Smooth muscle F-actin disassembly and RhoA/Rho-kinase signaling during endotoxin-induced alterations in pulmonary arterial compliance

2004 ◽  
Vol 287 (4) ◽  
pp. L649-L655 ◽  
Author(s):  
Christa Boer ◽  
Geerten P. van Nieuw Amerongen ◽  
A. B. Johan Groeneveld ◽  
Gert Jan Scheffer ◽  
Jaap J. de Lange ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Actin Cytoskeleton ◽  
Vascular Function ◽  
Kinase Inhibitor ◽  
Arterial Compliance ◽  
Pulmonary Arteries ◽  
Rho Kinase ◽  
Kinase Signaling ◽  
Arterial Mechanics ◽  
Pulmonary Arterial

Endotoxemia is associated with changed pulmonary vascular function with respect to vasoreactivity, endothelial permeability, and activation of inducible nitric oxide synthase II (NOSII). However, whether altered passive arterial wall mechanics contribute to this endotoxin-induced pulmonary vascular dysfunction is still unknown. Therefore, we investigated whether endotoxin affects the passive arterial mechanics and compliance of isolated rat pulmonary arteries. Pulmonary arteries of pentobarbital-anesthetized Wistar rats ( n = 55) were isolated and exposed to Escherichia coli endotoxin (50 μg/ml) for 20 h. Endotoxin increased pulmonary artery diameter and compliance (transmural pressure = 13 mmHg) in an endothelium-, Ca2+-, or NOSII-induced NO release-independent manner. Interestingly, the endotoxin-induced alterations in the passive arterial mechanics were accompanied by disassembly of the smooth muscle cell (SMC) F-actin cytoskeleton. Disassembly of F-actin by incubation of control arteries with the cytoskeleton-disrupting agent cytochalasin B or the Rho-kinase inhibitor Y-27632 induced a similar increase in passive arterial diameter and compliance. In contrast, RhoA activation by lysophosphatidic acid prevented the endotoxin-induced alterations in the pulmonary SMC F-actin cytoskeleton and passive mechanics. In conclusion, these findings indicate that disassembly of the SMC F-actin cytoskeleton and RhoA/Rho-kinase signaling act as mediators of endotoxin-induced changes in the pulmonary arterial mechanics. They imply the involvement of F-actin rearrangement and RhoA/Rho-kinase signaling in endotoxemia-induced vascular lung injury.

Involvement of RhoA/Rho kinase signaling in pulmonary hypertension of the fawn-hooded rat

2006 ◽  
Vol 100 (3) ◽  
pp. 996-1002 ◽  
Author(s):  
Tetsutaro Nagaoka ◽  
Sarah A. Gebb ◽  
Vijaya Karoor ◽  
Noriyuki Homma ◽  
Kenneth G. Morris ◽  
...  
Keyword(s):  
Pulmonary Hypertension ◽  
Sea Level ◽  
Kinase Inhibitor ◽  
Pulmonary Arteries ◽  
Rho Kinase ◽  
Kinase Signaling ◽  
Rhoa Activity ◽  
Pulmonary Arterial ◽  
Mild Hypoxia

The fawn-hooded rat (FHR) develops severe pulmonary hypertension (PH) when raised for the first 3–4 wk of life in the mild hypoxia of Denver’s altitude (5,280 ft.). The PH is associated with sustained pulmonary vasoconstriction and pulmonary artery remodeling. Furthermore, lung alveolarization and vascularization are reduced in the Denver FHR. We have recently shown that RhoA/Rho kinase signaling is involved in both vasoconstriction and vascular remodeling in animal models of hypoxic PH. In this study, we investigated the role of RhoA/Rho kinase signaling in the PH of Denver FHR. In α-toxin permeabilized pulmonary arteries from Denver FHR, the contractile sensitivity to Ca2+was increased compared with those from sea-level FHR. RhoA activity and Rho kinase I protein expression in pulmonary arteries of Denver FHR (10-wk-old) were higher than in those of sea-level FHR. Acute inhalation of the Rho kinase inhibitor fasudil selectively reduced the elevated pulmonary arterial pressure in Denver FHR in vivo. Chronic fasudil treatment (30 mg·kg−1·day−1, from birth to 10 wk old) markedly reduced the development of PH and improved lung alveolarization and vascularization in Denver FHR. These results suggest that Rho kinase-mediated sustained vasoconstriction, through increased Ca2+sensitivity, plays an important role in the established PH and that RhoA/Rho kinase signaling contributes significantly to the development of PH and lung dysplasia in mild hypoxia-exposed FHR.

Effect of changes in pH on wall tension in isolated rat pulmonary artery: role of the RhoA/Rho-kinase pathway

2004 ◽  
Vol 287 (4) ◽  
pp. L673-L684 ◽  
Author(s):  
Jean-Marc Hyvelin ◽  
Clare O’Connor ◽  
Paul McLoughlin
Keyword(s):  
Smooth Muscle ◽  
Vascular Smooth Muscle ◽  
Kinase Inhibitor ◽  
Pulmonary Arteries ◽  
Rho Kinase ◽  
Systemic Circulation ◽  
Pulmonary Vasculature ◽  
Tension Development ◽  
Kinase Pathway

Pulmonary arteries (PA) are resistant to the vasodilator effects of extracellular acidosis in systemic vessels; the mechanism underlying this difference between systemic and pulmonary circulations has not been elucidated. We hypothesized that RhoA/Rho-kinase-mediated Ca2+ sensitization pathway played a greater role in tension development in pulmonary than in systemic vascular smooth muscle and that this pathway was insensitive to acidosis. In arterial rings contracted with the α1-agonist phenylephrine (PE), the Rho-kinase inhibitor Y-27632 (≤3 μM) induced greater relaxation in precontracted PA rings than in aortic rings. In PA rings stimulated by PE, the activation of RhoA was greater than in aorta. Normocapnic acidosis (NA) induced a smaller relaxation in precontracted PA than in aorta. However, in the presence of nifedipine and thapsigargin, when PE-induced contraction was predominantly mediated by Rho-kinase, the relaxant effect of NA was reduced and similar in both vessel types. Furthermore, in the presence of Y-27632, NA induced a greater relaxation in both PA and aorta, which was similar in both vessels. Finally, in α-toxin-permeabilized smooth muscle, PE-induced contraction at constant Ca2+ activity was inhibited by Y-27632 and unaffected by acidosis. These results indicate that Ca2+ sensitization induced by the RhoA/Rho-kinase pathway played a greater role in agonist-induced vascular smooth muscle contraction in PA than in aorta and that tension mediated by this pathway was insensitive to acidosis. The predominant role of the RhoA/Rho-kinase pathway in the pulmonary vasculature may account for the resistance of this circulation to the vasodilator effect of acidosis observed in the systemic circulation.

Abstract 59: Loss of Cullin-3 in Smooth Muscle Increases RhoA, Vascular Contraction and Arterial Pressure

Hypertension ◽  
2012 ◽  
Vol 60 (suppl_1) ◽  
Author(s):  
Christopher Pelham ◽  
Pimonrat Ketsawatsomkron ◽  
Justin Grobe ◽  
Frank Faraci ◽  
Curt Sigmund
Keyword(s):  
Smooth Muscle ◽  
Arterial Pressure ◽  
Vascular Function ◽  
Enzyme Inhibitor ◽  
Cyclin E ◽  
Rho Kinase ◽  
Dominant Negative ◽  
Dark Phase ◽  
Kinase Signaling ◽  
Cullin 3

Mutations in Cullin-3 result in early-onset hypertension in humans through an undefined mechanism. Cullin-3 RING E3 ubiquitin ligase complex mediates proteasomal degradation of RhoA, a key mediator of vascular smooth muscle tone. We reported that smooth muscle-specific expression of dominant negative PPARγ (S-P467L) in transgenic mice causes hypertension and aortic dysfunction via increased RhoA/Rho-kinase signaling. Dominant negative PPARγ caused decreased Cullin-3 protein (0.31±0.04-fold of Non-Transgenic (NT); P<0.05) and decreased ratio of active Nedd8-Cullin-3 / Cullin-3 (0.06±0.04 vs 0.51±0.18 NT; P<0.05), which correlated with increased protein levels of Cullin-3 substrates RhoA (2.6±0.2-fold of NT; P<0.05) and Cyclin E (2.0±0.1-fold of NT; P<0.05) in S-P467L medial aorta. We tested the hypothesis that Cullin-3 controls arterial pressure by regulating vascular function. siRNA-mediated knockdown of Cullin-3 (72 hr), confirmed by Western blot, increased RhoA (2.5±0.1-fold of NC1 negative control siRNA; P<0.05) and Cyclin E (1.9±0.2-fold of NC1; P<0.05) protein in primary rat aortic smooth muscle cells. Inhibition of cullin-RING ligase activity using the Nedd8-activating enzyme inhibitor, MLN4924 (1μM, 16 hr), also increased RhoA (4.3±1.0-fold of DMSO vehicle). Treatment of aortic rings from control mice with MLN4924 (1μM in DMEM/F12, 16 hr) resulted in enhanced agonist-mediated contraction in response to endothelin 1 (30nM: 0.35±0.05g vs 0.19±0.02g DMSO vehicle; P<0.05), serotonin (3μM: 1.2±0.07g vs 0.9±0.08g; P<0.05), and phenylephrine (3μM: 1.1±0.05g vs 0.7±0.05g, P<0.05) that was Rho-kinase-dependent, despite decreased contraction to KCl (100mM: 0.7±0.03g vs 1.0±0.03g; P<0.05). Finally, administration of MLN4924 (30mg/kg TID, 2 days) to control mice in vivo increased mean arterial pressure during the light phase (121±4 vs. 108±3mmHg baseline, P<0.05; dark phase: 122±4 vs. 123±4mmHg baseline, NS) as measured by radiotelemetry. Our results demonstrate that interference with PPARγ in smooth muscle causes vascular dysfunction via impaired Cullin-3-mediated regulation of RhoA/Rho-kinase signaling and provide a mechanistic link between mutations in Cullin-3 and hypertension.

Rho/Rho kinase signaling mediates increased basal pulmonary vascular tone in chronically hypoxic rats

2004 ◽  
Vol 287 (4) ◽  
pp. L665-L672 ◽  
Author(s):  
Tetsutaro Nagaoka ◽  
Yoshiteru Morio ◽  
Nina Casanova ◽  
Natalie Bauer ◽  
Sarah Gebb ◽  
...  
Keyword(s):  
Salt Solution ◽  
Vascular Tone ◽  
Kinase Inhibitor ◽  
Pulmonary Arteries ◽  
Rho Kinase ◽  
Vascular Diseases ◽  
Physiological Salt Solution ◽  
Kinase Signaling ◽  
Arterial Blood ◽  
Rho Kinase Inhibitor

Recent evidence suggests that Rho/Rho kinase signaling plays an important role in the sustained vasoconstriction induced by many agonists and is involved in the pathogenesis of systemic vascular diseases. However, little is known about its role in increased vascular tone in hypoxic pulmonary hypertension (PH). The purpose of this study was to examine whether Rho/Rho kinase-mediated Ca2+ sensitization contributed to sustained vasoconstriction and increased vasoreactivity in hypoxic PH in rats. Acute intravenous administration of Y-27632, a Rho kinase inhibitor, nearly normalized the high pulmonary arterial blood pressure and total pulmonary resistance in chronically hypoxic rats. In contrast to nifedipine, Y-27632 also markedly decreased elevated basal vascular tone in hypertensive blood-perfused lungs and isolated pulmonary arteries. Y-27632 and another Rho kinase inhibitor, HA-1077, completely reversed nitro-l-arginine-induced vasoconstriction in physiological salt solution-perfused hypertensive lungs, whereas inhibitors of myosin light chain kinase (ML-9), protein kinase C (GF-109203X), phosphatidylinositol 3-kinase (LY-294002), and tyrosine kinase (tyrphostin A23) caused only partial or no reversal of the vasoconstriction. Vasoconstrictor responses to KCl were augmented in hypertensive physiological salt solution-perfused lungs and pulmonary arteries, and the augmentation was eliminated by Y-27632. These results suggest that Rho/Rho kinase-mediated Ca2+ sensitization plays a central role in mediating sustained vasoconstriction and increased vasoreactivity in hypoxic PH.

Abstract 1633: RhoA and Rho Kinase Activation in Human Pulmonary Hypertension - Role of 5-HT Signaling

Circulation ◽  
2008 ◽  
Vol 118 (suppl_18) ◽  
Author(s):  
Christophe Guilluy ◽  
Saadia Eddahibi ◽  
Christian Agard ◽  
Laurent Savale ◽  
Elie Fadel ◽  
...  
Keyword(s):  
Pulmonary Hypertension ◽  
Smooth Muscle ◽  
Kinase Activity ◽  
Kinase Inhibitor ◽  
Rho Kinase ◽  
Strong Increase ◽  
Kinase Signaling ◽  
Kinase Activation ◽  
Promising Target ◽  
New Treatments

Background- The complex and multifactorial pathogenesis of pulmonary hypertension (PH) involves constriction, remodeling, and in situ thrombosis of pulmonary vessels. Both serotonin (5-HT) and Rho kinase signaling may contribute to these alterations. Here, we investigated possible links between the 5-HT transporter (5-HTT) and RhoA/Rho kinase pathways, as well as their involvement in the progression of human and experimental PH. Methods and Results- Lungs, platelets, and quiescent cultured pulmonary-artery smooth-muscle cells (PA-SMCs) from patients with idiopathic PH (iPH) were characterized by marked elevations in RhoA and Rho kinase activity and by a strong increase in serotonin binding to RhoA, compared to controls. The 5-HTT inhibitor fluoxetine and the type 2 transglutaminase inhibitor monodansylcadaverin prevented 5-HT-induced RhoA serotonylation and RhoA/Rho kinase activation, as well as 5-HT-induced proliferation of PA-SMCs from iPH patients, that was also inhibited by the Rho kinase inhibitor fasudil. Increased Rho kinase activity, RhoA activation, and RhoA serotonylation were also observed in lungs from SM22–5-HTT+ mice, which overexpress 5-HTT in smooth muscle and spontaneously develop PH. Treatment of SM22–5-HTT+ mice with either fasudil or fluoxetine limited both PH progression and RhoA/Rho kinase activation. Conclusions- RhoA and Rho kinase activities are increased in iPH, as a result of transglutaminase-mediated transamidation of RhoA by 5-HT internalized via 5-HTT. Direct involvement of this pathway in 5-HT-mediated PA-SMC proliferation and platelet activation during PH progression identify RhoA/Rho kinase signaling as a promising target for new treatments against PH.

Vasoconstrictor effect of endothelin-1 on hypertensive pulmonary arterial smooth muscle involves Rho-kinase and protein kinase C

2007 ◽  
Vol 293 (2) ◽  
pp. L472-L479 ◽  
Author(s):  
Scott A. Barman
Keyword(s):  
Pulmonary Hypertension ◽  
Smooth Muscle ◽  
Protein Kinase C ◽  
Pulmonary Arteries ◽  
Rho Kinase ◽  
Vascular Response ◽  
Arterial Smooth Muscle ◽  
Kinase C ◽  
Pulmonary Arterial ◽  
Pulmonary Arterial Smooth Muscle

Although one of the common characteristics of pulmonary hypertension is abnormal sustained vasoconstriction, the signaling pathways that mediate this heightened pulmonary vascular response are still not well defined. Protein kinase C (PKC) and Rho-kinase are regulators of smooth muscle contraction induced by G protein-coupled receptor agonists including endothelin-1 (ET-1), which has been implicated as a signaling pathway in pulmonary hypertension. Toward this end, it was hypothesized that both Rho-kinase and PKC mediate the pulmonary vascular response to ET-1 in hypertensive pulmonary arterial smooth muscle, and therefore, the purpose of this study was to determine the role of PKC and Rho-kinase signaling in ET-1-induced vasoconstriction in both normotensive (Sprague-Dawley) and hypertensive (Fawn-Hooded) rat pulmonary arterial smooth muscle. Results indicate that ET-1 caused greater vasoconstriction in hypertensive pulmonary arteries compared with the normal vessels, and treatment with the PKC antagonists chelerythrine, rottlerin, and Gö 6983 inhibited the vasoconstrictor response to ET-1 in the hypertensive vessels. In addition, the specific Rho-kinase inhibitor Y-27632 significantly attenuated the effect of ET-1 in both normotensive and hypertensive phenotypes, with greater inhibition occurring in the hypertensive arteries. Furthermore, Western blot analysis revealed that ET-1 increased RhoA expression in both normotensive and hypertensive pulmonary arteries, with expression being greater in the hypertensive state. These results suggest that both PKC and Rho/Rho-kinase mediate the heightened pulmonary vascular response to ET-1 in hypertensive pulmonary arterial smooth muscle.

Abstract 641: Expression Of GLUT4 In Vascular Smooth Muscle Affects Endothelial Function In Hypertension.

Hypertension ◽  
2013 ◽  
Vol 62 (suppl_1) ◽  
Author(s):  
Kevin B Atkins ◽  
Jharna Saha ◽  
Frank C Brosius
Keyword(s):  
Smooth Muscle ◽  
Vascular Smooth Muscle ◽  
Kinase Inhibitor ◽  
Rho Kinase ◽  
Interesting Phenomenon ◽  
Glut4 Expression ◽  
Endothelial Denudation ◽  
Rho Kinase Inhibitor ◽  
Total Body ◽  
Rats And Mice

Expression of GLUT4 is decreased in arterial smooth muscle of hypertensive rats and mice, and total body overexpression of GLUT4 in mice prevents enhanced arterial reactivity. To demonstrate that the effect on vascular response to GLUT4 overexpression is vascular rather than systemic in origin we utilized smooth muscle-specific GLUT4 transgenic mice (SMG4). GLUT4 expression in aortae of SMG4 compared to WT mice was increased 2-3 fold. Adult wild-type (WT) and SMG4 mice were made hypertensive or not through implantation of angiotensin II (AngII; 1.4mg/kg/d for 2 wks) or vehicle containing osmotic mini-pumps. Both WT and SMG4 mice AngII-treated mice exhibited significantly increased systolic blood pressure. In AngII-treated WT mice (WT-AngII) aortic GLUT4 expression was significantly decreased, whereas GLUT4 expression in aortae of AngII-treated SMG4 mice (SMG4-AngII) was maintained. The phosphorylation of ERM and MYPT1(Thr850) were significantly increased in aortae of WT-AngII compared to WT-Sham and SMG4-AngII mice. Responsiveness to the contractile agonists, phenylephrine, 5-HT, and PGF 2 was significantly increased in endothelium-intact aortic rings from WT-AngII mice, but remained normal in aortae of SMG4-AngII mice. Following pretreatment with Rho-kinase inhibitor Y-27632, relative inhibition of contractility to 5-HT was equal in aortae from WT-AngII and SMG4-AngII-treated mice. With endothelial denudation, contractility to 5-HT was equally enhanced in aortae of WT-AngII and SMG4-AngII-treated mice. Interestingly, whereas acetylcholine stimulated relaxation was significantly decreased in aortic rings of WT-AngII mice, relaxation in rings from SMG4-AngII mice was not significantly different from WT or SMG4. These results demonstrate an interesting phenomenon whereby decreased expression of GLUT4 in vascular smooth muscle leads to an endothelial dysfunction that not only impairs relaxation, but also enhances contractility.

scholarly journals Effects of acute Rho kinase inhibition on chronic hypoxia-induced changes in proximal and distal pulmonary arterial structure and function

2011 ◽  
Vol 110 (1) ◽  
pp. 188-198 ◽  
Author(s):  
Rebecca R. Vanderpool ◽  
Ah Ram Kim ◽  
Robert Molthen ◽  
Naomi C. Chesler
Keyword(s):  
Pulmonary Artery ◽  
Pulmonary Arteries ◽  
Rho Kinase ◽  
Pressure Flow ◽  
Kinase Inhibition ◽  
Pulmonary Vasoconstriction ◽  
Pulsatile Pressure ◽  
Arterial Structure ◽  
Pulmonary Arterial ◽  
Induced Changes

Hypoxic pulmonary hypertension (HPH) is initially a disease of the small pulmonary arteries. Its severity is usually quantified by pulmonary vascular resistance (PVR). Acute Rho kinase inhibition has been found to reduce PVR toward control values in animal models, suggesting that persistent pulmonary vasoconstriction is the dominant mechanism for increased PVR. However, HPH may also cause proximal arterial changes, which are relevant to right ventricular (RV) afterload. RV afterload can be quantified by pulmonary vascular impedance, which is obtained via spectral analysis of pulsatile pressure-flow relationships. To determine the effects of HPH independent of persistent pulmonary vasoconstriction in proximal and distal arteries, we quantified pulsatile pressure-flow relationships before and after acute Rho kinase inhibition and measured pulmonary arterial structure with microcomputed tomography. In control lungs, Rho kinase inhibition decreased 0 Hz impedance (Z0), which is equivalent to PVR, from 2.1 ± 0.4 to 1.5 ± 0.2 mmHg·min·ml−1 ( P < 0.05) and tended to increase characteristic impedance (ZC) from 0.21 ± 0.01 to 0.22 ± 0.01 mmHg·min·ml−1. In HPH lungs, Rho kinase inhibition decreased Z0 ( P < 0.05) without affecting ZC. Microcomputed tomography measurements performed on lungs after acute Rho kinase inhibition demonstrated that HPH significantly decreased the unstressed diameter of the main pulmonary artery (760 ± 60 vs. 650 ± 80 μm; P < 0.05), decreased right pulmonary artery compliance, and reduced the frequency of arteries of diameter 50–100 μm (both P < 0.05). These results demonstrate that acute Rho kinase inhibition reverses many but not all HPH-induced changes in distal pulmonary arteries but does not affect HPH-induced changes in the conduit arteries that impact RV afterload.

Acute hypoxia increases intracellular [Ca2+] in pulmonary arterial smooth muscle by enhancing capacitative Ca2+ entry

2005 ◽  
Vol 288 (6) ◽  
pp. L1059-L1069 ◽  
Author(s):  
Jian Wang ◽  
Larissa A. Shimoda ◽  
Letitia Weigand ◽  
Wenqian Wang ◽  
Dejun Sun ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Hypoxic Pulmonary Vasoconstriction ◽  
Acute Hypoxia ◽  
Primary Cultures ◽  
Fluorescent Microscopy ◽  
Pulmonary Arteries ◽  
Arterial Smooth Muscle ◽  
Fura 2 ◽  
Pulmonary Arterial ◽  
Pulmonary Arterial Smooth Muscle

Hypoxic pulmonary vasoconstriction (HPV) requires influx of extracellular Ca2+ in pulmonary arterial smooth muscle cells (PASMCs). To determine whether capacitative Ca2+ entry (CCE) through store-operated Ca2+ channels (SOCCs) contributes to this influx, we used fluorescent microscopy and the Ca2+-sensitive dye fura-2 to measure effects of 4% O2 on intracellular [Ca2+] ([Ca2+]i) and CCE in primary cultures of PASMCs from rat distal pulmonary arteries. In PASMCs perfused with Ca2+-free Krebs Ringer bicarbonate solution (KRBS) containing cyclopiazonic acid to deplete Ca2+ stores in sarcoplasmic reticulum and nifedipine to prevent Ca2+ entry through L-type voltage-operated Ca2+ channels (VOCCs), hypoxia markedly enhanced both the increase in [Ca2+]i caused by restoration of extracellular [Ca2+] and the rate at which extracellular Mn2+ quenched fura-2 fluorescence. These effects, as well as the increased [Ca2+]i caused by hypoxia in PASMCs perfused with normal salt solutions, were blocked by the SOCC antagonists SKF-96365, NiCl2, and LaCl3 at concentrations that inhibited CCE >80% but did not alter [Ca2+]i responses to 60 mM KCl. In contrast, the VOCC antagonist nifedipine inhibited [Ca2+]i responses to hypoxia by only 50% at concentrations that completely blocked responses to KCl. The increased [Ca2+]i caused by hypoxia was completely reversed by perfusion with Ca2+-free KRBS. LaCl3 increased basal [Ca2+]i during normoxia, indicating effects other than inhibition of SOCCs. Our results suggest that acute hypoxia enhances CCE through SOCCs in distal PASMCs, leading to depolarization, secondary activation of VOCCs, and increased [Ca2+]i. SOCCs and CCE may play important roles in HPV.

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