Hypoxic constriction and reactive oxygen species in porcine distal  pulmonary arteries

To determine whether reactive oxygen species (ROS) play an essential role in hypoxic pulmonary vasoconstriction (HPV) and the cellular locus of ROS production and action during HPV, we measured internal diameter (ID) at constant transmural pressure, lucigenin-derived chemiluminescence (LDCL), and electron paramagnetic resonance (EPR) spin adduct spectra in small distal porcine pulmonary arteries, and dichlorofluorescein (DCF) fluorescence in myocytes isolated from these arteries. Hypoxia (4% O2) decreased ID, increased DCF fluorescence, tended to increase LDCL, and in some preparations produced EPR spectra consistent with hydroxyl and alkyl radicals. Superoxide dismutase (SOD, 150 U/ml) or SOD + catalase (CAT, 200 U/ml) did not alter ID during normoxia but reduced or abolished the constriction induced by hypoxia. SOD also blocked HPV in endotheliumdenuded arteries after restoration of the response by exposure to 10-10 M endothelin-1. Confocal fluorescence microscopy demonstrated that labeled SOD and CAT entered pulmonary arterial myocytes. SOD, SOD + CAT, and CAT blocked the increase in DCF fluorescence induced by hypoxia, but SOD + CAT and CAT also caused a stable increase in fluorescence during normoxia, suggesting that CAT diminished efflux of DCF from cells or oxidized the dye directly. We conclude that HPV required increased concentrations of ROS produced by and acting on pulmonary arterial smooth muscle rather than endothelium.

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Actin polymerization contributes to enhanced pulmonary vasoconstrictor reactivity after chronic hypoxia

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00664.2017 ◽

2018 ◽

Vol 314 (5) ◽

pp. H1011-H1021 ◽

Cited By ~ 7

Author(s):

Laura Weise-Cross ◽

Michelle A. Sands ◽

Joshua R. Sheak ◽

Brad R. S. Broughton ◽

Jessica B. Snow ◽

...

Keyword(s):

Reactive Oxygen Species ◽

Actin Polymerization ◽

Chronic Hypoxia ◽

Pulmonary Arteries ◽

Rho Kinase ◽

Reactive Oxygen ◽

Dependent Manner ◽

Oxygen Species ◽

Pulmonary Arterial ◽

Arterial Constriction

Chronic hypoxia (CH) augments basal and endothelin-1 (ET-1)-induced pulmonary vasoconstrictor reactivity through reactive oxygen species (ROS) generation and RhoA/Rho kinase (ROCK)-dependent myofilament Ca2+ sensitization. Because ROCK promotes actin polymerization and the actin cytoskeleton regulates smooth muscle tension, we hypothesized that actin polymerization is required for enhanced basal and ET-1-dependent vasoconstriction after CH. To test this hypothesis, both end points were monitored in pressurized, endothelium-disrupted pulmonary arteries (fourth-fifth order) from control and CH (4 wk at 0.5 atm) rats. The actin polymerization inhibitors cytochalasin and latrunculin attenuated both basal and ET-1-induced vasoconstriction only in CH vessels. To test whether CH directly alters the arterial actin profile, we measured filamentous actin (F-actin)-to-globular actin (G-actin) ratios by fluorescent labeling of F-actin and G-actin in fixed pulmonary arteries and actin sedimentation assays using homogenized pulmonary artery lysates. We observed no difference in actin polymerization between groups under baseline conditions, but ET-1 enhanced actin polymerization in pulmonary arteries from CH rats. This response was blunted by the ROS scavenger tiron, the ROCK inhibitor fasudil, and the mDia (RhoA effector) inhibitor small-molecule inhibitor of formin homology domain 2. Immunoblot analysis revealed an effect of CH to increase both phosphorylated (inactive) and total levels of the actin disassembly factor cofilin but not phosphorylated cofilin-to-total cofilin ratios. We conclude that actin polymerization contributes to increased basal pulmonary arterial constriction and ET-1-induced vasoconstrictor reactivity after CH in a ROS- and ROCK-dependent manner. Our results further suggest that enhanced ET-1-mediated actin polymerization after CH is dependent on mDia but independent of changes in the phosphorylated cofilin-to-total cofilin ratio. NEW & NOTEWORTHY This research is the first to demonstrate a role for actin polymerization in chronic hypoxia-induced basal pulmonary arterial constriction and enhanced agonist-induced vasoconstrictor activity. These results suggest that a reactive oxygen species-Rho kinase-actin polymerization signaling pathway mediates this response and may provide a mechanistic basis for the vasoconstrictor component of pulmonary hypertension.

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Recent advances in oxygen sensing and signal transduction in hypoxic pulmonary vasoconstriction

Journal of Applied Physiology ◽

10.1152/japplphysiol.00103.2017 ◽

2017 ◽

Vol 123 (6) ◽

pp. 1647-1656 ◽

Cited By ~ 4

Author(s):

Ievgen Strielkov ◽

Oleg Pak ◽

Natasha Sommer ◽

Norbert Weissmann

Keyword(s):

Reactive Oxygen Species ◽

Transient Receptor Potential ◽

Oxygen Sensing ◽

Hypoxic Pulmonary Vasoconstriction ◽

Reactive Oxygen ◽

Receptor Potential ◽

Complex Iii ◽

Oxygen Species ◽

Pulmonary Vasoconstriction ◽

Transient Receptor

Hypoxic pulmonary vasoconstriction (HPV) is a physiological reaction, which adapts lung perfusion to regional ventilation and optimizes gas exchange. Impaired HPV may cause systemic hypoxemia, while generalized HPV contributes to the development of pulmonary hypertension. The triggering mechanisms underlying HPV are still not fully elucidated. Several hypotheses are currently under debate, including a possible decrease as well as an increase in reactive oxygen species as a triggering event. Recent findings suggest an increase in the production of reactive oxygen species in pulmonary artery smooth muscle cells by complex III of the mitochondrial electron transport chain and occurrence of oxygen sensing at complex IV. Other essential components are voltage-dependent potassium and possibly L-type, transient receptor potential channel 6, and transient receptor potential vanilloid 4 channels. The release of arachidonic acid metabolites appears also to be involved in HPV regulation. Further investigation of the HPV mechanisms will facilitate the development of novel therapeutic strategies for the treatment of HPV-related disorders.

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Cyclic stretch increases VEGF expression in pulmonary arterial smooth muscle cells via TGF-β1 and reactive oxygen species: a requirement for NAD(P)H oxidase

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00417.2004 ◽

2005 ◽

Vol 289 (2) ◽

pp. L288-L289 ◽

Cited By ~ 68

Author(s):

Eugenia Mata-Greenwood ◽

Albert Grobe ◽

Sanjiv Kumar ◽

Yelina Noskina ◽

Stephen M. Black

Keyword(s):

Reactive Oxygen Species ◽

Blood Flow ◽

Smooth Muscle ◽

Reactive Oxygen ◽

Cyclic Stretch ◽

Oxygen Species ◽

Vegf Expression ◽

Pulmonary Vessels ◽

Pulmonary Arterial ◽

Tgf Β1

Our previous studies have indicated that transforming growth factor (TGF)-β1 and VEGF expression are increased in the smooth muscle cell (SMC) layer of the pulmonary vessels of lambs with pulmonary hypertension secondary to increased pulmonary blood flow. Furthermore, we found that TGF-β1 expression increased before VEGF. Because of the increased blood flow in the shunt lambs, the SMC in the pulmonary vessels are exposed to increased levels of the mechanical force, cyclic stretch. Thus, in this study, using primary cultures of pulmonary arterial SMC isolated from pulmonary arteries of 4-wk-old lambs, we investigated the role of cyclic stretch in the apparent coordinated regulation of TGF-β1 and VEGF. Our results demonstrated that cyclic stretch induced a significant increase in VEGF expression both at the mRNA and protein levels ( P < 0.05). The increased VEGF mRNA was preceded by both an increased expression and secretion of TGF-β1 and an increase in reactive oxygen species (ROS) generation. In addition, a neutralizing antibody against TGF-β1 abolished the cyclic stretch-dependent increases in both superoxide generation and VEGF expression. Our data also demonstrated that cyclic stretch activated an NAD(P)H oxidase that was TGF-β1 dependent and that NAD(P)H oxidase inhibitors abolished the cyclic stretch-dependent increase in VEGF expression. Therefore, our results indicate that cyclic stretch upregulates VEGF expression via the TGF-β1-dependent activation of NAD(P)H oxidase and increased generation of ROS.

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