RESUMEN

PURPOSE: Chronic granulomatous disease (CGD) is an inherited immunodeficiency caused by pathogenic variants of genes encoding the enzyme complex NADPH oxidase. In countries where tuberculosis (TB) is endemic and the Bacillus Calmette-Guérin (BCG) vaccine is routinely administered, mycobacteria are major disease-causing pathogens in CGD. However, information on the clinical evolution and treatment of mycobacterial diseases in patients with CGD is limited. The present study describes the adverse reactions to BCG and TB in Mexican patients with CGD. METHODS: Patients with CGD who were evaluated at the Immunodeficiency Laboratory of the National Institute of Pediatrics between 2013 and 2024 were included. Medical records were reviewed to determine the clinical course and treatment of adverse reactions to BCG and TB disease. RESULTS: A total of 79 patients with CGD were included in this study. Adverse reactions to BCG were reported in 55 (72%) of 76 patients who received the vaccine. Tuberculosis was diagnosed in 19 (24%) patients. Relapse was documented in three (10%) of 31 patients with BGC-osis and six (32%) of 19 patients with TB, despite antituberculosis treatment. There was no difference in the frequency of BCG and TB disease between patients with pathogenic variants of the X-linked CYBB gene versus recessive variants. CONCLUSIONS: This report highlights the importance of considering TB in endemic areas and BCG complications in children with CGD to enable appropriate diagnostic and therapeutic approaches to improve prognosis and reduce the risk of relapse.

Asunto(s)

Vacuna BCG , Enfermedad Granulomatosa Crónica , NADPH Oxidasa 2 , Tuberculosis , Humanos , Enfermedad Granulomatosa Crónica/diagnóstico , Enfermedad Granulomatosa Crónica/epidemiología , Enfermedad Granulomatosa Crónica/complicaciones , Vacuna BCG/efectos adversos , Masculino , Femenino , Niño , Tuberculosis/epidemiología , Tuberculosis/inmunología , Preescolar , Lactante , Adolescente , NADPH Oxidasa 2/genética , Estudios de Cohortes , Mycobacterium bovis , México/epidemiología , Antituberculosos/uso terapéutico , NADPH Oxidasas/genética

RESUMEN

BACKGROUND: This study evaluated the effects of concurrent isolated training (T) or training combined with the antioxidant N-acetylcysteine (NAC) on cardiac remodeling and oxidative stress in spontaneously hypertensive rats (SHR). METHODS: Six-month-old male SHR were divided into sedentary (S, n = 12), concurrent training (T, n = 13), sedentary supplemented with NAC (SNAC, n = 13), and concurrent training with NAC supplementation (TNAC, n = 14) groups. T and TNAC rats were trained three times a week on a treadmill and ladder; NAC supplemented groups received 120 mg/kg/day NAC in rat chow for eight weeks. Myocardial antioxidant enzyme activity and lipid hydroperoxide concentration were assessed by spectrophotometry. Gene expression of NADPH oxidase subunits Nox2, Nox4, p22 phox, and p47 phox was evaluated by real time RT-PCR. Statistical analysis was performed using ANOVA and Bonferroni or Kruskal-Wallis and Dunn. RESULTS: Echocardiogram showed concentric remodeling in TNAC, characterized by increased relative wall thickness (S 0.40 ± 0.04; T 0.39 ± 0.03; SNAC 0.40 ± 0.04; TNAC 0.43 ± 0.04 *; * p < 0.05 vs T and SNAC) and diastolic posterior wall thickness (S 1.50 ± 0.12; T 1.52 ± 0.10; SNAC 1.56 ± 0.12; TNAC 1.62 ± 0.14 * mm; * p < 0.05 vs T), with improved contractile function (posterior wall shortening velocity: S 39.4 ± 5.01; T 36.4 ± 2.96; SNAC 39.7 ± 3.44; TNAC 41.6 ± 3.57 * mm/s; * p < 0.05 vs T). Myocardial lipid hydroperoxide concentration was lower in NAC treated groups (S 210 ± 48; T 182 ± 43; SNAC 159 ± 33 *; TNAC 110 ± 23 *# nmol/g tissue; * p < 0.05 vs S, # p < 0.05 vs T and SNAC). Nox 2 and p22 phox expression was higher and p47 phox lower in T than S [S 1.37 (0.66-1.66); T 0.78 (0.61-1.04) *; SNAC 1.07 (1.01-1.38); TNAC 1.06 (1.01-1.15) arbitrary units; * p < 0.05 vs S]. NADPH oxidase subunits did not differ between TNAC, SNAC, and S groups. CONCLUSION: N-acetylcysteine supplementation alone reduces oxidative stress in untreated spontaneously hypertensive rats. The combination of N-acetylcysteine and concurrent exercise further decreases oxidative stress. However, the lower oxidative stress does not translate into improved cardiac remodeling and function in untreated spontaneously hypertensive rats.

Asunto(s)

Acetilcisteína , Hipertensión , NADPH Oxidasas , Estrés Oxidativo , Ratas Endogámicas SHR , Remodelación Ventricular , Animales , Masculino , Estrés Oxidativo/efectos de los fármacos , Acetilcisteína/farmacología , Remodelación Ventricular/efectos de los fármacos , Hipertensión/fisiopatología , Hipertensión/tratamiento farmacológico , Hipertensión/metabolismo , NADPH Oxidasas/metabolismo , NADPH Oxidasas/genética , Ratas , Antioxidantes/farmacología , Condicionamiento Físico Animal , Modelos Animales de Enfermedad , NADPH Oxidasa 2/metabolismo , NADPH Oxidasa 2/genética , NADPH Oxidasa 4/metabolismo , NADPH Oxidasa 4/genética , Glicoproteínas de Membrana/metabolismo , Glicoproteínas de Membrana/genética , Miocardio/metabolismo , Miocardio/patología , Peróxidos Lipídicos/metabolismo , Función Ventricular Izquierda/efectos de los fármacos , Suplementos Dietéticos , Hipertrofia Ventricular Izquierda/fisiopatología , Hipertrofia Ventricular Izquierda/prevención & control , Hipertrofia Ventricular Izquierda/metabolismo

RESUMEN

Paraquat (1,1'-dimethyl-4,4'-bipyridyl dichloride) is an herbicide widely used worldwide and officially banned in Brazil in 2020. Kidney lesions frequently occur, leading to acute kidney injury (AKI) due to exacerbated reactive O2 species (ROS) production. However, the consequences of ROS exposure on ionic transport and the regulator local renin-angiotensin-aldosterone system (RAAS) still need to be elucidated at a molecular level. This study evaluated how ROS acutely influences Na+-transporting ATPases and the renal RAAS. Adult male Wistar rats received paraquat (20 mg/kg; ip). After 24 h, we observed body weight loss and elevation of urinary flow and serum creatinine. In the renal cortex, paraquat increased ROS levels, NADPH oxidase and (Na++K+)ATPase activities, angiotensin II-type 1 receptors, tumor necrosis factor-α (TNF-α), and interleukin-6. In the medulla, paraquat increased ROS levels and NADPH oxidase activity but inhibited (Na++K+)ATPase. Paraquat induced opposite effects on the ouabain-resistant Na+-ATPase in the cortex (decrease) and medulla (increase). These alterations, except for increased serum creatinine and renal levels of TNF-α and interleukin-6, were prevented by 4-hydroxy-2,2,6,6-tetramethylpiperidin-1-oxyl (tempol; 1 mmol/L in drinking water), a stable antioxidant. In summary, after paraquat poisoning, ROS production culminated with impaired medullary function, urinary fluid loss, and disruption of Na+-transporting ATPases and angiotensin II signaling.

Asunto(s)

Paraquat , Sistema Renina-Angiotensina , Ratas , Animales , Masculino , Especies Reactivas de Oxígeno/metabolismo , Paraquat/metabolismo , Paraquat/farmacología , Angiotensina II/metabolismo , Angiotensina II/farmacología , Creatinina/metabolismo , Creatinina/orina , Interleucina-6 , Factor de Necrosis Tumoral alfa/metabolismo , Ratas Wistar , Riñón , Adenosina Trifosfatasas/metabolismo , Adenosina Trifosfatasas/farmacología , Sodio/metabolismo , Sodio/farmacología , NADPH Oxidasas/metabolismo , NADPH Oxidasas/farmacología

RESUMEN

The endogenous metabolite of estradiol, estradiol 17ß-D-glucuronide (E17G), is considered the main responsible of the intrahepatic cholestasis of pregnancy. E17G alters the activity of canalicular transporters through a signaling pathway-dependent cellular internalization, phenomenon that was attributed to oxidative stress in different cholestatic conditions. However, there are no reports involving oxidative stress in E17G-induced cholestasis, representing this the aim of our work. Using polarized hepatocyte cultures, we showed that antioxidant compounds prevented E17G-induced Mrp2 activity alteration, being this alteration equally prevented by the NADPH oxidase (NOX) inhibitor apocynin. The model antioxidant N-acetyl-cysteine prevented, in isolated and perfused rat livers, E17G-induced impairment of bile flow and Mrp2 activity, thus confirming the participation of reactive oxygen species (ROS) in this cholestasis. In primary cultured hepatocytes, pretreatment with specific inhibitors of ERK1/2 and p38MAPK impeded E17G-induced ROS production; contrarily, NOX inhibition did not affect ERK1/2 and p38MAPK phosphorylation. Both, knockdown of p47phox by siRNA and preincubation with apocynin in sandwich-cultured rat hepatocytes significantly prevented E17G-induced internalization of Mrp2, suggesting a crucial role for NOX in this phenomenon. Concluding, E17G-induced cholestasis is partially mediated by NOX-generated ROS through internalization of canalicular transporters like Mrp2, being ERK1/2 and p38MAPK necessary for NOX activation.

Asunto(s)

Estradiol , Hepatocitos , NADPH Oxidasas , Especies Reactivas de Oxígeno , Animales , NADPH Oxidasas/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Ratas , Hepatocitos/metabolismo , Hepatocitos/efectos de los fármacos , Estradiol/farmacología , Estradiol/metabolismo , Estradiol/análogos & derivados , Femenino , Colestasis/inducido químicamente , Colestasis/metabolismo , Colestasis/patología , Ratas Wistar , Acetofenonas/farmacología , Estrés Oxidativo/efectos de los fármacos , Acetilcisteína/farmacología , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/metabolismo , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Células Cultivadas , Antioxidantes/farmacología , Antioxidantes/metabolismo , Colestasis Intrahepática , Complicaciones del Embarazo , Transportadoras de Casetes de Unión a ATP

RESUMEN

NADPH oxidase enzymes (NOX) are involved in all stages of carcinogenesis, but their expression levels and prognostic value in breast cancer (BC) remain unclear. Thus, we aimed to assess the expression and prognostic value of NOX enzymes in BC samples using online databases. For this, mRNA expression from 290 normal breast tissue samples and 1904 BC samples obtained from studies on cBioPortal, Kaplan-Meier Plotter, and The Human Protein Atlas were analyzed. We found higher levels of NOX2, NOX4, and Dual oxidase 1 (DUOX1) in normal breast tissue. NOX1, NOX2, and NOX4 exhibited higher expression in BC, except for the basal subtype, where NOX4 expression was lower. DUOX1 mRNA levels were lower in all BC subtypes. NOX2, NOX4, and NOX5 mRNA levels increased with tumor progression stages, while NOX1 and DUOX1 expression decreased in more advanced stages. Moreover, patients with low expression of NOX1, NOX4, and DUOX1 had lower survival rates than those with high expression of these enzymes. In conclusion, our data suggest an overexpression of NOX enzymes in breast cancer, with certain isoforms showing a positive correlation with tumor progression.

Asunto(s)

Neoplasias de la Mama , NADPH Oxidasas , Humanos , Femenino , NADPH Oxidasas/genética , NADPH Oxidasas/metabolismo , Oxidasas Duales/genética , Neoplasias de la Mama/genética , Pronóstico , Especies Reactivas de Oxígeno/metabolismo , ARN Mensajero/genética , Expresión Génica , NADPH Oxidasa 4/genética , NADPH Oxidasa 1/genética

RESUMEN

Quiescin/sulfhydryl oxidase (QSOX1) is a secreted flavoprotein that modulates cellular proliferation, migration and adhesion, roles attributed to its ability to organize the extracellular matrix. We previously showed that exogenously added QSOX1b induces smooth muscle cells migration in a process that depends on its enzymatic activity and that is mediated by hydrogen peroxide derived from Nox1, a catalytic subunit of NAD(P)H oxidases. Here, we report that exogenous QSOX1b also stimulates the migration of L929 fibroblasts and that this effect is regulated by its endocytosis. The use of endocytosis inhibitors and caveolin 1-knockdown demonstrated that this endocytic pathway is caveola-mediated. QSOX1b colocalized with Nox1 in intracellular vesicles, as detected by confocal fluorescence, suggesting that extracellular QSOX1b is endocytosed with the transmembrane Nox1. These results reveal that endosomal QSOX1b is a novel intracellular redox regulator of cell migration.

Asunto(s)

Caveolas , NADPH Oxidasas , Fibroblastos , Endocitosis , Proliferación Celular

RESUMEN

Reactive oxygen species (ROS) function as signaling molecules in several physiologic and pathologic processes. In central nervous system, ROS are critical for differentiation, migration, polarization, and neurite growth. These actions are mediated by reversible oxidation of target proteins. On the other hand, PI3K/Akt signaling pathway is susceptible to be modulated by ROS and it has been implicated in neurite growth. In this study, we evaluated the participation of ROS in the neurite growth of cultured rat cerebellar granule neurons (CGN), as well as the possible regulation of the PI3K/Akt pathway by ROS during neurite outgrowth. For this purpose, CGN were treated with cellular or mitochondrial antioxidants, or an NOX inhibitor and neurite growth was evaluated. Moreover, to assess the participation Akt in this process, the p-Akt levels were measured in CGN treated with antioxidants or a NOX inhibitor. The effect of antioxidants on the neurite growth in the presence of a PI3K inhibitor was also measured. We found that cellular antioxidants and the NOX inhibitor decreased the neurite growth, but not the mitochondrial antioxidant. Interestingly, the antioxidants increased the p-Akt levels; however, the effect of antioxidants on neurite growth was no dependent on the Akt activity since the inhibitor of PI3K did not modify the antioxidant action on neurite growth. Our results show that the PI3K/Akt pathway participates in neurite growth and that ROS produced by NOX could function as signals in this process; however, this action is not mediated by a redox regulation of Akt activity.

Asunto(s)

Antioxidantes , Proteínas Proto-Oncogénicas c-akt , Ratas , Animales , Antioxidantes/farmacología , Proteínas Proto-Oncogénicas c-akt/metabolismo , Neuritas , NADPH Oxidasas/metabolismo , NADPH Oxidasas/farmacología , Especies Reactivas de Oxígeno/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo

RESUMEN

Complex regional pain syndrome type I (CRPS-I) is a disabling pain condition without adequate treatment. Chronic post-ischemia pain injury (CPIP) is a model of CRPS-I that causes allodynia, spontaneous pain, inflammation, vascular injury, and oxidative stress formation. Antioxidants, such as alpha lipoic acid (ALA), have shown a therapeutic potential for CRPS-I pain control. Thus, we aim to evaluate if ALA repeated treatment modulates neuroinflammation in a model of CRPS-I in mice. We used male C57BL/6 mice to induce the CPIP model (O-ring torniquet for 2 h in the hindlimb). For the treatment with ALA or vehicle (Veh) mice were randomly separated in four groups and received 100 mg/kg orally once daily for 15 days (CPIP-ALA, CPIP-Veh, Control-ALA, and Control-Veh). We evaluated different behavioral tests including von Frey (mechanical stimulus), acetone (cold thermal stimulus), rotarod, open field, hind paw edema determination, and nest-building (spontaneous pain behavior). Also, hydrogen peroxide (H2O2) levels, NADPH oxidase and superoxide dismutase (SOD) activity in the sciatic nerve and spinal cord, and Iba1, Nrf2, and Gfap in spinal cord were evaluated at 16 days after CPIP or sham induction. Repeated ALA treatment reduced CPIP-induced mechanical and cold allodynia and restored nest-building capacity without causing locomotor or body weight alteration. ALA treatment reduced SOD and NADPH oxidase activity, and H2O2 production in the spinal cord and sciatic nerve. CPIP-induced neuroinflammation in the spinal cord was associated with astrocyte activation and elevated Nfr2, which were reduced by ALA. ALA repeated treatment prevents nociception by reducing oxidative stress and neuroinflammation in a model of CRPS-I in mice.

Asunto(s)

Dolor Crónico , Distrofia Simpática Refleja , Ácido Tióctico , Ratones , Masculino , Animales , Hiperalgesia , Ácido Tióctico/farmacología , Enfermedades Neuroinflamatorias , Nocicepción , Peróxido de Hidrógeno , Ratones Endogámicos C57BL , Distrofia Simpática Refleja/tratamiento farmacológico , Distrofia Simpática Refleja/complicaciones , Estrés Oxidativo , Isquemia , NADPH Oxidasas/uso terapéutico , Superóxido Dismutasa , Modelos Animales de Enfermedad

RESUMEN

BACKGROUND: the antihypertensive drug α-methyldopa (MD) stands as one of the extensively used medications for managing hypertension during pregnancy. Zinc deprivation has been associated with many diseases. In this context, the synthesis of a Zn coordination complex [Zn(MD)(OH)(H2O)2]·H2O (ZnMD) provide a promising alternative pathway to improve the biological properties of MD. METHODS: ZnMD was synthesized and physicochemically characterized. Fluorescence spectral studies were conducted to examine the binding of both, the ligand and the metal with bovine serum albumin (BSA). MD, ZnMD, and ZnCl2 were administered to spontaneous hypertensive rats (SHR) rats during 8 weeks and blood pressure and echocardiographic parameters were determined. Ex vivo assays were conducted to evaluate levels of reactive oxygen species (ROS), thiobarbituric acid reactive substances (TBARS), and nitric oxide (NO). Cross-sectional area (CSA) and collagen levels of left ventricular cardiomyocytes were also assessed. Furthermore, the expression of NAD(P)H oxidase subunits (gp91phox and p47phox) and Superoxide Dismutase 1 (SOD1) was quantified through western blot analysis. RESULTS: The complex exhibited a moderate affinity for binding with BSA showing a spontaneous interaction (indicated by negative ΔG values) and moderate affinity (determined by affinity constant values). The binding process involved the formation of Van der Waals forces and hydrogen bonds. Upon treatment with MD and ZnMD, a reduction in the systolic blood pressure in SHR was observed, being ZnMD more effective than MD (122 ± 8.1 mmHg and 145 ± 5.6 mmHg, at 8th week of treatment, respectively). The ZnMD treatment prevented myocardial hypertrophy, improved the heart function and reduced the cardiac fibrosis, as evidenced by parameters such as left ventricular mass, fractional shortening, and histological studies. In contrast, MD did not show noticeable differences in these parameters. ZnMD regulates negatively the oxidative damage by reducing levels of ROS and lipid peroxidation, as well as the cardiac NAD(P)H oxidase, and increasing SOD1 expression, while MD did not show significant effect. Moreover, cardiac nitric oxide levels were greater in the ZnMD therapy compared to MD treatment. CONCLUSION: Both MD and ZnMD have the potential to be transported by albumin. Our findings provide important evidence suggesting that this complex could be a potential therapeutic drug for the treatment of hypertension and cardiac hypertrophy and dysfunction.

Asunto(s)

Antihipertensivos , Hipertensión , Ratas , Animales , Antihipertensivos/uso terapéutico , Metildopa/farmacología , Metildopa/uso terapéutico , Especies Reactivas de Oxígeno/metabolismo , Superóxido Dismutasa-1 , Óxido Nítrico/metabolismo , Hipertensión/tratamiento farmacológico , Presión Sanguínea , Ratas Endogámicas SHR , Miocitos Cardíacos/metabolismo , Cardiomegalia , NADPH Oxidasas , Zinc/farmacología , Zinc/uso terapéutico

RESUMEN

PURPOSE: Cerebral ischemia-reperfusion (I/R) is a neurovascular disorder that leads to brain injury. In mice, Fasudil improves nerve injury induced by I/R. However, it is unclear if this is mediated by increased peroxisome proliferator-activated receptor-α (PPARα) expression and reduced oxidative damage. This study aimed to investigate the neuroprotective mechanism of action of Fasudil. METHODS: MCAO (Middle cerebral artery occlusion) was performed in male C57BL/6J wild-type and PPARα KO mice between September 2021 to April 2023. Mice were treated with Fasudil and saline; 2,3,5-Triphenyltetrazolium chloride (TTC) staining was performed to analyze cerebral infarction. PPARα and Rho-associated protein kinase (ROCK) expression were detected using Western blot, and the expression of NADPH subunit Nox2 mRNA was detected using real-time polymerase chain reaction. The NADPH oxidase activity level and reactive oxygen species (ROS) content were also investigated. RESULTS: After cerebral ischemia, the volume of cerebral necrosis was reduced in wild-type mice treated with Fasudil. The expression of PPARα was increased, while ROCK was decreased. Nox2 mRNA expression, NADPH oxidase activity, and ROS content decreased. There were no significant changes in cerebral necrosis volumes, NADPH oxidase activity, and ROS content in the PPARα KO mice treated with Fasudil. CONCLUSIONS: In mice, the neuroprotective effect of Fasudil depends on the expression of PPARα induced by ROCK-PPARα-NOX axis-mediated reduction in ROS and associated oxidative damage.

Asunto(s)

Isquemia Encefálica , Daño por Reperfusión , Ratones , Masculino , Animales , PPAR alfa/fisiología , Especies Reactivas de Oxígeno/metabolismo , Neuroprotección , Daño por Reperfusión/tratamiento farmacológico , Daño por Reperfusión/prevención & control , Daño por Reperfusión/genética , Ratones Endogámicos C57BL , Isquemia , Isquemia Encefálica/tratamiento farmacológico , Isquemia Encefálica/prevención & control , NADPH Oxidasas/genética , NADPH Oxidasas/metabolismo , Reperfusión , Necrosis , ARN Mensajero

Asunto(s)

Enfermedad Granulomatosa Crónica , Humanos , Enfermedad Granulomatosa Crónica/diagnóstico , Enfermedad Granulomatosa Crónica/genética , Penetrancia , NADPH Oxidasas/genética , Neutrófilos , Exones , Mutación/genética

RESUMEN

PURPOSE: This study aimed to evaluate the role of NADPH in pancreatic ductal adenocarcinoma using bioinformatic analyses and experimental validations. METHODS: We compared the expression levels, performed GO and KEGG analysis of NADPH oxidase family and its regulatory subunits, and determined the survival of patients with pancreatic ductal adenocarcinoma by GEPIA, David and KM plotter. The relationship between their expression with immune infiltration levels, phagocytotic/NK cell immune checkpoints, recruitment-related molecules were detected by Timer 2.0 and TISIDB, respectively. Subsequently, their correlation with NK cell infiltration level was verified by immunohistochemistry. RESULTS: The expression of some members of the NADPH oxidase family and its regulatory subunits was significantly increased in pancreatic ductal adenocarcinoma tissues compared to that in normal tissues and was positively correlated with natural killer (NK) cell infiltration. Furthermore, the NADPH oxidase family and its regulatory subunits were associated with survival and immune status in patients with pancreatic ductal adenocarcinoma, including chemokines, immune checkpoints, and immune infiltration levels of NK cells, monocytes, and myeloid-derived suppressor cells. CONCLUSIONS: These results suggest the NADPH oxidase family and its regulatory subunits might serve as indicators for predicting the responsiveness to immunotherapy and outcome of patients with pancreatic ductal adenocarcinoma, providing a new perspective or strategy for immunotherapy in pancreatic ductal adenocarcinoma.

Asunto(s)

Adenocarcinoma , Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Humanos , Pronóstico , NADPH Oxidasas/uso terapéutico , Adenocarcinoma/patología , Carcinoma Ductal Pancreático/tratamiento farmacológico , Neoplasias Pancreáticas/patología , Inmunoterapia , Biomarcadores

RESUMEN

Oxidative stress participates at the baseline of different non-communicable pathologies such as cardiovascular diseases. Excessive formation of reactive oxygen species (ROS), above the signaling levels necessary for the correct function of organelles and cells, may contribute to the non-desired effects of oxidative stress. Platelets play a relevant role in arterial thrombosis, by aggregation triggered by different agonists, where excessive ROS formation induces mitochondrial dysfunction and stimulate platelet activation and aggregation. Platelet is both a source and a target of ROS, thus we aim to analyze both the platelet enzymes responsible for ROS generation and their involvement in intracellular signal transduction pathways. Among the proteins involved in these processes are Protein Disulphide Isomerase (PDI) and NADPH oxidase (NOX) isoforms. By using bioinformatic tools and information from available databases, a complete bioinformatic analysis of the role and interactions of PDI and NOX in platelets, as well as the signal transduction pathways involved in their effects was performed. We focused the study on analyzing whether these proteins collaborate to control platelet function. The data presented in the current manuscript support the role that PDI and NOX play on activation pathways necessary for platelet activation and aggregation, as well as on the platelet signaling imbalance produced by ROS production. Our data could be used to design specific enzyme inhibitors or a dual inhibition for these enzymes with an antiplatelet effect to design promising treatments for diseases involving platelet dysfunction.

Asunto(s)

NADPH Oxidasas , Proteína Disulfuro Isomerasas , Humanos , NADPH Oxidasas/metabolismo , Proteína Disulfuro Isomerasas/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Amigos , Transducción de Señal , Oxidación-Reducción

RESUMEN

INTRODUCTION: Chronic granulomatous disease (CGD) is an inborn error of immunity, characterized by abnormal susceptibility to bacterial and fungal infections and a lack of systemic inflammatory regulation. Pathogenic variants in the CYBB gene are transmitted in an X-linked pattern of inheritance; while the pathogenic variants present in the EROS, NCF1, NCF2, NCF4, or CYBA genes are transmitted with an autosomal recessive inheritance pattern. OBJETIVES: To describe the clinical, immunological, and genetic characteristics of two patients with CGD and BCG infection. METHODS: In peripheral blood neutrophils, H2O2 production and the expression of NADPH oxidase subunits were measured. Detection of pathogenic variants was by Sanger sequencing of the NCF2 gene. The clinical information was extracted from the records by the treating physicians. RESULTS: We present two male infants from two unrelated families of Mayan ethnicity, with CGD and BCG vaccine infection. Three different pathogenic variants in the NCF2 gene were identified; on the one hand, c.304 C>T (p.Arg102*) has already been reported, on the other hand, c.1369 A>T (p.Lys457*) and c.979 G>T (p.Gly327*) not reported. CONCLUSIONS: In patients with mycobacterial infection with BCG, we should suspect an inborn error of immunity, such as CGD. The diagnosis of CGD is made through the detection of a lack of radical oxygen species in neutrophils. The reported patients had pathogenic variants in the NCF2 gene, two of which have not been previously reported in the literature.

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INTRODUCCIÓN: La enfermedad granulomatosa crónica (EGC) es un error innato de la inmunidad, se caracteriza por una susceptibilidad a padecer infecciones bacterianas y fúngicas y a una falta de regulación inflamatoria sistémica. Las variantes patogénicas en el gen CYBB se trasmiten con un patrón de herencia ligada al X; mientras que las variantes patogénicas presentes en los genes EROS, NCF1, NCF2, NCF4 o CYBA se trasmiten con un patrón de herencia autosómico recesivo. OBJETIVOS: Describir las características clínicas, inmunológicas y genéticas de dos pacientes con EGC e infección por BCG. MÉTODOS: En neutrófilos de sangre periférica se midió la producción de H2O2 y la expresión de las subunidades de la NADPH oxidasa. La detección de las variantes patogénicas fue por secuenciación Sanger del gen NCF2. La información clínica fue extraída de los expedientes por los médicos tratantes. RESULTADOS: Presentamos a dos lactantes masculinos de dos familias no relacionadas de la etnia maya, con EGC e infección por la vacuna de BCG. Se identificaron tres diferentes variantes patogénicas en el gen NCF2; por un lado, c.304 C>T (p.Arg102*) ya reportada, por otro lado, c.1369 A>T (p.Lys457*) y c.979 G>T (p.Gly327*) no reportadas. CONCLUSIONES: En pacientes con infección micobacteriana por BCG debemos sospechar en un error innato de la inmunidad, como la EGC. El diagnóstico de EGC se realiza a través de la detección de una falta de producción de radicales libres en los neutrófilos. Los pacientes reportados tuvieron variantes patogénicas en el gen NCF2, dos de ellas no han sido reportadas previamente en la literatura.

Asunto(s)

Enfermedad Granulomatosa Crónica , Mycobacterium bovis , Humanos , Lactante , Masculino , Vacuna BCG/efectos adversos , Etnicidad , Enfermedad Granulomatosa Crónica/genética , Enfermedad Granulomatosa Crónica/diagnóstico , Peróxido de Hidrógeno , Mutación , NADPH Oxidasas/genética , Indígenas Centroamericanos

RESUMEN

The modulation of plant growth and development through reactive oxygen species (ROS) is a hallmark during the interactions with microorganisms, but how fungi and their molecules influence endogenous ROS production in the root remains unknown. In this report, we correlated the biostimulant effect of Trichoderma atroviride with Arabidopsis root development via ROS signaling. T. atroviride enhanced ROS accumulation in primary root tips, lateral root primordia, and emerged lateral roots as revealed by total ROS imaging through the fluorescent probe H2DCF-DA and NBT detection. Acidification of the substrate and emission of the volatile organic compound 6-pentyl-2H-pyran-2-one appear to be major factors by which the fungus triggers ROS accumulation. Besides, the disruption of plant NADPH oxidases, also known as respiratory burst oxidase homologs (RBOHs) including ROBHA, RBOHD, but mainly RBOHE, impaired root and shoot fresh weight and the root branching enhanced by the fungus in vitro. RbohE mutant plants displayed poor lateral root proliferation and lower superoxide levels than wild-type seedlings in both primary and lateral roots, indicating a role for this enzyme for T. atroviride-induced root branching. These data shed light on the roles of ROS as messengers for plant growth and root architectural changes during the plant-Trichoderma interaction.

Asunto(s)

Proteínas de Arabidopsis , Arabidopsis , Trichoderma , Trichoderma/genética , Especies Reactivas de Oxígeno/metabolismo , Proteínas de Arabidopsis/metabolismo , NADPH Oxidasas/genética , NADPH Oxidasas/metabolismo , Raíces de Plantas , Regulación de la Expresión Génica de las Plantas

RESUMEN

Capacitation is a series of physiological, biochemical, and metabolic changes experienced by mammalian spermatozoa. These changes enable them to fertilize eggs. The capacitation prepares the spermatozoa to undergo the acrosomal reaction and hyperactivated motility. Several mechanisms that regulate capacitation are known, although they have not been fully disclosed; among them, reactive oxygen species (ROS) play an essential role in the normal development of capacitation. NADPH oxidases (NOXs) are a family of enzymes responsible for ROS production. Although their presence in mammalian sperm is known, little is known about their participation in sperm physiology. This work aimed to identify the NOXs related to the production of ROS in guinea pig and mouse spermatozoa and define their participation in capacitation, acrosomal reaction, and motility. Additionally, a mechanism for NOXs' activation during capacitation was established. The results show that guinea pig and mouse spermatozoa express NOX2 and NOX4, which initiate ROS production during capacitation. NOXs inhibition by VAS2870 led to an early increase in the capacitation and intracellular concentration of Ca2+ in such a way that the spermatozoa also presented an early acrosome reaction. In addition, the inhibition of NOX2 and NOX4 reduced progressive motility and hyperactive motility. NOX2 and NOX4 were found to interact with each other prior to capacitation. This interaction was interrupted during capacitation and correlated with the increase in ROS. Interestingly, the association between NOX2-NOX4 and their activation depends on calpain activation, since the inhibition of this Ca2+-dependent protease prevents NOX2-NOX4 from dissociating and ROS production. The results indicate that NOX2 and NOX4 could be the most important ROS producers during guinea pig and mouse sperm capacitation and that their activation depends on calpain.

Asunto(s)

Calpaína , Especies Reactivas de Oxígeno , Semen , Capacitación Espermática , Animales , Cobayas , Masculino , Ratones , Calpaína/metabolismo , NADPH Oxidasa 4/metabolismo , NADPH Oxidasas/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Semen/metabolismo , Espermatozoides/metabolismo

RESUMEN

Rhinella marina toad is abundant in Brazil. Its poison contains cardiac glycosides called bufadienolides, which are extensively investigated for their bioactivity. Our aim was to characterize the vasoactivity of Rhinella marina poison (RmP) on the aorta of male Wistar rats. For this, the RmP was first collected and processed to obtain an alcoholic extract. To determine cardiovascular effects of RmP, we performed in vivo tests by administering RmP intravenously in doses of 0.1-0.8 mg/kg. Vascular reactivity was also performed through concentration-response curves to RmP (10 ng/mL to 200 µg/mL) in aortic segments with and without endothelium. RmP induced a concentration-dependent contraction in rat aorta which was partly endothelium-mediated. Nitric oxide contributes with this response in view that incubation with L-NAME increased the contractile response. Additionally, treatment with indomethacin [cyclooxygenase, (COX) inhibitor], nifedipine (L-type voltage-gated calcium channels blocker), and BQ-123 (ETA receptors antagonist) decreased maximum response, and ketanserin (5-HT2 receptors antagonist) decreased pEC50, suggesting active participation of these pathways in the contractile response. On the other hand, apocynin (NADPH oxidase inhibitor) did not alter contractility. Incubation with prazosin (α1-adrenergic receptor antagonist) abolished the contractile response, suggesting that the RmP-induced contraction is dependent on the adrenergic pathway. In the Na+/K+ ATPase protocol, a higher Emax was observed in the RmP experimental group, suggesting that RmP potentiated Na+/K+ATPase hyperpolarizing response. When this extract was injected (i.v.) in vivo, increase in blood pressure and decrease in heart rate were observed. The results were immediate and transitory, and occurred in a dose-dependent manner. Overall, these data suggest that the poison extract of R. marina toad has an important vasoconstrictor action and subsequent vasopressor effects, and its use can be investigated to some cardiovascular disorders.

Asunto(s)

Bufanólidos , Venenos , Adenosina Trifosfatasas/metabolismo , Adenosina Trifosfatasas/farmacología , Adrenérgicos/farmacología , Antagonistas Adrenérgicos/farmacología , Animales , Bufanólidos/toxicidad , Bufo marinus/metabolismo , Canales de Calcio , Endotelio Vascular , Hemodinámica , Indometacina/farmacología , Ketanserina/farmacología , Masculino , Metanol/farmacología , NADPH Oxidasas , NG-Nitroarginina Metil Éster , Nifedipino/farmacología , Óxido Nítrico/metabolismo , Prazosina/farmacología , Prostaglandina-Endoperóxido Sintasas/metabolismo , Ratas , Ratas Wistar , Serotonina/farmacología , Vasoconstrictores

RESUMEN

l-Amino acid oxidase isolated from Calloselasma rhodostoma (Cr-LAAO) snake venom is a potent stimulus for neutrophil activation and production of inflammatory mediators, contributing to local inflammatory effects in victims of envenoming. Cr-LAAO triggered the activation of nicotinamide adenine dinucleotide phosphatase (NADPH) oxidase complex and protein kinase C (PKC)-α signaling protein for reactive oxygen species (ROS) production. This study aims to evaluate the ROS participation in the NLRP3 inflammasome complex activation in human neutrophil. Human neutrophils were isolated and stimulated for 1 or 2 h with RPMI (negative control), LPS (1 µg/mL, positive control) or Cr-LAAO (50 µg/mL). The neutrophil transcriptome was examined using the microarray technique, and RT-qPCR for confirmation of gene expression. Immunofluorescence assays for NLRP3, caspase-1, IL-1ß and GSDMD proteins was performed by Western blot in the presence and/or absence of Apocynin, an inhibitor of NADPH oxidase. IL-1ß release was also detected in the presence and/or absence of NLRP3, caspase-1 and NADPH oxidase inhibitors. Results showed that Cr-LAAO upregulated the expression of genes that participate in the NADPH oxidase complex formation and inflammasome assembly. NLRP3 was activated and accumulated in the cytosol forming punctas, indicating its activation. Gasdermin D was not cleaved but lactate dehydrogenase was released. Furthermore, ROS inhibition decreased the expression of NLRP3 inflammasome complex proteins, as observed by protein expression in the presence and/or absence of apocynin, an NADPH oxidase inhibitor. IL-1ß was also released, and pharmacological inhibition of NLRP3, caspase-1, and ROS reduced the amount of released cytokine. This is the first report demonstrating the activation of the NLRP3 inflammasome complex via ROS generation by Cr-LAAO, which may lead to the development of local inflammatory effects observed in snakebite victims.

Asunto(s)

Inflamasomas , L-Aminoácido Oxidasa , Acetofenonas , Caspasa 1/metabolismo , Citocinas/metabolismo , Humanos , Inflamasomas/metabolismo , Mediadores de Inflamación/metabolismo , Interleucina-1beta/metabolismo , L-Aminoácido Oxidasa/metabolismo , L-Aminoácido Oxidasa/farmacología , Lactato Deshidrogenasas/metabolismo , Lipopolisacáridos/farmacología , NAD/metabolismo , NADP/metabolismo , NADPH Oxidasas/metabolismo , Proteína con Dominio Pirina 3 de la Familia NLR/genética , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Neutrófilos/metabolismo , Monoéster Fosfórico Hidrolasas/metabolismo , Proteína Quinasa C/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Venenos de Serpiente/metabolismo , Venenos de Serpiente/farmacología

RESUMEN

Iron [Fe(II)] and copper [Cu(II)] overloads in rat brain are associated with oxidative stress and damage. The purpose of this research is to study whether brain antioxidant enzymes are involved in the control of intracellular redox homeostasis in the brain of rats male Sprague-Dawley rats (80-90 g) that received drinking water supplemented with either 1.0 g/L of ferrous chloride (n = 24) or 0.5 g/L cupric sulfate (n = 24) for 42 days. Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, superoxide dismutase (SOD), catalase, glutathione peroxidase (GPx) and glutathione transferase (GT) activities in brain were determined by spectrophotometric methods and NO production by the content of nitrite concentration in the organ. Chronic treatment with Fe(II) and Cu(II) led to a significant decrease of nitrite content and SOD activity in brain. Activity of NADPH oxidase increased with Cu(II) treatment. Concerning Fe(II), catalase and GT activities increased in brain after 28 and 4 days of treatment, respectively. In the case of Cu(II), catalase activity decreased whereas GT activity increased after 2 and 14 days, respectively. The regulation of redox homeostasis in brain involves changes of the activity of these enzymes to control the steady state of oxidant species related to redox signaling pathways upon Cu and Fe overload. NO may serve to detoxify cells from superoxide anion and hydrogen peroxide with the concomitant formation of peroxynitrite. However, the latest is a powerful oxidant which leads to oxidative modifications of biomolecules. These results suggest a common pathway to oxidative stress and damage in brain for Cu(II) and Fe(II).

Asunto(s)

Antioxidantes , Agua Potable , Animales , Antioxidantes/química , Encéfalo/metabolismo , Catalasa/metabolismo , Cobre/metabolismo , Sulfato de Cobre , Compuestos Ferrosos/metabolismo , Glutatión Peroxidasa/metabolismo , Glutatión Transferasa/metabolismo , Peróxido de Hidrógeno/metabolismo , Hierro/metabolismo , Masculino , NADP/metabolismo , NADPH Oxidasas/metabolismo , Nitritos , Oxidantes/metabolismo , Ácido Peroxinitroso/metabolismo , Ratas , Ratas Sprague-Dawley , Superóxido Dismutasa , Superóxidos/metabolismo

RESUMEN

Parkinson's Disease (PD) is a neurogenerative disorder characterized by the death of dopaminergic neurons in the Substantia Nigra pars compacta (SNpc), leading to motor, cognitive, learning, and respiratory dysfunctions. New evidence revealed that breathing impairment in PD mainly results from oxidative stress (OS) that initiates apoptotic signaling in respiratory neurons. Here, we investigated the role of OS inhibition using apocynin (non-specific NADPH oxidase inhibitor) in a 6-OHDA PD animal model in the neural control of breathing. The PD model was confirmed with a 70% reduction in TH-expressing neurons within the SNpc. After 20 and 40 days of PD induction, no differences were observed in superoxide anion levels in any respiratory nuclei. At 30 days after PD induction, 6-OHDA animals presented OS that was prevented in all respiratory nuclei by adding apocynin to the drinking water for 10 days. Forty days after PD animal model induction, impaired motor and breathing function, reduced Phox2b and NK1 receptors-expressing neurons in the medullary respiratory areas; decreased latency to fall in the rotarod motor test; and attenuated respiratory frequency and minute ventilation parameters at rest and under hypercapnia conditions were observed. After 20 days of apocynin treatment, neurodegeneration of respiratory nuclei and breathing dysfunction in 6-OHDA animals were prevented. Thus, OS contributes to respiratory neuron death, consequently leading to breathing dysfunction in the 6-OHDA PD animal model. Furthermore, these results present a new perspective for preventing the onset and progression of PD-related respiratory impairments.

Asunto(s)

Agua Potable , Enfermedad de Parkinson , Animales , Oxidopamina/toxicidad , Superóxidos , Neuronas Dopaminérgicas , Modelos Animales de Enfermedad , NADPH Oxidasas , Estrés Oxidativo , Sustancia Negra