Glucagon induces airway smooth muscle relaxation by nitric oxide and prostaglandin E2

Submitted by sandra infurna ([email protected]) on 2016-07-07T16:52:39Z No. of bitstreams: 1 julio_dalleprane_etal_IOC_2015.pdf: 587356 bytes, checksum: 37345750ef0f8fa52a04dc27261e4b50 (MD5)Approved for entry into archive by sandra infurna ([email protected]) on 2016-07-07T17:06:25Z (GMT) No. of bitstreams: 1 julio_dalleprane_etal_IOC_2015.pdf: 587356 bytes, checksum: 37345750ef0f8fa52a04dc27261e4b50 (MD5)Made available in DSpace on 2016-07-07T17:06:25Z (GMT). No. of bitstreams: 1 julio_dalleprane_etal_IOC_2015.pdf: 587356 bytes, checksum: 37345750ef0f8fa52a04dc27261e4b50 (MD5) Previous issue date: 2015Made available in DSpace on 2016-07-08T18:49:09Z (GMT). No. of bitstreams: 3 julio_dalleprane_etal_IOC_2015.pdf.txt: 59513 bytes, checksum: 559c7c6ee5be85cfd601b6e5f9a4a61a (MD5) julio_dalleprane_etal_IOC_2015.pdf: 587356 bytes, checksum: 37345750ef0f8fa52a04dc27261e4b50 (MD5) license.txt: 2991 bytes, checksum: 5a560609d32a3863062d77ff32785d58 (MD5) Previous issue date: 2015Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Inflamação. Rio de Janeiro, RJ, Brasil.Universidade do Estado do Rio de Janeiro. Instituto de Nutrição. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Inflamação. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Imunofarmacologia. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Inflamação. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Inflamação. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Inflamação. Rio de Janeiro, RJ, Brasil.Glucagon is a hyperglycemic pancreatic hormone that has been shown to provide a beneficial effect against asthmatic bronchospasm. We investigated the role of this hormone on airway smooth muscle contraction and lung inflammation using both in vitro and in vivo approaches. The action of glucagon on mouse cholinergic tracheal contraction was studied in a conventional organ bath system, and its effect on airway obstruction was also investigated using the whole-body pletysmographic technique in mice. We also tested the effect of glucagon on lipopolysaccharide (LPS)-induced airway hyperreactivity (AHR) and inflammation. The expression of glucagon receptor (GcgR), CREB, phospho-CREB, nitric oxide synthase (NOS)-3, pNOS-3 and cyclooxygenase (COX)-1 was evaluated by western blot, while prostaglandin E2 (PGE2) and tumour necrosis factor-a were quantified by enzyme-linked immunoassay and ELISA respectively. Glucagon partially inhibited carbacholinduced tracheal contraction in a mechanism clearly sensitive to des-His1-[Glu9]-glucagon amide, a GcgR antagonist. Remarkably, GcgR was more expressed in the lung and trachea with intact epithelium than in the epithelium-denuded trachea. In addition, the glucagon-mediated impairment of carbachol-induced contraction was prevented by either removing epithelial cells or blocking NOS (L-NAME), COX (indomethacin) or COX-1 (SC-560). In contrast, inhibitors of either heme oxygenase or COX-2 were inactive. Intranasal instillation of glucagon inhibited methacholine-induced airway obstruction by a mechanism sensitive to pretreatment with L-NAME, indomethacin and SC-560. Glucagon induced CREB and NOS-3 phosphorylation and increased PGE2 levels in the lung tissue without altering COX-1 expression. Glucagon also inhibited LPS-induced AHR and bronchoalveolar inflammation. These findings suggest that glucagon possesses airway-relaxing properties that are mediated by epithelium-NOS-3-NO- and COX-1-PGE2- dependent mechanisms

Mangiferin Prevents Guinea Pig Tracheal Contraction via Activation of the Nitric Oxide-Cyclic GMP Pathway

Made available in DSpace on 2015-09-28T13:02:38Z (GMT). No. of bitstreams: 2 license.txt: 1914 bytes, checksum: 7d48279ffeed55da8dfe2f8e81f3b81f (MD5) aline_vieira_etal_IOC_2013.pdf: 2770092 bytes, checksum: 918f487813f46e76854091dffeabd7a0 (MD5) Previous issue date: 2013Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Inflamação. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Inflamação. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Inflamação. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Inflamação. Rio de Janeiro, RJ, Brasil.Universidade Federal de Alfenas. Departamento de Farmácia. Laboratório de Fitoquímica e Química Medicinal. Alfenas, MG, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Inflamação. Rio de Janeiro, RJ, Brasil.Previous studies have described the antispasmodic effect of mangiferin, a natural glucoside xanthone (2-C-b-Dglucopyranosyl-1,3,6,7-tetrahydroxyxanthone) that is present in mango trees and other plants, but its mechanism of action remains unknown. The aim of this study was to examine the potential contribution of the nitric oxide-cyclic GMP pathway to the antispasmodic effect of mangiferin on isolated tracheal rings preparations. The functional effect of mangiferin on allergic and non-allergic contraction of guinea pig tracheal rings was assessed in conventional organ baths. Cultured tracheal rings were exposed to mangiferin or vehicle, and nitric oxide synthase (NOS) 3 and cyclic GMP (cGMP) levels were quantified using western blotting and enzyme immunoassays, respectively. Mangiferin (0.1–10 mM) inhibited tracheal contractions induced by distinct stimuli, such as allergen, histamine, 5-hydroxytryptamine or carbachol, in a concentrationdependent manner. Mangiferin also caused marked relaxation of tracheal rings that were precontracted by carbachol, suggesting that it has both anti-contraction and relaxant properties that are prevented by removing the epithelium. The effect of mangiferin was inhibited by the nitric oxide synthase inhibitor, Nv-nitro-L-arginine methyl ester (L-NAME) (100 mM), and the soluble guanylate cyclase inhibitor, 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) (10 mM), but not the adenylate cyclase inhibitor, 9-(tetrahydro-2-furyl)adenine (SQ22536) (100 mM). The antispasmodic effect of mangiferin was also sensitive to K+ channel blockers, such as tetraethylammonium (TEA), glibenclamide and apamin. Furthermore, mangiferin inhibited Ca2+ -induced contractions in K+ (60 mM)-depolarised tracheal rings preparations. In addition, mangiferin increased NOS3 protein levels and cGMP intracellular levels in cultured tracheal rings. Finally, mangiferininduced increase in cGMP levels was abrogated by co-incubation with either ODQ or L-NAME. These data suggest that the antispasmodic effect of mangiferin is mediated by epithelium-nitric oxide- and cGMP-dependent mechanisms

The Neuropeptides Vasoactive Intestinal Peptide and Pituitary Adenylate Cyclase-Activating Polypeptide Control HIV-1 Infection in Macrophages Through Activation of Protein Kinases A and C

Submitted by Sandra Infurna ([email protected]) on 2018-10-04T13:28:05Z No. of bitstreams: 1 raissa_vieira_etal_IOC_2081.pdf: 2115333 bytes, checksum: 85546e2e12a7e1dc32b0da5a6546bb92 (MD5)Approved for entry into archive by Sandra Infurna ([email protected]) on 2018-10-04T13:40:58Z (GMT) No. of bitstreams: 1 raissa_vieira_etal_IOC_2081.pdf: 2115333 bytes, checksum: 85546e2e12a7e1dc32b0da5a6546bb92 (MD5)Made available in DSpace on 2018-10-04T13:41:00Z (GMT). No. of bitstreams: 1 raissa_vieira_etal_IOC_2081.pdf: 2115333 bytes, checksum: 85546e2e12a7e1dc32b0da5a6546bb92 (MD5) Previous issue date: 2018Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Pesquisa sobre o Timo. Rio de Janeiro, RJ. Brasil / Fundação Oswaldo Cruz. Instituto Oswaldo Cruz.. Instituto Nacional de Ciência e Tecnologia em Neuroimunomodulação. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de AIDS e Imunologia Molecular. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Inflamação. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Pesquisa sobre o Timo. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Pesquisa sobre o Timo. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz.. Instituto Nacional de Ciência e Tecnologia em Neuroimunomodulação. Rio de Janeiro, RJ, Brasil / Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Inflamação. Rio de Janeiro, RJ. Brasil..Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de AIDS e Imunologia Molecular. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Pesquisa sobre o Timo. Rio de Janeiro, RJ. Brasil / Fundação Oswaldo Cruz. Instituto Oswaldo Cruz.. Instituto Nacional de Ciência e Tecnologia em Neuroimunomodulação. Rio de Janeiro, RJ, Brasil.Vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase-activating polypeptide (PACAP) are highly similar neuropeptides present in several tissues, endowed with immunoregulatory functions and other systemic effects. We previously reported that both neuropeptides reduce viral production in HIV-1-infected primary macrophages, with the participation of β-chemokines and IL-10, and now we describe molecular mechanisms engaged in this activity. Macrophages exposed to VIP or PACAP before HIV-1 infection showed resistance to viral replication, comparable to that observed when the cells were treated after infection. Also, multiple treatments with a suboptimal dose of VIP or PACAP after macrophage infection resulted in a decline of virus production similar to the inhibition promoted by a single exposure to the optimal inhibitory concentration. Cellular signaling pathways involving cAMP production and activation of protein kinases A and C were critical components of the VIP and PACAP anti-HIV-1 effects. Analysis of the transcription factors and the transcriptional/cell cycle regulators showed that VIP and PACAP induced cAMP response element-binding protein activation, inhibited NF-kB, and reduced Cyclin D1 levels in HIV-1-infected cells. Remarkably, VIP and PACAP promoted G-to-A mutations in the HIV-1 provirus, matching those derived from the activity of the APOBEC family of viral restriction factors, and reduced viral infectivity. In conclusion, our findings strengthen the antiretroviral potential of VIP and PACAP and point to new therapeutic approaches to control the progression of HIV-1 infection

Potency (<i>p</i>EC₅₀) and maximal response (E_MAX) values obtained from concentration-response curves of allergen (ovalbumin, 0.001–100 µg/ml), histamine (0.1–3000 µM), 5-HT (0.01–30 µM) or carbachol challenge (0.01–100 µM) in guinea pig tracheal rings, following co-incubation with mangiferin (0.1–10 µM) or vehicle (0.1% DMSO).

<p>Data represent the mean ± S.E.M. of 4 to 8 tracheal segments.</p>*<p><i>p</i><0.05,</p>**<p><i>p</i><0.01 and.</p>***<p><i>p</i><0.001 compared with the tracheal responses of vehicle-treated tissues.</p

Mangiferin inhibits allergen- and spasmogen-induced tracheal contractions.

<p>The antispasmodic effects of mangiferin (0.1–10 µM) on guinea pig tracheal contraction induced by ovalbumin (0.001–100 µg/ml) (A), histamine (0.1–3000 µM) (B), 5-HT (0.01–30 µM) (C) or carbachol (0.01–100 µM) (D). Each point represents the mean ± S.E.M. of 4 to 8 segments. All results are expressed as a percentage of the contractile response induced by 2.5 µM carbachol. * <i>p</i><0.05 compared with the tracheal responses of vehicle-treated tissues (open circles).</p

Mangiferin increases cGMP levels in cultured tracheal rings.

<p>Effect of mangiferin on tracheal cGMP levels in the presence or absence of L-NAME or ODQ. Each bar represents the mean ± S.E.M. of 4 segments.*<i>p</i><0.05 compared with vehicle-treated tissues. +<i>p</i><0.05 compared with 10 µM mangiferin-treated tissues.</p

Proposed mechanism of action of mangiferin on guinea pig tracheal epithelium and smooth muscle cells.

<p>Mangiferin activates nitric oxide synthase 3 (NOS3) isoform that up-regulates the production of nitric oxide (NO) in the epithelial cell. Nitric oxide activates guanylate cyclase (GC), which increases the level of intracellular cyclic GMP (cGMP). Increased cGMP then activates the protein kinase G (PKG) cascade, enhancing K⁺ efflux and attenuating Ca²⁺ influx-associated smooth muscle cell contractility. GTP (guanosine triphosphate); L-NAME (N-nitro-L-arginine methyl ester); ODQ (1H-<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0071759#pone.0071759-Vyas1" target="_blank">[1]</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0071759#pone.0071759-Viswanadh1" target="_blank">[2]</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0071759#pone.0071759-Bowen1" target="_blank">[4]</a>oxadiazolo[4,3-a]quinoxalin-1-one); TEA (tetraethylammonium).</p

The Neuropeptides Vasoactive Intestinal Peptide and Pituitary Adenylate Cyclase-Activating Polypeptide Control HIV-1 Infection in Macrophages Through Activation of Protein Kinases A and C

Vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase-activating polypeptide (PACAP) are highly similar neuropeptides present in several tissues, endowed with immunoregulatory functions and other systemic effects. We previously reported that both neuropeptides reduce viral production in HIV-1-infected primary macrophages, with the participation of β-chemokines and IL-10, and now we describe molecular mechanisms engaged in this activity. Macrophages exposed to VIP or PACAP before HIV-1 infection showed resistance to viral replication, comparable to that observed when the cells were treated after infection. Also, multiple treatments with a suboptimal dose of VIP or PACAP after macrophage infection resulted in a decline of virus production similar to the inhibition promoted by a single exposure to the optimal inhibitory concentration. Cellular signaling pathways involving cAMP production and activation of protein kinases A and C were critical components of the VIP and PACAP anti-HIV-1 effects. Analysis of the transcription factors and the transcriptional/cell cycle regulators showed that VIP and PACAP induced cAMP response element-binding protein activation, inhibited NF-kB, and reduced Cyclin D1 levels in HIV-1-infected cells. Remarkably, VIP and PACAP promoted G-to-A mutations in the HIV-1 provirus, matching those derived from the activity of the APOBEC family of viral restriction factors, and reduced viral infectivity. In conclusion, our findings strengthen the antiretroviral potential of VIP and PACAP and point to new therapeutic approaches to control the progression of HIV-1 infection