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@#Infectious diseases with complications of sepsis are still public health concern in both developed and developing countries. Sepsis is a potentially life-threatening systemic immune response to infection that can lead to end- stage organ failure and death. Sepsis involves multiple mechanisms such as neuroendocrine, complement activation, blood coagulation, and fibrinolytic system. Reactive oxygen species (ROS) is an inflammatory mediator produced by NADPH oxidase activation. This study aimed to investigate the effects of ethanol extract of E. elatior fruits on NADPH oxidase activity. Forty Mus musculus mice were randomized divided into five groups (n=8), with the intervention group receiving an intraperitoneal injection of 0.3 mg/kg BW lipopolysaccharide (LPS). There was a normal group without LPS injection (N-1), LPS injection only (N-2), and those that received LPS injection and ethanol extracts of E. elatior fruits containing 2.1 mg/20 g (N-3), 4.2 mg/20 g (N-4), and 8.4 mg/20 g (N-5). NADPH oxidase activity were measured using ELISA. The oneway ANOVA was used to investigate the differences between the groups. After administration of the extract at a varied dose, N-5 group the lowest NADPH oxidase activity (p=0.001). The ethanol extract of E. elatior fruit has antioxidant effects. In this study, a dose of 8.4 mg/20 g of extract significantly reduced NADPH oxidase activity. The ethanol extract of E. elatior might be considered a treatment in sepsis.
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OBJECTIVE@#To identify phytochemical constituents present in the extract of flowers of Xanthoceras sorbifolia and evaluate their anti-oxidant and anti-hyperglycemic capacities.@*METHODS@#The AlCl3 colorimetric method and Prussian Blue assay were used to determine the contents of total flavonoids and total phenolic acids in extraction layers, and the bioactive layers was screened through anti - oxidative activity in vitro. The Waters ACQUITY UPLC system and a Waters ACQUITY UPLC BEH C18 column (2.0 mm × 150 mm, 5 μm) were used to identify the ingredients. And anti-oxidative ingredients were screened by off-line UPLC-QTOF-MS/MS-free radical scavenging. The ameliorative role of it was further evaluated in a high-fat, streptozotocin-induced type 2 diabetic rat model and the study was carried out on NADPH oxidase (PDB ID: 2CDU) by molecular docking.@*RESULTS@#Combined with the results of activity screening in vitro, the anti - oxidative part was identified as the ethyl acetate layer. A total of 24 chemical constituents were identified by liquid chromatography-mass spectrometry in the ethyl acetate layer and 13 main anti-oxidative active constituents were preliminarily screened out through off-line UPLC-QTOF-MS/MS-free radical scavenging. In vivo experiments showed that flowers of X. sorbifolia could significantly reduce the blood glucose level of diabetic mice and alleviate liver cell damage. Based on the results of docking analysis related to the identified phytocompounds and oxidase which involved in type 2 diabetes, quercetin 3-O-rutinoside, kaempferol-3-O-rhamnoside, isorhamnetin-3-O-glucoside, and isoquercitrin showed a better inhibitory profile.@*CONCLUSION@#The ethyl acetate layer was rich in flavonoids and phenolic acids and had significant anti-oxidant activity, which could prevent hyperglycemia. This observed activity profile suggested X. sorbifolia flowers as a promising new source of tea to develop alternative natural anti-diabetic products with a high safety margin.
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ObjectiveTo explore the underlying mechanism of modified Zhenwutang in delaying renal interstitial fibrosis in chronic renal failure (CRF) by observing the effects of modified Zhenwutang on the expression of angiotensin Ⅱ (Ang Ⅱ), angiotensin Ⅱ type 1 receptor (AT1R), nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 4 (NOX4), transforming growth factor-β1 (TGF-β1), type I collagen (COL1A1), and type Ⅲ collagen (COL3A1) in the serum and renal tissues of adenine-induced CRF rats. MethodFifty male SPF-grade SD rats were randomly divided into a normal group (n=10) and an experimental group (n=40) using a random number table. After one week of adaptive feeding, the experimental CRF model was established in rats by administering adenine at 150 mg·kg-1·d-1 orally. Three rats from each group were randomly selected to evaluate the model induction. After successful modeling, rats in the experimental group were randomly divided into a model group, low-, medium, and high-dose modified Zhenwutang groups, and a benazepril hydrochloride group, with six rats in each group. The rats were orally administered the corresponding drugs once daily for four weeks. At the end of the first week, 13th week, and 17th week of the experiment, 24 hour urinary protein quantification (24 h-UTP) was measured. At the end of the 17th week, the rats were euthanized, and blood samples were collected from the abdominal aorta for the measurement of total protein (TP), albumin (ALB), creatinine (Cr), and blood urea nitrogen (BUN) in the serum. Enzyme-linked immunosorbent assay (ELISA) was used to measure the expression levels of serum Ang Ⅱ. Hematoxylin-eosin (HE) staining and Masson's trichrome staining were performed to observe the pathological changes in renal tissues. Immunohistochemistry (IHC) was performed to observe the expression of AT1R, NOX4, TGF-β1, COL1A1, and COL3A1. Real-time fluorescence-based quantitative polymerase chain reaction (Real-time PCR) was used to observe the mRNA expression levels of AT1R, NOX4, and TGF-β1. Western blot was conducted to measure the protein expression levels of AT1R, NOX4, and TGF-β1. Result① Compared with the normal group, the model group showed a significant increase in 24 h-UTP (P<0.01). The levels of Cr and BUN in the model group were significantly higher (P<0.01), while the levels of TP and ALB were significantly lower (P<0.01). The serum Ang Ⅱ level in the model group was significantly elevated (P<0.01). The model group exhibited widening of the renal glomerular mesangial space, necrotic glomeruli, increased interstitial width with extensive inflammatory cell infiltration, brownish precipitates blocking the renal tubular lumens, irregular renal tubules, and significant deposition of collagen fibers in the renal interstitium. Additionally, the collagen fibers around the renal vessels, outside the parietal layer of the renal sacs, glomerular basement membrane, and tubular basement membrane increased significantly. The expression of AT1R and NOX4 in the glomeruli and renal tubules of the model group was significantly enhanced, and TGF-β1 expression also significantly increased in the renal tubules. The expression of COL1A1 and COL3A1 in the renal interstitium significantly increased. The mRNA expression of AT1R and TGF-β1 in the model group significantly increased (P<0.01), while NOX4 mRNA expression significantly decreased (P<0.01). The protein expression of AT1R, NOX4, and TGF-β1 was significantly enhanced (P<0.01). ② Compared with the model group, modified Zhenwutang significantly reduced 24h-UTP (P<0.01), decreased levels of Cr and BUN (P<0.01), increased levels of TP and ALB (P<0.01), reduced serum Ang Ⅱ level (P<0.01), alleviated renal pathological damage, reduced expression of AT1R, NOX4, TGF-β1, COL1A1, and COL3A1 in the glomeruli, renal tubules, and renal interstitium, reduced mRNA expression of AT1R and TGF-β1 (P<0.01), increased NOX4 mRNA expression (P<0.01), and weakened protein expression of AT1R, NOX4, and TGF-β1 (P<0.01). The modified Zhenwutang groups showed a significant dose-effect trend. ConclusionModified Zhenwutang may delay renal interstitial fibrosis in CRF rats by reducing the expression of Ang Ⅱ, AT1R, NOX4, and TGF-β1 in the serum and renal tissues, thereby alleviating renal pathological damage, reducing proteinuria, protecting renal function, and delaying the progression of CRF. The modified Zhenwutang group exhibited a dose-effect trend.
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Reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 2 (Nox2) acts as a source of reactive oxygen species, which participates in and influences normal physiological function of human body. Nowadays, many studies have found that Nox2 is related to prognosis of patients, drug resistance and molecular targeted therapy in various malignant tumors, such as acute myeloid leukemia, gastric cancer, colorectal cancer, ovarian cancer, lung cancer and esophageal cancer. What's more, it may be a novel biomarker and a potential therapeutic target for malignant tumors.
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Objective:To observe the influence of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) 4 inhibitors on epithelial-mesenchymal transition (EMT) of human retinal pigment epithelial (RPE) cells induced by bevacizumab.Methods:The cultured ARPE-19 cells were divided into blank control group, bevacizumab group, bevacizumab+ VAS2870 group and bevacizumab+ GKT137831 group.Cells were cultured with 0.25 g/L bevacizumab, 0.25 g/L bevacizumab plus 3 μmol/L VAS2870 (a NOX4 inhibitor), 0.25 g/L bevaczumab plus 20 μmol/L GKT137831 (a NOX4 inhibitor) for 72 hours according to grouping.No intervention was administered to the blank control group.The mRNA and protein expression levels of NOX4 and EMT markers including fibronectin (FN), vimentin, α-smooth muscle actin (α-SMA) and tight junction related protein zonula occludens-1 (ZO-1) were measured by real-time PCR and Western blot assay, and the expression levels in different intervention groups were compared.The expressions of NOX4 and EMT markers were verified by immunofluorescence staining.Results:There were statistically significant differences in the relative mRNA and protein expression levels of FN, vimentin, α-SMA, ZO-1 and NOX4 among blank control group, bevacizumab group, bevacizumab+ VAS2870 group and bevacizumab+ GKT137831 group (mRNA: F=97.07, 195.40, 722.40, 38.56, 70.81; all at P<0.001.Protein: F=23.09, 64.58, 58.19, 26.97, 63.19; all at P<0.001). The relative mRNA and protein expression levels of FN, vimentin, α-SMA and NOX4 were significantly higher and the relative mRNA and protein expression level of ZO-1 was significantly lower in bevacizumab group than those in blank control group (all at P<0.05). The relative mRNA and protein expression levels of FN, vimentin, α-SMA and NOX4 were significantly lower and the relative mRNA and protein expression levels of ZO-1 were significantly higher in bevacizumab+ VAS2870 and bevacizumab+ GKT137831 groups than those in bevacizumab group (all at P<0.05). The immunofluorescence intensity of FN, vimentin and α-SMA was stronger and the immunofluorescence intensity of ZO-1 was weaker in bevacizumab group than blank control group.The immunofluorescence intensity of FN, vimentin and α-SMA were weaker and the immunofluorescence intensity of ZO-1 was stronger in bevacizumab+ VAS2870 group and bevacizumab+ GKT137831 group than those in bevacizumab group. Conclusions:NOX4 is involved in the bevacizumab-induced EMT of human RPE cells, the degree of which can be reduced by NOX4 inhibitors.
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ABSTRACT Background: In Philadelphia chromosome-negative myeloproliferative neoplasm (MPN) models, reactive oxygen species (ROS) are elevated and have been implicated in genomic instability, JAK2/STAT signaling amplification, and disease progression. Although the potential effects of ROS on the MPN phenotype, the effects of ruxolitinib treatment on ROS regulation have been poorly explored. Herein, we have reported the impact of ruxolitinib on redox signaling transcriptional network, and the effects of diphenyleneiodonium (DPI), a pan NOX inhibitor, in JAK2V617F-driven cellular models. Method: Redox signaling-related genes were investigated in SET2 cells upon ruxolitinib treatment by RNA-seq (GEO accession GSE69827). SET2 and HEL cells, which represent JAK2V617F-positive MPN cellular models with distinct sensitivity to apoptosis induced by ruxolitinib, were used. Cell viability was evaluated by MTT, apoptosis by annexin V/PI and flow cytometry, and cell signaling by quantitative PCR and Western blot. Main results: Ruxolitinib impacted on a network composed of redox signaling-related genes, and DUOX1 and DUOX2 were identified as potential modulators of ruxolitinib response. In SET2 and HEL cells, DPI reduced cell viability and, at low doses, it significantly potentiated ruxolitinib-induced apoptosis. In the molecular scenario, DPI inhibited STAT3, STAT5 and S6 ribosomal protein phosphorylation and induced PARP1 cleavage in JAK2V617F-positive cells. DPI combined with ruxolitinib increased PARP1 cleavage in SET2 cells and potentiated ruxolitinib-reduced STAT3, STAT5 and S6 ribosomal protein in HEL cells. Conclusion: Our study reveals a potential adaptation mechanism for resistance against ruxolitinib by transcriptionally reprogramming redox signaling in JAK2V617F cells and exposes redox vulnerabilities with therapeutic value in MPN cellular models.
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Janus Quinasa 2 , Enfermedades Mielodisplásicas-Mieloproliferativas/tratamiento farmacológico , Oxidación-Reducción , NADPH Oxidasas , Oxidasas Duales , Trastornos MieloproliferativosRESUMEN
Introducción: La enfermedad granulomatosa crónica es una inmunodeficiencia primaria congénita del sistema inmune innato, originada por defectos en el complejo enzimático nicotinamida adenina dinucleótido fosfato oxidasa presente en células fagocíticas. Estos defectos funcionales causan incapacidad para producir especies reactivas del oxígeno en los fagocitos, que afectan la eliminación de algunos microorganismos patógenos dentro del fagolisosoma. El diagnóstico de esta enfermedad se realiza actualmente mediante la prueba de 1,2,3-dihidrorodamina asistida por citometría de flujo multiparamétrica, o la tinción de fagocitos con nitroazul de tetrazolio asistida por microscopio óptico. Objetivos: Describir los aspectos fisiopatológicos y moleculares de la enfermedad granulomatosa crónica; y discutir aspectos relacionados con las pruebas de diagnóstico antes mencionadas. Métodos: Se realizó una investigación bibliográfica-documental a partir de artículos científicos publicados desde 1933 hasta 2018, para ello fueron consultadas las bases de datos SciELO, PubMed y Springer. Desarrollo: Se exponen las características fisiopatológicas de la enfermedad granulomatosa crónica, así como la relación entre las mutaciones genéticas más abundantes en la población afectada y la gravedad de las manifestaciones clínicas que presentan los pacientes. Además, se analizan críticamente los beneficios y las deficiencias de dos técnicas que se utilizan actualmente para diagnosticar la enfermedad. Conclusiones: La enfermedad granulomatosa crónica puede generar consecuencias inmunológicas e inflamatorias graves, que se hallan en consonancia con las características genéticas expresadas en el complejo enzimático dañado. El diagnóstico de la enfermedad resulta más confiable, exhaustivo y específico, mediante la citometría de flujo y su prueba de 1,2,3-dihidrorodamina(AU)
Introduction: Chronic granulomatous disease is a congenital primary immunodeficiency of the innate immune system, caused by defects in the nicotinamide adenine dinucleotide phosphate oxidase enzyme complex present in phagocytic cells. These functional defects cause inability to produce reactive oxygen species in phagocytes, affecting the elimination of some pathogenic microorganisms within the phagolysosome. The diagnosis of this disease is currently made by means of the 1,2,3-dihydrorodamine test assisted by multiparametric flow cytometry, or the staining of phagocytes with nitro-blue tetrazolium assisted by light microscopy. Objectives: To characterize molecular and pathophysiologically the chronic granulomatous disease; and to discuss aspects related to the aforementioned diagnostic tests. Methods: In this work, a bibliographic-documentary research was carried out from scientific articles published from 1933 to 2018, for which the SciELO, PubMed and Springer databases were consulted. Development: The pathophysiological characteristics of chronic granulomatous disease are exposed, as well as the relationship between the most abundant genetic mutations in the affected population, and the severity of the clinical manifestations presented by the patients. In addition, the benefits and deficiencies of two techniques currently used to diagnose the disease are critically analyzed. Conclusions: Chronic granulomatous disease can generate severe immunological and inflammatory consequences, which are in line with the genetic characteristics expressed in the damaged enzyme complex. The diagnosis of the disease is more reliable, exhaustive and specific, using flow cytometry and its 1,2,3-dihydrorodamine test(AU)
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Humanos , Especies Reactivas de Oxígeno , Pruebas Diagnósticas de Rutina , Nitroazul de Tetrazolio/uso terapéutico , Técnicas y Procedimientos Diagnósticos , Citometría de Flujo/métodos , Enfermedad Granulomatosa Crónica/fisiopatología , Enfermedad Granulomatosa Crónica/genéticaRESUMEN
Resumo Fundamento: O exercício físico tem sido considerado uma importante terapia não farmacológica para a prevenção e tratamento das doenças cardiovasculares. No entanto, seus efeitos na remodelação cardíaca leve não são claros. Objetivo: Avaliar a influência do exercício aeróbico sobre a capacidade funcional, estrutura cardíaca, função ventricular esquerda (VE) e expressão gênica das subunidades da NADPH oxidase em ratos com infarto do miocárdio pequeno (IM). Métodos: Três meses após a indução do IM, ratos Wistar foram divididos em três grupos: Sham; IM sedentário (IM-SED); e IM exercício aeróbico (IM-EA). Os ratos se exercitaram em uma esteira três vezes por semana durante 12 semanas. Um ecocardiograma foi realizado antes e após o treinamento. O tamanho do infarto foi avaliado por histologia e a expressão gênica por RT-PCR. O nível de significância para análise estatística foi estabelecido em 5%. Resultados: Ratos com IM menor que 30% da área total do VE foram incluídos no estudo. A capacidade funcional foi maior no IM-EA do que nos ratos Sham e IM-SED. O tamanho do infarto não diferiu entre os grupos. Ratos infartados apresentaram aumento do diâmetro diastólico e sistólico do VE, diâmetro do átrio esquerdo e massa do VE, com disfunção sistólica. A espessura relativa da parede foi menor no grupo IM-SED do que nos grupos IM-EA e Sham. A expressão gênica das subunidades NADPH oxidase NOX2, NOX4, p22phox e p47phox não diferiu entre os grupos. Conclusão: Infarto do miocárdio pequeno altera a estrutura cardíaca e a função sistólica do VE. O exercício aeróbico tardio pode melhorar a capacidade funcional e a remodelação cardíaca por meio da preservação da geometria ventricular esquerda. A expressão gênica das subunidades da NADPH oxidase não está envolvida na remodelação cardíaca, nem é modulada pelo exercício aeróbico em ratos com infarto do miocárdio pequeno.
Abstract Background: Physical exercise has been considered an important non-pharmacological therapy for the prevention and treatment of cardiovascular diseases. However, its effects on minor cardiac remodeling are not clear. Objective: To evaluate the influence of aerobic exercise on the functional capacity, cardiac structure, left ventricular (LV) function, and gene expression of NADPH oxidase subunits in rats with small-sized myocardial infarction (MI). Methods: Three months after MI induction, Wistar rats were divided into three groups: Sham; sedentary MI (MI-SED); and aerobic exercised MI (MI-AE). The rats exercised on a treadmill three times a week for 12 weeks. An echocardiogram was performed before and after training. The infarction size was evaluated by histology, and gene expression was assessed by RT-PCR. The significance level for statistical analysis was set at 5%. Results: Rats with MI lower than 30% of the LV total area were included in the study. Functional capacity was higher in MI-AE than in Sham and MI-SED rats. The infarction size did not differ between groups. Infarcted rats had increased LV diastolic and systolic diameter, left atrial diameter, and LV mass, with systolic dysfunction. Relative wall thickness was lower in MI-SED than in the MI-AE and Sham groups. Gene expression of the NADPH oxidase subunits NOX2, NOX4, p22phox, and p47phox did not differ between groups. Conclusion: Small-sized MI changes cardiac structure and LV systolic function. Late aerobic exercise is able to improve functional capacity and cardiac remodeling by preserving the left ventricular geometry. NADPH oxidase subunits gene expression is not involved in cardiac remodeling or modulated by aerobic exercise in rats with small-sized MI.
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Animales , Ratas , Remodelación Ventricular , Infarto del Miocardio/terapia , Ejercicio Físico , Ratas Wistar , CorazónRESUMEN
OBJECTIVE@#To explore the protective effect and the underlying mechanism of Hu-Lu-Ba-Wan (, HLBW) on the testis of diabetic rats.@*METHODS@#Twenty-four male Wistar rats (160-180 g) were randomly divided into 3 groups according to a random number table, including a control group (n=8), diabetic group (n=8), and HLBW group (n=8). Diabetic rat model was established by high-fat-diet administration and single intravenous injection of streptozotocin (26 mg/kg). Then HLBW granule was administrated for 12 weeks. Fasting blood glucose and insulin levels as well as serum total testosterone level and testicular testosterone content were examined. Oxidative stress markers in both serum and testis were tested. Meanwhile, testicular morphology was observed under hematoxylin and eosin (HE) and the ultrastructure of Leydig cell was observed by electron microscope. The superoxide anion level was detected by DHE, and TUNEL-positive cells of testis was evaluated by TUNEL assay. The gene and protein expression of protein kinase C (PKCα), phosphorylated PKCα (P-PKCα) and P47phox in testicular tissues were determined by quantitative RT-PCR analysis and Western bolt analysis.@*RESULTS@#Compared with the diabetic group, HLBW treatment significantly reduced the fasting glucose levels and increased the levels of fasting insulin and testosterone in serum (P<0.01). HLBW administration also reduced the levels of reactive oxygen species (ROS) in plasma and alleviated the damage of oxidative stress in the testis of diabetic rats. Additionally, HLBW down-regulated the protein and mRNA levels of PKCα, P-PKCα and P47phox in testicular tissues.@*CONCLUSION@#HLBW may attenuate the oxidative stress in the testis of diabetic rats via PKCα /NAPDH oxidase signaling pathway.
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Aim To investigate the effect of ginsenoside Rgl on PC 12 cell hypoxia-reoxygenation injury and its possible mechanism. Methods PC 12 cells were randomly divided into six groups. Except for the blank control group, all the other groups were hypoxia and hypoglycemia for 6 hours, and then reoxygenated and glycosylated for 24 hours to make OGD/R models. Each drug group was given corresponding drugs 2 hours before modeling pretreatment. DCFH-DA method was used to detect the ROS production in cells, Annexin V- FITC/PI double staining method was performed to detect cell apoptosis rate, ELISA method was used to detect LDH activity and IL-1 (3 content in cell supernatant, and Western blot was applied to detect the ex- pression of proteins of N0X2, p22phox, p47phox, NLRPl, ASC, Caspase-1, PSD95, Tau, p-Tau and observe the intervention effect of ginsenoside Rgl. Re sults Tempol, Apocynin and Rgl (5, 10 jjLmol • L"1) groups could significantly inhibit ROS production and apoptosis, reduce LDH release and IL-1 (3 content in cell supernatant; Apocynin and Rgl (5, 10 |xmol • L"1) groups could significantly down-regulate the expression of N0X2, p22phox and p47phox in cells. The Tempol, Apocynin and Rgl (5, 10 jxmol • L"1 ) groups could significantly decrease the protein expression of NLRP1, Caspase-1, ASC, IL-1 (3 and p-Tau, and markedly down-regulate the expression of PSD95 protein. Conclusion Rgl is likely to reduce the is- chemia-reperfusion injury of PC 12 cells by inhibiting the NOX2-NLRP1 pathway.
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Dendritic cells (DCs) are specialized antigen-presenting cells. Immature dendritic cells can be activated into mature dendritic cells by recognizing antigens, then the antigens are processed and pres-ented to T lymphocytes. DCs play a vital role in initiating immune response, regulating immune response and maintaining immune tolerance. Therefore, regulating the immune function of DCs can be used to treat diseases such as autoimmune diseases and tumors. With the deepening of research on the regulation of DCs, people have gradually realized that the existence of reactive oxygen species (ROS) in DCs is of great significance. ROS is a term of strong oxidizing reactive species, and the dynamic balance of its pro-duction and removal is the key to maintaining cell redox homeostasis. ROS of physiological level is an im-portant molecule involved in a variety of signal pathways, which can regulate cell growth, differentiation and different physiological and biochemical reactions. The changes in the level of ROS affect the state and function of cells. In addition, because there are many ways to produce ROS in the cell, the effects of dif-ferent sources of ROS on DCs are not usually the same; and even the same source of ROS may have dif-ferent effects when cells are in different states. This article summarized the influence of intracellular ROS changes and different sources of ROS on the differentiation, maturation and function of DCs, aiming to re-veal how ROS plays an important role in regulating the immune function of DCs. At the same time, this article also showed the urgent need for the in-depth study of ROS regulating the function of DCs, which may help the application of DCs immune regulation to a wider range of disease treatments.
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Neutrophils play a pivotal role in innate immunity and in the inflammatory response. Neutrophils are very motile cells that are rapidly recruited to the inflammatory site as the body first line of defense. Their bactericidal activity is due to the release into the phagocytic vacuole, called phagosome, of several toxic molecules directed against microbes. Neutrophil stimulation induces release of this arsenal into the phagosome and induces the assembly at the membrane of subunits of the NAPDH oxidase, the enzyme responsible for the production of superoxide anion that gives rise to other reactive oxygen species (ROS), a process called respiratory burst. Altogether, they are responsible for the bactericidal activity of the neutrophils. Excessive activation of neutrophils can lead to extensive release of these toxic agents, inducing tissue injury and the inflammatory reaction. Envenomation, caused by different animal species (bees, wasps, scorpions, snakes etc.), is well known to induce a local and acute inflammatory reaction, characterized by recruitment and activation of leukocytes and the release of several inflammatory mediators, including prostaglandins and cytokines. Venoms contain several molecules such as enzymes (phospholipase A2, L-amino acid oxidase and proteases, among others) and peptides (disintegrins, mastoporan, parabutoporin etc.). These molecules are able to stimulate or inhibit ROS production by neutrophils. The present review article gives a general overview of the main neutrophil functions focusing on ROS production and summarizes how venoms and venom molecules can affect this function.(AU)
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Animales , Venenos/administración & dosificación , Especies Reactivas de Oxígeno , NADPH Oxidasas , L-Aminoácido Oxidasa , Neutrófilos , AntiinflamatoriosRESUMEN
SUMMARY OBJECTIVES Lymphomas are a heterogeneous set of malignant neoplasias of lymphoid B and NK/T mature and immature cells at various stages of differentiation. Genetic and molecular biology tools are used to appropriately classify the type and prognosis of the lymphomas, which have implications in therapeutic effectiveness. Among them, the nicotinamide adenine dinucleotide phosphate-oxidase (NADPH) oxidase (NOX5) enzymes have been explored. This study analyzed the expression of NADPH oxidase 5 in lymphoma tissue according to the degree of tumor aggressiveness. METHODS Slides from 64 patients with lymphoma who had paraffin-embedded tissue available were reviewed by two independent, experienced pathologists. They classified tumors according to the WHO classification (2017). NOX5 expression in tissues was assessed by immunohistochemical staining using a tissue microarray. The assay was interpreted using a scoring system of 0, 1, 2, and 3, for cytoplasmic staining of NOX5 corresponding to negative, weak, intermediate, and strong staining, respectively. We compared the expression of NOX5 in patients with aggressive versus non-aggressive lymphomas. RESULTS NOX5 expression was positive in 100% (27/27) of aggressive lymphomas and in 19% (7/37) of non-aggressive ones. The seven patients with positive expression of NOX5 presented intermediate staining (2); strong staining (3) was observed only in tissues of aggressive lymphomas, and negative and weak staining (0 and 1) were observed only in non-aggressive lymphomas. CONCLUSIONS Aggressive lymphomas overexpress NOX5 protein. The higher NOX5 expression in aggressive lymphomas can suggest an involvement of this enzyme on the acquisition of an aggressive phenotype in lymphoid neoplasia.
RESUMO OBJETIVOS Os linfomas são um grupo heterogêneo de neoplasias malignas de células linfoides B e NK/T maduras e imaturas em vários estágios de diferenciação. Ferramentas de biologia molecular e genética são usadas para classificar adequadamente o tipo e o prognóstico dos linfomas, os quais têm implicações na eficácia terapêutica. Entre eles, as enzimas nicotinamida adenina dinucleótido fosfato oxidase (NADPH) oxidase (NOX5) foram exploradas. Este estudo analisou a expressão da NADPH oxidase 5 em linfomas de acordo com o grau de agressividade tumoral. MÉTODOS As lâminas de 64 pacientes com linfoma, que tinham tecido embebido em parafina disponível, foram revisadas por dois patologistas experientes independentemente. Eles utilizaram a classificação da OMS (2017). A expressão de NOX5 nos tecidos foi avaliada por coloração imuno-histoquímica utilizando microarray de tecido. O ensaio foi interpretado com um sistema de pontuação de 0, 1, 2 e 3, para coloração citoplasmática de NOX5 correspondente à coloração negativa, fraca, intermediária e forte, respectivamente. Comparamos a expressão de NOX5 em pacientes com linfomas agressivos versus não agressivos. RESULTADOS A expressão de NOX5 foi positiva em 100% (27/27) dos linfomas agressivos e em 19% (7/37) dos linfomas não agressivos. Os sete pacientes com expressão positiva de NOX5 apresentaram coloração intermediária (2); coloração forte (3) foi observada apenas em tecidos de linfomas agressivos, e negativos e fracos (0 e 1) observados apenas em linfomas não agressivos. CONCLUSÕES Linfomas agressivos superexpressam a proteína NOX5. A expressão aumentada de NOX5 em linfomas agressivos pode sugerir um envolvimento dessa enzima na aquisição de um fenótipo agressivo na neoplasia linfoide.
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Humanos , Regulación hacia Arriba , NADPH Oxidasa 5/análisis , Linfoma/patología , Pronóstico , Inmunohistoquímica , Estudios Retrospectivos , Adhesión en Parafina , Invasividad NeoplásicaRESUMEN
The present study analyzed the in vivo effects of drinking caffeinated and decaffeinated instant coffee (8% w/v) by adult male Wistar rats submitted to high-intensity exercises. The parameters used in the evaluation were the determination of the activities of NADPH oxidase, myeloperoxidase and other antioxidant enzymes present in neutrophils of rats. It was observed that exercise-induced superoxide anion production depends on the NADPH oxidase activity (estimated by the cytochrome C reduction test) in peritoneal neutrophils (p < 0.05). The intake of caffeinated and decaffeinated instant coffee beverages and of a caffeine solution to 1.67% did not induced changes in the activities of the enzymes myeloperoxidase, superoxide dismutase and glutathione peroxidase (p < 0.05). But consumption of caffeinated instant coffee drink prevented an increase in NADPH oxidase-mediated superoxide production induced by highly intense exercise in rat neutrophils. While the decaffeinated instant coffee drink or caffeine solution alone did not affect NADPH oxidase-mediated superoxide production. We suggest that this activity is associated with the chemical composition and concentration of phenolic compounds and other antioxidant substances formed during roasting. From the obtained results, it was concluded that moderate intake of caffeinated instant coffee (equivalent to a daily human consumption of 4 50-mLcups of coffee) may have beneficial effects on health, contributing to a reduction in superoxide anion generation. Therefore, more research must be conducted to elucidate the mechanism of action of caffeinated coffee on NADPH oxidase in neutrophils.
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Análisis de MediaciónRESUMEN
Objective:From a new perspective,to explore therapeutic effect of Huidouba (HDB) on alleviating kidney oxidative damage in rats with diabetic nephropathy (DN) and provide a scientific basis for developing HDB as a potential Tibetan medicine for treatment of DN. Method:Rats were fed with high-fat diet (HFD) and injected with streptozocin (STZ, 65 mg·kg-1) intraperitoneally to induce DN model, while rats in Blank group were injected with an equal volume of vehicle and fed with normal chow. The successfully modeling DN rats were randomly divided into three groups, 8 rats per group, DN model group (10 mL·kg-1·d-1), Metformin group (0.045 g·kg-1·d-1) and HDB group (0.18 g·kg-1·d-1). Monitor body weight (BW) and fasting blood glucose (FBG) weekly, and collect 24 hours urine before and after medication to examine microalbuminuria (mAlb). Calculate kidney index (KI) after sacrificing, analyze mAlb, serum creatinine (SCr) and blood urea nitrogen (BUN) with a fully automatic biochemical analyzer. Histopathology of kidney was observed by Masson staining. Lipid peroxidation malondialdehyde (MDA) assay kit was used to examine MDA content in kidney tissue. Nox4, as a subtype of triphosphopyridine nucleotide (NADPH) oxidase family was determined by Western blot and immunofluorescence assay of kidney tissue. Result:Compared with blank group, levels of FBG, 24 h mAlb, SCr, BUN and MDA in DN model group were increased (P<0.01), tissue damage was obvious and Nox4 expression in glumeruli was increased significantly (P<0.01). Compared with DN model group, levels of FBG, 24 h mAlb, SCr, BUN and MDA in drug administration groups were decreased (P<0.01), kidney injury was alleviated and Nox4 expression was down-regulated(P<0.01). Conclusion:HDB as a Yiqiyangyin Tibetan medicine, could ease oxidative stress injury of kidney and reduce proteinuria in DN rats, thus prevent the development of DN. Its mechanism is closely related to down-regulating Nox4 expression of kidney tissue in DN rats.
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AIM: To study the effect and mechanism of glx351322, an inhibitor of NADPH oxidase (NOX4), on the formation of tumor associated fibroblasts (CAFs). METHODS: NIH3T3 cells were co-cultured with H22 cells. 5 μm (IC50) GLX351322 cells were pretreated with NIH3T3. After PI staining, flow cytometry was used to detect the change of cell cycle, immunofluorescence staining was used to detect the expression of CAFs markers α-SMA and FAP, Western blot was used to detect the expression of CAFs markers α-SMA, Desmin, FAP, TSP-1, FSP, CyclinD, TGF-β1 and Smad1.After H22 was used to construct tumor bearing mice model, GLX351322 was used for treatment,Immunohistochemical staining was used to detect the expression of α-SMA and CAFs in tumor tissue. RESULTS:GLX351322 pretreatment could inhibit the proliferation of NIH3T3, decrease cell viability and change cell cycle. At the same time, it could down-regulate the expression of α-SMA, desmin, FAP, TSP-1, FSP, CyclinD, and TGF-β signal was inhibited. GLX351322 also significantly inhibited the expression of α-SMA and CAFs markers in tumor mice. CONCLUSION: GLX351322, a NOX4 inhibitor, can inhibit the formation of tumor-related fibroblasts, which is related to the inhibition of TGF-β signal.
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Objective: To study the effects of cadmium stress on mycelial growth and accumulation of metabolites in Ganoderma lucidum, and to explore the mechanisms affecting growth and accumulation of metabolites, and to provide evidence for controlling cadmium in the production and cultivation of G. lucidum. Methods: The mycelium of G. lucidum was cultured under the conditions of heavy metal ion cadmium concentration of 0, 0.5, 1, 4, 10, and 40 mg/L, and its biomass accumulation, intracellular ROS level, membrane oxidative damage, anti-oxidant enzyme activity, and ROS regulation related enzyme expression were analyzed. Results: When the concentration of cadmium reached 4 mg/L, the mycelial growth was inhibited. The levels of intracellular ROS, H2O2, and MDA increased significantly, increasing by 76%, 46% and 325%, respectively, and increased with the increase of cadmium concentration; The NADPH expression levels of oxidase gene (NOXA), superoxide dismutase gene (SOD1 and SOD4), and CATalase gene (CAT) were significantly up-regulated. When the cadmium concentration reached 10 mg/L, the inhibitory effect was significant. The colony growth diameter and the dry weight inhibition rate of fermentation mycelium were 26.15% and 32.78%, respectively. The total triterpenoid inhibition rate of G. lucidum was 33.7%, and the inhibition rate of total protein synthesis was 30.3%. Inhibition of polysaccharides was not significant. When the cadmium concentration reached 40 mg/L, the expression levels of Ascorbate peroxidase gene (APX) and Glutathione peroxidase gene (GPX) were significantly up-regulated. With the increase of cadmium concentration, the activities of SOD, CAT, APX, and GPX increased first and then decreased. When the concentration of cadmium reached 1 mg/L, the activity of GPX decreased and the activity of APX increased significantly. Exogenous addition of diphenyleneiodonium chloride (DPI), N-acetyl-L-cysteine (NAC) and vitamin C (VC) had significant effects on cadmium-induced G. lucidum clearance of ROS and reduction of MDA content. Conclusion: Cadmium stress causes the decrease of mycelial production and metabolite accumulation of G. lucidum, which may be due to the inhibition of GPX activity by cadmium ions, resulting in the accumulation of H2O2, causing the increase of ROS level and membrane oxidative damage, inhibiting mycelial growth and accumulation of metabolites, and regulating NOX. Up-regulation of gene expression results in an increase in anti-oxidant enzyme activity and expression to increase the clearance of reactive oxygen species. Therefore, the cadmium content should be controlled within the range of 1 mg/L during the production process.
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Objective@#To investigate the aggravation of pancreatic tissue injury in rats with acute hypertriglyceridemic pancreatitis and the possible role of NADPH oxidase (NOX).@*Methods@#Thirty SPF rats were randomly (random number)divided into five groups: N group, H group, NLAP group, HLAP group and HAPO group. AMY, TG, TC and FFA levels were detected. The pathological changes of pancreas were observed under light microscope and the ultrastructural changes of pancreatic acinar cells were observed by TEM. Serum levels of MDA, SOD, IL-1β, TNF-α and LDH were detected. The expression of NOX4, p-Akt and p-GSK3β in pancreas was detected by immunofluorescence, and the expression of NF-κB and TNF-α in pancreas was detected by immunohistochemistry.@*Results@#Intraperitoneal injection of P-407 could significantly increase the levels of serum TG, TC and FFA in rats. After acute pancreatitis induced by L-Arg, the levels of serum AMY in the NLAP and HLAP groups were significantly increased, while Apocynin could significantly decrease the level of serum AMY. Compared with the NLAP group, the pathological injury of pancreatic tissue in the HLAP group was more serious, the level of inflammatory mediators was significantly increased, and the cell necrosis was more serious. After inhibiting NOX, the activation of Akt/GSK3β pathway was regulated and the pancreatic injury was improved.@*Conclusion@#In HTGP, NOX aggravates pancreatic injury by regulating the activation of Akt/GSK3 β pathway. Inhibition of NOX expression can play a protective role in pancreas injury of HTGP..
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Objective To detect the expression levels of collagen1 (colla-1),transforming growth factor-β1 (TGF-β1),a-smooth muscle actin (α-SMA) and nicotinamide adenine dinucleotide phosphate oxidase 4 (NOX-4) in mouse esophagus submitted to chronic restraint stress (CRS),in order to discuss stress-induced esophageal fibrosis and the role of oxidative stress.Methods 20 male Kunming mice were randomly divided into two groups,CRS and normal control (NC).The mice in CRS group were submitted to 2 h per day of restraint stress using home-made device for a period of 14 days,and the mice in both group were treated the same at rest of the time.Fibrotic changes of esophageal tissue were observed using Masson staining.The expression levels of NOX-4 and related fibrotic cytokines in esophageal tissues were detected by several methods such as immunohistochemistry,enzyme-linked immunosorbent assay (ELISA) and realtime polymerase chain reaction (qRT-PCR).Results Body weight in CRS group was significantly lower than NC group (8.75 ± 1.69 vs 12.69 ± 3.16),with statistically significant difference (t =3.11,P < 0.05).Masson staining revealed that CRS mice showed distinct fibrosis of epithelial interstitium,while there was no distinct changes observed in NC mice.Immunohistochemical staining revealed intense staining for NOX-4 in epithelial,mucosal and submucosal layers of esophagi in CRS mice.ELISA showed that the serum level of NOX-4 in CRS mice was higher than NC mice (1.442 ± 0.05 vs 0.449 ± 0.08),with statistically significant difference (t =-27.32,P < 0.01).Real-time PCR results showed that the expression of colla-1,TGF-β1,α-SMA and NOX-4 in CRS mice were as (2.443 ±0.36,2.78 ±0.13,2.244 ±0.18,2.448 ±0.440) times higher than NC mice,with statistically significant difference (t =-11.19,-38.86,-19.90,-10.37,P < 0.01).Conclusions Fibrotic cytokines such as colla-1,TGF-β1 and α-SMA may participate in formation of stress induced esophageal fibrosis,and oxidative stress may play crucial role in the process of esophageal fibrosis.
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Objective To investigate the aggravation of pancreatic tissue injury in rats with acute hypertriglyceridemic pancreatitis and the possible role of NADPH oxidase(NOX).Methods Thirty SPF rats were randomly(random number)divided into five groups: N group,H group,NLAP group,HLAP group and HAPO group.AMY,TG,TC and FFA levels were detected.The pathological changes of pancreas were observed under light microscope and the ultrastructural changes of pancreatic acinar cells were observed by TEM.Serum levels of MDA,SOD,LM-1β,TNF-α and LDH were detected.The expression of NOX4,p-Akt and p-GSK3β in pancreas was detected by immunofluorescence,and the expression of NF-κB and TNF-α in pancreas was detected by immunohistochemistry.Results Intraperitoneal injection of P-407 could significantly increase the levels of serum TG,TC and FF A in rats.After acute pancreatitis induced by L-Arg,the levels of serum AMY in the NLAP and HLAP groups were significantly increased,while Apocynin could significantly decrease the level of serum AMY.Compared with the NLAP group,the pathological injury of pancreatic tissue in the HLAP group was more serious,the level of inflammatoty mediators was significantly increased,and the cell necrosis was more serious.After inhibiting NOX,the activation of Akt/GSK3β pathway was regulated and the pancreatic injury was improved.Conclusion In HTGP,NOX aggravates pancreatic injury by regulating the activation of Akt/GSK3 β pathway.Inhibition of NOX expression can play a protective role in pancreas injury of HTGP..