scholarly journals Soluble guanylate cyclase modulators blunt hyperoxia effects on calcium responses of developing human airway smooth muscle

2015 ◽  
Vol 309 (6) ◽  
pp. L537-L542 ◽  
Author(s):  
Rodney D. Britt ◽  
Michael A. Thompson ◽  
Ine Kuipers ◽  
Alecia Stewart ◽  
Elizabeth R. Vogel ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Airway Smooth Muscle ◽  
Guanylate Cyclase ◽  
Therapeutic Strategy ◽  
Soluble Guanylate Cyclase ◽  
Novel Drugs ◽  
Intracellular Ca ◽  
Recurrent Wheeze ◽  
Airway Tone ◽  
Sgc Stimulators

Exposure to moderate hyperoxia in prematurity contributes to subsequent airway dysfunction and increases the risk of developing recurrent wheeze and asthma. The nitric oxide (NO)-soluble guanylate cyclase (sGC)-cyclic GMP (cGMP) axis modulates airway tone by regulating airway smooth muscle (ASM) intracellular Ca2+ ([Ca2+]i) and contractility. However, the effects of hyperoxia on this axis in the context of Ca2+/contractility are not known. In developing human ASM, we explored the effects of novel drugs that activate sGC independent of NO on alleviating hyperoxia (50% oxygen)-induced enhancement of Ca2+ responses to bronchoconstrictor agonists. Treatment with BAY 41–2272 (sGC stimulator) and BAY 60-2770 (sGC activator) increased cGMP levels during exposure to 50% O2. Although 50% O2 did not alter sGCα1 or sGCβ1 expression, BAY 60-2770 did increase sGCβ1 expression. BAY 41-2272 and BAY 60-2770 blunted Ca2+ responses to histamine in cells exposed to 50% O2. The effects of BAY 41-2272 and BAY 60-2770 were reversed by protein kinase G inhibition. These novel data demonstrate that BAY 41-2272 and BAY 60-2770 stimulate production of cGMP and blunt hyperoxia-induced increases in Ca2+ responses in developing ASM. Accordingly, sGC stimulators/activators may be a useful therapeutic strategy in improving bronchodilation in preterm infants.

Role of Ca2+ entry in the modulation of airway tone by hypoxia

1993 ◽  
Vol 264 (3) ◽  
pp. L284-L289 ◽  
Author(s):  
L. B. Fernandes ◽  
K. Stuart-Smith ◽  
T. L. Croxton ◽  
C. A. Hirshman
Keyword(s):  
Smooth Muscle ◽  
Airway Smooth Muscle ◽  
Soluble Guanylate Cyclase ◽  
Muscle Tone ◽  
Maintenance Phase ◽  
Dependent Manner ◽  
Cellular Mechanisms ◽  
Concentration Dependent Manner ◽  
Initiation Phase ◽  
Airway Tone

To evaluate the cellular mechanisms involved in hypoxic relaxation of airway smooth muscle, we investigated the effects of hypoxia on the behavior of third- and fourth-order porcine bronchial rings contracted with either carbachol or KCl. In one series of experiments, hypoxia (95% N2-5% CO2) was imposed and rings were then exposed to increasing concentrations of carbachol or KCl. In separate experiments, rings were first contracted with carbachol (10(-6) M) or KCl (40 mM) and were then exposed to solutions bubbled with decreasing concentrations of O2. The CO2 concentration was maintained constant at 5% in all experiments. The initial magnitude of KCl-induced but not carbachol-induced contractions was profoundly reduced by 95% N2-5% CO2. The sensitivity of the airway to carbachol was unchanged. In rings precontracted with either carbachol or KCl, hypoxia caused similar losses of airway smooth muscle tone in a reversible and concentration-dependent manner. The effects of hypoxia were independent of the presence of an intact epithelium and were not inhibited by the cyclooxygenase inhibitor indomethacin (5 microM), the soluble guanylate cyclase inhibitor methylene blue (50 microM), or the beta-adrenoceptor antagonist propranolol (1 microM). The impairment by hypoxia of the initiation phase of KCl-induced contractions and of the maintenance phase of both KCl- and carbachol-induced contractions, but not the initiation phase of carbachol-induced contractions, suggests that changes in O2 tension modulate airway tone by altering the entry of extracellular calcium into the airway smooth muscle.

scholarly journals Rapid effects of estrogen on intracellular Ca2+ regulation in human airway smooth muscle

2010 ◽  
Vol 298 (4) ◽  
pp. L521-L530 ◽  
Author(s):  
Elizabeth A. Townsend ◽  
Michael A. Thompson ◽  
Christina M. Pabelick ◽  
Y. S. Prakash
Keyword(s):  
Smooth Muscle ◽  
Airway Smooth Muscle ◽  
Sex Steroids ◽  
Airway Hyperresponsiveness ◽  
Clinical Findings ◽  
Human Airway Smooth Muscle ◽  
Selective Agonist ◽  
Intracellular Ca ◽  
17Β Estradiol ◽  
Airway Tone

The severity of asthma, a disease characterized by airway hyperresponsiveness and inflammation, is enhanced in some women during the menstrual cycle and during pregnancy but relieved in others. These clinical findings suggest that sex steroids modulate airway tone. Based on well-known relaxant effects of estrogens on vascular smooth muscle, we hypothesized that estrogens relax airway smooth muscle (ASM), thus facilitating bronchodilation. In ASM tissues from female patients, Western and immunocytochemical analyses confirmed the presence of both estrogen receptor (ER) isoforms, ERα and ERβ. In fura 2-loaded, dissociated ASM cells maintained in culture, acute exposure to physiological concentrations of 17β-estradiol (E2; 100 pM to 10 nM) decreased the intracellular Ca2+ ([Ca2+]i) response to 1 μM histamine, an effect reversed by the ER antagonist ICI-182,780. The ERα-selective agonist (R,R)-THC had a greater reducing effect on [Ca2+]i responses to histamine and 1 μM ACh compared with the ERβ-selective agonist (DPN). The effects of E2 on [Ca2+]i were mediated, at least in part, via decreased Ca2+ influx through l-type channels and store-operated Ca2+ entry but not via Ca2+-activated K+ channels, receptor-operated entry, or sarcoplasmic reticulum reuptake. Overall, these data support our hypothesis that estrogens relax ASM and suggest a potentially novel therapeutic target in airway hyperresponsiveness.

scholarly journals Soluble guanylate cyclase as an alternative target for bronchodilator therapy in asthma

2016 ◽  
Vol 113 (17) ◽  
pp. E2355-E2362 ◽  
Author(s):  
Arnab Ghosh ◽  
Cynthia J. Koziol-White ◽  
Kewal Asosingh ◽  
Georgina Cheng ◽  
Lisa Ruple ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Airway Smooth Muscle ◽  
Guanylate Cyclase ◽  
Human Lung ◽  
Soluble Guanylate Cyclase ◽  
Alternative Pathway ◽  
Allergic Inflammation ◽  
Normal Lung ◽  
Airway Smooth Muscle Cells ◽  
Muscle Enzyme

Asthma is defined by airway inflammation and hyperresponsiveness, and contributes to morbidity and mortality worldwide. Although bronchodilation is a cornerstone of treatment, current bronchodilators become ineffective with worsening asthma severity. We investigated an alternative pathway that involves activating the airway smooth muscle enzyme, soluble guanylate cyclase (sGC). Activating sGC by its natural stimulant nitric oxide (NO), or by pharmacologic sGC agonists BAY 41–2272 and BAY 60–2770, triggered bronchodilation in normal human lung slices and in mouse airways. Both BAY 41–2272 and BAY 60–2770 reversed airway hyperresponsiveness in mice with allergic asthma and restored normal lung function. The sGC from mouse asthmatic lungs displayed three hallmarks of oxidative damage that render it NO-insensitive, and identical changes to sGC occurred in human lung slices or in human airway smooth muscle cells when given chronic NO exposure to mimic the high NO in asthmatic lung. Our findings show how allergic inflammation in asthma may impede NO-based bronchodilation, and reveal that pharmacologic sGC agonists can achieve bronchodilation despite this loss.

Abstract 107: Cytochrome B5 Reductase 3 Sensitizes Soluble Guanylate Cyclase to Nitric Oxide in Vascular Smooth Muscle

Hypertension ◽  
2015 ◽  
Vol 66 (suppl_1) ◽  
Author(s):  
Adam C Straub ◽  
Anh T Nguyen ◽  
Mizanur Rahaman ◽  
Stephanie M Mutchler ◽  
Megan Miller ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Blood Pressure ◽  
Smooth Muscle ◽  
Vascular Smooth Muscle ◽  
Guanylate Cyclase ◽  
Soluble Guanylate Cyclase ◽  
Cytochrome B5 ◽  
Heme Iron ◽  
Cytochrome B5 Reductase ◽  
Arterial Blood

The inability nitric oxide (NO) to stimulate soluble guanylate cyclase (sGC) has been linked to numerous cardiovascular diseases (CVD) including hypertension. While several studies have defined the importance of sGC expression in the cardiovascular system, the basic mechanisms that regulate sGC activity remain incompletely understood. Here, we report for the first time that sGC heme iron redox state, which is essential for NO-induced sGC activation, is regulated by cytochrome B5 reductase 3 (CyB5R3). Genetic knockdown and pharmacological inhibition of CyB5R3 in primary rat vascular smooth muscle cells resulted in a 60% loss in cGMP production. Conversely, the sGC activator Bay 58-2667, which activates oxidized or heme free sGC, reversed these effects. Consistent with our cell culture work, purified protein studies demonstrate that CyB5R3 can directly reduce oxidized sGC heme iron and sensitize sGC to NO. To test the functional importance of Cyb5R3 activity, we cultured mouse thoracodorsal arteries with a pharmacological inhibitor of Cyb5R3 (ZINC 747) and performed vascular reactivity studies using pressure myography. Arteries treated with ZINC 747 showed decreased responsiveness the NO donor DETA-NONOate but increase sensitivity to Bay 58-2667. We then treated mice with 10mg/kg/day of ZINC 747 using osmotic mini pumps, which caused an increase in mean arterial blood pressure (107.5±3.4 vs 131±13.16) measured via radio telemetry. Lastly, translational studies reveal that the CyB5R3 T116S polymorphism with allele frequency 0.23 only in African Americans is unable to reduce sGC and correlates with increased blood pressure. Considering the defining role of sGC in NO signaling and the fact that the oxidation state of sGC may predict responses to NO therapies and new classes of sGC activator medications, we anticipate that these studies may significantly impact our understanding of biology, precision therapeutics (right drug for the right patient) and pharmacogenetics (T117S SNP based drug selection).

scholarly journals cGMP modulation therapeutics for sickle cell disease

2019 ◽  
Vol 244 (2) ◽  
pp. 132-146 ◽  
Author(s):  
Nicola Conran ◽  
Lidiane Torres
Keyword(s):  
Sickle Cell Disease ◽  
Sickle Cell ◽  
Guanylate Cyclase ◽  
Soluble Guanylate Cyclase ◽  
Cell Transplant ◽  
Cell Disease ◽  
Chronic Complications ◽  
Intracellular Cgmp ◽  
No Bioavailability ◽  
Sgc Stimulators

Sickle cell disease (SCD) is an inherited disease caused by the production of abnormal hemoglobin (Hb) S, whose deoxygenation-induced polymerization results in red blood cell (RBC) sickling and numerous pathophysiological consequences. SCD affects approximately 300,000 newborns worldwide each year and is associated with acute and chronic complications, including frequent painful vaso-occlusive episodes that often require hospitalization. Chronic intravascular hemolysis in SCD significantly reduces vascular nitric oxide (NO) bioavailability, consequently decreasing intracellular signaling via cyclic guanosine monophosphate (cGMP), in turn diminishing vasodilation and contributing to the inflammatory mechanisms that trigger vaso-occlusive processes. Oxidative stress may further reduce NO bioavailability in SCD and can oxidize the intracellular enzyme target of NO, soluble guanylate cyclase (sGC), rendering it inactive. Increasing intracellular cGMP-dependent signaling constitutes an important pharmacological therapeutic approach for SCD with a view to augmenting vasodilation, and reducing inflammatory mechanisms, as well as for increasing the production of anti-polymerizing fetal Hb in erythroid cells. Pharmacological agents under pre-clinical and clinical investigation for SCD include NO-based therapeutics to augment NO bioavailability, as well as heme-dependent sGC stimulators and heme-independent sGC activators that directly stimulate native and oxidized sGC, respectively, therefore bypassing the need for vascular NO delivery. Additionally, the phosphodiesterases (PDEs) that degrade intracellular cyclic nucleotides with specific cellular distributions are attractive drug targets for SCD; PDE9 is highly expressed in hematopoietic cells, making the use of PDE9 inhibitors, originally developed for use in neurological diseases, a potential approach that could rapidly amplify intracellular cGMP concentrations in a relatively tissue-specific manner. Impact statement Sickle cell disease (SCD) is one of the most common inherited diseases and is associated with a reduced life expectancy and acute and chronic complications, including frequent painful vaso-occlusive episodes that often require hospitalization. At present, treatment of SCD is limited to hematopoietic stem cell transplant, transfusion, and limited options for pharmacotherapy, based principally on hydroxyurea therapy. This review highlights the importance of intracellular cGMP-dependent signaling pathways in SCD pathophysiology; modulation of these pathways with soluble guanylate cyclase (sGC) stimulators or phosphodiesterase (PDE) inhibitors could potentially provide vasorelaxation and anti-inflammatory effects, as well as elevate levels of anti-sickling fetal hemoglobin.

scholarly journals Airway smooth muscle CXCR3 ligand production: regulation by JAK-STAT1 and intracellular Ca2+

2013 ◽  
Vol 304 (11) ◽  
pp. L790-L802 ◽  
Author(s):  
X. Tan ◽  
Y. A. Alrashdan ◽  
H. Alkhouri ◽  
B. G. G. Oliver ◽  
C. L. Armour ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Airway Smooth Muscle ◽  
Immunoblot Analysis ◽  
Interferon Γ ◽  
Protein Activation ◽  
Early Activation ◽  
Plasmic Reticulum ◽  
Cxcr3 Ligand ◽  
Intracellular Ca ◽  
Jnk Activation

In asthma, airway smooth muscle (ASM) chemokine (C-X-C motif) receptor 3 (CXCR3) ligand production may attract mast cells or T lymphocytes to the ASM, where they can modulate ASM functions. In ASM cells (ASMCs) from people with or without asthma, we aimed to investigate JAK-STAT1, JNK, and Ca2+ involvement in chemokine (C-X-C motif) ligand (CXCL)10 and CXCL11 production stimulated by interferon-γ, IL-1β, and TNF-α combined (cytomix). Confluent, growth-arrested ASMC were treated with inhibitors for pan-JAK (pyridone-6), JAK2 (AG-490), JNK (SP-600125), or the sarco(endo)plasmic reticulum Ca2+ATPase (SERCA) pump (thapsigargin), Ca2+ chelator (BAPTA-AM), or vehicle before and during cytomix stimulation for up to 24 h. Signaling protein activation as well as CXCL10/CXCL11 mRNA and protein production were examined using immunoblot analysis, real-time PCR, and ELISA, respectively. Cytomix-induced STAT1 activation was lower and CXCR3 ligand mRNA production was more sensitive to pyridone-6 and AG-490 in asthmatic than nonasthmatic ASMCs, but CXCL10/CXCL11 release was inhibited by the same proportion. Neither agent caused additional inhibition of release when used in combination with the JNK inhibitor SP-600125. Conversely, p65 NF-κB activation was higher in asthmatic than nonasthmatic ASMCs. BAPTA-AM abolished early CXCL10/CXCL11 mRNA production, whereas thapsigargin reduced it in asthmatic cells and inhibited CXCL10/CXCL11 release by both ASMC types. Despite these inhibitory effects, neither Ca2+ agent affected early activation of STAT1, JNK, or p65 NF-κB. In conclusion, intracellular Ca2+ regulated CXCL10/CXCL11 production but not early activation of the signaling molecules involved. In asthma, reduced ASM STAT1-JNK activation, increased NF-κB activation, and altered Ca2+ handling may contribute to rapid CXCR3 ligand production and enhanced inflammatory cell recruitment.

scholarly journals Effect of proinflammatory cytokines on regulation of sarcoplasmic reticulum Ca2+ reuptake in human airway smooth muscle

2009 ◽  
Vol 297 (1) ◽  
pp. L26-L34 ◽  
Author(s):  
Venkatachalem Sathish ◽  
Michael A. Thompson ◽  
Jeffrey P. Bailey ◽  
Christina M. Pabelick ◽  
Y. S. Prakash ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Sarcoplasmic Reticulum ◽  
Airway Inflammation ◽  
Airway Smooth Muscle ◽  
Proinflammatory Cytokines ◽  
Cell Types ◽  
Human Airway Smooth Muscle ◽  
Inhibitory Protein ◽  
Intracellular Ca ◽  
Additional Exposure

Airway inflammation leads to increased intracellular Ca2+ ([Ca2+]i) levels in airway smooth muscle (ASM) cells. Sarcoplasmic reticulum Ca2+ release and reuptake are key components of ASM [Ca2+]i regulation. Ca2+ reuptake occurs via sarcoendoplasmic reticulum Ca2+ ATPase (SERCA) and is regulated by the inhibitory protein phospholamban (PLB) in many cell types. In human ASM, we tested the hypothesis that inflammation increases PLB, thus inhibiting SERCA function, and leading to maintained [Ca2+]i levels. Surprisingly, we found that human ASM does not express PLB protein (although mRNA is detectable). Overnight exposure to the proinflammatory cytokines TNFα and IL-13 did not induce PLB expression, raising the issue of how SERCA is regulated. We then found that direct SERCA phosphorylation (via CaMKII) occurs in human ASM. In fura-2-loaded human ASM cells, we found that the CaMKII antagonist KN-93 significantly slowed the rate of fall of [Ca2+]i transients induced by ACh or bradykinin (in zero extracellular Ca2+), suggesting a role for CaMKII-mediated SERCA regulation. SERCA expression was decreased by cytokine exposure, and the rate of fall of [Ca2+]i transients was slowed in cells exposed to TNFα and IL-13. Cytokine effects on Ca2+ reuptake were unaffected by additional exposure to KN-93. These data indicate that in human ASM, SERCA is regulated by mechanisms such as CaMKII and that airway inflammation maintains [Ca2+]i levels by decreasing SERCA expression and slowing Ca2+ reuptake.

Selected Contribution: Tryptase-induced PAR-2-mediated Ca2+signaling in human airway smooth muscle cells

2001 ◽  
Vol 91 (2) ◽  
pp. 995-1003 ◽  
Author(s):  
Patrick Berger ◽  
J. Manuel Tunon-De-Lara ◽  
Jean-Pierre Savineau ◽  
Roger Marthan
Keyword(s):  
Smooth Muscle ◽  
Airway Inflammation ◽  
Airway Smooth Muscle ◽  
Airway Smooth Muscle Cells ◽  
Atpase Inhibitor ◽  
Human Airway Smooth Muscle ◽  
Human Airway ◽  
Transient Rise ◽  
Intracellular Ca ◽  
Protein Kinase C Inhibitor

Tryptase, the major mast cell product, is considered to play an important role in airway inflammation and hyperresponsiveness. Tryptase produces different, sometimes opposite, effects on airway responsiveness (bronchoprotection and/or airway contraction). This study was designed to examine the effect of human lung tryptase and activation of protease-activated receptor (PAR)-2 by synthetic activated peptide (AP) SLIGKV-NH2 on Ca2+ signaling in human airway smooth muscle (HASM) cells. Immunocytochemistry revealed that PAR-2 was expressed by HASM cells. Tryptase (7.5–30 mU/ml) induced a concentration-dependent transient relative rise in cytoplasmic Ca2+ concentration ([Ca2+]i) that reached 207 ± 32 nM ( n = 10) measured by indo 1 spectrofluorometry. The protease inhibitors leupeptin or benzamidine (100 μM) abolished tryptase-induced [Ca2+]iincrease. Activation of PAR-2 by AP (1–100 μM) also induced a concentration-dependent transient rise in [Ca2+]i, whereas the reverse peptide produced no effect. There was a homologous desensitization of the [Ca2+]i response on repeated stimulation with tryptase or AP. U-73122, a specific phospholipase C (PLC) antagonist, xestospongin, an inositol trisphosphate (IP3)-receptor antagonist, or thapsigargin, a sarcoplamic Ca2+-ATPase inhibitor, abolished tryptase-induced [Ca2+]iresponse, whereas Ca2+ removal, in the additional presence of EGTA, had no effect. Calphostin C, a protein kinase C inhibitor, increased PAR-2 [Ca2+]i response. Our results indicate that tryptase activates a [Ca2+]iresponse, which appears as PAR-2 mediated in HASM cells. Signal transduction implicates the intracellular Ca2+ store via PLC activation and thus via the IP3 pathway. This study provides evidence that tryptase, which is increasingly recognized as an important mediator in airway inflammation and hyperresponsiveness, is also a potent direct agonist at the site of airway smooth muscle.

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