Chronic TGF-β1 Signaling in Pulmonary Arterial Hypertension Induces Sustained Canonical Smad3 Pathways in Vascular Smooth Muscle Cells

(A) The mRNA and microRNA (miR) expression of TGF-β1 targets was evaluated by qPCR in starved (48 h) human pulmonary artery smooth muscle cells (HPASMCs) stimulated with TGF-β1 (5 ng/ml, 8 hours, n = 3), as well as in the main pulmonary artery (PA) and total lung of transgenic mice overexpressing TGF-β1 in the circulation (TG-TGFβ1) versus wild-type control (males, n = 8–16). (C) Adult male Sprague Dawley rats (∼200 g body weight) were divided into three age-matched groups: control normoxia, control hypoxia (i.e., rats injected once s.c. with vehicle [DMSO; vol/vol] and then exposed to hypoxia [FiO2 = 0.1] for 3 weeks, followed by 6 weeks in room air [FiO2 = 0.21]), and Sugen hypoxia (SuHx, i.e., rats injected with SU5416 20 mg/kg/dose s.c. and then exposed to hypoxia [3 wk], followed by 6 weeks in room air). mRNA expression was evaluated in total lungs (n = 6–11). Here, we found that the TGF-β1 pathway was indeed activated in the hypertensive SuHx rat lung (increased TGF-β1 mRNA expression; Figure 1C), whereas Smad3 mRNA expression remained unchanged (Figure 1C). [...]based on our SuHx data, the canonical TGF-β1/Smad3 pathway is not decreased in the rat lung with severe PAH. Smad3 mRNA (trend, statistically not significant) and miR-130a/301b (P < 0.01) expression was higher in the PAs and plexiform lesions of patients with idiopathic PAH compared with control subjects (Figure 1E). [...]our human data support the notion that boosted signaling of TGF-β1 and its canonical downstream effector Smad3 in human PAs is crucial for PAH development and severity.