Abstract 320: Increased Expression and Activity of Phosphodiesterase-1 Isoforms in Pulmonary Artery Smooth Muscle Cells from Patients with Pulmonary Hypertension and Role in Proliferation
Pulmonary artery smooth muscle cells (PASMC) from patients with idiopathic pulmonary arterial hypertension (IPAH) or secondary pulmonary hypertension (SPH) have reduced agonist-induced cAMP and increased serum-induced proliferation. We tested the hypothesis that PASMC isolated from such patients have increased expression of PDE isoforms and that inhibition of specific PDEs would increase zcAMP levels and inhibit proliferation. Real-time PCR demonstrated a significant increase in PDE1A (40 ± 12; 75 ± 10 fold increase) and PDE1C (277 ± 21; 96 ± 19 fold increase) mRNA in both IPAH- and SPH-PASMC respectively, elevated protein expression of these isoforms and a ~2-fold increase in PDE1 enzymatic activity. Consistent with those results, PDE1 inhibitors (Vinpocetine and 8-MM-IBMX, 30μM) reduced proliferation and calcium-induced calcium entry, and increased cAMP generation to 10μM forskolin more effectively in PASMC from IPAH and SPH patients than from controls. To assess the role of PDE1C, we used siRNAs, which decreased PDE1C expression by 60–70% in both normal and patient PASMC. Incubation of PASMC with PDE1C siRNA helped restore forskolin-induced cAMP levels from 8.8 ± 2.3 to 27.8 ± 5.1 pmol/min/mg in IPAH- and 8.3 ± 2.9 to 28.6 ± 2.9 pmol/min/mg in SPH-PASMC and inhibited serum-stimulated proliferation ~2-fold more than did the siRNA-negative control. Increasing serum concentration (up to 20%) increased PDE1C expression in PASMC up to 350 ± 20% and increased [3H]thymidine incorporation; treatment with the co-mitogens (e.g., angiotensin II and endothelin-1) also increased PDE1C expression. We conclude that PDE1C is a PDE isoform that contributes to decreased agonist-promoted cAMP levels and increased proliferation PASMC from patients with both IPAH and SPH and thus PDE1C-specific inhibitors may be a useful therapy in this disease.
Supported by ALA/NIH.