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[Display omitted] • Androgen-dependent benign prostatic hyperplasia (BPH) model exerts excessive ROS, which was revealed by the involvement of the AR and NOX4 networks. • In BPH in vitro and in vivo model, the NADPH oxidase inhibitor Apocynin (Apo) suppresses inflammation and proliferation to suppress BPH through the AR/NOX4/TGF-β signaling. • Apo exerts anti-proliferative, anti-inflammatory, and anti-oxidant effects, highlighting the potential of Apo as a therapeutic agent for treatment of BPH. Apocynin (Apo), an NADPH oxidase (NOX) inhibitor, has been widely used to treat various inflammatory diseases. However, the therapeutic effects of Apo on benign prostatic hyperplasia (BPH), a multifactorial disease associated with chronic inflammation and hormone imbalance, remain unknown. The link between androgen signaling, reactive oxygen species (ROS), and prostate cell proliferation may contribute to the pathogenesis of BPH; therefore, the aim of this study was to identify the specific signaling pathway involved and to demonstrate whether the anti-oxidant Apo plays a role in the prevention and treatment of BPH. Ingenuity pathway analysis and si-RNA transfection were conducted to demonstrate the androgen receptor (AR) and NOX4 linkage in BPH. Pathological markers of BPH were measured by H&E staining, immunoblotting, ELISA, qRT-PCR, and immunofluorescence to examine the effect of Apo. Rats stimulated with testosterone and BPH-1 cells were used as BPH models. AR and NOX4 network-mediated oxidative stress was upregulated in the BPH model. Next, we examined the effects of Apo on oxidative stress and chronic prostatic inflammation in BPH mouse models. In a testosterone-induced BPH rat model, Apo alleviated pathological prostate enlargement and suppressed androgen/AR signaling. Apo suppressed the upregulation of proinflammatory markers and promoted the expression of anti-oxidant factors. Furthermore, Apo regulated the TGF-β/Glut9/activin pathway and macrophage programming. In BPH-1 cells, Apo suppressed AR-mediated proliferation and upregulation of TGFB and NOX4 expression by alleviating oxidative stress. Apo activated anti-oxidant and anti-inflammatory systems and regulated macrophage polarization in BPH-1 cells. AR knockdown partially abolished the beneficial effects of Apo in prostate cells, indicating AR-dependent effects of Apo. In contrast with existing BPH therapies, Apo may provide a new application for prostatic disease treatment, especially for BPH, by targeting the AR/TGF-β/NOX4 signaling pathway. [ABSTRACT FROM AUTHOR]

Background: The purpose of this study was to prepare neutrophil membrane-engineered Panax ginseng root-derived exosomes (N-exo) and investigate the effects of N-exo microRNA (miRNA) 182-5p (N-exo-miRNA 182-5p) on acute lung injury (ALI) in sepsis. Methods: Panax ginseng root-derived exosomes were separated by differential centrifugation. Neutrophil membrane engineering was performed on exo to obtain N-exo. miRNA182-5p was transmitted into N-exo by electroporation technology to obtain N-exo-miRNA 182-5p. LPS was used to establish an in-vivo and in-vitromodel of ALI of sepsis to evaluate the anti-inflammatory effect of N-exo-miRNA 182-5p. Results: The results of transmission electron microscope showed that exo was a double-layer membrane structure like a saucer. Nanoparticle size analysis showed that the average particle size of exo was 129.7 nm. Further, compared with exo, the level of miRNA182-5p was significantly increased in N-exo. The experimental results showed that N-exo-miRNA 182-5p significantly improved ALI via target regulation of NOX4/Drp-1/NLRP3 signal pathway in vivo and in vitro. Conclusion: In conclusion, this study prepared a novel engineered exosome (N-exo and N-exo-miRNA 182-5p significantly improved ALI in sepsis via target regulation of NOX4/Drp-1/NLRP3 signal pathway, providing new ideas and methods for treatment of ALI in sepsis. [ABSTRACT FROM AUTHOR]

Oxidative stress closely related to the progression and severity of myocardial infarction (MI). Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 4 (NOX4) is one of the major enzymes that generate reactive oxygen species (ROS) in cardiovascular system. Here, we aim to elucidate the pathological role of NOX4 in MI. MI mouse model was created by the coronary artery ligation. NOX4 was specifically knocked down in heart through intramyocardial injection of siRNA. NOX4 expression and oxidative stress indicators were determined at different time points using qRT-PCR, Western blot, and ELISA, and then analyzed by Pearson's correlation. Cardiac function was evaluated by using echocardiographic technique. NOX4 was upregulated in myocardial tissues of MI mice, which positively correlated with the elevation of oxidative stress indicators. Knockdown of NOX4 in heart significantly reduced the production of ROS and the level of oxidative stress in left ventricle tissues, which was accompanied by significant improvement of cardiac function in MI mice. Selective knockdown of NOX4 in heart attenuates MI-induced oxidative stress and improves cardiac function, suggesting inhibition of NOX4/ROS axis in heart using siRNA is a potential therapeutic treatment for MI-induced cardiac dysfunction. [ABSTRACT FROM AUTHOR]

• Trichodelphinine A targets NOX4 for anti-fibrotic effects. • Inhibition of NOX4 reduces ARG1/OAT and TGF-β/Smads activation, then decreases collagen synthesis. • The PHE354 and THR355 residues of NOX4 are essential for its interaction with trichodelphinine A. Pulmonary fibrosis, a progressive and fatal lung disease with no effective treatment medication, is characterized by lung remodeling and fibroblastic foci caused by an oxidative imbalance with an overloading deposition of collagen. Trichodelphinine A, a hetisine-type C20-diterpenoid alkaloid, was found anti-fibrotic activity in vitro, but its effect and mechanism on pulmonary fibrosis still unknown. Our study aimed to investigate and validate the anti-fibrotic properties of trichodelphinine A in pulmonary fibrosis animals induced by bleomycin (BLM), and its mechanism whether via NOX4-ARG1/TGF-β signaling pathway. The anti-fibrotic effects of trichodelphinine A were evaluated using BLM-induced rats through indicators of lung histopathology and collagen synthesis. Dynamic metabolomics evaluated the metabolic disorder and therapeutic effect of trichodelphinine A. The interaction between trichodelphinine A and NOX4 receptor was confirmed using CETSA and molecular dynamics experiments. Molecular biology experiments were conducted in NOX4 gene knockout mice to investigate the intervention effect of trichodelphinine A. Trichodelphinine A could suppress histopathologic changes, collagen deposition and proinflammatory cytokine release pulmonary fibrosis in bleomycin induced rats. Dynamic metabolomics studies revealed that trichodelphinine A could correct endogenous metabolic disorders of arachidonic acid, arginine and proline during fibrosis development, which revealed that the regulation of oxidative stress and amino acid metabolism targeting NOX4 and ARG1 may be the main pharmacological mechanisms of trichodelphinine A on pulmonary fibrosis. We further determined that trichodelphinine A inhibited over oxidative stress and collagen deposition by suppressing Nrf2-keap1 and ARG1-OAT signaling pathways, respectively. Molecular dynamics studies showed that trichodelphinine A was directly binds with NOX4, in which PHE354 and THR355 residues of NOX4 are critical binding sites for trichodelphinine A. Mechanistic validation in cells or mice with NOX4 knockout or silencing suggested that the anti-fibrotic effects of trichodelphinine A depended on inhibition of NOX4 to suppress ARG1/OAT activation and TGF-β/Smads signaling pathway. Collectively, our findings indicate a powerful anti-fibrotic function of trichodelphinine A in pulmonary fibrosis via targeting NOX4. NOX4 mediates the activation of ARG1/OAT to regulate arginase-proline metabolism, and promotes TGF-β/Smads signaling pathway, thereby affecting the collagen synthesis in pulmonary fibrosis, which is a novel finding and indicates that inhibition of NOX4 is a novel therapeutic strategy for pulmonary fibrosis. [ABSTRACT FROM AUTHOR]

Glioblastoma (GBM) is the most aggressive and common glioma subtype, with a median survival of 15 months after diagnosis. Current treatments have limited therapeutic efficacy; thus, more effective approaches are needed. The glioblastoma tumoural mass is characterised by a small cellular subpopulation - glioblastoma stem cells (GSCs) - that has been held responsible for glioblastoma initiation, cell invasion, proliferation, relapse and resistance to chemo- and radiotherapy. Targeted therapies against GSCs are crucial, as is understanding the molecular mechanisms that govern the GSCs. Transforming growth factor β (TGFβ) signalling and reactive oxygen species (ROS) production are known to govern and regulate cancer stem cell biology. Among the differentially expressed genes regulated by TGFβ in a transcriptomic analysis of two different patient-derived GSCs, we found NADPH oxidase 4 (NOX4) as one of the top upregulated genes. Interestingly, when patient tissues were analysed, NOX4 expression was found to be higher in GSCs versus differentiated cells. A functional analysis of the role of NOX4 downstream of TGFβ in several patient-derived GSCs showed that TGFβ does indeed induce NOX4 expression and increases ROS production in a NOX4-dependent manner. NOX4 downstream of TGFβ regulates GSC proliferation, and NOX4 expression is necessary for TGFβ-induced expression of stem cell markers and of the transcription factor nuclear factor erythroid 2-related factor 2 (NRF2), which in turn controls the cell's antioxidant and metabolic responses. Interestingly, overexpression of NOX4 recapitulates the effects induced by TGFβ in GSCs: enhanced proliferation, stemness and NRF2 expression. In conclusion, this work functionally establishes NOX4 as a key mediator of GSC biology. [ABSTRACT FROM AUTHOR]

Breast cancer arises as a result of multiple interactions between environmental and genetic factors. Conventionally, breast cancer is treated based on histopathological and clinical features. DNA technologies like the human genome microarray are now partially integrated into clinical practice and are used for developing new "personalized medicines" and "pharmacogenetics" for improving the efficiency and safety of cancer medications. We investigated the effects of four established therapies—for ER+ ductal breast cancer—on the differential gene expression. The therapies included single agent tamoxifen, two-agent docetaxel and capecitabine, or combined three-agents CAF (cyclophosphamide, doxorubicin, and fluorouracil) and CMF (cyclophosphamide, methotrexate, and fluorouracil). Genevestigator 8.1.0 was used to compare five datasets from patients with infiltrating ductal carcinoma, untreated or treated with selected drugs, to those from the healthy control. We identified 74 differentially expressed genes involved in three pathways, i.e., apoptosis (extrinsic and intrinsic), oxidative signaling, and PI3K/Akt signaling. The treatments affected the expression of apoptotic genes (TNFRSF10B [ TRAIL ], FAS , CASP3/6/7/8 , PMAIP1 [ NOXA ], BNIP3L , BNIP3 , BCL2A1 , and BCL2), the oxidative stress-related genes (NOX4 , XDH , MAOA , GSR , GPX3 , and SOD3), and the PI3K/Akt pathway gene (ERBB2 [ HER2 ]). Breast cancer treatments are complex with varying drug responses and efficacy among patients. This necessitates identifying novel biomarkers for predicting the drug response, using available data and new technologies. GSR , NOX4 , CASP3 , and ERBB2 are potential biomarkers for predicting the treatment response in primary ER+ ductal breast carcinoma. [ABSTRACT FROM AUTHOR]

• Hyperglycemic-induced oxidative stress upregulates NF-κB and downregulate Nrf2 pathways. • NF-κB/SOD2 participates in an early adaptive response to oxidative stress. • rWPP reduce endothelial oxidative stress through Nrf2, SIRT1 and NF-κB. • rWPP increases antioxidant and antiinflamatory cell response. Wine by-products show great potential as source of bioactive compounds that protect the vascular endothelial function by the modulation of both Nrf2 and Nf-κB pathways. This study investigates the pathways involved in the effects of a red wine pomace product (rWPP) against inflammatory and oxidative damage in hyperglycemic EA.hy926 endothelial cells. rWPP-digested fractions showed an inhibitory effect on IKK/IκBα/NF-κB pathway and a stimulatory effect on Akt-p38-MAPK/Nrf2 pathway with an impact on their antioxidant and anti-inflammatory downstream targets. In addition, the pathways regulation was also accompanied by downregulation of the gene expression of superoxide dismutase 2, cyclooxygenase 2 and NADPH oxidase 4. These results suggest the expression of SOD2 as an early adaptive response to the inflammatory effect mediated by NF-κB in hyperglycemic cells, and the treatment with the rWPP-digested fractions regulate this inflammatory process by Nrf2 pathway increased expression of antioxidant enzymes. [ABSTRACT FROM AUTHOR]

OBJECTIVE: To elucidate the effect of estrogen on right ventricular remodeling of pulmonary arterial hypertension (PAH) induced by monocrotaline (MCT) in rats and its underlying mechanism. MATERIALS AND METHODS: Male Sprague-Dawley (SD) rats were adaptively fed for one week, and then, randomly divided into control group, MCT group, MCT+17-β estradiol (50 mg·kg− 1·d−1) group, and MCT+17-β estradiol (100 mg·kg−1·d−1) group, with 8 rats in each group. PAH rat model was constructed by subcutaneous injection of 60 mg·kg−1 MCT for consecutive 4 weeks. The right external jugular vein was intubated to monitor rat RVSP (right ventricular systolic pressure) and mPAP (mean pulmonary arterial pressure). After animal procedures, RV (right ventricle) and LV+S (left ventricle+ventricular septum) of rat were harvested and weighed. Rat tibia was collected and recorded for its length, followed by calculation of RV/(LV+S) and RV/length of the tibia. HE staining and Masson staining were conducted to observe the pathological change and collagen deposition in RV, respectively. RV was prepared for homogenate, followed by detection of total antioxidant capacity (T-AOC) and malondialdehyde (MDA) activities. Expression levels of collagen I, collagen III, NOX4, and NF-κB in RV of PAH rats were detected by qRT-PCR and Western blot. RESULTS: Lower RVSP, mPAP, RV/(LV+S) and RV/length of the tibia were observed in rats of MCT+17-β estradiol (50 mg·kg−1·d−1) group and MCT+17-β estradiol (100 mg·kg−1·d−1) group compared with those of MCT group. Injection of 17-β estradiol (50 mg·kg−1·d−1 and 100 mg·kg−1·d−1) in PAH rats remarkably alleviated pathological changes and collagen deposition in RV. T-AOC activity increased, whereas MDA activity decreased in PAH rats injected with 17-β estradiol. Expression levels of collagen I, collagen III, NOX4, and NF-κB in RV of PAH rats were all downregulated by 17-β estradiol treatment. CONCLUSIONS: 17-β e stradiol could remarkably alleviate MCT-induced right ventricular remodeling of PAH rats. It is suggested that 17-β estradiol exerts its protective role in PAH by inhibiting NOX4-mediated oxidative stress and NF-κB-mediated collagen deposition. [ABSTRACT FROM AUTHOR]

OBJECTIVE: Nasopharyngeal carcinoma (NPC) is a malignancy caused by Epstein-Barr virus (EBV). NPC is radiosensitive and has a high frequency of treatment failure due to metastasis, which results in recurrent nasopharyngeal carcinoma (rNPC). PATIENTS AND METHODS: In the present study, nasopharyngeal carcinoma biopsies were obtained from NPC and rNPC patients, as well as healthy controls, and reverse transcription-polymerase chain reaction (RT-PCR), immunohistochemistry, and immunoblotting analyses were performed. RESULTS: The RTPCR data showed expression of CDX2 and NOX4 in rNPC biopsy samples but not in control or NPC samples. Immunohistochemical and immunoblotting analyses confirmed the expression of CDX2 and NOX4 in rNPC samples, but not in NPC biopsy samples. CONCLUSIONS: The finding concludes that an association of CDX2 and NOX4 expression with rNPC was noted; thus, these proteins may have value as prognostic indicators and may facilitate the development of novel therapeutics for rNPC patients. [ABSTRACT FROM AUTHOR]

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Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) have been widely used for human non-small-cell lung cancer (NSCLC) treatment. However, acquired resistance to EGFR-TKIs is the major barrier of treatment success, and new resistance mechanism remains to be elucidated. In this study, we found that elevated NADPH oxidase 4 (NOX4) expression was associated with acquired EGFR-TKIs resistance. Gefitinib is the first-generation FDA-approved EGFR-TKI, and osimertinib is the third-generation FDA-approved EGFR-TKI. We demonstrated that NOX4 knockdown in the EGFR-TKI resistant cells enabled the cells to become sensitive to gefitinib and osimertinib treatment, while forced expression of NOX4 in the sensitive parental cells was sufficient to induce resistance to gefitinib and osimertinib in the cells. To elucidate the mechanism of NOX4 upregulation in increasing TKIs resistance, we found that knockdown of NOX4 significantly down-regulated the expression of transcription factor YY1. YY1 bound directly to the promoter region of IL-8 to transcriptionally activate IL-8 expression. Interestingly, knockdown of NOX4 and IL-8 decreased programmed death ligand 1 (PD-L1) expression, which provide new insight on TKIs resistance and immune escape. We found that patients with higher NOX4 and IL-8 expression levels showed a shorter survival time compared to those with lower NOX4 and IL-8 expression levels in response to the anti-PD-L1 therapy. Knockdown of NOX4, YY1 or IL-8 alone inhibited angiogenesis and tumor growth. Furthermore, the combination of NOX4 inhibitor GKT137831 and gefitinib had synergistic effect to inhibit cell proliferation and tumor growth and to increase cellular apoptosis. These findings demonstrated that NOX4 and YY1 were essential for mediating the acquired EGFR-TKIs resistance. IL-8 and PD-L1 are two downstream targets of NOX4 to regulate TKIs resistance and immunotherapy. These molecules may be used as potential new biomarkers and therapeutic targets for overcoming TKIs resistance in the future. [Display omitted] [ABSTRACT FROM AUTHOR]

Background: Psoralidin (PSO), a natural phenolic coumarin, was reported to have anti-cancer activities. PSO induced reactive oxygen species (ROS) generation in cancer cells. The role of ROS in its anti-cancer effect remains unclear.Purpose: This study was designed to investigate the potential roles of ROS in PSO-induced anti-cancer effect in MCF-7 breast cancer cells.Methods: Effect of PSO on cancer cell proliferation was determined by MTT assay. Comet assay was used to determine DNA damage. Protein expression was detected by Western blotting. Autophagic vacuoles were detected by monodansylcadaverine (MDC) staining. ROS generation was measured by fluorescent probe. NOX4 localization was determined by immunofluorescence staining.Results: PSO treatment caused proliferation inhibition in time- and dose- dependent manners, which was partially reversed by N-acetyl cysteine (NAC) and diphenyleneiodonium (DPI). PSO induced DNA damage and increased protein expression of γ-H2AX, phosphorylation of ATM, ATR, Chk1, and Chk2. PSO induced autophagy as evidenced by the accumulation of autophagic vacuoles and alterations of autophagic protein expression. PSO-induced cell death was enhanced by autophagy inhibitor chloroquine (CQ). Furthermore, PSO treatment induced ROS formation, which was reversed by NAC or DPI pretreatment. The expression of NOX4 was significantly enhanced by PSO. Both NAC and DPI could reverse PSO-induced DNA damage and autophagic responses. In addition, silencing NOX4 by siRNA inhibited PSO-induced ROS generation, DNA damage, and autophagy.Conclusions: Taken together, these results showed that PSO induced DNA damage and protective autophagy mediated by ROS generation in a NOX4-dependent manner in MCF-7 cells. [ABSTRACT FROM AUTHOR]

This study reports the consequences of knocking out NADPH (nicotinamide adenine dinucleotide phosphate) oxidase 4 (Nox4) on the development of hypertension and kidney injury in the Dahl salt-sensitive (SS) rat. Zinc finger nuclease injection of single-cell SS embryos was used to create an 8 base-pair frame-shift deletion of Nox4, resulting in a loss of the ≈68 kDa band in Western blot analysis of renal cortical tissue of the knock out of Nox4 in the SS rat (SS(Nox4-/-)) rats. SS(Nox4-/-) rats exhibited a significant reduction of salt-induced hypertension compared with SS rats after 21 days of 4.0% NaCl diet (134±5 versus 151±3 mm Hg in SS) and a significant reduction of albuminuria, tubular casts, and glomerular injury. Optical fluorescence 3-dimensional cryoimaging revealed significantly higher redox ratios (NADH/FAD [reduced nicotinamide adenine dinucleotide/flavin adenine dinucleotide]) in the kidneys of SS(Nox4-/-) rats even when fed the 0.4% NaCl diet, indicating greater levels of mitochondrial electron transport chain metabolic activity and reduced oxidative stress compared with SS rats. Before the development of hypertension, RNA expression levels of Nox subunits Nox2, p67(phox), and p22(phox) were found to be significantly lower (P<0.05) in SS(Nox4-/-) compared with SS rats in the renal cortex. Thus, the mutation of Nox4 seems to modify transcription of several genes in ways that contribute to the protective effects observed in the SS(Nox4-/-) rats. We conclude that the reduced renal injury and attenuated blood pressure response to high salt in the SS(Nox4-/-) rat could be the result of multiple pathways, including gene transcription, mitochondrial energetics, oxidative stress, and protein matrix production impacted by the knock out of Nox4. [ABSTRACT FROM AUTHOR]

To date, DNA methylation has been regarded as the most promising age-predictive biomarker. In support of this, several researchers have reported age predictive models based on the use of blood or even across a broad spectrum of tissues. However, there have been no publications that report epigenetic age signatures from semen, one of the most forensically relevant body fluids. In genome-wide DNA methylation profiles of 36 body fluids including blood, saliva, and semen, the previous age predictive models showed considerable prediction accuracy in blood and saliva but not in semen. Therefore, we selected CpG sites, whose methylation levels are strongly correlated with age in 12 semen profiles obtained from individuals of different ages, and investigated DNA methylation changes at these CpGs in 68 additional semen samples obtained from individuals aged 20 to 73 years using methylation SNaPshot reaction. Among the selected age-related CpG candidates, outstanding age correlation was obtained at cg06304190 in the TTC7B gene. Interestingly, the region around the TTC7B gene has been reported to show age-related DNA methylation alteration in the sperm methylome of 2 samples collected from individuals at certain time intervals. The age-predictive linear regression model trained with 3 CpGs (cg06304190 in the TTC7B gene, cg06979108 in the NOX4 gene and cg12837463) showed a high correlation between the predicted age and the chronological age, with an average absolute difference of approximately 5 years. These selected epigenetic age signatures are expected to be useful for considerably accurate age estimation in the forensically relevant body fluid of semen. However, because the findings were limited by small sample size, it will be necessary to further evaluate the age correlation of the selected CpGs and to encourage further investigation. [ABSTRACT FROM AUTHOR]

Enzymes synthesizing reactive oxygen (Nox family) have recently been identified. Elucidation of the production mechanism has been initiated, and the involvement of reactive oxygen in metabolism, intracellular transport, signal transmission and apoptosis has been reported. We immunohistochemically investigated expression and localization of the Nox family in endochondral ossification using a normal mouse femur. Weakly positive reactions with Nox1, Noxa1, and Noxo1 were observed in the zones of proliferative and prehypertrophic chondrocytes at 3 weeks of age. Nox4 was widely positive from the resting over the hypertrophic cell zone. At 18 weeks of age, none of the Nox types was expressed in chondrocytes as the zones disappeared. On the other hand, positive reactions with Nox1, Noxa1, Noxo1, and Nox4 were observed in osteoblasts in the zone of ossification at 3 weeks of age, and each Nox was also positive in osteoblasts arranged on the bone marrow side in the epiphyseal cartilage at 18 weeks of age. In addition, a reactive oxygen-eliminating enzyme, Mn-SOD, was observed only in prehypertrophic chondrocytes at 3 weeks of age, and not detected in osteoblasts. It was suggested that the Nox family is closely associated with endochondral ossification of the mouse femur, and Nox1 and Nox4 are closely involved in the chondrocyte maturation process and bone matrix formation. [ABSTRACT FROM AUTHOR]

Abstract: The functional significance and regulation of NAD(P)H oxidase (Nox) isoforms by angiotensin II (Ang II) and endothelin-1 (ET-1) in vascular smooth muscle cells (VSMCs) from normotensive Wistar-Kyoto (WKY) and spontaneously hypertensive rats (SHR) was studied. Expression of Nox1, Nox2, and Nox4 (gene and protein) and NAD(P)H oxidase activity were increased in SHR. Basal NAD(P)H oxidase activity was blocked by GKT136901 (Nox1/4 inhibitor) and by Nox1 siRNA in WKY cells and by siNOX1 and siNOX2 in SHR. Whereas Ang II increased expression of all Noxes in WKY, only Nox1 was influenced in SHR. Ang II–induced NAD(P)H activity was inhibited by siNOX1 in WKY and by siNOX1 and siNOX2 in SHR. ET-1 upregulated Nox expression only in WKY and increased NAD(P)H oxidase activity, an effect inhibited by siNOX1 and siNOX2. Nox1 co-localized with Nox2 but not with Nox4, implicating association between Nox1 and Nox2 but not between Nox1 and Nox4. These data highlight the complexity of Nox biology in VSMCs, emphasising that more than one Nox member, alone or in association, may be involved in NAD(P)H oxidase-mediated •O2 − production. Nox1 regulation by Ang II, but not by ET-1, may be important in •O2 − formation in VSMCs from SHR. [Copyright &y& Elsevier]

The article presents a study that aims to identify Nicotinamide Adenosine Dinucleotide Phosphate (NADPH) oxidase (NOX) regulatory proteins. The study used cell culture from rat aortic vascular smooth muscle cells (VSMC) and human aortic smooth muscle cells grown in DMEM. Through the use of yeast-2 hybrid screen, a p22phox binding partner and poldip2 were identified. Moreover, poldip2 facilitates Nox4 enzymatic activity by 3-fold and regulates basal reactive oxygen species production in VSMCs.

Background: The purpose of this study was to investigate the expression and distribution of superoxide anion, NADPH oxidase (NOX)1, and NOX4 in healthy, allergic nasal mucosa and nasal polyps to evaluate the possible influence of oxidative stress on the development of allergic rhinitis and nasal polyps. Methods: The expression and distribution of superoxide anion, NOX1 and NOX4 were evaluated in healthy and allergic nasal mucosa and nasal polyps, using dihydroethidium fluorescence, semiquantitative reverse transcriptase-polymerase chain reaction, immunohistochemistry, and Western blot. Results: NOX1 and NOX4 were localized mainly in the epithelial layer, submucosal glands, vascular endothelium, and inflammatory cells in healthy and allergic nasal mucosa and nasal polyps. The cellular source that generated superoxide anion is also localized in the epithelial cells, submucosal glands, vascular endothelium, and inflammatory cells, demonstrating the similar sites of expression of NOX1 and NOX4 in healthy and allergic nasal mucosa and nasal polyps. NOX1 and NOX4 mRNA and proteins and superoxide anions had increased levels of expression in allergic nasal mucosa and nasal polyps compared with healthy nasal mucosa. Conclusions: These results indicate that NOX1 and NOX4 may play an important role in reactive oxygen species production, contributing to the oxidative stress in allergic rhinitis and nasal polyp tissues. [ABSTRACT FROM AUTHOR]

A large body of literature suggest that vascular reduced nicotinamide-adenine dinucleotide phosphate (NADPH) oxidases are important sources of reactive oxygen species. Many studies, however, relied on data obtained with the inhibitor apocynin (4'-hydroxy-3'methoxyacetophenone). Because the mode of action of apocynin, however, is elusive, we determined its mechanism of inhibition on vascular NADPH oxidases. In HEK293 cells overexpressing NADPH oxidase isoforms (Nox1, Nox2, or Nox4), apocynin failed to inhibit superoxide anion generation detected by lucigenin chemiluminescence. In contrast, apocynin interfered with the detection of reactive oxygen species in assay systems selective for hydrogen peroxide or hydroxyl radicals. Importantly, apocynin interfered directly with the detection of peroxides but not superoxide, if generated by xanthine/xanthine oxidase or nonenzymatic systems. In leukocytes, apocynin is a prodrug that is activated by myeloperoxidase, a process that results in the formation of apocynin dimers. Endothelial cells and smooth muscle cells failed to form these dimers and, therefore, are not able to activate apocynin. Dimer formation was, however, observed in Nox-overexpressing HEK293 cells when myeloperoxidase was supplemented. As a consequence, apocynin should only inhibit NADPH oxidase in leukocytes, whereas in vascular cells, the compound could act as an antioxidant. Indeed, in vascular smooth muscle cells, the activation of the redox-sensitive kinases p38-mitogen-activate protein kinase, Akt, and extracellular signal-regulated kinase 1/2 by hydrogen peroxide and by the intracellular radical generator menadione was prevented in the presence of apocynin. These observations indicate that apocynin predominantly acts as an antioxidant in endothelial cells and vascular smooth muscle cells and should not be used as an NADPH oxidase inhibitor in vascular systems. [ABSTRACT FROM AUTHOR]

[Display omitted] Clitoria ternatia L. (CT) has been reported to have anti-inflammatory and antioxidant effects. This study investigated the effect of CT aqueous flower extract on blood pressure and renal alterations in N ω-nitro- l -arginine methyl ester hydrochloride (l -NAME)-induced hypertensive rats. Male Sprague Dawley rats received l -NAME in drinking water and were treated with CT flower extract or lisinopril. CT aqueous flower extract and lisinopril alleviated l -NAME-induced hypertension (p < 0.05). Glomerular extracellular matrix accumulation, renal fibrosis, and increased serum creatinine levels were observed in l -NAME-induced hypertensive rats and attenuated by CT flower extract or lisinopril co-treatment (p < 0.05). High levels of plasma angiotensin II (Ang II) and upregulated nicotinamide adenine dinucleotide phosphate oxidase 4 (Nox4) protein expression in the kidneys induced by l -NAME were alleviated by CT flower extract or lisinopril co-treatment (p < 0.05). Furthermore, CT flower extract and lisinopril treatment reduced lipid peroxidation and elevated plasma and kidney malondialdehyde levels in l -NAME-induced hypertensive rats (p < 0.05). In conclusion, CT flower extract prevented l -NAME-induced renal injury and dysfunction in rats. The possible mechanism may be related to the suppression of Ang II-mediated Nox4 expression and the oxidative stress cascade in rats. [ABSTRACT FROM AUTHOR]

Both Ca2+ and reactive oxygen species (ROS) are double face entities, acting as signaling messengers or cell fate determinants according to their concentration and to spatial temporal restrictions. Recently, Beretta and colleagues found that ROS generated at ER-mitochondria contact sites (MAMs) support cell survival in stress conditions by decreasing inter-organelle Ca2+ transfer. [ABSTRACT FROM AUTHOR]

The immediate signals that couple exercise to metabolic adaptations are incompletely understood. Nicotinamide adenine dinucleotide phosphate oxidase 4 (Nox4) produces reactive oxygen species (ROS) and plays a significant role in metabolic and vascular adaptation during stress conditions. Our objective was to determine the role of Nox4 in exercise-induced skeletal muscle metabolism. Mice were subjected to acute exercise to assess their immediate responses. mRNA and protein expression responses to Nox4 and hydrogen peroxide (H 2 O 2) were measured by qPCR and immunoblotting. Functional metabolic flux was measured via ex vivo fatty acid and glucose oxidation assays using 14C-labeled palmitate and glucose, respectively. A chronic exercise regimen was also utilized and the time to exhaustion along with key markers of exercise adaptation (skeletal muscle citrate synthase and beta-hydroxyacyl-coA-dehydrogenase activity) were measured. Endothelial-specific Nox4-deficient mice were then subjected to the same acute exercise regimen and their subsequent substrate oxidation was measured. We identified key exercise-responsive metabolic genes that depend on H 2 O 2 and Nox4 using catalase and Nox4-deficient mice. Nox4 was required for the expression of uncoupling protein 3 (Ucp3), hexokinase 2 (Hk2), and pyruvate dehydrogenase kinase 4 (Pdk4), but not the expression of peroxisome proliferator-activated receptor gamma coactivator 1-alpha (Pgc-1α). Global Nox4 deletion resulted in decreased UCP3 protein expression and impaired glucose and fatty acid oxidization in response to acute exercise. Furthermore, Nox4-deficient mice demonstrated impaired adaptation to chronic exercise as measured by the time to exhaustion and activity of skeletal muscle citrate synthase and beta-hydroxyacyl-coA-dehydrogenase. Importantly, mice deficient in endothelial-Nox4 similarly demonstrated attenuated glucose and fatty acid oxidation following acute exercise. We report that H 2 O 2 and Nox4 promote immediate responses to exercise in skeletal muscle. Glucose and fatty acid oxidation were blunted in the Nox4-deficient mice post-exercise, potentially through regulation of UCP3 expression. Our data demonstrate that endothelial-Nox4 is required for glucose and fatty acid oxidation, suggesting inter-tissue cross-talk between the endothelium and skeletal muscle in response to exercise. • Nox4 and hydrogen peroxide (H 2 O 2) activate genes involved in skeletal muscle metabolism after acute exercise. • Global deletion of Nox4 attenuates muscle glucose and fatty acid oxidation post-acute exercise. • Endothelial-specific deletion of Nox4 blunts acute adaptation to exercise in muscle suggesting inter-tissue cross-talk. • Nox4 is required for chronic exercise adaptation. [ABSTRACT FROM AUTHOR]

The article presents a study which examines the patho-physiological role of aldosterone-mineralocorticoid receptor (MR) in the development of retinal vasculopathy. It cites vascular endothelial growth factor (VEGF) implicated in vascular permeability and angiogenesis. It reveals that MR antagonism has vasculoprotective in the retina and serves as treatment for vascular retinopathies.