Oxidase Nox Research Articles

Blocking endothelial TRPV4-Nox2 interaction helps reduce ROS production and inflammation, and improves vascular function in obese mice

Obesity induces inflammation and oxidative stress, and ultimately leads to vasodilatory dysfunction in which Transient receptor potential vanilloid type 4 (TRPV4) and Nicotinamide Adenine Dinucleotide Phosphate Oxidase (Nox2) have been reported to be involved. However, little attention has been paid to the role of the TRPV4-Nox2 complex in these problems. The purpose of this study was to figure out the role of the TRPV4-Nox2 complex in obesity-induced inflammation, oxidative stress, and vasodilatory dysfunction. Using fluorescence resonance energy transfer and immunoprecipitation assays, we found enhanced TRPV4 and Nox2 interactions in obese mice. Using q-PCR, fluorescent dye dihydroethidium staining, and myotonic techniques, we found that obesity caused inflammation, oxidative stress, and vasodilatory dysfunction. Using adeno-associated viruses, we found that enhancement or attenuation of TRPV4-Nox2 interaction altered the vaso-function. Based on these findings, we found a small-molecule drug, M12, that interrupted the TRPV4-Nox2 interaction, thereby reducing inflammatory factors and reactive oxygen species production and helping to restore the vasodilatory function. In summary, our results revealed a new mechanism by which obesity-induced inflammation, oxidative stress, and vasodilatory dysfunction is caused by enhanced TRPV4-Nox2 interactions. Using M12 to interrupt the TRPV4-Nox2 interaction may have anti-inflammatory and anti-oxidative stress effects and help restore vasodilatory function and thus provide a new therapeutic approach to obesity.

Microglial Nox2 Plays a Key Role in the Pathogenesis of Experimental Autoimmune Encephalomyelitis.

While oxidative stress has been linked to multiple sclerosis (MS), the role of superoxide-producing phagocyte NADPH oxidase (Nox2) in central nervous system (CNS) pathogenesis remains unclear. This study investigates the impact of Nox2 gene ablation on pro- and anti-inflammatory cytokine and chemokine production in a mouse experimental autoimmune encephalomyelitis (EAE) model. Nox2 deficiency attenuates EAE-induced neural damage and reduces disease severity, pathogenic immune cells infiltration, demyelination, and oxidative stress in the CNS. The number of autoreactive T cells, myeloid cells, and activated microglia, as well as the production of cytokines and chemokines, including GM-CSF, IFNγ, TNFα, IL-6, IL-10, IL-17A, CCL2, CCL5, and CXCL10, were much lower in the Nox2−/− CNS tissues but remained unaltered in the peripheral lymphoid organs. RNA-seq profiling of microglial transcriptome identified a panel of Nox2 dependent proinflammatory genes: Pf4, Tnfrsf9, Tnfsf12, Tnfsf13, Ccl7, Cxcl3, and Cxcl9. Furthermore, gene ontology and pathway enrichment analyses revealed that microglial Nox2 plays a regulatory role in multiple pathways known to be important for MS/EAE pathogenesis, including STAT3, glutathione, leukotriene biosynthesis, IL-8, HMGB1, NRF2, systemic lupus erythematosus in B cells, and T cell exhaustion signaling. Taken together, our results provide new insights into the critical functions performed by microglial Nox2 during the EAE pathogenesis, suggesting that Nox2 inhibition may represent an important therapeutic target for MS.

RESVERATROL PROTECT AGAINST HOMOCYSTEINE-INDUCED RENAL DAMAGE THROUGH ANTIOXIDANT ACTION IN SPONTANEOUS HYPERTENSIVE RATS

Objective: The aim of this study was to investigate the effect of resveratrol in renal damage and the underlying mechanism induced by Hhcy in SHRs. Design and method: Twenty-four SHRs were divided into control group, Hhcy group, Hhcy + resveratrol (Hhcy+Res) group. Hhcy groups were given intraperitoneal injection of homocysteine. In addition, the Hhcy + Res group was given resveratrol by gavage. The Systolic blood pressure (SBP), diastolic blood pressure (DBP), plasma Hcy, serum malondialdehyde (MDA), superoxide dismutase (SOD) and the urinary microalbumin (UACR) were measured. To quantify RNA and protein expression levels, the mRNA and the protein expression of nephrin and NADPH oxidase subunits NOX2 and NOX4 in the kidney were examined by quantitative Real-time PCR and western blot. Renal histological or pathological changes were observed under light and electron microscope. Results: There was no difference in blood pressure before and after Hcy or Res intervention among groups. The homocysteine levels in Hhcy and Res group were significantly higher than that in control group, which showed that the model was successfully built by intraperitoneal injection. The serum SOD level was significantly decreased in Hhcy group, while it was increased in Res group. On the contrary, the serum MDA and UACR levels of model group were significantly increased. after the intervention of resveratrol, the MDA and UACR levels were reduced. Furthermore, the expression levels of mRNA and the protein of NOX2 and NOX4 were elevated while those of the nephrin were reduced in Hhcy group. Resveratrol significantly reduced the expressions of NOX2 and NOX4, and increased the expression of nephrin in renal tissue of SHRs with Hhcy. Conclusions: These results suggested that resveratrol may improve renal damage in hypertensive rats with Hhcy by exerting oxidative stress beyond blood pressure.

The NADPH oxidase NOX4 regulates metabolism in hepatocellular carcinoma cells

The NADPH oxidase NOX4 regulates metabolism in hepatocellular carcinoma cells

LRRC8A contributes to angiotensin II-induced cardiac hypertrophy by interacting with NADPH oxidases via the C-terminal leucine-rich repeat domain

Cardiac hypertrophy, an important cause of heart failure, is characterized by an increase in heart weight, the ventricular wall, and cardiomyocyte volume. The volume regulatory anion channel (VRAC) is an important regulator of cell volume. However, its role in cardiac hypertrophy remains unclear. The purpose of this study was to investigate the effect of leucine-rich repeat-containing 8A (LRRC8A), an essential component of the VRAC, on angiotensin II (AngII)-induced cardiac hypertrophy. Our results showed that LRRC8A expression, NADPH oxidase activity, and reactive oxygen species (ROS) production were increased in AngII-induced hypertrophic neonatal mouse cardiomyocytes and the myocardium of C57/BL/6 mice. In addition, AngII activated VRAC currents in cardiomyocytes. The delivery of adeno-associated viral (AAV9) bearing siRNA against mouse LRRC8A into the left ventricular wall inhibited AngII-induced cardiac hypertrophy and fibrosis. Accordingly, the knockdown of LRRC8A attenuated AngII-induced cardiomyocyte hypertrophy and VRAC currents in vitro. Furthermore, knockdown of LRRC8A suppressed AngII-induced ROS production, NADPH oxidase activity, the expression of NADPH oxidase membrane-bound subunits Nox2, Nox4, and p22phox, and the translocation of NADPH oxidase cytosolic subunits p47phox and p67phox. Immunofluorescent staining showed that LRRC8A co-localized with NADPH oxidase membrane subunits Nox2, Nox4, and p22phox. Co-immunoprecipitation and analysis of a C-terminal leucine-rich repeat domain (LRRD) mutant showed that LRRC8A physically interacts with Nox2, Nox4, and p22phox via the LRRD. Taken together, the results of this study suggested that LRRC8A might play an important role in promoting AngII-induced cardiac hypertrophy by interacting with NADPH oxidases via the LRRD.

Knockdown of Salusin-β Improves Cardiovascular Function in Myocardial Infarction-Induced Chronic Heart Failure Rats.

Salusin-β is a biologically active peptide with 20 amino acids that exerts several cardiovascular activity-regulating effects, such as regulating vascular endothelial function and the proliferation of vascular smooth muscle cells. However, the regulatory effects of salusin-β in myocardial infarction-induced chronic heart failure (CHF) are still unknown. The current study is aimed at investigating the effects of silencing salusin-β on endothelial function, cardiac function, vascular and myocardial remodeling, and its underlying signaling pathways in CHF rats induced by coronary artery ligation. CHF and sham-operated (Sham) rats were subjected to tail vein injection of adenoviral vectors encoding salusin-β shRNA or a control-shRNA. The coronary artery (CA), pulmonary artery (PA), and mesenteric artery (MA) were isolated from rats, and isometric tension measurements of arteries were performed. Compared with Sham rats, the plasma salusin-β, leptin and visfatin levels and the salusin-β protein expression levels of CA, PA, and MA were increased, while the acetylcholine- (ACh-) induced endothelium-dependent vascular relaxation of CA, PA, and MA was attenuated significantly in CHF rats and was improved significantly by salusin-β gene knockdown. Salusin-β knockdown also improved cardiac function and vascular and myocardial remodeling, increased endothelial nitric oxide synthase (eNOS) activity and nitric oxide (NO) levels, and decreased NAD(P)H oxidase activity, NOX-2 and NOX-4 expression, and reactive oxygen species (ROS) levels in arteries in CHF rats. The effects of salusin-β knockdown in CHF rats were attenuated significantly by pretreatment with the NOS inhibitor L-NAME. These results indicate that silencing salusin-β contributes to the improvement of endothelial function, cardiac function, and cardiovascular remodeling in CHF by inhibiting NAD(P)H oxidase-ROS generation and activating eNOS-NO production.

NADPH oxidase 4 (Nox4) deletion accelerates liver regeneration in mice

Liver is a unique organ in displaying a reparative and regenerative response after acute/chronic damage or partial hepatectomy, when all the cell types must proliferate to re-establish the liver mass. The NADPH oxidase NOX4 mediates Transforming Growth Factor-beta (TGF-β) actions, including apoptosis in hepatocytes and activation of stellate cells to myofibroblasts. Aim of this work was to analyze the impact of NOX4 in liver regeneration by using two mouse models where Nox4 was deleted: 1) general deletion of Nox4 (NOX4−/−) and 2) hepatocyte-specific deletion of Nox4 (NOX4hepKO). Liver regeneration was analyzed after 2/3 partial hepatectomy (PH). Results indicated an earlier recovery of the liver-to-body weight ratio in both NOX4−/− and NOX4hepKO mice and an increased survival, when compared to corresponding WT mice. The regenerative hepatocellular fat accumulation and the parenchyma organization recovered faster in NOX4 deleted livers. Hepatocyte proliferation, analyzed by Ki67 and phospho-Histone3 immunohistochemistry, was accelerated and increased in NOX4 deleted mice, coincident with an earlier and increased Myc expression. Primary hepatocytes isolated from NOX4 deleted mice showed higher proliferative capacity and increased expression of Myc and different cyclins in response to serum. Transcriptomic analysis through RNA-seq revealed significant changes after PH in NOX4−/− mice and support a relevant role for Myc in a node of regulation of proliferation-related genes. Interestingly, RNA-seq also revealed changes in the expression of genes related to activation of the TGF-β pathway. In fact, levels of active TGF-β1, phosphorylation of Smads and levels of its target p21 were lower at 24 h in NOX4 deleted mice. Nox4 did not appear to be essential for the termination of liver regeneration in vivo, neither for the in vitro hepatocyte response to TGF-β1 in terms of growth inhibition, which suggest its potential as therapeutic target to improve liver regeneration, without adverse effects.

ABCG1 Attenuates Oxidative Stress Induced by H2O2 through the Inhibition of NADPH Oxidase and the Upregulation of Nrf2-Mediated Antioxidant Defense in Endothelial Cells

Summary. Oxidative stress is an important factor that is related to endothelial dysfunction. ATP-binding cassette transporter G1 (ABCG1), a regulator of intracellular cholesterol efflux, has been found to prevent endothelial activation in vessel walls. To explore the role of ABCG1 in oxidative stress production in endothelial cells, HUAECs were exposed to H2O2 and transfected with the specific ABCG1 siRNA or ABCG1 overexpression plasmid. The results showed that overexpression of ABCG1 by ABCG1 plasmid or liver X receptor (LXR) agonist T0901317 treatment inhibited ROS production and MDA content induced by H2O2 in HUAECs. Furthermore, ABCG1 upregulation blunted the activity of prooxidant NADPH oxidase and the expression of Nox4, one of the NADPH oxidase subunits. Moreover, the increased migration of Nrf2 from the cytoplasm to the nucleus and antioxidant HO-1 expression were detected in HUAECs with upregulation of ABCG1. Conversely, ABCG1 downregulation by ABCG1 siRNA increased NADPH oxidase activity and Nox4 expression and abrogated the increase at Nrf2 nuclear protein levels. In addition, intracellular cholesterol load interfered with the balance between NADPH oxidase activity and HO-1 expression. It was suggested that ABCG1 attenuated oxidative stress induced by H2O2 in endothelial cells, which might be involved in the balance between decreased NADPH oxidase activity and increased Nrf2/OH-1 antioxidant defense signaling via its regulation for intracellular cholesterol accumulation.

Sparged but not stirred: Rapid, ADH-NADH oxidase catalyzed deracemization of alcohols in a bubble column

We investigate in detail both the deracemization reaction rate and maximum conversion of (R,S)-1-phenylethanol in a bubble column and the deactivation rates of both enzymes involved, alcohol dehydrogenase (ADH) and NADH oxidase (nox2). Instead of the predicted ellipsoidal shape of the bubbles, we observe a spherical shape in all our experiments. Calculated values for the area-specific mass transfer coefficient kLa in sparged, but not stirred, solution ranged from 17 to 87 h−1. We find that minimizing the air–liquid interface over time results in the lowest rate of nox2 deactivation. The onset of nox2 dissociation beyond the initial 50 h correlates with a tenfold increase in the observed deactivation rate constant. The highest conversion (99%) for deracemizing 50 mM (R,S)-1-phenylethanol was reached in the shortest reaction time (6 h) if large air bubbles (diameter 2.6 mm) at slow flowrates (5 bubbles/s) were employed. A faster flowrate, which delivers more oxygen per unit time, increased the initial reaction rate but resulted in a slower rate at higher conversions and ultimately incomplete maximum conversion (94%). Reaction rates under sparging were 4–8 times faster than in quiescent solution. Lastly, stirring did not significantly aid conversion (41% vs 37% in quiescent solution after 12 h) but caused increased enzyme deactivation (44 h vs 136 h half-life quiescent solution).

Protective effect of carnosine on hydrogen peroxide–induced oxidative stress in human kidney tubular epithelial cells

Diabetic nephropathy (DN) endangers health and is a high financial public burden worldwide. Risk of DN is positively correlated with high levels of reactive oxygen species (ROS). Carnosine, an antioxidant, actively regulates cell function and has the potential to reduce the occurrence of DN. Here, we explored whether carnosine could prevent oxidative stress in human kidney tubular epithelial (HK2) cells and, if so, the mechanisms underlying this effect. HK2 cells were cultured with the ROS hydrogen peroxide (H2O2) for 24 h and then treated with carnosine. In H2O2-damaged HK2 cells, carnosine significantly increased cell viability, assessed using a Cell Counting Kit 8, increased total superoxide dismutase (T-SOD) activity, assessed using a T-SOD activity detection kit, but decreased ROS levels, assessed using a ROS-sensitive fluorescent probe. Western blotting analyses to determine the protein expression levels of BAX, BCL-2, caspase 3, and the NADPH oxidase isoforms NOX2 and NOX4, as well as confocal laser scanning microscopy to assess changes in the mitochondrial membrane potential and the relative position of mitochondria to cytochrome c, indicated that carnosine inhibited apoptosis via the mitochondrial pathway in H2O2-damaged HK2 cells. Significantly decreased NOX4 expression and increased T-SOD activity in the presence of carnosine reduced the production of intracellular ROS, relieving oxidative stress to inhibit apoptosis via the mitochondrial pathway. These findings provide molecular mechanistic insights underlying the effects of carnosine, particularly as a potential therapeutic in DN.

Anaerobic Respiration of NOX1-Derived Hydrogen Peroxide Licenses Bacterial Growth at the Colonic Surface

The colonic microbiota exhibits cross-sectional heterogeneity, but the mechanisms that govern its spatial organization remain incompletely understood. Here we used Citrobacter rodentium, a pathogen that colonizes the colonic surface, to identify microbial traits that license growth and survival in this spatial niche. Previous work showed that during colonic crypt hyperplasia, type III secretion system (T3SS)-mediated intimate epithelial attachment provides C.rodentium with oxygen for aerobic respiration. However, we find that prior to the development of colonic crypt hyperplasia, T3SS-mediated intimate attachment is not required for aerobic respiration but for hydrogen peroxide (H2O2) respiration using cytochrome c peroxidase (Ccp). The epithelial NADPH oxidase NOX1 is the primary source of luminal H2O2 early after C.rodentium infection and is required for Ccp-dependent growth. Our results suggest that NOX1-derived H2O2 is a resource that governs bacterial growth and survival in close proximity to the mucosal surface during gut homeostasis.

Idelalisib Rescues Natural Killer Cells from Monocyte-Induced Immunosuppression by Inhibiting NOX2-Derived Reactive Oxygen Species.

The phosphatidylinositol-4,5-bisphosphate-3 kinase-δ (PI3Kδ) inhibitor idelalisib, used alone or in combination with anti-CD20, is clinically efficacious in B-cell lymphoma and chronic lymphocytic leukemia (CLL) by promoting apoptosis of malignant B cells. PI3K regulates the formation of reactive oxygen species (ROS) by the myeloid NADPH oxidase NOX2, but the role of PI3Kδ in myeloid cell-induced immunosuppression is unexplored. We assessed the effects of idelalisib on the spontaneous and IgG antibody-induced ROS production by human monocytes, on ROS-induced cell death of human natural killer (NK) cells, and on tumor cell clearance in an NK cell-dependent mouse model of metastasis. Idelalisib potently and efficiently inhibited the formation of NOX2-derived ROS from monocytes and rescued NK cells from ROS-induced cell death. Idelalisib also promoted NK cell cytotoxicity against anti-CD20-coated primary human CLL cells and cultured malignant B cells. Experiments using multiple PI3K inhibitors implicated the PI3Kδ isoform in regulating NOX2-induced ROS formation and immunosuppression. In B6 mice, systemic treatment with idelalisib significantly reduced the formation of lung metastases from intravenously injected melanoma cells but did not affect metastasis in B6.129S6-Cybbtm1Din (Nox2 -/-) mice or in NK cell-deficient mice. Our results imply that idelalisib rescues NK cells from NOX2/ROS-dependent immunosuppression and thus exerts antineoplastic efficacy beyond B-cell inhibition.

NOX2-Derived Reactive Oxygen Species in Cancer.

The formation of reactive oxygen species (ROS) by the myeloid cell NADPH oxidase NOX2 is critical for the destruction of engulfed microorganisms. However, recent studies imply that ROS, formed by NOX2+ myeloid cells in the malignant microenvironment, exert multiple actions of relevance to the growth and spread of neoplastic cells. By generating ROS, tumor-infiltrating myeloid cells and NOX2+ leukemic myeloid cells may thus (i) compromise the function and viability of adjacent cytotoxic lymphocytes, including natural killer (NK) cells and T cells, (ii) oxidize DNA to trigger cancer-promoting somatic mutations, and (iii) affect the redox balance in cancer cells to control their proliferation and survival. Here, we discuss the impact of NOX2-derived ROS for tumorigenesis, tumor progression, regulation of antitumor immunity, and metastasis. We propose that NOX2 may be a targetable immune checkpoint in cancer.

MiR-673/menin/JunD axis modulates hyperglycemia-induced oxidative stress and inflammation in the diabetic heart

Abstract Background Hyperglycemia-induced reactive oxygen species generation in diabetic heart contributes to myocardial dysfunction. JunD, a member of the activated protein 1 (AP-1) family of transcription factors, is emerging as a major gatekeeper against oxidative stress. Previous studies have shown that downregulation of AP-1 transcription factor JunD is involved in vascular aging and heart failure. However, the role of JunD in diabetes-induced myocardial dysfunction is unknown. Purpose The present study was designed to investigate whether hyperglycemia-driven epigenetic regulation of JunD contributes to oxidative stress, inflammation and myocardial dysfunction in the diabetic heart. Methods Diabetes (DB) was induced in C57BL/6 wild-type (WT) mice by streptozotocin. After four weeks of DB, left ventricular (LV) function was assessed by standard and 2D speckle-tracking echocardiography in both groups (n=10). Then, the animals were euthanized and LV specimens were collected to determine JunD mRNA and protein expression as well as superoxide anion production by ESR spectroscopy. Chromatin modifications of JunD gene promoter were assessed by chromatin immunoprecipitation. Isolated DNA was analyzed for promoter methylation following Methylminer kit. Cardiac biopsies were collected from age-matched patients with and without diabetes. Results DB mice showed LV dysfunction with reduced ejection fraction and fractional shortening. JunD mRNA and protein expression were reduced in the myocardium of DB as compared to control mice. JunD downregulation was associated with oxidative stress, increased NF-kB binding activity and expression of inflammatory mediators. Accordingly, expression of free radical scavenger superoxide dismutase 1 and aldehyde dehydrogenase 2 was reduced, whereas nicotinamide adenine dinucleotide phosphate oxidase subunits NOX2 and NOX4 were upregulated in DB. A reduction of JunD mRNA and protein expression was confirmed in LV specimens obtained from patients with diabetes. The downregulation of JunD was epigenetically regulated by promoter hypermethylation and histone modifications. Post-translational repression by tumor suppressor menin also contributed to JunD downregulation. Indeed, menin was significantly upregulated in DB hearts and co-immunoprecipitation experiments confirmed the binding of menin to JunD. Furthermore, rat ventricular myocytes exposed to high glucose (HG) showed increased menin expression. We found that miR-673 targeting menin was downregulated in hearts of DB mice. Reprogramming miR-673 in HG-treated myocytes was able to restore both menin and JunD expression to control levels. Conclusions Our findings show that downregulation of AP-1 transcription factor JunD contributes to diabetes-induced myocardial dysfunction and miR-673/menin/JunD represents a novel molecular axis involved in hyperglycemia-induced ROS-driven cardiac damage. Funding Acknowledgement Type of funding source: Foundation. Main funding source(s): European Society of Cardiology (ESC) Research Grant 2017

Effects of late aerobic exercise on cardiac remodeling of rats with small myocardial infarction

Abstract Background Physical exercise has been highlighted as an important non-pharmacological therapy for prevention and treatment of several cardiovascular diseases. However, its effects on hearts with minor cardiac remodeling are not clear. Purpose To evaluate the influence of aerobic physical exercise on 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-EA). Rats exercised three times a week for 12 weeks on a treadmill. Echocardiogram was performed before and after experimental period. Infarction size and cardiomyocyte diameters were evaluated by histology. Gene expression was assessed by RT-PCR. Results Only rats with MI size lower than 30% of LV total area were included in the study. Functional capacity was higher in MI-AE than the other groups. 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. LV diastolic posterior wall thickness was higher in MI-AE than Sham, and relative wall thickness was lower in MI-SED than MI-AE and Sham groups. Cardiomyocyte diameter was smaller in infarcted groups than Sham. Myocardial gene expression of the NADPH oxidase subunits NOX2, NOX4, p22phox, and p47phox did not differ between groups. Conclusion Small-sized myocardial infarction changes cardiac structures and left ventricular systolic function. Late aerobic physical exercise improves functional capacity and cardiac remodeling by preserving left ventricular geometry. NADPH oxidase subunits gene expression is not involved in cardiac remodeling or modulated by aerobic exercise in rats with small myocardial infarction. Funding Acknowledgement Type of funding source: Public Institution(s). Main funding source(s): CNPq; CAPES

Alpha lipoic acid attenuates iron induced oxidative acute kidney injury in rats

ABSTRACT Iron has been implicated in oxidative tissue injury owing to its ability to generate reactive oxygen species (ROS). We investigated the reno-protective effects of alpha lipoic acid (ALA) by investigating its effects on the kidney isoform of NADPH oxidase (Nox4) and the specific signaling pathways, p38 MAPK and PI3K/Akt, which participate in apoptosis and survival, respectively. We established four groups of seven rats: control, 100 mg/kg ALA, 80 mg/kg iron sucrose (IS) and IS + ALA. IS and ALA were injected intravenously and rats were sacrificied after 6 h. The mRNA expression of the subunits of NADPH oxidase, Nox4 and p22phox; tumor necrosis factor-alpha (TNF-α); and kidney injury molecule-1 (KIM-1) were measured using quantitative real time polymerase chain reaction (qRT-PCR). Active caspase-3 protein expression was evaluated by immunostaining. Also, p38 MAPK and PI3K/Akt signaling pathways were analyzed using western blot. ALA suppressed the mRNA expression of Nox4, p22phox, TNF-α and KIM-1. Active caspase-3 protein expression induced by IS was decreased by ALA. ALA also suppressed p38 MAPK and activated the PI3K/Akt signaling pathway following IS administration. We found that ALA may be an effective strategy for preventing oxidative acute kidney injury caused by IS.

1α,25(OH)2D3 Radiosensitizes Cancer Cells by Activating the NADPH/ROS Pathway.

The radioresistance of tumors affect the outcome of radiotherapy. Accumulating data suggest that 1α,25(OH)2D3 is a potential anti-oncogenic molecule in various cancers. In the present study, we investigated the radiosensitive effects and underlying mechanisms of 1α,25(OH)2D3 in vitro and in vivo. We found that 1α,25(OH)2D3 enhanced the radiosensitivity of lung cancer and ovarian cancer cells by promoting the NADPH oxidase-ROS-apoptosis axis. Compared to the group that only received radiation, the survival fraction and self-renewal capacity of cancer cells treated with a combination of 1α,25(OH)2D3 and radiation were decreased. Both apoptosis and ROS were significantly increased in the combination group compared with the radiation only group. Moreover, N-acetyl-L-cysteine, a scavenger of intracellular ROS, reversed the apoptosis and ROS induced by 1α,25(OH)2D3, indicating that 1α,25(OH)2D3 enhanced the radiosensitivity of cancer cells in vitro by promoting ROS-induced apoptosis. Moreover, our results demonstrated that 1α,25(OH)2D3 promoted the ROS level via activating NADPH oxidase complexes, NOX4, p22phox, and p47phox. In addition, knockdown of the vitamin D receptor (VDR) abolished the radiosensitization of 1α,25(OH)2D3, which confirmed that 1α,25(OH)2D3 radiosensitized tumor cells that depend on VDR. Similarly, our study also evidenced that vitamin D3 enhanced the radiosensitivity of cancer cells in vivo and extended the overall survival of mice with tumors. In summary, these results demonstrate that 1α,25(OH)2D3 enhances the radiosensitivity depending on VDR and activates the NADPH oxidase-ROS-apoptosis axis. Our findings suggest that 1α,25(OH)2D3 in combination with radiation enhances lung and ovarian cell radiosensitivity, potentially providing a novel combination therapeutic strategy.

AS128 - The NADPH oxidase NOX4 regulates lipid metabolism, which contributes to its tumour suppressor actions in hepatocellular carcinoma

AS128 - The NADPH oxidase NOX4 regulates lipid metabolism, which contributes to its tumour suppressor actions in hepatocellular carcinoma

ROS Regulate Caspase-Dependent Cell Delamination without Apoptosis in the Drosophila Pupal Notum.

SummaryThorax fusion occurs in the midline of the Drosophila pupal notum and involves epithelial cell delamination requiring apoptotic signaling. By genetic screening, we found that NADPH oxidases (Nox and Duox) associated with superoxide anion (O˙-2) are responsible for caspase-3 activation and delamination. We observed that Nox is upregulated in cells that undergo delamination and that delamination depends on caspase activation. However, the cell morphology and the almost complete lack of propidium iodide incorporation suggested little membrane disruption and signified apoptotic modulation. These results demonstrate that most delaminating cells undergo caspase activation, but this activation is not sufficient for apoptosis. We showed that the expression of Catalase, encoding an H2O2 scavenger in the cytosol, increases delamination and induces apoptotic nuclear fragmentation in caspase-3-activated cells. These findings suggest that the roles of O˙-2 and intracellular H2O2 for delamination differs before and after caspase-3 activation, which involves live cell delamination.

Nox2 dependent redox-regulation of Akt and ERK1/2 to promote left ventricular hypertrophy in dietary obesity of mice

BackgroundA Nox2 containing NADPH oxidase (Nox2) is involved in the global oxidative stress found in dietary obesity and metabolic disorders. However, the effects of high fat diet (HFD) on cardiac Nox2 activation and signaling in left ventricular hypertrophy (LVH) remain unknown. MethodsLeft ventricular (LV) tissues isolated from C57BL/6J wild-type (WT) and Nox2 knockout (Nox2KO) mice (11 months old, n = 6 per group) after 4 months of HFD treatment were used. Cardiomyocyte sizes were measured digitally on LV cross-sections. The levels of cardiac reactive oxygen species (ROS) production was determined using lucigenin-chemiluminescence and in situ dihydroethidium (DHE) fluorescence. The levels of Nox subunit expression and redox signaling were examined by immunoblotting and immunofluorescence. ResultsIn comparison to WT normal chow diet control hearts, WT HFD hearts had 1.8-fold increases in cardiomyocyte size, a sign of cardiac hypertrophy, and this was accompanied with ≥2-fold increase in the levels of ROS production, Nox2 expression and the phosphorylation of Akt and ERK1/2. Increased ROS production measured in HFD heart homogenates was inhibited to control levels by Tiron (a cell membrane permeable O2•−scavenger), diphenyleneiodonium (DPI, a flavohaemoprotein inhibitor) and Nox2 ds-tat (a Nox2 assembly inhibitor). However, all of these abnormalities were significantly reduced or absent in Nox2KO hearts under the same HFD. ConclusionsNox2 activation in response to dietary obesity and metabolic disorders plays a key role in cardiac oxidative stress, aberrant redox signaling and cardiomyocyte hypertrophy. Knockout of Nox2 protects hearts from oxidative damage associated with obesity and metabolic disorders.