PGE2 potentiates tonicity-induced COX-2 expression in renal medullary cells in a positive feedback loop involving EP2-cAMP-PKA signaling.
Cyooxygenase-2 (COX-2)-derived PGE2 is critical for the integrity and function of renal medullary cells during antidiuresis. The present study extended our previous finding that tonicity-induced COX-2 expression is further stimulated by the major COX-2 product PGE2 and investigated the underlying signaling pathways and the functional relevance of this phenomenon. Hyperosmolality stimulated COX-2 expression and activity in Madin-Darby canine kidney (MDCK) cells, a response that was further increased by PGE2-cAMP signaling, suggesting the existence of a positive feedback loop. This effect was diminished by AH-6809, an EP2 antagonist, and by the PKA inhibitor H-89, but not by AH-23848, an EP4 antagonist. The effect of PGE2 was mimicked by forskolin and dibutyryl-cAMP, suggesting that the stimulatory effect of PGE2 on COX-2 is mediated by a cAMP-PKA-dependent mechanism. Accordingly, cAMP-responsive element (CRE)-driven reporter activity paralleled the effects of PGE2, AH-6809, AH-23848, H-89, forskolin, and dibutyryl-cAMP on COX-2 expression. In addition, the stimulatory effect of PGE2 on tonicity-induced COX-2 expression was blunted in cells transfected with dominant-negative CRE binding (CREB) protein, as was the case in a COX-2 promoter reporter construct in which a putative CRE was deleted. Furthermore, PGE2 resulted in PKA-dependent phosphorylation of the pro-apoptotic protein Bad at Ser155, a mechanism that is known to inactivate Bad, which coincided with reduced caspase-3 activity during osmotic stress. Conversely, pharmacological interruption of the PGE2-EP2-cAMP-PKA pathway abolished Ser155 phosphorylation of Bad and blunted the protective effect of PGE2 on cell survival during osmotic stress. These observations indicate the existence of a positive feedback loop of PGE2 on COX-2 expression during osmotic stress, an effect that apparently is mediated by EP2-cAMP-PKA signaling, and that contributes to cell survival under hypertonic conditions.
Steinert
Daniela
D
Department of Physiology, University of Munich, Pettenkoferstrasse 12, 80336 Munich, Germany.
Küper
Christoph
C
Bartels
Helmut
H
Beck
Franz-X
FX
Neuhofer
Wolfgang
W
D000818:Animals; D017209:Apoptosis; D053148:Caspase 3 / Q000378:metabolism; D002460:Cell Line; D000242:Cyclic AMP / Q000378:metabolism*; D017362:Cyclic AMP Response Element-Binding Protein / Q000235:genetics / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051546:Cyclooxygenase 2 / Q000235:genetics / Q000378:metabolism*; D052246:Cyclooxygenase 2 Inhibitors / Q000494:pharmacology; D015232:Dinoprostone / Q000378:metabolism*; D004285:Dogs; D007679:Kidney Medulla / Q000187:drug effects / Q000201:enzymology* / Q000473:pathology; D009997:Osmotic Pressure; D010766:Phosphorylation; D011401:Promoter Regions, Genetic; D011448:Prostaglandin Antagonists / Q000494:pharmacology; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D018078:Receptors, Prostaglandin E / Q000037:antagonists & inhibitors / Q000378:metabolism*; D058307:Receptors, Prostaglandin E, EP2 Subtype; D012462:Saline Solution, Hypertonic / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D013312:Stress, Physiological; D013997:Time Factors; D014162:Transfection; D015854:Up-Regulation; D051025:bcl-Associated Death Protein / Q000378:metabolism
D017362:Cyclic AMP Response Element-Binding Protein; D052246:Cyclooxygenase 2 Inhibitors; D011448:Prostaglandin Antagonists; D047428:Protein Kinase Inhibitors; D012333:RNA, Messenger; D018078:Receptors, Prostaglandin E; D058307:Receptors, Prostaglandin E, EP2 Subtype; D012462:Saline Solution, Hypertonic; D051025:bcl-Associated Death Protein; D000242:Cyclic AMP; D051546:Cyclooxygenase 2; D017868:Cyclic AMP-Dependent Protein Kinases; D053148:Caspase 3; D015232:Dinoprostone