Figure 1.
Activation of FoxM1 (forkhead box M1) signaling in lungs of patients with idiopathic pulmonary arterial hypertension (IPAH) and pulmonary hypertension mice and rats. (A) qRT-PCR analysis demonstrating upregulation of FOXM1 mRNA expression in human IPAH lungs. (B) Representative micrographs showing prominent FOXM1 mRNA expression in smooth muscle cells (SMCs) of remodeled vessel (middle) and plexiform lesion (right) of patients with IPAH. RNA expression was fluorescently labeled by RNAscope reagent (RNA in situ hybridization) (red). SMCs were immunostained with anti–α-SMA (green). Arrows point to FoxM1-expressing SMCs. (C) Immunofluorescent staining demonstrating marked increase of FoxM1 protein expression in SMCs of vascular lesions in patients with IPAH. Lung tissue sections were immunostained with anti-FoxM1 (red) and anti–α-SMA (green). Arrows point to FoxM1-expressing SMCs. (D and E) Upregulation of FoxM1 in pulmonary arterial smooth muscle cells isolated from patients with IPAH analyzed by qRT-PCR and Western blotting. The intensity of FoxM1 protein band was normalized to the loading control of β-actin. (F) Similar levels of FoxM1 expression in pulmonary endothelial arterial cells from patients with IPAH and normal subjects. (G) Induced FoxM1 expression in lungs of Egln1Tie2Cre mice. Lung tissues were collected from Egln1Tie2Cre mice and Egln1f/f littermate control animals at the age of 2 months. (H) Western blotting analysis showing increased FoxM1 protein levels in Sugen 5416/hypoxia rats compared with basal. (I) Increased FoxM1 protein expression in monocrotaline-exposed rats compared with basal rats. *P < 0.05, **P < 0.01, and ***P < 0.001. (A) Welch t test. (D–I) Student’s t test. Horizontal bars in dot plot panels represent the mean. Scale bars, 50 μm. A.U. = arbitrary units; MCT = monocrotaline; hPAEC = human pulmonary endothelial arterial cell; hPASMC = human pulmonary arterial smooth muscle cell; SMA = smooth muscle actin; SuHx = Sugen 5416/hypoxia.