Role of Epac1 in mediating anti-proliferative effects of prostanoid EP(2) receptors and cAMP in human lung fibroblasts.
In lung fibroblasts, proliferation is inhibited by activation of EP(2) prostanoid receptors which are known to couple to adenylyl cyclase. Beside the classic target of cAMP, protein kinase A (PKA), alternative cAMP effectors have been identified, among them Epac (exchange protein activated by cAMP). The present study aimed to illuminate transduction pathways mediating the anti-proliferative effects of EP(2) receptors in lung fibroblasts. Proliferative activity of human lung fibroblasts was determined by measuring [(3)H]-thymidine incorporation. The selective EP(2) receptor agonist butaprost inhibited [(3)H]-thymidine incorporation by 75%, an effect mimicked by forskolin, the phosphodiesterase inhibitor IBMX, the stable cAMP analogues dibutyryl-cAMP and bromo-cAMP, as well as by the Epac selective cAMP analogues 8-pCPT-2'-O-Me-cAMP and Sp-8-pCPT-2'-O-Me-cAMPS, whereas the PKA selective agonist 6-Bnz-cAMP was inactive. The PKA inhibitor Rp-8-Br-cAMPS inhibited butaprost-induced phosphorylation of CREB (cAMP response element-binding protein), but did not affect butaprost-induced inhibition of [(3)H]-thymidine incorporation. Partial knockdown of Epac1 by specific siRNA transfection resulted in a marked attenuation of the inhibitory potency of butaprost, whereas transfection of Epac2 siRNA or non-silencing siRNA did not affect the effectiveness of butaprost to inhibit [(3)H]-thymidine incorporation. In conclusion, Epac1 rather than the classic cAMP effector PKA is a crucial element in the signal transduction pathway mediating anti-proliferative effects of EP(2) receptor activation.
Haag
S
S
Institute of Pharmacology and Toxicology, University of Bonn, Reuterstrasse 2b, 53113, Bonn, Germany.
Warnken
M
M
Juergens
U R
UR
Racké
K
K
D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D000527:Alprostadil / Q000031:analogs & derivatives / Q000494:pharmacology; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D003094:Collagen / Q000378:metabolism; D000242:Cyclic AMP / Q000502:physiology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D004789:Enzyme Activation; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D055785:Gene Knockdown Techniques; D020662:Guanine Nucleotide Exchange Factors / Q000819:agonists / Q000378:metabolism / Q000502:physiology*; D006801:Humans; D008168:Lung / Q000166:cytology*; D010766:Phosphorylation; D034741:RNA, Small Interfering / Q000235:genetics; D018078:Receptors, Prostaglandin E / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D013873:Thionucleotides / Q000494:pharmacology
C105593:8-bromoadenosine-3',5'-cyclic monophosphorothioate; D020662:Guanine Nucleotide Exchange Factors; C116122:RAPGEF3 protein, human; D034741:RNA, Small Interfering; D018078:Receptors, Prostaglandin E; D013873:Thionucleotides; D015124:8-Bromo Cyclic Adenosine Monophosphate; D003094:Collagen; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D000527:Alprostadil; C048491:butaprost