Induction of Serotonin Transporter by Hypoxia in Pulmonary Vascular Smooth Muscle Cells

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Abstract
—The increased delivery of serotonin (5-hydroxytryptamine, 5-HT) to the lung aggravates the development of hypoxia-induced pulmonary hypertension in rats, possibly through stimulation of the proliferation of pulmonary artery smooth muscle cells (PA-SMCs). In cultured rat PA-SMCs, 5-HT (10–8 to 10–6 mol/L) induced DNA synthesis and potentiated the mitogenic effect of platelet-derived growth factor-BB (10 ng/mL). This effect was dependent on the 5-HT transporter (5-HTT), since it was prevented by the 5-HTT inhibitors fluoxetine (10–6 mol/L) and paroxetine (10–7 mol/L), but it was unaltered by ketanserin (10–6 mol/L), a 5-HT2A receptor antagonist. In PA-SMCs exposed to hypoxia, the levels of 5-HTT mRNA (measured by competitive reverse transcriptase–polymerase chain reaction) increased by 240% within 2 hours, followed by a 3-fold increase in the uptake of [3H]5-HT at 24 hours. Cotransfection of the cells with a construct of human 5-HTT promoter-luciferase gene reporter and of pCMV-β-galactosidase gene allowed the demonstration that exposure of cells to hypoxia produced a 5.5-fold increase in luciferase activity, with no change in β-galactosidase activity. The increased expression of 5-HTT in hypoxic cells was associated with a greater mitogenic response to 5-HT (10–8 to 10–6 mol/L) in the absence as well as in the presence of platelet-derived growth factor-BB. 5-HTT expression assessed by quantitative reverse transcriptase–polymerase chain reaction and in situ hybridization in the lungs was found to predominate in the media of pulmonary artery, in which a marked increase was noted in rats that had been exposed to hypoxia for 15 days. These data show that in vitro and in vivo exposure to hypoxia induces, via a transcriptional mechanism, 5-HTT expression in PA-SMCs, and that this effect contributes to the stimulatory action of 5-HT on PA-SMC proliferation. In vivo expression of 5-HTT by PA-SMC may play a key role in serotonin-mediated pulmonary vascular remodeling.