELISA for estimation of corresponding PDGF-R subunits. On the mRNA level, the overall PDGF-R beta expression was approximately 100 times lower than that of PDGF-R alpha. Furthermore, although PDGF-R alpha mRNA levels were high irrespective of hASMC phenotype, PDGF-R beta mRNA was influenced by serum stimulation with lower copy numbers in proliferating and confluent cells compared with quiescent cells. On the protein level, quiescent hASMCs expressed 10 times more PDGF-R beta than PDGF-R alpha. Serum stimulation decreased cell surface PDGF-Rs, with most prominent loss of PDGF-R alpha (ELISA and immunohistochemistry). Our results suggest a differential regulatory pattern for PDGF-R alpha and -R beta and are compatible with the usage of alternative promoters for regulation of -R alpha expression. Further, it seems that the number of available receptor subunits is not the only determinant of variations in cell stimulation with different PDGF isoforms.
Krettek
A
A
Wallenberg Laboratory for Cardiovascular Research, Göteborg University, Sweden. alexandra.krettek@wlab.wall.gu.se
Fager
G
G
Jernberg
P
P
Ostergren-Lundén
G
G
Lustig
F
F
D002455:Cell Division; D002478:Cells, Cultured; D019281:Dimerization; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D019084:Fluorescent Antibody Technique, Indirect; D005786:Gene Expression Regulation; D006801:Humans; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000378:metabolism*; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic / Q000235:genetics; D055503:Protein Multimerization; D012333:RNA, Messenger / Q000096:biosynthesis; D020796:Receptor, Platelet-Derived Growth Factor alpha; D020797:Receptor, Platelet-Derived Growth Factor beta; D017479:Receptors, Platelet-Derived Growth Factor / Q000032:analysis* / Q000096:biosynthesis / Q000235:genetics; D011994:Recombinant Proteins / Q000494:pharmacology; D013698:Templates, Genetic; D014158:Transcription, Genetic
D010982:Platelet-Derived Growth Factor; D012333:RNA, Messenger; D011994:Recombinant Proteins; D020796:Receptor, Platelet-Derived Growth Factor alpha; D020797:Receptor, Platelet-Derived Growth Factor beta; D017479:Receptors, Platelet-Derived Growth Factor