PROLONGED BETA-ADRENOCEPTOR STIMULATION UP-REGULATES CAMP-PHOSPHODIESTERASE ACTIVITY IN HUMAN MONOCYTES BY INCREASING MESSENGER-RNA AND PROTEIN FOR PHOSPHODIESTERASES 4A AND 4B

Human peripheral blood monocytes were treated for 4 h with a combinati on of the beta-agonist salbutamol (3 mu M) and the low-K-m cAMP-specif ic phosphodiesterase (PDE4) inhibitor rolipram (30 mu M) to produce a prolonged elevation of cAMP and consequent increase in PDE activity. A fter this treatment, isozyme-selective PDE inhibitors were used to cha racterize the cAMP PDE profiles of high-speed supernatants before and after DEAE-Sepharose column chromatography. These experiments, in whic h total soluble PDE activity was increased by 58%, showed that the inc reased PDE activity is due to up-regulation of PDE4 and that at least two of the four subtypes are up-regulated. Experiments in whole cells demonstrated that this relatively modest increase in PGE4 activity has significant functional consequences, reducing cAMP accumulation in re sponse to both PGE(2) and lower, though not maximal, concentrations of rolipram. Further characterization of PDE4 subtype expression in cont rol and treated monocytes, using polymerase chain reaction and Western blotting with subtype-specific peptide antibodies, showed that restin g monocytes express both mRNA and protein for PDE4A, PDE4B and PDE4D. The amount of message for PDE4A and PDE4B appeared to increase upon up -regulation, whereas mRNA for PDE4D was not detected in treated cells. Western blots showed increases in the amount of protein for both PDE4 A and PDE4B after treatment. We conclude that the PDE4 subtypes are di fferentially regulated upon prolonged exposure to elevated cAMP, with the consequence that the PDE4 profiles of control and treated cells di ffer not only in total activity but also in the relative proportions o f the subtypes represented.