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Intrauterine Pulmonary Hypertension Impairs Angiogenesis In Vitro

Role of Vascular Endothelial Growth Factor–Nitric Oxide Signaling

¹ Section of Neonatology and ² Section of Pulmonary Medicine, Pediatric Heart Lung Center, Department of Pediatrics, University of Colorado School of Medicine, Denver, Colorado

Correspondence and requests for reprints should be addressed to Jason Gien, M.D., P18-4402K, Mail Stop 8317, University of Colorado Health Sciences Center, 12800 East 19th Avenue, P.O. Box 6511, Aurora, CO 80045. E-mail: jason.gien{at}uchsc.edu

Rationale: Mechanisms that impair angiogenesis in neonatal persistent pulmonary hypertension (PPHN) are poorly understood.

Objectives: To determine if PPHN alters fetal pulmonary artery endothelial cell (PAEC) phenotype and impairs growth and angiogenesis in vitro, and if altered vascular endothelial growth factor–nitric oxide (VEGF–NO) signaling contributes to this abnormal phenotype.

Methods: Proximal PAECs were harvested from fetal sheep that had undergone partial ligation of the ductus arteriosus in utero (PPHN) and age-matched control animals. Growth and tube formation ± VEGF and NO stimulation and inhibition were studied in normal and PPHN PAECs. Western blot analysis was performed for VEGF, VEGF receptor-2 (VEGF-R2), and endothelial NO synthase (eNOS) protein content. NO production with VEGF administration was measured in normal and PPHN PAECs.

Measurements and Main Results: PPHN PAECs demonstrate decreased growth and tube formation in vitro. VEGF and eNOS protein expression were decreased in PPHN PAECs, whereas VEGF-R2 protein expression was not different. VEGF and NO increased PPHN PAEC growth and tube formation to values achieved in normal PAECs. VEGF inhibition decreased growth and tube formation in normal and PPHN PAECs. NOS inhibition decreased growth in normal and PPHN PAECs, but tube formation was only reduced in normal PAECs. NO reversed the inhibitory effects of VEGF-R2 inhibition on tube formation in normal and PPHN PAECs. VEGF increased NO production in normal and PPHN PAECs.

Conclusions: PPHN in utero causes sustained impairment of PAEC phenotype in vitro, with reduced PAEC growth and tube formation and down-regulation of VEGF and eNOS protein. VEGF and NO enhanced growth and tube formation of PPHN PAECs.

Key Words: persistent pulmonary hypertension of the newborn • angiogenesis • vascular endothelial growth factor • nitric oxide • endothelial cells

AT A GLANCE COMMENTARY Scientific Knowledge on the Subject In severe neonatal pulmonary hypertension, lung vascular growth is impaired. Mechanisms that regulate vascular growth during normal lung development and impair angiogenesis in severe neonatal persistent pulmonary hypertension (PPHN) are poorly understood. What This Study Adds to the Field Impaired VEGF–NO signaling within the endothelial cell itself appears to contribute to abnormal vascular growth in PPHN.

 

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				J. Gien, G. J. Seedorf, V. Balasubramaniam, N. Tseng, N. Markham, and S. H. Abman Chronic intrauterine pulmonary hypertension increases endothelial cell Rho kinase activity and impairs angiogenesis in vitro Am J Physiol Lung Cell Mol Physiol, October 1, 2008; 295(4): L680 - L687. [Abstract] [Full Text] [PDF]