Sildenafil potentiates the proliferative effect of porcine pulmonary artery smooth muscle cells induced by serotonin in vitro

Background Sildenafil is one of the selective phosphodiesterase 5 inhibitors that has been proven by manyinvestigators to suppress growth factor stimulated (e.g. platelet-derived growth factor (PDGF) or epidermal growth factor(EGF)) proliferation and hypertrophy of pulmonary artery smooth muscle cells (PASMCs) via cGMP/cGKIα pathway.Serotonin promotes cell cycle progression leading to cell mitogenesis and plays a key role in the pathogenesis ofpulmonary artery hypertension. The role of sildenafil in proliferation of PASMCs induced by serotonin has not beeninvestigated so far. In this study we explored the underlying mechanism of the effect of sildenafil on serotonin inducedproliferation of porcine PASMCs.Methods PASMCs were cells from primary cultures by the explant method from the pulmonary artery of swine and cellsat passage 3-5 were used in this study. MTT colorimetric assay and flow cytometry analysis were used to evaluate thecell proliferation and alterations in cell cycle progression respectively. Western blotting analysis was applied to determinethe expression of phosphorylated extracellular signal-regulated kinase (ERK), proliferating cell nuclear antigen (PCNA)and mitogen activated protein kinase (MAPK) phosphatase-1 (MKP-1).Results Serotonin (10 μmol/L) induced the upregulation of phosphorylation of ERK1/ERK2 and PCNA, an increase inthe percentage of cells in S phase and subsequent cell proliferation. Pretreatment with 1 μmol/L sildenafil potentiated thephosphorylation of ERK1/ERK2, an increase in the percentage of cells in S phase and cell proliferation, compared withserotonin stimulation alone (P<0.05). Furthermore, 30-minute pretreatment with 10 μmol/L U0126, specific antagonist forERK kinase (MEK) prevented the increase in phosphorylation of ERK1/ERK2 and abolished cell cycle progression andthe proliferation of PASMCs induced by sildenafil.Conclusion This study shows that sildenafil potentiated the proliferative effect of serotonin on PASMCs viaphosphorylation of ERK1/ERK2.