The effect of nonsteroidal anti-inflammatory drugs on cAMP-dependent protein kinase-mediated phosphorylation by human chondrocytes in culture.
Although a principal pharmacological action of nonsteroidal anti-inflammatory drugs (NSAIDs) is to blunt eicosanoid synthesis by inhibiting cyclooxygenase, recent evidence indicates that NSAIDs may also interact directly with proteins that control the activity of adenylyl cyclase. Since the only physiological mechanism governing the action of cyclic AMP occurs via activation of its receptor, cyclic AMP-dependent protein kinase (cAMPPk; Kinase A), we determined whether NSAIDs affect intracellular substrate phosphorylation dependent on cAMPPk. The incorporation of 32Pi into cellular proteins that are substrates for cAMPPk was determined in intact human non-arthritic, aged non-arthritic and osteoarthritic chondrocytes in the presence or absence of NSAIDs, namely, sodium meclofenamate, indomethacin, tiaprofenic acid and sodium salicyclate. The transfer of [32P]-ATP was employed to identify phosphoproteins in a membrane fraction prepared from chondrocyte homogenates in the presence or absence of these NSAIDs. The lowest concentration of NSAID was similar to NSAID concentrations achieved during therapy for the arthritides. In intact human chondrocyte strains, activation of cAMPPk by dibutyryl cAMP (dBcAMP) resulted in the phosphorylation of intracellular substrates with an apparent M(r) of 55kD, 42kD, 26kD, 25kD, 22kD, 21.5kD, 20.5kD, and 17kD when examined by autoradiography after 12.5% SDS/PAGE. The NSAIDs augmented or potentiated phosphorylation of these proteins which were cAMPPk-dependent. In the chondrocyte membrane fraction, protein phosphorylation by cAMP was mimicked by isobutylmethylxanthine or by the purified catalytic subunit of bovine cAMPPk. NSAIDs augmented chondrocyte phosphorylation in the chondrocyte membrane fraction.(ABSTRACT TRUNCATED AT 250 WORDS)
Malemud
C J
CJ
Department of Medicine, Biomedical Research Building, Western Reserve University School of Medicine, Cleveland, Ohio 44106-4946.
Papay
R S
RS
Hasler
P
P
Kammer
G M
GM
D000293:Adolescent; D000328:Adult; D000368:Aged; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000494:pharmacology*; D002358:Cartilage, Articular / Q000166:cytology / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D017868:Cyclic AMP-Dependent Protein Kinases / Q000502:physiology*; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D010766:Phosphorylation / Q000187:drug effects
D000894:Anti-Inflammatory Agents, Non-Steroidal; D017868:Cyclic AMP-Dependent Protein Kinases