PDE1B2 regulates cGMP and a subset of the phenotypic characteristics acquired upon macrophage differentiation from a monocyte

Abstract

Monocyte-to-macrophage differentiation with the cytokine granulocyte-macrophage colony-stimulating factor induces expression of the cyclic nucleotide phosphodiesterase PDE1B2. However, what role PDE1B2 plays in macrophage biology has not been elucidated. We have addressed this question by inhibiting PDE1B2 induction by using RNA interference. Using a retrovirus-based system, we created HL-60 stable cell lines that express a short-hairpin RNA targeting PDE1B2. HL-60 cells treated with phorbol-12-myristate-13-acetate differentiate to a macrophage-like phenotype and upregulate PDE1B2. However, expression of PDE1B2 short hairpin RNA effectively suppresses PDE1B2 mRNA, protein, and activity up-regulation. Using the HL-60 PDE1B2 knockdown cells and agonists for either adenylyl or guanylyl cyclase, it was found that PDE1B2 predominantly regulates cGMP and plays a lesser role in cAMP regulation in response to cyclase agonists. Furthermore, in intact HL-60 cells, PDE1B2 activity can be regulated by changes in Ca ؉2 levels. Inhibiting PDE1B2 up-regulation does not prevent HL-60 cell differentiation, because several markers of macrophage differentiation are unaffected. However, suppression of PDE1B2 expression alters some aspects of the macrophage-like phenotype, because cell spreading, phagocytic ability, and CD11b expression are augmented. The cAMP analog 8-Bromo-cAMP reverses the changes caused by PDE1B2 knockdown. Also, PDE1B2 knockdown cells have lower basal levels of cAMP and alterations in the phosphorylation state of several probable PKA substrate proteins. Thus, the effects of PDE1B2 on differentiation may ultimately be mediated through decreased cAMP. In conclusion, PDE1B2 regulates a subset of phenotypic changes that occur upon phorbol-12-myristate-13-acetate-induced differentiation and likely also plays a role in differentiated macrophages by regulating agonist-stimulated cGMP levels.cyclic AMP ͉ cyclic GMP ͉ phosphodiesterase

Role of cAMP-Phosphodiesterase 1C Signaling in Regulating Growth Factor Receptor Stability, Vascular Smooth Muscle Cell Growth, Migration, and Neointimal Hyperplasia

“…We failed to detect PDE1C in mouse peritoneal macrophages and found that PDE1C deficiency does not alter LPS-stimulated cytokine expression in macrophages nor circulating inflammatory molecule levels in mice (our unpublished observations). This is also consistent with the previous findings that the PDE1B isozyme represents the major PDE1 activity in macrophages 62, 63 . Together, these suggest that PDE1C does not regulate macrophage function and systemic inflammation in murine animals.…”

cGMP-Dependent Protein Kinases and cGMP Phosphodiesterases in Nitric Oxide and cGMP Action

“…Overall, little is known about regulation of the levels of PDE proteins, although changes in both the activities and levels of particular cGMP-hydrolyzing PDEs have been demonstrated in association with cellular proliferation and differentiation, as well as with tissue dysfunction (Maclean et al, 1997;Kotera et al, 1999a;Palmer and Maurice, 2000;Bender et al, 2004Bender et al, , 2005Champion et al, 2005;Wharton et al, 2005;Bender and Beavo, 2006b;Nagel et al, 2006;Schermuly et al, 2007;Farrow et al, 2008;Miller et al, 2009). Both short-term and long-term regulation of the action of PDEs allows for fine control of the amplitude and duration of cGMP elevation and provides for refined blunting or enhancement of the signal (Gopal et al, 2001;Mullershausen et al, 2001Mullershausen et al, , 2003Rybalkin et al, 2002Rybalkin et al, , 2003Bender et al, 2004Bender et al, , 2005Champion et al, 2005;Takimoto et al, 2005a;Fischmeister et al, 2006;Kass et al, 2007a;Omori and Kotera, 2007;Zhu and Strada, 2007;Muzaffar et al, 2008;Pokreisz et al, 2009;Zhu et al, 2009).…”

Cardiac Cyclic Nucleotide Phosphodiesterases: Function, Regulation, and Therapeutic Prospects

“…In cardiac fibroblasts, PDE1A appears to regulate both cGMP and cAMP [19]. PDE1B has been shown to predominantly regulate cGMP levels in macrophages [20], and also appears to play a role in regulating cAMP levels in certain types of cells [21,22]. PDE1C has the highest affinity for cAMP, and appears to regulate cellular cAMP in multiple cell types, including VSMCs [23], βTC3 insulinoma cells, pancreatic islet cells [24], and A172 glioblastoma cells [25].…”