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1.
Redox Biol ; 60: 102614, 2023 04.
Artigo em Inglês | MEDLINE | ID: mdl-36717349

RESUMO

Supraphysiological concentrations of oxygen (hyperoxia) can compromise host defense and increase susceptibility to bacterial and viral infections, causing ventilator-associated pneumonia (VAP). Compromised host defense and inflammatory lung injury are mediated, in part, by high extracellular concentrations of HMGB1, which can be decreased by GTS-21, a partial agonist of α7 nicotinic acetylcholine receptor (α7nAChR). Here, we report that a novel α7nAChR agonistic positive allosteric modulator (ago-PAM), GAT107, at 3.3 mg/kg, i.p., significantly decreased animal mortality and markers of inflammatory injury in mice exposed to hyperoxia and subsequently infected with Pseudomonas aeruginosa. The incubation of macrophages with 3.3 µM of GAT107 significantly decreased hyperoxia-induced extracellular HMGB1 accumulation and HMGB1-induced macrophage phagocytic dysfunction. Hyperoxia-compromised macrophage function was correlated with impaired mitochondrial membrane integrity, increased superoxide levels, and decreased manganese superoxide dismutase (MnSOD) activity. This compromised MnSOD activity is due to a significant increase in its level of glutathionylation. The incubation of hyperoxic macrophages with 3.3 µM of GAT107 significantly decreases the levels of glutathionylated MnSOD, and restores MnSOD activity and mitochondrial membrane integrity. Thus, GAT107 restored hyperoxia-compromised phagocytic functions by decreasing HMGB1 release, most likely via a mitochondrial-directed pathway. Overall, our results suggest that GAT107 may be a potential treatment to decrease acute inflammatory lung injury by increasing host defense in patients with VAP.


Assuntos
Lesão Pulmonar Aguda , Proteína HMGB1 , Hiperóxia , Pneumonia Associada à Ventilação Mecânica , Animais , Camundongos , Pneumonia Associada à Ventilação Mecânica/tratamento farmacológico , Pneumonia Associada à Ventilação Mecânica/metabolismo , Pneumonia Associada à Ventilação Mecânica/microbiologia , Receptor Nicotínico de Acetilcolina alfa7 , Proteína HMGB1/metabolismo , Hiperóxia/metabolismo , Macrófagos/metabolismo , Lesão Pulmonar Aguda/metabolismo , Superóxido Dismutase/metabolismo , Estresse Oxidativo
4.
Mol Med ; 27(1): 79, 2021 07 16.
Artigo em Inglês | MEDLINE | ID: mdl-34271850

RESUMO

BACKGROUND: High mobility group box 1 protein (HMGB1) is an alarmin following its release by immune cells upon cellular activation or stress. High levels of extracellular HMGB1 play a critical role in impairing the clearance of invading pulmonary pathogens and dying neutrophils in the injured lungs of cystic fibrosis (CF) and acute respiratory distress syndrome (ARDS). A heparin derivative, 2-O, 3-O desulfated heparin (ODSH), has been shown to inhibit HMGB1 release from a macrophage cell line and is efficacious in increasing bacterial clearance in a mouse model of pneumonia. Thus, we hypothesized that ODSH can attenuate the bacterial burden and inflammatory lung injury in CF and we conducted experiments to determine the underlying mechanisms. METHODS: We determined the effects of ODSH on lung injury produced by Pseudomonas aeruginosa (PA) infection in CF mice with the transmembrane conductance regulator gene knockout (CFTR-/-). Mice were given ODSH or normal saline intraperitoneally, followed by the determination of the bacterial load and lung injury in the airways and lung tissues. ODSH binding to HMGB1 was determined using surface plasmon resonance and in silico docking analysis of the interaction of the pentasaccharide form of ODSH with HMGB1. RESULTS: CF mice given 25 mg/kg i.p. of ODSH had significantly lower PA-induced lung injury compared to mice given vehicle alone. The CF mice infected with PA had decreased levels of nitric oxide (NO), increased levels of airway HMGB1 and HMGB1-impaired macrophage phagocytic function. ODSH partially attenuated the PA-induced alteration in the levels of NO and airway HMGB1 in CF mice. In addition, ODSH reversed HMGB1-impaired macrophage phagocytic function. These effects of ODSH subsequently decreased the bacterial burden in the CF lungs. In a surface plasmon resonance assay, ODSH interacted with HMGB1 with high affinity (KD = 3.89 × 10-8 M) and induced conformational changes that may decrease HMGB1's binding to its membrane receptors, thus attenuating HMGB1-induced macrophage dysfunction. CONCLUSIONS: The results suggest that ODSH can significantly decrease bacterial infection-induced lung injury in CF mice by decreasing both HMGB1-mediated impairment of macrophage function and the interaction of HMGB1 with membrane receptors. Thus, ODSH could represent a novel approach for treating CF and ARDS patients that have HMGB1-mediated lung injury.


Assuntos
Fibrose Cística/complicações , Fibrose Cística/metabolismo , Proteína HMGB1/genética , Heparina/análogos & derivados , Macrófagos/imunologia , Macrófagos/metabolismo , Pneumonia Bacteriana/etiologia , Pneumonia Bacteriana/metabolismo , Animais , Carga Bacteriana , Biomarcadores , Regulador de Condutância Transmembrana em Fibrose Cística/genética , Regulador de Condutância Transmembrana em Fibrose Cística/metabolismo , Modelos Animais de Doenças , Suscetibilidade a Doenças , Proteína HMGB1/química , Proteína HMGB1/metabolismo , Heparina/química , Heparina/metabolismo , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Imuno-Histoquímica , Masculino , Camundongos , Camundongos Knockout , Modelos Moleculares , Óxido Nítrico/metabolismo , Fagocitose/imunologia , Pneumonia Bacteriana/patologia , Ligação Proteica , Células RAW 264.7 , Relação Estrutura-Atividade
5.
J Immunotoxicol ; 18(1): 23-29, 2021 12.
Artigo em Inglês | MEDLINE | ID: mdl-33860730

RESUMO

The coronavirus SARS-CoV-2 of 2019 (COVID-19) causes a pandemic that has been diagnosed in more than 70 million people worldwide. Mild-to-moderate COVID-19 symptoms include coughing, fever, myalgia, shortness of breath, and acute inflammatory lung injury (ALI). In contrast, acute respiratory distress syndrome (ARDS) and respiratory failure occur in patients diagnosed with severe COVID-19. ARDS is mediated, at least in part, by a dysregulated inflammatory response due to excessive levels of circulating cytokines, a condition known as the "cytokine-storm syndrome." Currently, there are FDA-approved therapies that attenuate the dysregulated inflammation that occurs in COVID-19 patients, such as dexamethasone or other corticosteroids and IL-6 inhibitors, including sarilumab, tocilizumab, and siltuximab. However, the efficacy of these treatments have been shown to be inconsistent. Compounds that activate the vagus nerve-mediated cholinergic anti-inflammatory reflex, such as the α7 nicotinic acetylcholine receptor agonist, GTS-21, attenuate ARDS/inflammatory lung injury by decreasing the extracellular levels of high mobility group box-1 (HMGB1) in the airways and the circulation. It is possible that HMGB1 may be an important mediator of the "cytokine-storm syndrome." Notably, high plasma levels of HMGB1 have been reported in patients diagnosed with severe COVID-19, and there is a significant negative correlation between HMGB1 plasma levels and clinical outcomes. Nicotine can activate the cholinergic anti-inflammatory reflex, which attenuates the up-regulation and the excessive release of pro-inflammatory cytokines/chemokines. Therefore, we hypothesize that low molecular weight compounds that activate the cholinergic anti-inflammatory reflex, such as nicotine or GTS-21, may represent a potential therapeutic approach to attenuate the dysregulated inflammatory responses in patients with severe COVID-19.


Assuntos
Compostos de Benzilideno/farmacologia , Tratamento Farmacológico da COVID-19 , Colinérgicos/farmacologia , Inflamação/tratamento farmacológico , Nicotina/metabolismo , Piridinas/farmacologia , SARS-CoV-2/fisiologia , Tabagismo/tratamento farmacológico , Anticorpos Monoclonais Humanizados/uso terapêutico , Fumar Cigarros/efeitos adversos , Dexametasona/uso terapêutico , Proteína HMGB1/sangue , Humanos , Pandemias , Receptor Nicotínico de Acetilcolina alfa7/agonistas
6.
Antioxidants (Basel) ; 10(1)2021 Jan 19.
Artigo em Inglês | MEDLINE | ID: mdl-33477969

RESUMO

Supplemental oxygen therapy with supraphysiological concentrations of oxygen (hyperoxia; >21% O2) is a life-saving intervention for patients experiencing respiratory distress. However, prolonged exposure to hyperoxia can compromise bacterial clearance processes, due to oxidative stress-mediated impairment of macrophages, contributing to the increased susceptibility to pulmonary infections. This study reports that the activation of the α7 nicotinic acetylcholine receptor (α7nAChR) with the delete allosteric agonistic-positive allosteric modulator, GAT107, decreases the bacterial burden in mouse lungs by improving hyperoxia-induced lung redox imbalance. The incubation of RAW 264.7 cells with GAT107 (3.3 µM) rescues hyperoxia-compromised phagocytic functions in cultured macrophages, RAW 264.7 cells, and primary bone marrow-derived macrophages. Similarly, GAT107 (3.3 µM) also attenuated oxidative stress in hyperoxia-exposed macrophages, which prevents oxidation and hyper-polymerization of phagosome filamentous actin (F-actin) from oxidation. Furthermore, GAT107 (3.3 µM) increases the (1) activity of superoxide dismutase 1; (2) activation of Nrf2 and (3) the expression of heme oxygenase-1 (HO-1) in macrophages exposed to hyperoxia. Overall, these data suggest that the novel α7nAChR compound, GAT107, could be used to improve host defense functions in patients, such as those with COVID-19, who are exposed to prolonged periods of hyperoxia.

7.
Free Radic Biol Med ; 164: 34-43, 2021 02 20.
Artigo em Inglês | MEDLINE | ID: mdl-33418109

RESUMO

Fuchs endothelial corneal dystrophy (FECD) is an age-related disease whereby progressive loss of corneal endothelial cells (CEnCs) leads to loss of vision. There is currently a lack of therapeutic interventions as the etiology of the disease is complex, with both genetic and environmental factors. In this study, we have provided further insights into the pathogenesis of the disease, showing a causal relationship between senescence and endothelial-mesenchymal transition (EMT) using in vitro and in vivo models. Ultraviolet A (UVA) light induced EMT and senescence in CEnCs. Senescent cells were arrested in G2/M phase of the cell cycle and responsible for the resulting profibrotic phenotype. Inhibiting ATR signaling and subsequently preventing G2/M arrest attenuated EMT. In vivo, UVA irradiation induced cell cycle re-entry in post mitotic CEnCs, resulting in senescence and fibrosis at 1- and 2-weeks post-UVA. Selectively eliminating senescent cells using the senolytic cocktail of dasatinib and quercetin attenuated UVA-induced fibrosis, highlighting the potential for a new therapeutic intervention for FECD.


Assuntos
Distrofia Endotelial de Fuchs , Apoptose , Divisão Celular , Linhagem Celular Tumoral , Células Endoteliais , Endotélio Corneano/metabolismo , Fibrose , Distrofia Endotelial de Fuchs/genética , Pontos de Checagem da Fase G2 do Ciclo Celular , Humanos , Estresse Oxidativo
8.
Mol Med ; 26(1): 98, 2020 10 30.
Artigo em Inglês | MEDLINE | ID: mdl-33126860

RESUMO

BACKGROUND: Mechanical ventilation, in combination with supraphysiological concentrations of oxygen (i.e., hyperoxia), is routinely used to treat patients with respiratory distress, such as COVID-19. However, prolonged exposure to hyperoxia compromises the clearance of invading pathogens by impairing macrophage phagocytosis. Previously, we have shown that the exposure of mice to hyperoxia induces the release of the nuclear protein high mobility group box-1 (HMGB1) into the pulmonary airways. Furthermore, extracellular HMGB1 impairs macrophage phagocytosis and increases the mortality of mice infected with Pseudomonas aeruginosa (PA). The aim of this study was to determine whether GTS-21 (3-(2,4-dimethoxybenzylidene) anabaseine), an α7 nicotinic acetylcholine receptor (α7nAChR) agonist, could (1) inhibit hyperoxia-induced HMGB1 release into the airways; (2) enhance macrophage phagocytosis and (3) increase bacterial clearance from the lungs in a mouse model of ventilator-associated pneumonia. METHOD: GTS-21 (0.04, 0.4, and 4 mg/kg) or saline were administered by intraperitoneal injection to mice that were exposed to hyperoxia (≥ 99% O2) and subsequently challenged with PA. RESULTS: The systemic administration of 4 mg/kg i.p. of GTS-21 significantly increased bacterial clearance, decreased acute lung injury and decreased accumulation of airway HMGB1 compared to the saline control. To determine the mechanism of action of GTS-21, RAW 264.7 cells, a macrophage-like cell line, were incubated with different concentrations of GTS-21 in the presence of 95% O2. The phagocytic activity of macrophages was significantly increased by GTS-21 in a dose-dependent manner. In addition, GTS-21 significantly inhibited the cytoplasmic translocation and release of HMGB1 from RAW 264.7 cells and attenuated hyperoxia-induced NF-κB activation in macrophages and mouse lungs exposed to hyperoxia and infected with PA. CONCLUSIONS: Our results indicate that GTS-21 is efficacious in improving bacterial clearance and reducing acute lung injury via enhancing macrophage function by inhibiting the release of nuclear HMGB1. Therefore, the α7nAChR represents a possible pharmacological target to improve the clinical outcome of patients on ventilators by augmenting host defense against bacterial infections.


Assuntos
Compostos de Benzilideno/farmacologia , Hiperóxia/imunologia , Macrófagos Alveolares/efeitos dos fármacos , Infecções por Pseudomonas/tratamento farmacológico , Piridinas/farmacologia , Lesão Pulmonar Induzida por Ventilação Mecânica/tratamento farmacológico , Receptor Nicotínico de Acetilcolina alfa7/antagonistas & inibidores , Animais , Modelos Animais de Doenças , Proteína HMGB1/metabolismo , Hiperóxia/dietoterapia , Macrófagos Alveolares/imunologia , Macrófagos Alveolares/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fagocitose/efeitos dos fármacos , Pseudomonas aeruginosa , Células RAW 264.7
9.
Mol Med ; 26(1): 63, 2020 06 29.
Artigo em Inglês | MEDLINE | ID: mdl-32600307

RESUMO

BACKGROUND: Oxygen therapy, using supraphysiological concentrations of oxygen (hyperoxia), is routinely administered to patients who require respiratory support including mechanical ventilation (MV). However, prolonged exposure to hyperoxia results in acute lung injury (ALI) and accumulation of high mobility group box 1 (HMGB1) in the airways. We previously showed that airway HMGB1 mediates hyperoxia-induced lung injury in a mouse model of ALI. Cholinergic signaling through the α7 nicotinic acetylcholine receptor (α7nAChR) attenuates several inflammatory conditions. The aim of this study was to determine whether 3-(2,4 dimethoxy-benzylidene)-anabaseine dihydrochloride, GTS-21, an α7nAChR partial agonist, inhibits hyperoxia-induced HMGB1 accumulation in the airways and circulation, and consequently attenuates inflammatory lung injury. METHODS: Mice were exposed to hyperoxia (≥99% O2) for 3 days and treated concurrently with GTS-21 (0.04, 0.4 and 4 mg/kg, i.p.) or the control vehicle, saline. RESULTS: The systemic administration of GTS-21 (4 mg/kg) significantly decreased levels of HMGB1 in the airways and the serum. Moreover, GTS-21 (4 mg/kg) significantly reduced hyperoxia-induced acute inflammatory lung injury, as indicated by the decreased total protein content in the airways, reduced infiltration of inflammatory monocytes/macrophages and neutrophils into the lung tissue and airways, and improved lung injury histopathology. CONCLUSIONS: Our results indicate that GTS-21 can attenuate hyperoxia-induced ALI by inhibiting extracellular HMGB1-mediated inflammatory responses. This suggests that the α7nAChR represents a potential pharmacological target for the treatment regimen of oxidative inflammatory lung injury in patients receiving oxygen therapy.


Assuntos
Lesão Pulmonar Aguda/etiologia , Lesão Pulmonar Aguda/metabolismo , Compostos de Benzilideno/farmacologia , Proteína HMGB1/metabolismo , Hiperóxia/complicações , Agonistas Nicotínicos/farmacologia , Piridinas/farmacologia , Lesão Pulmonar Aguda/tratamento farmacológico , Lesão Pulmonar Aguda/patologia , Animais , Biomarcadores , Suscetibilidade a Doenças , Proteína HMGB1/sangue , Proteína HMGB1/genética , Imuno-Histoquímica , Masculino , Camundongos , Modelos Biológicos
10.
Int J Mol Sci ; 21(3)2020 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-32024151

RESUMO

Mechanical ventilation with hyperoxia is the major supportive measure to treat patients with acute lung injury and acute respiratory distress syndrome (ARDS). However, prolonged exposure to hyperoxia can induce oxidative inflammatory lung injury. Previously, we have shown that high levels of airway high-mobility group box 1 protein (HMGB1) mediate hyperoxia-induced acute lung injury (HALI). Using both ascorbic acid (AA, also known as vitamin C) and sulforaphane (SFN), an inducer of nuclear factor (erythroid-derived 2)-like 2 (Nrf2), we tested the hypothesis that dietary antioxidants can mitigate HALI by ameliorating HMGB1-compromised macrophage function in phagocytosis by attenuating hyperoxia-induced extracellular HMGB1 accumulation. Our results indicated that SFN, which has been shown to attenute HALI in mice exposed to hyperoxia, dose-dependently restored hyperoxia-compromised macrophage function in phagocytosis (75.9 ± 3.5% in 0.33 µM SFN versus 50.7 ± 1.8% in dimethyl sulfoxide (DMSO) control, p < 0.05) by reducing oxidative stress and HMGB1 release from cultured macrophages (47.7 ± 14.7% in 0.33 µM SFN versus 93.1 ± 14.6% in DMSO control, p < 0.05). Previously, we have shown that AA enhances hyperoxic macrophage functions by reducing hyperoxia-induced HMGB1 release. Using a mouse model of HALI, we determined the effects of AA on hyperoxia-induced inflammatory lung injury. The i.p. administration of 50 mg/kg of AA to mice exposed to 72 h of ≥98% O2 significantly decreased hyperoxia-induced oxidative and nitrosative stress in mouse lungs. There was a significant decrease in the levels of airway HMGB1 (43.3 ± 12.2% in 50 mg/kg AA versus 96.7 ± 9.39% in hyperoxic control, p < 0.05), leukocyte infiltration (60.39 ± 4.137% leukocytes numbers in 50 mg/kg AA versus 100 ± 5.82% in hyperoxic control, p < 0.05) and improved lung integrity in mice treated with AA. Our study is the first to report that the dietary antioxidants, ascorbic acid and sulforaphane, ameliorate HALI and attenuate hyperoxia-induced macrophage dysfunction through an HMGB1-mediated pathway. Thus, dietary antioxidants could be used as potential treatments for oxidative-stress-induced acute inflammatory lung injury in patients receiving mechanical ventilation.


Assuntos
Lesão Pulmonar Aguda/prevenção & controle , Antioxidantes/administração & dosagem , Suplementos Nutricionais , Proteína HMGB1/metabolismo , Hiperóxia/complicações , Macrófagos/metabolismo , Pneumonia/prevenção & controle , Lesão Pulmonar Aguda/etiologia , Lesão Pulmonar Aguda/metabolismo , Lesão Pulmonar Aguda/patologia , Animais , Proteína HMGB1/genética , Macrófagos/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fagocitose , Pneumonia/etiologia , Pneumonia/metabolismo , Pneumonia/patologia , Sistema Respiratório/efeitos dos fármacos , Sistema Respiratório/metabolismo
11.
Biochem Pharmacol ; 176: 113817, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-31972169

RESUMO

Mechanical ventilation (MV) with supraphysiological levels of oxygen (hyperoxia) is a life-saving therapy for the management of patients with respiratory distress. However, a significant number of patients on MV develop ventilator-associated pneumonia (VAP). Previously, we have reported that prolonged exposure to hyperoxia impairs the capacity of macrophages to phagocytize Pseudomonas aeruginosa (PA), which can contribute to the compromised innate immunity in VAP. In this study, we show that the high mortality rate in mice subjected to hyperoxia and PA infection was accompanied by a significant decrease in the airway levels of nitric oxide (NO). Decreased NO levels were found to be, in part, due to a significant reduction in NO release by macrophages upon exposure to PA lipopolysaccharide (LPS). Based on these findings, we postulated that NO supplementation should restore hyperoxia-compromised innate immunity and decrease mortality by increasing the clearance of PA under hyperoxic conditions. To test this hypothesis, cultured macrophages were exposed to hyperoxia (95% O2) in the presence or absence of the NO donor, (Z)-1-[N-(2-aminoethyl)-N-(2-ammonioethyl)amino]diazen-1-ium-1,2-diolate (DETA-NONOate/D-NO). Interestingly, D-NO (up to 37.5 µM) significantly attenuated hyperoxia-compromised macrophage migratory, phagocytic, and bactericidal function. To determine whether the administration of exogenous NO enhances the host defense in bacteria clearance, C57BL/6 mice were exposed to hyperoxia (99% O2) and intranasally inoculated with PA in the presence or absence of D-NO. D-NO (300 µM-800 µM) significantly increased the survival of mice inoculated with PA under hyperoxic conditions, and significantly decreased bacterial loads in the lung and attenuated lung injury. These results suggest the NO donor, D-NO, can improve the clinical outcomes in VAP by augmenting the innate immunity in bacterial clearance. Thus, provided these results can be extrapolated to humans, NO supplementation may represent a potential therapeutic strategy for preventing and treating patients with VAP.


Assuntos
Imunidade Inata/efeitos dos fármacos , Doadores de Óxido Nítrico/farmacologia , Compostos Nitrosos/farmacologia , Pneumonia Associada à Ventilação Mecânica/tratamento farmacológico , Infecções por Pseudomonas/prevenção & controle , Pseudomonas aeruginosa/efeitos dos fármacos , Animais , Modelos Animais de Doenças , Humanos , Hiperóxia/imunologia , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Óxido Nítrico/imunologia , Óxido Nítrico/metabolismo , Fagocitose/efeitos dos fármacos , Fagocitose/imunologia , Pneumonia Associada à Ventilação Mecânica/imunologia , Pneumonia Associada à Ventilação Mecânica/microbiologia , Infecções por Pseudomonas/imunologia , Infecções por Pseudomonas/microbiologia , Pseudomonas aeruginosa/imunologia , Pseudomonas aeruginosa/fisiologia , Células RAW 264.7
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