RESUMEN
Intestinal microbiota and their metabolites affect systemic inflammation and kidney disease outcomes. Here, we investigated the key metabolites associated with the acute kidney injury (AKI)-to chronic kidney disease (CKD) transition and the effect of antibiotic-induced microbiota depletion (AIMD) on this transition. In 61 patients with AKI, 59 plasma metabolites were assessed to determine the risk of AKI-to-CKD transition. An AKI-to-CKD transition murine model was established four weeks after unilateral ischemia-reperfusion injury (IRI) to determine the effects of AIMD on the gut microbiome, metabolites, and pathological responses related to CKD transition. Human proximal tubular epithelial cells were challenged with CKD transition-related metabolites, and inhibitory effects of NADPH oxidase 2 (NOX2) signals were tested. Based on clinical metabolomics, plasma trimethylamine N-oxide (TMAO) was associated with a significantly increased risk for AKI-to-CKD transition [adjusted odds ratio 4.389 (95% confidence interval 1.106-17.416)]. In vivo, AIMD inhibited a unilateral IRI-induced increase in TMAO, along with a decrease in apoptosis, inflammation, and fibrosis. The expression of NOX2 and oxidative stress decreased after AIMD. In vitro, TMAO induced fibrosis with NOX2 activation and oxidative stress. NOX2 inhibition successfully attenuated apoptosis, inflammation, and fibrosis with suppression of G2/M arrest. NOX2 inhibition (in vivo) showed improvement in pathological changes with a decrease in oxidative stress without changes in TMAO levels. Thus, TMAO is a key metabolite associated with the AKI-to-CKD transition, and NOX2 activation was identified as a key regulator of TMAO-related AKI-to-CKD transition both in vivo and in vitro.
Asunto(s)
Lesión Renal Aguda , Antibacterianos , Modelos Animales de Enfermedad , Microbioma Gastrointestinal , Metilaminas , NADPH Oxidasa 2 , Estrés Oxidativo , Insuficiencia Renal Crónica , Lesión Renal Aguda/inducido químicamente , Lesión Renal Aguda/microbiología , Lesión Renal Aguda/prevención & control , Lesión Renal Aguda/patología , Lesión Renal Aguda/tratamiento farmacológico , Metilaminas/sangre , Metilaminas/metabolismo , Animales , NADPH Oxidasa 2/antagonistas & inhibidores , NADPH Oxidasa 2/metabolismo , Humanos , Masculino , Microbioma Gastrointestinal/efectos de los fármacos , Insuficiencia Renal Crónica/microbiología , Insuficiencia Renal Crónica/complicaciones , Persona de Mediana Edad , Ratones , Estrés Oxidativo/efectos de los fármacos , Antibacterianos/efectos adversos , Antibacterianos/farmacología , Ratones Endogámicos C57BL , Femenino , Daño por Reperfusión/prevención & control , Anciano , Apoptosis/efectos de los fármacos , Progresión de la EnfermedadRESUMEN
Oxidants participate in lymphocyte activation and function. We previously demonstrated that eliminating the activity of NADPH oxidase 2 (NOX2) significantly impaired the effectiveness of autoreactive CD8+ CTLs. However, the molecular mechanisms impacting CD8+ T cell function remain unknown. In the present study, we examined the role of NOX2 in both NOD mouse and human CD8+ T cell function. Genetic ablation or chemical inhibition of NOX2 in CD8+ T cells significantly suppressed activation-induced expression of the transcription factor T-bet, the master transcription factor of the Tc1 cell lineage, and T-bet target effector genes such as IFN-γ and granzyme B. Inhibition of NOX2 in both human and mouse CD8+ T cells prevented target cell lysis. We identified that superoxide generated by NOX2 must be converted into hydrogen peroxide to transduce the redox signal in CD8+ T cells. Furthermore, we show that NOX2-generated oxidants deactivate the tumor suppressor complex leading to activation of RheB and subsequently mTOR complex 1. These results indicate that NOX2 plays a nonredundant role in TCR-mediated CD8+ T cell effector function.
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Linfocitos T CD8-positivos , NADPH Oxidasa 2 , Especies Reactivas de Oxígeno , Animales , Humanos , Ratones , Linfocitos T CD8-positivos/citología , Linfocitos T CD8-positivos/inmunología , Citocinas/inmunología , Granzimas/metabolismo , Peróxido de Hidrógeno/metabolismo , Inflamación/inmunología , Interferón gamma/metabolismo , Activación de Linfocitos , Ratones Endogámicos NOD , NADPH Oxidasa 2/antagonistas & inhibidores , NADPH Oxidasa 2/metabolismo , Oxidación-Reducción , Especies Reactivas de Oxígeno/metabolismo , Receptores de Antígenos de Linfocitos T/metabolismo , Transducción de Señal , Proteínas de Dominio T Box/metabolismo , Masculino , Femenino , Adulto JovenRESUMEN
Recent work by us and others has implicated NADPH oxidase (NOX) enzymes as main producers of reactive oxygen species (ROS) following a brain insult such as status epilepticus, contributing to neuronal damage and development of epilepsy. Although several NOX isoforms have been examined in the context of epilepsy, most attention has focused on NOX2. In this present study, we demonstrate the effect of gp91ds-tat, a specific competitive inhibitor of NOX2, in in vitro epileptiform activity model as well as in temporal lobe epilepsy (TLE) model in rats. We showed that in in vitro seizure model, gp91ds-tat modulated Ca2+ oscillation, prevented epileptiform activity-induced ROS generation, mitochondrial depolarization, and neuronal death. Administration of gp91ds-tat 1 h after kainic acid-induced status epilepticus significantly decreased the expression of NOX2, as well as the overall NOX activity in the cortex and the hippocampus. Finally, we showed that upon continuous intracerebroventricular administration to epileptic rats, gp91ds-tat significantly reduced the seizure frequency and the total number of seizures post-treatment compared to the scrambled peptide-treated animals. The results of the study suggest that NOX2 may have an important effect on modulation of epileptiform activity and has a critical role in mediating seizure-induced NOX activation, ROS generation and oxidative stress in the brain, and thus significantly contributes to development of epilepsy following a brain insult.
Asunto(s)
Epilepsia , NADPH Oxidasa 2 , Estado Epiléptico , Animales , Ratas , Epilepsia/tratamiento farmacológico , NADPH Oxidasa 2/antagonistas & inhibidores , Especies Reactivas de Oxígeno/metabolismo , ConvulsionesRESUMEN
The present study aimed to determine the antioxidant effect of Citrus unshiu Markovich (CUM) extract in neuronal cell lines under oxidative stress and to investigate the effect of chemotherapy-induced peripheral neuropathy (CIPN) on the nociceptive response in a preclinical mice model. We tested the inhibition of H2 O2 in Neuro2A cells treated with CUM. Experimental animals were treated with oxaliplatin to induce CINP, and then administered oral CUM for 4 weeks in order to observe the effect of CUM. Animals were evaluated weekly for thermal hyperalgesia and digital motor nerve conduction velocity (NCV). Lumbar dorsal root ganglia (DRG) isolated from each animal were evaluated through immunochemical and western blot analysis for nerve damage, inflammatory response, and expression of redox signaling factors. The main mechanisms were determined to be decreased inducible nitric oxide synthase (iNOS) production due to the inhibition of NADPH oxidase 2 (NOX2). To determine the functional role of NOX2 in CINP, we administrated CUM into NOX2-deficient mice with neuropathic pain. Therefore, we suggest that CUM controls the expression levels of inflammatory factors in CINP via NOX2 inactivation. This study demonstrated that a complementary medicine such as CUM might be a potential novel therapeutic agent for the treatment of CINP.
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Antineoplásicos , Citrus , Hiperalgesia , NADPH Oxidasa 2/antagonistas & inhibidores , Neuralgia , Fármacos Neuroprotectores/farmacología , Extractos Vegetales , Animales , Antineoplásicos/efectos adversos , Citrus/química , Hiperalgesia/inducido químicamente , Hiperalgesia/tratamiento farmacológico , Ratones , Modelos Animales , Neuralgia/inducido químicamente , Neuralgia/tratamiento farmacológico , Extractos Vegetales/farmacologíaRESUMEN
A predominant trigger and driver of sporadic Alzheimer's disease (AD) is the synergy of brain oxidative stress and glucose hypometabolism starting at early preclinical stages. Oxidative stress damages macromolecules, while glucose hypometabolism impairs cellular energy supply and antioxidant defense. However, the exact cause of AD-associated glucose hypometabolism and its network consequences have remained unknown. Here we report NADPH oxidase 2 (NOX2) activation as the main initiating mechanism behind Aß1-42-related glucose hypometabolism and network dysfunction. We utilize a combination of electrophysiology with real-time recordings of metabolic transients both ex- and in-vivo to show that Aß1-42 induces oxidative stress and acutely reduces cellular glucose consumption followed by long-lasting network hyperactivity and abnormalities in the animal behavioral profile. Critically, all of these pathological changes were prevented by the novel bioavailable NOX2 antagonist GSK2795039. Our data provide direct experimental evidence for causes and consequences of AD-related brain glucose hypometabolism, and suggest that targeting NOX2-mediated oxidative stress is a promising approach to both the prevention and treatment of AD.
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Aminopiridinas/farmacología , Péptidos beta-Amiloides/farmacología , Encéfalo/metabolismo , Glucosa/metabolismo , Hipercinesia/inducido químicamente , NADPH Oxidasa 2/antagonistas & inhibidores , NADPH Oxidasa 2/genética , Estrés Oxidativo , Sulfonamidas/farmacología , Animales , Masculino , Ratones , NADPH Oxidasa 2/metabolismoRESUMEN
Repeated morphine administration results in analgesic tolerance. However, the underlying mechanism of morphine analgesic tolerance remains unclear. NADPH-oxidase 2 (NOX2) is the first discovered NADPH oxidase, which mainly functions to produce reactive oxygen species. Its specific role in morphine tolerance has not been fully investigated. In this work, we found that chronic morphine administration significantly increased the expression of NOX2 in spinal cord. Pretreatment of NOX2 inhibitor blocked the upregulation of NOX2 and autophagy markers, including LC3B and P62, and consequently the development of morphine tolerance. NOX2 and LC3B were both colocalized with NeuN in spinal dorsal horn in morphine-tolerant rats. Our results suggest that the increased autophagy activity in spinal neurons promoted by NOX2 activation contributes to the development of morphine tolerance. NOX2 may be considered as a new therapeutic target for morphine tolerance.
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Analgésicos Opioides/farmacología , Autofagia/efectos de los fármacos , Tolerancia a Medicamentos/fisiología , Morfina/farmacología , NADPH Oxidasa 2/metabolismo , Neuronas/efectos de los fármacos , Animales , Masculino , Proteínas Asociadas a Microtúbulos/metabolismo , NADPH Oxidasa 2/antagonistas & inhibidores , Compuestos Onio/farmacología , Ratas Sprague-Dawley , Asta Dorsal de la Médula Espinal/citologíaRESUMEN
In ventilation-induced lung injury (VILI), prolonged nonpathogen-mediated inflammation is triggered as a result of alveolar hyperinflation. In our previous study, we suggested that endoplasmic reticulum (ER) stress-mediated inflammation was involved in VILI, but how ER stress is triggered remains unknown. Toll-like receptor 4 (TLR4) activation plays an important role in mechanical ventilation (MV)-induced lung inflammation, however, it is unknown whether ER stress is activated by TLR4 to participate in VILI. In this study, C57BL/6 mice were exposed to MV with high tidal volumes (HTV 20 ml/kg). Mice were pretreated with TAK-242 the TLR4 inhibitor, C25-140, the TRAF6 inhibitor, or GSK2795039, the NOX2 inhibitor. Lung tissue and bronchoalveolar lavage fluid (BALF) were collected to measure lung injury, inflammatory responses and mRNA/protein expression associated with ER stress and the TLR4/TRAF6/NOX2 signaling pathway. Our results indicate that MV with HTV caused the TLR4/TRAF6/NOX2 signaling pathway activation and production of large amounts of ROS, which led to ER stress and NF-κB mediated inflammation in VILI. Furthermore, TLR4/TRAF6/NOX2 signaling pathway inhibition attenuated ER stress response and alleviate lung injury in mice.
Asunto(s)
Estrés del Retículo Endoplásmico , Inflamación/patología , NADPH Oxidasa 2/metabolismo , Factor 6 Asociado a Receptor de TNF/metabolismo , Receptor Toll-Like 4/metabolismo , Lesión Pulmonar Inducida por Ventilación Mecánica/metabolismo , Animales , Modelos Animales de Enfermedad , Inflamación/etiología , Inflamación/inmunología , Masculino , Ratones , Ratones Endogámicos C57BL , NADPH Oxidasa 2/antagonistas & inhibidores , Transducción de Señal , Factor 6 Asociado a Receptor de TNF/antagonistas & inhibidores , Receptor Toll-Like 4/antagonistas & inhibidores , Lesión Pulmonar Inducida por Ventilación Mecánica/etiología , Lesión Pulmonar Inducida por Ventilación Mecánica/patologíaRESUMEN
Insufficient stress response and elevated oxidative stress can contribute to skeletal muscle atrophy during mechanical unloading (e.g., spaceflight and bedrest). Perturbations in heat shock proteins (e.g., HSP70), antioxidant enzymes, and sarcolemmal neuronal nitric oxidase synthase (nNOS) have been linked to unloading-induced atrophy. We recently discovered that the sarcolemmal NADPH oxidase-2 complex (Nox2) is elevated during unloading, downstream of angiotensin II receptor 1, and concomitant with atrophy. Here, we hypothesized that peptidyl inhibition of Nox2 would attenuate disruption of HSP70, MnSOD, and sarcolemmal nNOS during unloading, and thus muscle fiber atrophy. F344 rats were divided into control (CON), hindlimb unloaded (HU), and hindlimb unloaded +7.5 mg/kg/day gp91ds-tat (HUG) groups. Unloading-induced elevation of the Nox2 subunit p67phox-positive staining was mitigated by gp91ds-tat. HSP70 protein abundance was significantly lower in HU muscles, but not HUG. MnSOD decreased with unloading; however, MnSOD was not rescued by gp91ds-tat. In contrast, Nox2 inhibition protected against unloading suppression of the antioxidant transcription factor Nrf2. nNOS bioactivity was reduced by HU, an effect abrogated by Nox2 inhibition. Unloading-induced soleus fiber atrophy was significantly attenuated by gp91ds-tat. These data establish a causal role for Nox2 in unloading-induced muscle atrophy, linked to preservation of HSP70, Nrf2, and sarcolemmal nNOS.
Asunto(s)
Fibras Musculares Esqueléticas/metabolismo , Fibras Musculares Esqueléticas/patología , Atrofia Muscular/etiología , Atrofia Muscular/metabolismo , NADPH Oxidasa 2/antagonistas & inhibidores , Estrés Fisiológico , Ingravidez/efectos adversos , Animales , Biomarcadores , Proteínas del Choque Térmico HSP72/metabolismo , Modelos Biológicos , Complejos Multiproteicos/metabolismo , Óxido Nítrico Sintasa de Tipo I/metabolismo , Estrés Oxidativo , Unión Proteica , RatasRESUMEN
Rationale: Mechanisms underlying the compromised bone formation in type 1 diabetes mellitus (T1DM), which causes bone fragility and frequent fractures, remain poorly understood. Recent advances in organ-specific vascular endothelial cells (ECs) identify type H blood vessel injury in the bone, which actively direct osteogenesis, as a possible player. Methods: T1DM was induced in mice by streptozotocin (STZ) injection in two severity degrees. Bony endothelium, the coupling of angiogenesis and osteogenesis, and bone mass quality were evaluated. Insulin, antioxidants, and NADPH oxidase (NOX) inhibitors were administered to diabetic animals to investigate possible mechanisms and design therapeutic strategies. Results: T1DM in mice led to the holistic abnormality of the vascular system in the bone, especially type H vessels, resulting in the uncoupling of angiogenesis and osteogenesis and inhibition of bone formation. The severity of osteopathy was positively related to glycemic levels. These pathological changes were attenuated by early-started, but not late-started, insulin therapy. ECs in diabetic bones showed significantly higher levels of reactive oxygen species (ROS) and NOX 1 and 2. Impairments of bone vessels and bone mass were effectively ameliorated by treatment with anti-oxidants or NOX2 inhibitors, but not by a NOX1/4 inhibitor. GSK2795039 (GSK), a NOX2 inhibitor, significantly supplemented the insulin effect on the diabetic bone. Conclusions: Diabetic osteopathy could be a chronic microvascular complication of T1DM. The impairment of type H vessels by NOX2-mediated endothelial oxidative stress might be an important contributor that can serve as a therapeutic target for T1DM-induced osteopathy.
Asunto(s)
Huesos/irrigación sanguínea , Diabetes Mellitus Experimental/patología , Diabetes Mellitus Experimental/fisiopatología , NADPH Oxidasa 2/metabolismo , Animales , Antioxidantes/farmacología , Fenómenos Biomecánicos , Huesos/patología , Huesos/fisiopatología , Diabetes Mellitus Experimental/complicaciones , Diabetes Mellitus Tipo 1/patología , Diabetes Mellitus Tipo 1/fisiopatología , Células Endoteliales/fisiología , Insulina/administración & dosificación , Insulina/uso terapéutico , Masculino , Ratones , Ratones Endogámicos C57BL , Terapia Molecular Dirigida , NADPH Oxidasa 2/antagonistas & inhibidores , Neovascularización Fisiológica/efectos de los fármacos , Osteogénesis/efectos de los fármacos , Osteogénesis/fisiología , Osteoporosis/etiología , Osteoporosis/patología , Osteoporosis/fisiopatología , Estrés Oxidativo , Medicina de PrecisiónRESUMEN
BACKGROUND: Perinatal hypoxia is a universal cause of death and neurological deficits in neonates worldwide. Activation of microglial NADPH oxidase 2 (NOX2) leads to oxidative stress and neuroinflammation, which may contribute to hypoxic damage in the developing brain. Dexmedetomidine has been reported to exert potent neuroprotection in several neurological diseases, but the mechanism remains unclear. We investigated whether dexmedetomidine acts through microglial NOX2 to reduce neonatal hypoxic brain damage. METHODS: The potential role of microglial NOX2 in dexmedetomidine-mediated alleviation of hypoxic damage was evaluated in cultured BV2 microglia and neonatal rats subjected to hypoxia. In vivo, neonatal rats received dexmedetomidine (25 µg/kg, i.p.) 30 min before or immediately after hypoxia (5% O2, 2 h). Apocynin-mediated NOX inhibition and lentivirus-mediated NOX2 overexpression were applied to further assess the involvement of microglial NOX2 activation. RESULTS: Pre- or posttreatment with dexmedetomidine alleviated hypoxia-induced cognitive impairment, restored damaged synapses, and increased postsynaptic density-95 and synaptophysin protein expression following neonatal hypoxia. Importantly, dexmedetomidine treatment suppressed hypoxia-induced microglial NOX2 activation and subsequent oxidative stress and the neuroinflammatory response, as reflected by reduced 4-hydroxynonenal and ROS accumulation, and decreased nuclear NF-κB p65 and proinflammatory cytokine levels in cultured BV2 microglia and the developing hippocampus. In addition, treating primary hippocampal neurons with conditioned medium (CM) from hypoxia-activated BV2 microglia resulted in neuronal damage, which was alleviated by CM from dexmedetomidine-treated microglia. Moreover, the neuroprotective effect of dexmedetomidine was reversed in NOX2-overexpressing BV2 microglia and diminished in apocynin-pretreated neonatal rats. CONCLUSION: Dexmedetomidine targets microglial NOX2 to reduce oxidative stress and neuroinflammation and subsequently protects against hippocampal synaptic loss following neonatal hypoxia.
Asunto(s)
Disfunción Cognitiva/enzimología , Disfunción Cognitiva/etiología , Dexmedetomidina/farmacología , Hipocampo/patología , Hipoxia/complicaciones , Microglía/enzimología , NADPH Oxidasa 2/metabolismo , Sinapsis/patología , Acetofenonas/farmacología , Animales , Animales Recién Nacidos , Células Cultivadas , Disfunción Cognitiva/patología , Citocinas/metabolismo , Activación Enzimática/efectos de los fármacos , Hipocampo/ultraestructura , Mediadores de Inflamación/metabolismo , Microglía/efectos de los fármacos , Microglía/patología , Microglía/ultraestructura , Modelos Biológicos , NADPH Oxidasa 2/antagonistas & inhibidores , FN-kappa B/metabolismo , Neuroprotección/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Ratas Sprague-Dawley , Transducción de Señal/efectos de los fármacos , Sinapsis/efectos de los fármacos , Sinapsis/ultraestructuraRESUMEN
AIMS: Gp91-containing NADPH oxidases (NOX2) are a significant source of myocardial superoxide production. An increase in NOX2 activity accompanies atrial fibrillation (AF) induction and electrical remodelling in animal models and predicts incident AF in humans; however, a direct causal role for NOX2 in AF has not been demonstrated. Accordingly, we investigated whether myocardial NOX2 overexpression in mice (NOX2-Tg) is sufficient to generate a favourable substrate for AF and further assessed the effects of atorvastatin, an inhibitor of NOX2, on atrial superoxide production and AF susceptibility. METHODS AND RESULTS: NOX2-Tg mice showed a 2- to 2.5-fold higher atrial protein content of NOX2 compared with wild-type (WT) controls, which was associated with a significant (twofold) increase in NADPH-stimulated superoxide production (2-hydroxyethidium by HPLC) in left and right atrial tissue homogenates (P = 0.004 and P = 0.019, respectively). AF susceptibility assessed in vivo by transoesophageal atrial burst stimulation was modestly increased in NOX2-Tg compared with WT (probability of AF induction: 88% vs. 69%, respectively; P = 0.037), in the absence of significant alterations in AF duration, surface ECG parameters, and LV mass or function. Mechanistic studies did not support a role for NOX2 in promoting electrical or structural remodelling, as high-resolution optical mapping of atrial tissues showed no differences in action potential duration and conduction velocity between genotypes. In addition, we did not observe any genotype difference in markers of fibrosis and inflammation, including atrial collagen content and Col1a1, Il-1ß, Il-6, and Mcp-1 mRNA. Similarly, NOX2 overexpression did not have consistent effects on RyR2 Ca2+ leak nor did it affect PKA or CaMKII-mediated RyR2 phosphorylation. Finally, treatment with atorvastatin significantly inhibited atrial superoxide production in NOX2-Tg but had no effect on AF induction in either genotype. CONCLUSION: Together, these data indicate that while atrial NOX2 overexpression may contribute to atrial arrhythmogenesis, NOX2-derived superoxide production does not affect the electrical and structural properties of the atrial myocardium.
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Fibrilación Atrial/enzimología , Atrios Cardíacos/enzimología , Frecuencia Cardíaca , Miocitos Cardíacos/enzimología , NADPH Oxidasa 2/biosíntesis , Potenciales de Acción , Animales , Antiarrítmicos/farmacología , Atorvastatina/farmacología , Fibrilación Atrial/genética , Fibrilación Atrial/fisiopatología , Fibrilación Atrial/prevención & control , Modelos Animales de Enfermedad , Inducción Enzimática , Inhibidores Enzimáticos/farmacología , Atrios Cardíacos/efectos de los fármacos , Atrios Cardíacos/fisiopatología , Ratones Transgénicos , Miocitos Cardíacos/efectos de los fármacos , NADPH Oxidasa 2/antagonistas & inhibidores , NADPH Oxidasa 2/genética , Transducción de Señal , Superóxidos/metabolismo , Factores de TiempoRESUMEN
Following traumatic brain injury (TBI), increased production of reactive oxygen species (ROS) and the ensuing oxidative stress promotes the secondary brain damage that encompasses both grey matter and white matter. As this contributes to the long-term neurological deficits, decreasing oxidative stress during the acute period of TBI is beneficial. While NADPH oxidase (NOX2) is the major producer of ROS, transcription factor Nrf2 that induces antioxidant enzymes promotes efficient ROS disposal. We recently showed that treatment with an antioxidant drug combo of apocynin (NOX2 inhibitor) and TBHQ (Nrf2 activator) protects the grey matter in adult mice subjected to TBI. We currently show that this antioxidant combo therapy given at 2 h and 24 h after TBI also protects white matter in mouse brain. Thus, the better functional outcomes after TBI in the combo therapy treated mice might be due to a combination of sparing both grey matter and white matter. Hence, the antioxidant combo we tested is a potent therapeutic option for translation in future.
Asunto(s)
Acetofenonas/uso terapéutico , Antioxidantes/uso terapéutico , Lesiones Traumáticas del Encéfalo/tratamiento farmacológico , Hidroquinonas/uso terapéutico , Sustancia Blanca/efectos de los fármacos , Acetofenonas/administración & dosificación , Animales , Antioxidantes/administración & dosificación , Lesiones Traumáticas del Encéfalo/patología , Esquema de Medicación , Evaluación Preclínica de Medicamentos , Sinergismo Farmacológico , Quimioterapia Combinada , Sustancia Gris/efectos de los fármacos , Sustancia Gris/patología , Hidroquinonas/administración & dosificación , Masculino , Ratones , Ratones Endogámicos C57BL , NADPH Oxidasa 2/antagonistas & inhibidores , Factor 2 Relacionado con NF-E2/agonistas , Estrés Oxidativo/efectos de los fármacos , Distribución Aleatoria , Sustancia Blanca/patologíaRESUMEN
Reduced mechanical loading results in atrophy of skeletal muscle fibers. Increased reactive oxygen species (ROS) are causal in sarcolemmal dislocation of nNOS and FoxO3a activation. The Nox2 isoform of NADPH oxidase and mitochondria release ROS during disuse in skeletal muscle. Activation of the angiotensin II type 1 receptor (AT1R) can elicit Nox2 complex formation. The AT1R blocker losartan was used to test the hypothesis that AT1R activation drives Nox2 assembly, nNOS dislocation, FoxO3a activation, and thus alterations in morphology in the unloaded rat soleus. Male Fischer 344 rats were divided into four groups: ambulatory control (CON), ambulatory + losartan (40 mg kg-1 day-1 ) (CONL), 7 days of tail-traction hindlimb unloading (HU), and HU + losartan (HUL). Losartan attenuated unloading-induced loss of muscle fiber cross-sectional area (CSA) and fiber-type shift. Losartan mitigated unloading-induced elevation of ROS levels and upregulation of Nox2. Furthermore, AT1R blockade abrogated nNOS dislocation away from the sarcolemma and elevation of nuclear FoxO3a. We conclude that AT1R blockade attenuates disuse remodeling by inhibiting Nox2, thereby lessening nNOS dislocation and activation of FoxO3a.
Asunto(s)
Losartán/farmacología , Fibras Musculares Esqueléticas/efectos de los fármacos , Atrofia Muscular/tratamiento farmacológico , NADPH Oxidasa 2/antagonistas & inhibidores , Especies Reactivas de Oxígeno/metabolismo , Animales , Antihipertensivos/farmacología , Modelos Animales de Enfermedad , Suspensión Trasera/efectos adversos , Suspensión Trasera/métodos , Masculino , Fibras Musculares Esqueléticas/metabolismo , Fibras Musculares Esqueléticas/patología , Atrofia Muscular/etiología , Atrofia Muscular/metabolismo , NADPH Oxidasa 2/metabolismo , Estrés Oxidativo/efectos de los fármacos , Ratas , Transducción de SeñalRESUMEN
Activation of the Nox2-dependent NADPH oxidase is the result of a conformational change in Nox2 induced by interaction with the cytosolic component p67phox . In preliminary work we identified a cluster of overlapping 15-mer synthetic peptides, corresponding to p67phox residues 259-279, which inhibited oxidase activity in an in vitro, cell-free assay, but the results did not point to a competitive mechanism. We recently identified an auto-inhibitory intramolecular bond in p67phox , one extremity of which was located within the 259-279 sequence, and we hypothesized that inhibition by exogenous peptides might mimic intrinsic auto-inhibition. In this study, we found that: (i) progressive N- and C-terminal truncation of inhibitory p67phox peptides, corresponding to residues 259-273 and 265-279, revealed that inhibitory ability correlated with the presence of residues 265 NIVFVL270 , exposed at either the N- or C-termini of the peptides; (ii) inhibition of oxidase activity was associated exclusively with self-assembled peptides, which pelleted upon centrifugation at 12,000 ×g; (iii) self-assembled p67phox peptides inhibited oxidase activity by specific binding of p67phox and the ensuing depletion of this component, essential for interaction with Nox2; and (iv) peptides subjected to scrambling or reversing the order of residues in NIVFVL retained the propensity for self-assembly, oxidase inhibitory ability, and specific binding of p67phox , indicating that the dominant parameter was the hydrophobic character of five of the six residues. This appears to be the first description of inhibition of oxidase activity by self-assembled peptides derived from an oxidase component, acting by an auto-inhibitory mechanism.
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NADPH Oxidasa 2/antagonistas & inhibidores , NADPH Oxidasa 2/metabolismo , Péptidos/farmacología , Fosfoproteínas/metabolismo , Secuencia de Aminoácidos , Animales , Activación Enzimática/efectos de los fármacos , Cobayas , Interacciones Hidrofóbicas e Hidrofílicas , Péptidos/química , Dominios ProteicosRESUMEN
Traumatic brain injury (TBI) is a subset of brain injury induced by external mechanical forces to the head or neck. TBI has been reported to be one of the leading causes of disability, and it causes a huge financial burden around the world. Aloin is the major anthraquinone glycoside extracted from Aloe species, and has presented anti-tumour, anti-oxidative and anti-inflammatory activities. However, few studies have focused the effect of aloin in treatment of TBI. Nicotinamide adenine dinucleotide phosphate oxidase (NOX) is the only subset of enzymes which produces solely the reactive oxygen species (ROS). A recent study showed that activation of NOX might aggravate the primary TBI, and among these members, NOX2 is the key member in regulation of uncontrolled ROS expression, and thus plays a critical role in development of inflammatory diseases. Here, we noticed that inhibition of NOX2 combined with aloin treatment promoted the recovery of brain function in a mice model as well as the viability rate in a cell model. A further study found that the inflammation response process was also inhibited after treatment. Then, we found that these effects might be mediated by the PI3K/AKT/mTOR signalling pathway and NOX2 might be a therapeutic target for TBI.
Asunto(s)
Lesiones Traumáticas del Encéfalo/tratamiento farmacológico , Encéfalo/efectos de los fármacos , Emodina/análogos & derivados , NADPH Oxidasa 2/antagonistas & inhibidores , Estrés Oxidativo/efectos de los fármacos , Animales , Encéfalo/metabolismo , Lesiones Traumáticas del Encéfalo/metabolismo , Citocinas/metabolismo , Emodina/farmacología , Emodina/uso terapéutico , Factor I del Crecimiento Similar a la Insulina/farmacología , Aprendizaje por Laberinto/efectos de los fármacos , Ratones , Células PC12 , Fosforilación/efectos de los fármacos , Proteínas Proto-Oncogénicas c-akt/metabolismo , Ratas , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal/efectos de los fármacos , Superóxido Dismutasa/metabolismoRESUMEN
Clinical studies implicated an increased risk of intestinal fibrosis in patients with nonalcoholic fatty liver disease (NAFLD). Our previous studies have shown that microcystin-LR (MC-LR) exposure led to altered gut microbiome and increased abundance of lactate producing bacteria and intestinal inflammation in underlying NAFLD. This led us to further investigate the effects of the MC-LR, a PP2A inhibitor in activating the TGF-ß fibrotic pathway in the intestines that might be mediated by increased lactate induced redox enzyme NOX2. Exposure to MC-LR led to higher lactate levels in circulation and in the intestinal content. The higher lactate levels were associated with NOX2 activation in vivo that led to increased Smad2/3-Smad4 co-localization and high alpha-smooth muscle actin (α-SMA) immunoreactivity in the intestines. Mechanistically, primary mouse intestinal epithelial cells treated with lactate and MC-LR separately led to higher NOX2 activation, phosphorylation of TGFßR1 receptor and subsequent Smad 2/3-Smad4 co-localization inhibitable by apocynin (NOX2 inhibitor), FBA (a peroxynitrite scavenger) and DMPO (a nitrone spin trap), catalase and superoxide dismutase. Inhibition of NOX2-induced redox signaling also showed a significant decrease in collagen protein thus suggesting a strong redox signaling induced activation of an ectopic fibrotic manifestation in the intestines. In conclusion, the present study provides mechanistic insight into the role of microcystin in dysbiosis-linked lactate production and subsequently advances our knowledge in lactate-induced NOX2 exacerbation of the cell differentiation and fibrosis in the NAFLD intestines.
Asunto(s)
Fibrosis/patología , Mucosa Intestinal/metabolismo , Intestinos/efectos de los fármacos , Ácido Láctico/metabolismo , Microcistinas/toxicidad , NADPH Oxidasa 2/metabolismo , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Animales , Línea Celular , Inhibidores Enzimáticos/toxicidad , Fibrosis/enzimología , Fibrosis/etiología , Mucosa Intestinal/efectos de los fármacos , Intestinos/enzimología , Intestinos/patología , Ácido Láctico/sangre , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , NADPH Oxidasa 2/antagonistas & inhibidores , Enfermedad del Hígado Graso no Alcohólico/sangre , Enfermedad del Hígado Graso no Alcohólico/enzimología , Enfermedad del Hígado Graso no Alcohólico/patología , FosforilaciónRESUMEN
Excessive fructose intake is a risk factor for liver oxidative stress injury. Magnesium isoglycyrrhizinate as a hepatoprotective agent is used to treat liver diseases in clinic. However, its antioxidant effects and the underlying potential mechanisms are still not clearly understood. In this study, magnesium isoglycyrrhizinate was found to alleviate liver oxidative stress and inflammatory injury in fructose-fed rats. Magnesium isoglycyrrhizinate suppressed hepatic reactive oxygen species overproduction (0.97 ± 0.04 a.u. versus 1.34 ± 0.07 a.u.) in fructose-fed rats by down-regulating mRNA and protein levels of nicotinamide adenine dinucleotide phosphate oxidase (NOX) 1, NOX2 and NOX4, resulting in reduction of interleukin-1ß (IL-1ß) levels (1.13 ± 0.09 a.u. versus 1.97 ± 0.12 a.u.). Similarly, magnesium isoglycyrrhizinate reduced reactive oxygen species overproduction (1.07 ± 0.02 a.u. versus 1.35 ± 0.06 a.u.) and IL-1ß levels (1.14 ± 0.09 a.u. versus 1.66 ± 0.07 a.u.) in fructose-exposed HepG2 cells. Furthermore, data from treatment of reactive oxygen species inhibitor N-acetyl-L-cysteine or NOXs inhibitor diphenyleneiodonium in fructose-exposed HepG2 cells showed that fructose enhanced NOX1, NOX2 and NOX4 expression to increase reactive oxygen species generation, causing oxidative stress and inflammation, more importantly, these disturbances were significantly attenuated by magnesium isoglycyrrhizinate. The molecular mechanisms underpinning these effects suggest that magnesium isoglycyrrhizinate may inhibit NOX1, NOX2 and NOX4 expression to reduce reactive oxygen species generation, subsequently prevent liver oxidative stress injury under high fructose condition. Thus, the blockade of NOX1, NOX2 and NOX4 expression by magnesium isoglycyrrhizinate may be the potential therapeutic approach for improving fructose-induced liver injury in clinic.
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Antiinflamatorios/farmacología , Antioxidantes/farmacología , Enfermedad Hepática Inducida por Sustancias y Drogas/prevención & control , Hígado/efectos de los fármacos , NADPH Oxidasas/antagonistas & inhibidores , Estrés Oxidativo/efectos de los fármacos , Saponinas/farmacología , Triterpenos/farmacología , Animales , Enfermedad Hepática Inducida por Sustancias y Drogas/enzimología , Enfermedad Hepática Inducida por Sustancias y Drogas/patología , Modelos Animales de Enfermedad , Fructosa , Células Hep G2 , Humanos , Mediadores de Inflamación/metabolismo , Interleucina-1beta/metabolismo , Hígado/enzimología , Hígado/patología , Masculino , NADPH Oxidasa 1/antagonistas & inhibidores , NADPH Oxidasa 1/metabolismo , NADPH Oxidasa 2/antagonistas & inhibidores , NADPH Oxidasa 2/metabolismo , NADPH Oxidasa 4/antagonistas & inhibidores , NADPH Oxidasa 4/metabolismo , NADPH Oxidasas/metabolismo , Ratas Sprague-Dawley , Transducción de SeñalRESUMEN
Myocardial infarction (M/I) is a common cause of mortality worldwide. Agomelatine (AGO), a potent melatonin receptor agonist, proved to have an anti-inflammatory and antioxidant effect. The present study aimed to explore the cardioprotective effect of AGO on isoproterenol (ISO)-induced myocardial injury in a rat model and determine the role of nitric oxide (NO) in mediating this beneficial effect. Rats were randomly divided into 6 groups and treated for 12 days. Group 1, control, received normal saline. Group 2, ISO group, received ISO (100 mg/kg, i.p.) in 11th and 12th days. Group 3, positive control group, received atenolol (100 mg/kg/day) + ISO. Group 4, AGO-treated group, received AGO (80 mg/kg/day) + ISO. Group 5, L-NNA + ISO, received L-NG-nitro arginine (L-NNA) (25 mg/kg, orally) + ISO. Group 6, AGO + L-NNA + ISO, co-treated with AGO + ISO + L-NNA. Serum cardiac enzymes and cardiac tissue oxidative stress parameters were assessed along with histopathological evaluation. Gene expression quantification of nuclear factor erythroid 2 (Nrf-2) and heme oxygenase-1 (HO-1) were assessed. Immunoexpression of inducible NO synthase (iNOS) and caspase-3 were evaluated. The outcomes proved that ISO significantly increased serum cardiac enzymes, with histopathological changes of myocardial tissue along with a major increase in oxidative, inflammatory, and nitrosative stress, besides a reduction in cardiac Nrf-2 and HO-1 gene expressions with marked myocardial cell apoptosis. However, pretreatment with AGO significantly reversed these profound ISO myocardial damaging effects. AGO protects against ISO-induced myocardial injury through its antioxidant, anti-inflammatory, and anti-apoptotic effects with modulation of NOS enzymes.
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Acetamidas/uso terapéutico , Cardiotónicos/uso terapéutico , Isoproterenol/toxicidad , Infarto del Miocardio/metabolismo , Infarto del Miocardio/prevención & control , Óxido Nítrico/metabolismo , Acetamidas/farmacología , Animales , Cardiotónicos/farmacología , Masculino , Infarto del Miocardio/inducido químicamente , Miocardio/metabolismo , NADPH Oxidasa 2/antagonistas & inhibidores , NADPH Oxidasa 2/metabolismo , Estrés Oxidativo/efectos de los fármacos , Estrés Oxidativo/fisiología , Ratas , Ratas WistarRESUMEN
Insulin resistance leads to excessive endothelial cell (EC) superoxide generation and accelerated atherosclerosis. The principal source of superoxide from the insulin-resistant endothelium is the Nox2 isoform of NADPH oxidase. Here we examine the therapeutic potential of Nox2 inhibition on superoxide generation in saphenous vein ECs (SVECs) from patients with advanced atherosclerosis and type 2 diabetes and on vascular function, vascular damage, and lipid deposition in apolipoprotein E-deficient (ApoE-/-) mice with EC-specific insulin resistance (ESMIRO). To examine the effect of genetic inhibition of Nox2, ESMIRO mice deficient in ApoE-/- and Nox2 (ESMIRO/ApoE-/-/Nox2-/y) were generated and compared with ESMIRO/ApoE-/-/Nox2+/y littermates. To examine the effect of pharmacological inhibition of Nox2, we administered gp91dstat or scrambled peptide to ESMIRO/ApoE-/- mice. SVECs from diabetic patients had increased expression of Nox2 protein with concomitant increase in superoxide generation, which could be reduced by the Nox2 inhibitor gp91dstat. After 12 wk Western diet, ESMIRO/ApoE-/-/Nox2-/y mice had reduced EC superoxide generation and greater aortic relaxation to acetylcholine. ESMIRO/ApoE-/-/Nox2-/y mice developed more lipid deposition in the thoraco-abdominal aorta with multiple foci of elastin fragmentation at the level of the aortic sinus and greater expression of intercellular adhesion molecule-1 (ICAM-1). Gp91dstat reduced EC superoxide and lipid deposition in the thoraco-abdominal aorta of ESMIRO/ApoE-/- mice without causing elastin fragmentation or increased ICAM-1 expression. These results demonstrate that insulin resistance is characterized by increased Nox2-derived vascular superoxide. Complete deletion of Nox2 in mice with EC insulin resistance exacerbates, whereas partial pharmacological Nox2 inhibition protects against, insulin resistance-induced vascular damage.
Asunto(s)
Diabetes Mellitus/metabolismo , Endotelio Vascular/metabolismo , Glicoproteínas/farmacología , Resistencia a la Insulina/fisiología , NADPH Oxidasa 2/antagonistas & inhibidores , NADPH Oxidasa 2/genética , Anciano , Anciano de 80 o más Años , Animales , Células Cultivadas , Diabetes Mellitus/genética , Diabetes Mellitus/patología , Endotelio Vascular/efectos de los fármacos , Endotelio Vascular/patología , Femenino , Humanos , Masculino , Ratones , Ratones Noqueados , Ratones Transgénicos , Persona de Mediana Edad , NADPH Oxidasa 2/deficiencia , Técnicas de Cultivo de ÓrganosRESUMEN
Sleep deprivation (SD) can induce cognitive and memory impairments. This impairment is in part due to oxidative stress damage in the hippocampus region of the brain. Corilagin (CL), a polyphenol belonging to the tannin family and extracted from Terminalia chebula and Phyllanthus emblica, shows strong antioxidant and neuroprotective effects. NF-E2-related factor (Nrf2)/heme oxygenase-1 (HO-1) and NADPH oxidase (NOX) are critical targets involved in cellular defense mechanisms against oxidative injury. Thus, we hypothesized that CL could be a preventive treatment for SD-induced memory impairments by inhibiting NOX2 and activating Nrf2. The results from behavioral tests showed that administration of CL resulted in significantly better performance compared to the SD mice. CL significantly normalized the elevated MDA level and the reduced activity of GPx and SOD (P <0.05, p<0.01) caused by SD. In hippocampal tissues, CL effectively activated Nrf2/HO-1 signaling and downregulated NOX2 protein expression compared with SD (P <0.05, P <0.01). Meanwhile, in vitro findings showed that knockdown of Nrf2 blocked the protective effect of CL versus Glu-induced toxicity, while the effect of CL was enhanced in NOX2 siRNA-transfected neurons. Overall, these findings provided evidence that CL ameliorates SD-induced memory impairments in mice by inhibiting NOX2 and activating Nrf2.