Contraction Of Pulmonary Artery Rings Research Articles

C60 fullerenes selectively inhibit BKCa but not Kv channels in pulmonary artery smooth muscle cells

Possessing unique physical and chemical properties, C60 fullerenes are arising as a potential nanotechnological tool that can strongly affect various biological processes. Recent molecular modeling studies have shown that C60 fullerenes can interact with ion channels, but there is lack of data about possible effects of C60 molecule on ion channels expressed in smooth muscle cells (SMC). Here we show both computationally and experimentally that water-soluble pristine C60 fullerene strongly inhibits the large conductance Ca2+-dependent K+ (BKCa), but not voltage-gated K+ (Kv) channels in pulmonary artery SMC. Both molecular docking simulations and analysis of single channel activity indicate that C60 fullerene blocks BKCa channel pore in its open state. In functional tests, C60 fullerene enhanced phenylephrine-induced contraction of pulmonary artery rings by about 25% and reduced endothelium-dependent acetylcholine-induced relaxation by up to 40%. These findings suggest a novel strategy for biomedical application of water-soluble pristine C60 fullerene in vascular dysfunction.

In vitro effect of endothelin-1 and nifedipine on pulmonary vascular contraction of pulmonary hypertensive and non-pulmonary hypertensive chickens

ABSTRACTEndothelin-1 (ET-1) is relevant to the pathogenesis of pulmonary arterial hypertension (PAH) in several species, including broiler chickens. In ET-1-induced vasoconstriction, L-type voltage-operated calcium channels (VOCCs) play a crucial role. In this study, the effect that ET-1 and the calcium channel blocker (CCB) nifedipine have on ET-1-induced vascular contraction in pulmonary artery rings of both, non-pulmonary hypertensive chickens (NPHC) and pulmonary hypertensive chickens (PHC), was evaluated. A significant increase response in ET-1 was observed for PHC vascular rings with respect to those of NPHC (p < .05). Incubation with 1 μM nifedipine did not completely reverse the contraction induced by 1 × 10−7 M ET-1 in both NPHC and PHC rings, suggesting the participation of additional calcium sources procured via influx through L-type VOCC. Nifedipine reduced, to a higher degree, the contraction induced by ET-1 in NPHC (50%) when compared with that observed in PHC rings (30.7%). Thus, the ET-1-induced vasoconstriction processes are more efficient in PHC, and the influx of calcium into the cell through the L-type VOCC is presumably only a fraction of the necessary calcium for ET-1-induced vasoconstriction. It is suggested that blocking calcium uptake through alternative pathways (receptor-operated calcium channels and storage-operated calcium channels) may produce a vasodilation effect of considerable therapeutic significance in PHC.

Inducible nitric oxide synthase inhibition reverses pulmonary arterial dysfunction in lung transplantation.

Ischemia-reperfusion injury (IRI) after lung transplantation remains a significant cause of morbidity and mortality. Lung IRI induces nitric oxide synthesis (iNOS) and reactive nitrogen species, decreasing nitric oxide bioavailability. We hypothesized that ischemia-induced iNOS intensifies with reperfusion and contributes to IRI-induced pulmonary arterial regulatory dysfunction, which may lead to early graft failure and cause pulmonary edema. The aim of this study was to determine whether ischemia-reperfusion alters inducible and endothelial nitric oxide synthase expression, potentially affecting pulmonary perfusion. We further evaluated the role of iNOS in post-transplantation pulmonary arterial disorder. We randomized 32 Sprague-Dawley rats into two groups. The control group was given a sham operation whilst the experimental group received orthotropic lung transplants with a modified three-cuff technique. Changes in lung iNOS, and endothelial nitric oxide synthase expression were measured after lung transplantation by enzyme-linked immunosorbent assay (ELISA). Vasoconstriction in response to exogenous phenylephrine and vasodilation in response to exogenous acetylcholine of pulmonary arterial rings were measured in vitro as a measure of vascular dysfunction. To elucidate the roles of iNOS in regulating vascular function, an iNOS activity inhibitor (N6-(1-iminoethyl)-L-lysine, L-NIL) was used to treat isolated arterial rings. In order to test whether iNOS inhibition has a therapeutic effect, we further used L-NIL to pre-treat transplanted lungs and then measured post-transplantation arterial responses. Lung transplantation caused upregulation of iNOS expression. This was also accompanied by suppression of both vasoconstriction and vasodilation of arterial rings from transplanted lungs. Removal of endothelium did not interfere with the contraction of pulmonary arterial rings from transplanted lungs. In contrast, iNOS inhibition rescued the vasoconstriction response to exogenous phenylephrine of pulmonary arterial rings from transplanted lungs. In addition, lung transplantation led to suppression of PaO2/FiO2 ratio, increased intrapulmonary shunt (Q s/Q t), and increase of lung wet to dry ratio (W/D), malondialdehyde and myeloperoxidase levels, all of which were reversed upon iNOS inhibition. Furthermore, inhibition of iNOS significantly rescued vascular function and alleviated edema and inflammatory cell infiltration in the transplanted lung. Our data suggest that lung transplantation causes upregulation of iNOS expression, and pulmonary vascular dysfunction. iNOS inhibition reverses the post-transplantational pulmonary vascular dysfunction.

Dehydroepiandrosterone (DHEA) inhibits voltage-gated T-type calcium channels

Background and purposeDehydroepiandrosterone (DHEA) and its sulfated form, DHEAS, are the most abundant steroid hormones in the mammalian blood flow. DHEA may have beneficial effects in various pathophysiological conditions such as cardiovascular diseases or deterioration of the sense of well-being. However to date, the cellular mechanism underlying DHEA action remains elusive and may involve ion channel modulation. In this study, we have characterized the effect of DHEA on T-type voltage-activated calcium channels (T-channels), which are involved in several cardiovascular and neuronal diseases. Key resultsUsing the whole-cell patch-clamp technique, we demonstrate that DHEA inhibits the three recombinant T-channels (CaV3.1, CaV3.2 and CaV3.3) expressed in NG108-15 cell line, as well as native T-channels in pulmonary artery smooth muscle cells. This effect of DHEA is both concentration (IC50 between 2 and 7μM) and voltage-dependent and results in a significant shift of the steady-state inactivation curves toward hyperpolarized potentials. Consequently, DHEA reduces window T-current and inhibits membrane potential oscillations induced by CaV3 channels. DHEA inhibition is not dependent on the activation of nuclear androgen or estrogen receptors and implicates a PTX-sensitive Gi protein pathway. Functionally, DHEA and the T-type inhibitor NNC 55-0396 inhibited KCl-induced contraction of pulmonary artery rings and their effect was not cumulative. ConclusionsAltogether, the present data demonstrate that DHEA inhibits T-channels by a Gi protein dependent pathway. DHEA-induced alteration in T-channel activity could thus account for its therapeutic action and/or physiological effects.

Fosinopril Prevents the Pulmonary Arterial Remodeling in Sinoaortic-denervated Rats by Regulating Phosphodiesterase

To study the effects of fosinopril on sinoaortic denervation (SAD)-induced pulmonary vascular remodeling and on phosphodiesterases (PDE) 1 in rats. SAD was performed in male Sprague-Dawley rats at the age of 10 weeks. The experiment included sham-operated (Sham), SAD, and fosinopril-treated SAD groups. Fosinopril (15 mg/kg/d) was given in rat chow. After 16 weeks of treatment, the pulmonary arteries were taken for investigations, including pharmacological study, measurement of cGMP, light microscopy, immunohistochemistry, Western blotting, and quantitative real-time RT-PCR. Compared with Sham rats, blood pressure variability (BPV) was significantly increased in the SAD group. However, the mean pulmonary artery pressure (mPAP) was not significant change among 3 groups. After SAD, maximal contraction of pulmonary artery rings to phenylephrine was markedly decreased; the most prominent morphological change in the lung included thickening vascular walls, increasing number of smooth muscle cells, and greater wall-to-lumen ratio; the tissue concentrations of cGMP was reduced significantly; PDE1A or PDE1C expression was upregulated significantly, and endothelial nitric oxide synthase (eNOS) expression was downregulated significantly. Fosinopril treatment prevented these changes induced by SAD. Pulmonary artery remodeling (structural and functional abnormalities) was induced by SAD. Fosinopril, an angiotensin-converting enzyme inhibitor, mainly via potentiating eNOS pathway and inhibiting AngII formation, effectively prevented increased blood pressure variability and vascular remodeling of the pulmonary artery after SAD by regulating the activity levels or expression of eNOS, cGMP, and PDE1s.

Homocysteine-induced changes in vascular reactivity of guinea-pig pulmonary arteries: Role of the oxidative stress and poly (ADP-ribose) polymerase activation

This study was aimed to examine the effects of homocysteine (Hcy) on vascular responsiveness of guinea-pig isolated pulmonary arteries and to investigate possible underlying mechanisms. In order to evaluate vascular reactivity, isometric tension studies were performed in response to potassium chloride (KCl), phenylephrine (Phe), acetylcholine (ACh), and sodium nitroprusside (SNP). Incubation of pulmonary artery rings with Hcy (10 −3 M, 180 min) resulted in significant inhibition of response to ACh (an endothelium-dependent vasodilator)( E max: 55.3±6.7 vs. 13.1±2.0 *, P<0.05) while SNP (an endothelium-independent vasodilator)-induced relaxation was not changed significantly. Furthermore, Hcy enhanced KCl- and Phe-induced contraction of pulmonary artery rings ( E max: 1568±81 vs. 2101±145 * mg for KCl and 1081±101 vs. 1544±117 * mg for Phe, P<0.05). Pulmonary artery ring contractions induced by stepwise addition to Ca 2+ to high KCl solution with no Ca 2+ were also significantly augmented by Hcy incubation ( E max: 1750±121 vs. 2295±134 * mg, P<0.05). To investigate mechanisms of Hcy action, additional sets of experiments involving rings incubation with Hcy alone or with addition of Tiron (an intracellular superoxide anion scavenger, 10 −2 M), PJ34 (an inhibitor of polyADP-ribose polymerase, 3×10 −6 M), and combination of two antioxidant enzymes superoxide dismutase (SOD, 100 U/ml) and catalase (CAT, 120 U/ml) for 180 min. The findings of our study clearly show that all these co-treatments significantly prevented the development of endothelial dysfunction induced by Hcy. Furthermore, the effect of Hcy on KCl- and Phe-induced contraction was significantly inhibited by the concomitant incubation with either SOD plus CAT, Tiron or PJ34. This study demonstrates that Hcy causes a significant alteration in vascular reactivity of pulmonary arteries, and this alteration seems to be via oxidative stress in pulmonary artery endothelium with subsequent DNA damage and activation of poly(ADP-ribose) polymerase (PARP) pathway.

Sildenafil prevents change in RhoA expression induced by chronic hypoxia in rat pulmonary artery.

Exposure to chronic hypoxia (CH) induces a sustained pulmonary hypertension associated with structural and functional changes in the pulmonary arterial bed, including alterations of contractile properties. The small G-protein RhoA and its effector Rho kinase play a major role in the sustained rise in tension induced by vasoconstrictors. The aim of this study was to analyze the effect of CH on the RhoA/Rho kinase signaling pathway in the rat pulmonary artery. Maximal contraction of pulmonary artery rings to endothelin-1, noradrenaline, and the thromboxane A2 analog U46619 was markedly decreased in rats exposed to CH (10% O2, 2 weeks). This CH-induced decrease response to agonists was attributable to the abolition of RhoA-mediated Ca2+ sensitization of the contraction. Real-time reverse transcriptase-polymerase chain reaction and Western blot analysis revealed a decrease in RhoA mRNA (79.4+/-6.0%, n=4) and RhoA (81.1+/-8.0%, n=4) expression in the main pulmonary artery from CH rats, whereas RhoA expression was not modified in arterial smooth muscle cells and arteries exposed to hypoxia and high intraluminal pressure, respectively. Treatment of rats with sildenafil (25 mg/kg per day) throughout 2 weeks of exposure to CH prevented CH-induced downregulation of RhoA, reduction of contraction, and pulmonary artery remodeling. These findings indicate that CH-induced downregulation of RhoA expression, leading to the abolition of RhoA/Rho kinase-mediated Ca2+ sensitization of contraction, is responsible for the decreased responses to contracting agonists in the pulmonary artery of CH rats. These alterations are prevented by sildenafil, indicating a major role of the NO/cyclic GMP pathway in CH-induced altered RhoA signaling in the pulmonary artery.

Delayed rectifier potassium channels contribute to the depressed pulmonary artery contractility in pneumonia.

We investigated the role of K(+) channels in the attenuated pulmonary artery (PA) contractility characteristic of acute Pseudomonas pneumonia. Contractility of PA rings from the lungs of control or pneumonia rats was assessed in vitro by obtaining cumulative concentration-response curves to the contractile agonists KCl, phenylephrine, or PGF(2 alpha) on PA rings before and after treatment with K(+) channel blockers. In rings from pneumonia rats, paxilline (10 microM), tetraethylammonium (2 mM) (blockers of large-conductance Ca(2+)-activated K(+) channels), and glybenclamide (ATP-sensitive K(+) channel blocker, 80 microM) had no significant effect on the attenuated contractile responses to KCl, phenylephrine, and PGF(2 alpha). However, 4-aminopyridine (2 mM), a blocker of voltage-gated K(+) channels (delayed rectifier K(+) channel) reversed this depressed contractility. Therefore, large-conductance Ca(2+)-activated K(+) and ATP-sensitive K(+) channels do not contribute to the attenuated PA contractility observed in this model of acute pneumonia. In contrast, 4-aminopyridine enhances contraction in PA rings from pneumonia lungs, consistent with involvement of a voltage-gated K(+) channel in the depressed PA contractility in acute pneumonia. Unraveling the precise mechanism of attenuated contractility in pneumonia could lead to innovative therapies for the pulmonary vascular abnormalities associated with this disease.

Role of GS Proteins in Hypoxic Constriction of Sheep Pulmonary Artery Rings

The introduction of hypoxia is well known to cause contraction of pulmonary artery rings in vitro. Despite intensive studies, the cellular mechanisms of hypoxic pulmonary vasoconstriction are still not well defined. In this study, we aimed to determine the contribution of G_S proteins in hypoxia-induced vasoconstriction in large-diameter sheep pulmonary arteries using cholera toxin (CT). Hypoxia caused further contractions in serotonin but not in NaF-precontracted pulmonary artery rings. However, hypoxic vasoconstriction due to lowering of pO₂ from 97 to 5 mm Hg was totally abolished by preincubation with CT in serotonin-precontracted arteries. These preliminary results indicate that signal transduction mediated by G_s proteins may be an important mechanism in the hypoxic vasoconstriction of isolated pulmonary arteries of sheep.

Leukotoxin, 9,10-epoxy-12-octadecenoate causes pulmonary vasodilation in rats

Leukotoxin (Lx), a cytochrome P-450-dependent metabolite of linoleate synthesized by neutrophils or synthesized by OH- and linoleate in neutrophil cell membranes, has been recovered in lung lavages of patients with the adult respiratory distress syndrome. We studied the direct vasoactive effects of Lx and linoleate, its parent compound, in the rat pulmonary circulation. In isolated rat lungs perfused at constant flow with a physiological salt solution, Lx (but not linoleate) caused a biphasic response, an initial transient vasoconstriction followed by a more prolonged vasodilation. The latter response was only evident when the pulmonary vascular tone was increased with either alveolar hypoxia (0% O2) or KCl (20 mM). The pressor response to angiotensin II was also attenuated in the presence of Lx. The vasodilatory response in perfused lungs was attenuated by methylene blue (2 x 10(-5) M), a putative inhibitor of the soluble guanylate cyclase but not by pretreatment with meclofenamate (10(-5) M), a cyclooxygenase inhibitor. In isolated pulmonary arterial (PA) rings preconstricted either with phenylephrine (5 x 10(-9) M), endothelin-1 (10(-8) M), or KCl (30 mM), Lx (but not linoleate) caused dose-dependent relaxation. The relaxing effect of Lx on endothelium-intact rings was attenuated by NG-monomethyl-L-arginine or methylene blue. The magnitude of the hypoxic contraction of PA rings was attenuated in the presence of Lx. Whereas the mechanism of Lx-induced vasoconstriction is not clear, we conclude that Lx causes vasodilation in rat lungs and that the vasodilatory component is to a large degree endothelium-derived relaxing factor-dependent.

Pharmacological and histological examinations of regional differences of guinea-pig lung: a role of pleural surface smooth muscle in lung strip contraction.

1. Parenchymal lung strip preparations have been widely used as an in vitro model of peripheral airway smooth muscle. The present study examined functional responses of 4 consecutive guinea-pig lung parenchymal strips isolated from the central region (segment 1) to the distal edge (segment 4) of the lower lung lobe. The middle two segments were designated as segments 2 and 3. 2. Lung segments 1 and 4 exhibited significantly greater contraction than the other 2 segments to KCl when responses were expressed as mg force per mg tissue weight. Contractile responses to bronchospastic agents including histamine, carbachol, endothelin-1, leukotrienes (LT) B4 and D4, and the thromboxane A2-mimetic U46619 demonstrated no significant difference in EC50 values among the 4 lung segments. 3. Contractile responses of segments 1 and 4 to antigen-challenge (ovalbumin), ionophore A23187 and substance P were significantly greater than the other 2 segments with respect to either sensitivity or maximum responsiveness. 4. U46619-induced contractions of the 4 lung segments were relaxed in similar manner by papaverine and theophylline up to 100%, salbutamol up to 80%, and sodium nitroprusside by only 20%. In contrast, sodium nitroprusside markedly reversed U46619-induced contraction of pulmonary arterial rings and bronchial rings. 5. Histological studies identified 2-4 layers of smooth muscle cells underlying the lung pleural surface. Mast cells were prominent in this area. Moreover, morphometric studies showed that segment 4 possessed the least amount of smooth muscle structures from bronchial/bronchiolar wall and vasculatures as compared to the other 3 segments, and a significant difference in this respect was evident between segment 1 and segment 4.6. Since lung segments 1 and 4 are covered with larger surface area of lung pleura, the present results suggest that the significantly greater intrinsic contractile responses of segments 1 and 4 are associated with the presence of increased lung pleural surface possibly together with more mast cells. Thus, a primary contribution to the net contraction of the lung parenchymal strips may be smooth muscle from the lung pleura, alveolar ducts and interstitial contractile cells rather than from bronchi/bronchioles and microvasculatures.

Effects of hypoxia on isolated intrapulmonary arteries from the sheep

The effects of hypoxia on contraction of sheep pulmonary artery rings (large= 2.2–4.1 mm diameter, small = 0.32–0.64 mm diameter) has been investigated following precontraction or with the artery rings set at their optimal resting force. Hypoxia (PO 2 4 mmHg) caused a marked contraction of pulmonary artery rings precontracted with 5-HT at its EC 85 (small arteries 40 ± 8 g cm −2) but not when precontracted with KCI. At optimal resting force hypoxia caused a small contraction (small arteries 11 ± 2 g cm −2). Large artery rings gave a smaller contraction in response to hypoxia at optimal resting force than did small artery rings (2 ± 0.2 g cm −2 at PO 2 = 4 mmHg). Large, unlike the small, artery rings did not contract in response to hypoxia when precontracted with 5-HT at its EC 35. Lowering the PO 2 to 40 mmHg caused contraction in arteries precontracted with 5-HT at its EC 85 but not in arteries at their optimal resting force. Removal of the endothelium abolished all hypoxia-induced contractile responses in sheep pulmonary artery rings. Hypoxia reversibly abolished acetylcholine-induced relaxation and augmented the 5-HT contraction (206 ± 28 to 255 ± 34 g cm −2) in small rings. It is concluded that hypoxia may produce contraction in sheep pulmonary artery rings at least, in part, by reducing the output of vasodilator mediators from the endothelium.