Endothelin 1 receptors, signal transduction and effects on DNA and proteoglycan synthesis in rat articular chondrocytes.
This study showed that endothelins (ETs) stimulate DNA and proteoglycan synthesis in monolayer culture of rat articular chondrocytes (AC) by interacting with specific cell surface receptors. The high affinity receptors bound [125I]ET-1 with a Kd of 0.54 nM and Bmax of 81.4 pM/microgram DNA (approximately 40 000 binding sites per cell) was demonstrated. [125I]ET-1 binding was completely inhibited by unlabelled ET-1 or ET-2, and by BQ123 (ETA receptor antagonist), whereas ET-3 and IRL1038 (ETB receptor antagonist) did so only weakly. SDS-PAGE of cell extracts containing [125I]ET-1 cross-linked to the receptors, followed by autoradiography of the gels revealed a single 50-kDa band. These findings indicate that most of the receptors are subtype ETA. Although mRNA transcripts specific for both ETA and ETB receptors were found by RT-PCR, the ETA mRNA was more abundant. ET-1 increased the production of cAMP, cGMP and prostaglandin E2 (PGE2) and protein kinase C (PKC) activity in a concentration- and time-dependent manner. ET-1, and to a lesser degree ET-2, stimulated DNA synthesis, whereas ET-3 was inactive. Stimulation of DNA synthesis by ET-1 was strongly inhibited in a concentration-dependent manner by BQ123 and, to a much lesser degree, by IRL1038, which is consistent with an ETA receptor. ET-1 also stimulated proteoglycan synthesis and increased the amount of mRNA specific for the aggrecan gene. These findings strongly suggest that ET-1 is involved in regulating chondrocyte proliferation and metabolism in health, and presumably in disease.
Khatib
A M
AM
INSERM Unit 349, Biologie Cellulaire et Moléculaire de l'Os et du Cartilage, Hôpital Lariboisière, 75475 Paris Cedex 10, France.
Lomri
A
A
Moldovan
F
F
Soliman
H
H
Fiet
J
J
Mitrovic
D R
DR
D000818:Animals; D002358:Cartilage, Articular / Q000187:drug effects* / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000096:biosynthesis; D006152:Cyclic GMP / Q000096:biosynthesis; D004261:DNA Replication / Q000187:drug effects* / Q000502:physiology; D015232:Dinoprostone / Q000096:biosynthesis; D019332:Endothelin-1 / Q000494:pharmacology* / Q000502:physiology; D019333:Endothelin-2 / Q000494:pharmacology; D019334:Endothelin-3 / Q000494:pharmacology; D016232:Endothelins / Q000494:pharmacology; D010446:Peptide Fragments / Q000494:pharmacology; D010456:Peptides, Cyclic / Q000494:pharmacology; D011493:Protein Kinase C / Q000378:metabolism; D011509:Proteoglycans / Q000096:biosynthesis*; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017208:Rats, Wistar; D044022:Receptor, Endothelin A; D044023:Receptor, Endothelin B; D017466:Receptors, Endothelin / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*
D019332:Endothelin-1; D019333:Endothelin-2; D019334:Endothelin-3; D016232:Endothelins; D010446:Peptide Fragments; D010456:Peptides, Cyclic; D011509:Proteoglycans; D012333:RNA, Messenger; D044022:Receptor, Endothelin A; D044023:Receptor, Endothelin B; D017466:Receptors, Endothelin; C077420:Cys(11)-Cys(15)-endothelin-1 (11-21); D000242:Cyclic AMP; D011493:Protein Kinase C; D006152:Cyclic GMP; D015232:Dinoprostone; C072247:cyclo(Trp-Asp-Pro-Val-Leu)