Dataset Information

Nogo-B Receptor Modulates Pulmonary Artery Smooth Muscle Cell Function in Developing Lungs.

ABSTRACT: Nogo-B and its receptor (NgBR) are involved in blood vessel growth in developing lungs, but their role in pulmonary artery smooth muscle cell (PASMC) growth is unknown. We hypothesized that NgBR regulates growth of PASMCs by modulating the function of endoplasmic reticulum (ER) and formation of reactive oxygen species (ROS). In utero constriction of the ductus arteriosus created pulmonary hypertension in fetal lambs (hypertensive fetal lamb [HTFL]). PASMCs isolated 8 days after surgery were assessed for the alteration of protein levels by immunoblots and ROS formation by dihydroethidium and Cell ROX deep red fluorescence. NgBR small interfering RNA and plasmid DNA were used to manipulate NgBR levels. Proliferation and wound healing were assessed by cell counts and scratch recovery assay, respectively. Acute ER stress was induced by tunicamycin. Differences of mitogen-activated protein kinase and Akt pathway activation in HTFL versus control PASMCs were evaluated. Results showed that HTFL PASMCs had decreased NgBR levels and increased proliferation, wound healing, ER stress, and ROS formation compared with controls. Knockdown of NgBR in control PASMCs generated a phenotype similar to HTFL, and overexpression in HTFL restored the defective phenotype to control. Decreased NgBR levels were associated with increased ROS formation in HTFL PASMCs. Subsequently, scavenging ROS decreased proliferation and wound healing. Mechanistically, ROS formation decreases NgBR expression, which induces ER stress. This leads to extracellular signal-regulated kinase pathway activation and PASMC phenotype alteration. Our data suggest that decreased NgBR expression in pulmonary hypertension of the newborn contributes to increased PASMC proliferation and oxidative stress, which lead to the pathogenesis of lung injury.

SUBMITTER: Tadokoro KS

PROVIDER: S-EPMC4942214 | biostudies-literature | 2016 Jun

REPOSITORIES: biostudies-literature

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Nogo-B Receptor Modulates Pulmonary Artery Smooth Muscle Cell Function in Developing Lungs.

Tadokoro Kent S KS Rana Ujala U Jing Xigang X Konduri G Ganesh GG Miao Qing R QR Teng Ru-Jeng RJ

American journal of respiratory cell and molecular biology 20160601 6

Nogo-B and its receptor (NgBR) are involved in blood vessel growth in developing lungs, but their role in pulmonary artery smooth muscle cell (PASMC) growth is unknown. We hypothesized that NgBR regulates growth of PASMCs by modulating the function of endoplasmic reticulum (ER) and formation of reactive oxygen species (ROS). In utero constriction of the ductus arteriosus created pulmonary hypertension in fetal lambs (hypertensive fetal lamb [HTFL]). PASMCs isolated 8 days after surgery were asse ...[more]

PMID: 26652754

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Project description:The chronic phase of pulmonary arterial hypertension (PAH) is associated with vascular remodeling, especially thickening of the smooth muscle layer of large pulmonary arteries and muscularization of small pulmonary vessels, which normally have no associated smooth muscle. Serotonin (5-hydroxytryptamine, 5-HT) has been shown to induce proliferation and hypertrophy of pulmonary artery smooth muscle cells (PASMC), and may be important for in vivo pulmonary vascular remodeling. Here, we show that 5-HT stimulates migration of pulmonary artery PASMC. Treatment with 5-HT for 16h increased migration of PASMC up to four-fold as monitored in a modified Boyden chamber assay. Increased migratory responses were associated with cellular morphological changes and reorganization of the actin cytoskeleton. 5-HT-induced alterations in morphology were previously shown in our laboratory to require cAMP [Lee SL, Fanburg BL. Serotonin produces a configurational change of cultured smooth muscle cells that is associated with elevation of intracellular cAMP. J Cell Phys 1992;150(2):396-405], and the 5-HT4 receptor was pharmacologically determined to be the primary activator of cAMP in bovine PASMC [Becker BN, Gettys TW, Middleton JP, Olsen CL, Albers FJ, Lee SL, et al. 8-Hydroxy-2-(di-n-propylamino)tetralin-responsive 5-hydroxytryptamine4-like receptor expressed in bovine pulmonary artery smooth muscle cells. Mol Pharmacol 1992;42(5):817-25]. We examined the role of the 5-HT4 receptor and cAMP in 5-HT-induced bovine PASMC migration. PASMC express 5-HT4 receptor mRNA, and a 5-HT4 receptor antagonist and a cAMP antagonist completely blocked 5-HT-induced cellular migration. Consistent with our previous report that a cAMP-dependent Cl(-) channel is required for 5-HT-induced morphological changes in PASMC, phenylanthranilic acid, a Cl(-) channel blocker, inhibited actin cytoskeletal reorganization and migration produced by 5-HT. We conclude that 5-HT stimulates PASMC migration and associated cytoskeletal reorganization through the 5-HT4 receptor and cAMP activation of a chloride channel.

Project description:Pulmonary arterial hypertension (PAH) is a cardiovascular disease that has high incidence and causes massive deaths. miR-155-5p/PYGL pathway was revealed to play a crucial role in PAH by weighted gene co-expression network analysis (WGCNA). The potential mechanism of miR-155-5p in regulating hypoxia-induced pulmonary artery smooth muscle cell (PASMC) function was analyzed through in vitro experiments. Hypoxia treatment stimulated the proliferation of PASMCs and increased the expression of vascular endothelial growth factor (VEGF) and hypoxia-inducible factor-1α (HIF-1α). At the same time, revealed by qRT-PCR and western blot, the level of miR-155-5p was raised, and the level of PYGL was decreased in hypoxia-induced PASMCs. Through CCK-8 assay, transwell assay and flow cytometry, it was revealed that miR-155-5p inhibitor remarkably inhibited the cell proliferation and migration and decreased the proportion of hypoxia-stimulated PASMCs in S and G2/M phases. Dual-luciferase reporter system was subsequently applied to validate the straight regulation of miR-155-5p on PYGL based on the analysis of online database. Furthermore, siPYGL was revealed to reverse the influence of miR-155-5p inhibitor on hypoxia-induced PASMCs. These outcomes indicate that the increased level of miR-155-5p in hypoxia-stimulated PASMCs could enhance the cell proliferation, cell migration, and cell cycle progression by targeting PYGL directly. This study may supply novel treatment strategies for PAH.Abbreviations: PH, pulmonary hypertension; PAH, pulmonary arterial hypertension; WGCNA, weighted gene co-expression network analysis; PASMCs, pulmonary artery smooth muscle cells; VEGF, vascular endothelial growth factor; HIF-1α, hypoxia-inducible factor-1α; SMCs, smooth muscle cells; DEGs, differentially expressed genes; GEO, Gene Expression Omnibus; GO, Gene Ontology; KEGG, Kyoto Encyclopedia of Genes and Genomes; FBS, fetal bovine serum; OD, optical density; BCA, bicinchoninic acid; PVDF, polyvinylidene fluoride; PBS, phosphate-buffered saline; BP, biological process; MF, molecular function; CC, cell component.

Dataset Information

Nogo-B Receptor Modulates Pulmonary Artery Smooth Muscle Cell Function in Developing Lungs.

Publications

Nogo-B Receptor Modulates Pulmonary Artery Smooth Muscle Cell Function in Developing Lungs.

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