bims-noxint Biomed News
on NADPH oxidases in tumorigenesis
Issue of 2019‒12‒22
eleven papers selected by
Laia Caja Puigsubira
Uppsala University

  1. Free Radic Biol Med. 2019 Dec 11. pii: S0891-5849(19)32276-2. [Epub ahead of print]
    Xia D, Halder B, Godoy C, Chakraborty A, Singla B, Thomas E, Shuja JB, Kashif H, Miller L, Csanyi G, Sabbatini ME.
      Inflammatory disorders of the pancreas are divided into acute (AP) and chronic (CP) forms. Both states of pancreatitis are a result of pro-inflammatory mediators, including reactive oxygen species (ROS). One of the sources of ROS is NADPH oxidase (Nox). The rodent genome encodes Nox1-4, Duox1 and Duox2. Our purpose was to assess the extent to which Nox enzymes contribute to the pathogenesis of both AP and CP using Nox-deficient mice. Using RT-PCR, Nox1 was found in both isolated mouse pancreatic acini and pancreatic stellate cells (PaSCs). Subsequently, mice with genetically deleted Nox1 were further studied and showed that the histo-morphologic characteristics of caerulein-induced CP, but not caerulein-induced AP, was ameliorated in Nox1 KO mice. We also found that the lack of Nox1 impaired caerulein-induced ROS generation in PaSCs. Using Western blotting, we found that AKT mediates the fibrotic effect of Nox1 in a mouse model of CP. We also found a decrease in phospho-ERK and p38MAPK levels in Nox1 KO mice with CP, but not with AP. Both CP-induced TGF-β up-regulation and NF-ĸB activation were impaired in pancreas from Nox1 KO mice. Western blotting indicated increases in proteins involved in fibrosis and acinar-to-ductal metaplasia in WT mice with CP. No change in those proteins were observed in Nox1 KO mice. The lack of Nox1 lowered mRNA levels of CP-induced matrix metalloproteinase MMP-9 and E-cadherin repressor Twist in PaSCs. CONCLUSION: Nox1-derived ROS in PaSCs mediate the fibrotic process of CP by activating the downstream redox-sensitive signaling pathways AKT and NF-ĸB, up-regulating MMP-9 and Twist, and producing α-smooth muscle actin and collagen I and III.
    Keywords:  Chronic pancreatitis; MMP-9; NADPH oxidase 1; NF-ĸB; Twist
  2. Redox Biol. 2019 Dec 05. pii: S2213-2317(19)31342-4. [Epub ahead of print] 101399
    Dei Zotti F, Verdoy R, Brusa D, Lobysheva II, Balligand JL.
      Oxidative stress perturbs vascular homeostasis leading to endothelial dysfunction and cardiovascular diseases. Vascular reactive oxygen species (ROS) reduce nitric oxide (NO) bioactivity, a hallmark of cardiovascular and metabolic diseases. We measured steady-state vascular NO levels through the quantification of heme nitrosylated hemoglobin (5-coordinate-α-HbNO) in venous erythrocytes of healthy human subjects using electron paramagnetic resonance (EPR) spectroscopy. To examine how ROS may influence HbNO complex formation and stability, we identified the pro- and anti-oxidant enzymatic sources in human erythrocytes and their relative impact on intracellular redox state and steady-state HbNO levels. We demonstrated that pro-oxidant enzymes such as NADPH oxidases are expressed and produce a significant amount of ROS at the membrane of healthy erythrocytes. In addition, the steady-state levels of HbNO were preserved when NOX (e.g. NOX1 and NOX2) activity was inhibited. We next evaluated the impact of selective antioxidant enzymatic systems on HbNO stability. Peroxiredoxin 2 and catalase, in particular, played an important role in endogenous and exogenous H2O2 degradation, respectively. Accordingly, inhibitors of peroxiredoxin 2 and catalase significantly decreased erythrocyte HbNO concentration. Conversely, steady-state levels of HbNO were preserved upon supplying erythrocytes with exogenous catalase. These findings support HbNO measurements as indicators of vascular oxidant stress and of NO bioavailability and potentially, as useful biomarkers of early endothelial dysfunction.
    Keywords:  Endothelial dysfunction; Erythrocytes; Nitric oxide; Reactive oxygen species
  3. Chem Biol Interact. 2019 Dec 12. pii: S0009-2797(19)31182-2. [Epub ahead of print] 108921
    Chen Z, Gao H, Wang L, Ma X, Tian L, Zhao W, Li K, Zhang Y, Ma F, Lu J, Jia L, Yang Y, Fu R.
      Hyperproliferation and oxidative stress induced by hyperglycemia in mesangial cells plays crucial roles in the pathological process of diabetic nephropathy. Farrerol, isolated from rhododendron leaves, possesses broad anti-oxidative and anti-inflammatory properties towards several diseases, but its role in diabetic neuropathy remains unclear. The aim of this study was to evaluate the effects of farrerol in high glucose induced mesangial cell injury, and to explore underlying molecular mechanisms. Our results showed that high glucose in vitro conditions significantly stimulated cell proliferation, inflammatory cytokine secretion, extracellular matrix deposition, excessive oxidative stress, and NADPH oxidase activity in mesangial cells. Levels of NADPH oxidase 4 (Nox4) expression, ERK1/2 phosphorylation, and TGF-β1/Smad2 activation were significantly induced by high glucose conditions in mesangial cells. Inversely, farrerol treatments at 40, 60, and 80 μM concentrations, dose-dependently alleviated this molecular damage by high glucose in mesangial cells. We also found that restoration of Nox4 expression abolished the protective effects of farrerol on high glucose-induced proliferation and reactive oxygen species generation. Furthermore, pretreatment with the Nox4 inhibitor diphenyliodonium or the ERK1/2 pathway inhibitor PD98059, displayed similar ameliorated effects of farrerol on high glucose-induced mesangial cell damage. Taken together, these data suggest that farrerol displays protective effects on high glucose induced mesangial cell injury, partly through the Nox4-mediated ROS/ERK1/2 signaling pathway. These observations may provide novel insights into the application of farrerol as a diabetic neuropathy treatment.
    Keywords:  Diabetic neuropathy; Farrerol; High glucose; Mesangial cells; Nox4; ROS
  4. J Endocrinol. 2019 Dec 01. pii: JOE-19-0258.R2. [Epub ahead of print]
    Wang HF, Yu QQ, Zheng RF, Xu M.
      Cardiovascular complications of type 2 diabetes mellitus (T2DM) are associated with vascular remodeling in the arteries. Perivascular sympathetic neurons release an abundance of trophic factors to regulate vascular function via a paracrine signaling. Netrin-1, a diffusible protein that can be secreted outside the cell, is one of common signals of 'conversation' between nerve and vessel. The present study investigated whether netrin-1 is a novel modulator of sympathetic neurons paracrine signaling and played a critical role in vascular adventitial remodeling under T2DM. Vascular adventitial remodeling was observed in adventitial fibroblasts (AFs) responding to netrin-1 deficiency in the supernatant from primary rat superior cervical ganglia (SCG) neurons, shown as AFs proliferation, migration, and collagen deposition. Conditioned medium from the high glucose (HG)-treated SCG neurons contributed to AFs remodeling, which was effectively alleviated by exogenous netrin-1 supplementation. Further, it was found that uncoordinated-5-B (UNC5b) was mainly expressed in AFs among netrin-1 specific receptors. Treatment of netrin-1 inhibited H2O2 production derived from NADPH oxidase 4 (Nox4) through the UNC5b/cAMP/PKA signal pathway in AFs remodeling. In vivo, aorta adventitial remodeling was accompanied with the down-regulation of netrin-1 in the perivascular sympathetic nerve in T2DM rats. Such abnormalities were restored by netrin-1 intervention, which was associated with the inhibition of Nox4 expression in the aorta adventitia. In conclusion, netrin-1 is a novel modulator of sympathetic neurons paracrine signaling to maintain AFs function. Vascular adventitial remodeling was aggravated by sympathetic neurons paracrine signaling under hyperglycemia, which was ameliorated by netrin-1 treatment through the UNC5b/cAMP/PKA/Nox4 pathway.
  5. Cancer Res. 2019 Dec 20. pii: canres.1920.2019. [Epub ahead of print]
    Robinson AJ, Hopkins GL, Rastogi N, Hodges M, Doyle M, Davies S, Hole PS, Omidvar N, Darley RL, Tonks A.
      Acute myeloid leukemia (AML) is a heterogeneous clonal disorder with a poor clinical outcome. Previously we showed that overproduction of reactive oxygen species (ROS), arising from constitutive activation of NOX2 oxidase, occurs in >60% of AML patients and that ROS production promotes proliferation of AML cells. We show here that the process most significantly affected by ROS overproduction is glycolysis. Whole metabolome analysis of 20 human primary AML showed that blasts generating high levels of ROS have increased glucose uptake and correspondingly increased glucose metabolism. In support of this, exogenous ROS increased glucose consumption whilst inhibition of NOX2 oxidase decreased glucose consumption. Mechanistically, ROS promoted uncoupling protein 2 (UCP2) protein expression and phosphorylation of AMPK, upregulating the expression of a key regulatory glycolytic enzyme, 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (PFKFB3). Overexpression of PFKFB3 promoted glucose uptake and cell proliferation, whilst downregulation of PFKFB3 strongly suppressed leukemia growth both in vitro and in vivo in the NSG model. These experiments provide direct evidence that oxidase-derived ROS promotes the growth of leukemia cells via the glycolytic regulator PFKFB3. Targeting PFKFB3 may therefore present a new mode of therapy for this disease with a poor outcome.
  6. Brain Behav Immun. 2019 Dec 10. pii: S0889-1591(19)30648-8. [Epub ahead of print]
    Huang WY, Liu KH, Lin S, Chen TY, Tseng CY, Chen HY, Wu HM, Hsu KS.
      BACKGROUND: Research indicates that sepsis increases the risk of developing cognitive impairment. After systemic inflammation, a corresponding activation of microglia is rapidly induced in the brain, and multiple neurotoxic factors, including inflammatory mediators (e.g., cytokines) and reactive oxygen species (e.g., superoxide), are also released that contribute to neuronal injury. NADPH oxidase (NOX) enzymes play a vital role in microglial activation through the generation of superoxide anions. We hypothesized that NOX isoforms, particularly NOX2, could exhibit remarkable abilities in developing cognitive deficits induced by systemic inflammation.METHODS: Mice with deficits of NOX2 organizer p47phox (p47phox-/-) and wild-type (WT) mice treated with the NOX inhibitor diphenyleneiodonium (DPI) were used in this study. Intraperitoneal lipopolysaccharide (LPS) injection was used to induce systemic inflammation. Spatial learning and memory were compared among treatment groups using the radial arm maze task. Brain tissues were collected for evaluating the transcript levels of proinflammatory cytokines, whereas immunofluorescence staining and immunoblotting were conducted to determine the percentage of activated glia (microglia and astroglia) and damaged neurons and the expression of synaptic proteins and BDNF.
    RESULTS: Cognitive impairment induced by systemic inflammation was significantly attenuated in the p47phox-/- mice compared to that in the WT mice. The p47phox-/- mice exhibited reduced microglial and astroglial activation and neuronal damage and attenuated the induction of multiple proinflammatory cytokines, including tumor necrosis factor-α, interleukin (IL)-1β, IL-6, and CCL2. Similar to that observed in the p47phox-/- mice, the administration of DPI significantly attenuated the cognitive impairment, reduced the glial activation and brain cytokine concentrations, and restored the expression of postsynaptic proteins (PSD-95) and BDNF in neurons and astrocytes, compared to those in the vehicle-treated controls within 10 days after LPS injection.
    CONCLUSIONS: This study clearly demonstrates that NOX2 contributes to glial activation with subsequent reduction in the expression of BDNF, synaptic dysfunction, and cognitive deficits after systemic inflammation in an LPS-injected mouse model. Our results provide evidence that NOX2 might be a promising pharmacological target that could be used to protect against synaptic dysregulation and cognitive impairment following systemic inflammation.
    Keywords:  Brain-derived neurotrophic factor; Cognition; Cytokine; Diphenyleneiodonium; Lipopolysaccharide; Neuroinflammation; Nicotinamide adenine dinucleotide phosphate oxidase; Sepsis
  7. Neuropharmacology. 2019 Dec 06. pii: S0028-3908(19)30478-2. [Epub ahead of print] 107907
    Eastman CL, D'Ambrosio R, Ganesh T.
      Traumatic brain injury (TBI) is a leading cause of death and disability in young adults worldwide. TBI survival is associated with persistent neuropsychiatric and neurological impairments, including posttraumatic epilepsy (PTE). To date, no pharmaceutical treatment has been found to prevent PTE or ameliorate neurological/neuropsychiatric deficits after TBI. Brain trauma results in immediate mechanical damage to brain cells and blood vessels that may never be fully restored given the limited regenerative capacity of brain tissue. This primary insult unleashes cascades of events, prominently including neuroinflammation and massive oxidative stress that evolve over time, expanding the brain injury, but also clearing cellular debris and establishing homeostasis in the region of damage. Accumulating evidence suggests that oxidative stress and neuroinflammatory sequelae of TBI contribute to posttraumatic epileptogenesis. This review will focus on possible roles of reactive oxygen species (ROS), their interactions with neuroinflammation in posttraumatic epileptogenesis, and emerging therapeutic strategies after TBI. We propose that inhibitors of the professional ROS-generating enzymes, the NADPH oxygenases and myeloperoxidase alone, or combined with selective inhibition of cyclooxygenase mediated signaling may have promise for the treatment or prevention of PTE and other sequelae of TBI.
    Keywords:  Myeloperoxidase and traumatic brain injury; NADPH oxidase; Neuroinflammation; Oxidative-stress; Posttraumatic epilepsy; Redox-signaling
  8. Diabetes Metab Syndr Obes. 2019 ;12 2597-2608
    Ma TK, Xu L, Lu LX, Cao X, Li X, Li LL, Wang X, Fan QL.
      Purpose: This study aimed to investigate whether ursolic acid (UA) mitigates renal inflammation, oxidative stress and fibrosis by regulating the angiotensin II type 1 receptor-associated protein (ARAP1)/angiotensin II type 1 receptor (AT1R) signaling pathway and subsequently alleviating renal damage.Methods: db/db mice were divided randomly into a diabetic nephropathy (DN) group and a UA treatment group. Light microscopy and electron microscopy were used to observe pathological changes in renal tissues. Immunohistochemistry (IHC) was employed to examine changes in the expression of ARAP1, AT1R, 8-hydroxydeoxyguanosine (8-OHdG), NADPH oxidase 2 (NOX2), the extracellular matrix protein fibronectin (FN), IL-1β and IL-18 in renal tissues. Western blotting and RT-qPCR were used to detect the respective changes in the protein and mRNA levels of ARAP1, AT1R, NOX4, NOX2, transforming growth factor-β1 (TGF-β1), FN, collagen IV, IL-1β and IL-18 in renal tissues and mesangial cells. In addition, immunofluorescence staining was employed to examine changes in FN and NOX2 expression in mesangial cells.
    Results: UA treatment effectively reduced the body weights and blood glucose levels of db/db mice (p<0.05) as well as the urinary albumin/creatinine ratio (p<0.05). In addition, the renal tissue lesions and glomerulosclerosis index of the db/db mice were significantly improved after treatment (p<0.01). Histochemical analysis results showed significantly lower expression levels of ARAP1, AT1R, FN, NOX2, 8-OHdG, IL-1β and IL-18 in renal tissues in the UA treatment group than in the DN group. Western blotting and RT-qPCR data also revealed UA-induced decreases in the renal levels of the ARAP1, AT1, NOX4, NOX2, TGF-β1, FN, collagen IV, IL-1β and IL-18 proteins in vivo and/or in vitro (p<0.01). ARAP1 knockdown effectively reduced the expression of NOX2 and FN in vitro.
    Conclusion: UA alleviated renal damage in type 2 diabetic db/db mice by downregulating proteins in the ARAP1/AT1R signaling pathway to inhibit extracellular matrix accumulation, renal inflammation, fibrosis and oxidative stress.
    Keywords:  ARAP1; AT1R; diabetic nephropathy; oxidative stress; renal fibrosis; ursolic acid
  9. J Neurosurg. 2019 Dec 20. pii: 2019.9.JNS191789. [Epub ahead of print] 1-12
    Datzmann T, Kapapa T, Scheuerle A, McCook O, Merz T, Unmuth S, Hoffmann A, Mathieu R, Mayer S, Mauer UM, Röhrer S, Yilmazer-Hanke D, Möller P, Nussbaum BL, Calzia E, Gröger M, Hartmann C, Radermacher P, Wepler M.
      OBJECTIVE: Acute subdural hematoma (ASDH) is a leading entity in brain injury. Rodent models mostly lack standard intensive care, while large animal models frequently are only short term. Therefore, the authors developed a long-term, resuscitated porcine model of ASDH-induced brain injury and report their findings.METHODS: Anesthetized, mechanically ventilated, and instrumented pigs with human-like coagulation underwent subdural injection of 20 mL of autologous blood and subsequent observation for 54 hours. Continuous bilateral multimodal brain monitoring (intracranial pressure [ICP], cerebral perfusion pressure [CPP], partial pressure of oxygen in brain tissue [PbtO2], and brain temperature) was combined with intermittent neurological assessment (veterinary modified Glasgow Coma Scale [MGCS]), microdialysis, and measurement of plasma protein S100β, GFAP, neuron-specific enolase [NSE], nitrite+nitrate, and isoprostanes. Fluid resuscitation and continuous intravenous norepinephrine were targeted to maintain CPP at pre-ASDH levels. Immediately postmortem, the brains were taken for macroscopic and histological evaluation, immunohistochemical analysis for nitrotyrosine formation, albumin extravasation, NADPH oxidase 2 (NOX2) and GFAP expression, and quantification of tissue mitochondrial respiration.
    RESULTS: Nine of 11 pigs survived the complete observation period. While ICP significantly increased after ASDH induction, CPP, PbtO2, and the MGCS score remained unaffected. Blood S100β levels significantly fell over time, whereas GFAP, NSE, nitrite+nitrate, and isoprostane concentrations were unaltered. Immunohistochemistry showed nitrotyrosine formation, albumin extravasation, NOX2 expression, fibrillary astrogliosis, and microglial activation.
    CONCLUSIONS: The authors describe a clinically relevant, long-term, resuscitated porcine model of ASDH-induced brain injury. Despite the morphological injury, maintaining CPP and PbtO2 prevented serious neurological dysfunction. This model is suitable for studying therapeutic interventions during hemorrhage-induced acute brain injury with standard brain-targeted intensive care.
    Keywords:  ASDH = acute subdural hematoma; CPP = cerebral perfusion pressure; GFAP; I/E = inspiratory/expiratory; ICP = intracranial pressure; MAP-2; MGCS = modified Glasgow Coma Scale; NO = nitric oxide; NOX2 = NADPH oxidase 2; NSE = neuron-specific enolase; PEEP = positive end-expiratory pressure; PbtO2 = partial pressure of oxygen in brain tissue; ROS = reactive oxygen species; mitochondrial respiration; multimodal brain monitoring; oxidative stress; trauma
  10. Free Radic Biol Med. 2019 Dec 12. pii: S0891-5849(19)32193-8. [Epub ahead of print]
    Liu J, Iwata K, Zhu K, Matsumoto M, Matsumoto K, Asaoka N, Zhang X, Ibi M, Katsuyama M, Tsutsui M, Kato S, Yabe-Nishimura C.
      The involvement of reactive oxygen species (ROS) has been suggested in the development of inflammatory bowel disease (IBD). An impaired intestinal barrier function is common in IBD patients. Here, we report the central role of NOX1/NADPH oxidase, a major source of ROS in nonphagocytic cells, in intestinal barrier dysfunction. By in vivo imaging using L-012 as a probe, a time-dependent increase in ROS was demonstrated in the abdomen of wild-type mice (WT) administered lipopolysaccharide (LPS: 6 mg/kg i.p.), but it was almost completely abolished in mice deficient in Nox1 (Nox1-KO) or the inducible nitric oxide synthase gene (iNOS-KO). By ex vivo imaging, increased ROS production was mainly shown in the ileum, where enhanced immunostaining of NOX1 was observed on the apical side of the epithelium. On the other hand, a punctate staining pattern of 3-nitrotyrosine, a marker of peroxynitrite production, was demonstrated in the lamina propria. When LPS-induced intestinal hyperpermeability was assessed by the oral administration of fluorescein isothiocyanate-conjugated dextran (FD-4), it was significantly suppressed in Nox1-KO as well as iNOS-KO. When Nox1-KO adoptively transferred with WT bone marrow were treated with LPS, the serum level of FD-4 was significantly elevated, whereas it remained unchanged in WT receiving bone marrow derived from Nox1-KO. Concomitantly, the activation of matrix metalloproteinase-9 induced by LPS was alleviated not only in intestinal tissue but also in peritoneal macrophages of Nox1-KO. Up-regulation of iNOS by LPS was significantly inhibited in macrophages deficient in Nox1, illustrating a functional hierarchy in NOX1/iNOS signaling. Together, these findings suggest that NOX1 in bone marrow-derived cells, but not epithelial cells, perturbs intestinal barrier integrity during endotoxemia.
  11. Brain Sci. 2019 Dec 15. pii: E378. [Epub ahead of print]9(12):
    Tong Y, Elkin KB, Peng C, Shen J, Li F, Guan L, Ji Y, Wei W, Geng X, Ding Y.
      Phenothiazine treatment has been shown to reduce post-stroke ischemic injury, though the underlying mechanism remains unclear. This study sought to confirm the neuroprotective effects of phenothiazines and to explore the role of the NOX (nicotinamide adenine dinucleotide phosphate oxidase)/Akt/PKC (protein kinase C) pathway in cerebral apoptosis. Sprague-Dawley rats underwent middle cerebral artery occlusion (MCAO) for 2 h and were randomly divided into 3 different cohorts: (1) saline, (2) 8 mg/kg chlorpromazine and promethazine (C+P), and (3) 8 mg/kg C+P as well as apocynin (NOX inhibitor). Brain infarct volumes were examined, and cell death/NOX activity was determined by assays. Western blotting was used to assess protein expression of kinase C-δ (PKC-δ), phosphorylated Akt (p-Akt), Bax, Bcl-XL, and uncleaved/cleaved caspase-3. Both C+P and C+P/NOX inhibitor administration yielded a significant reduction in infarct volumes and cell death, while the C+P/NOX inhibitor did not confer further reduction. In both treatment groups, anti-apoptotic Bcl-XL protein expression generally increased, while pro-apoptotic Bax and caspase-3 proteins generally decreased. PKC protein expression was decreased in both treatment groups, demonstrating a further decrease by C+P/NOX inhibitor at 6 and 24 h of reperfusion. The present study confirms C+P-mediated neuroprotection and suggests that the NOX/Akt/PKC pathway is a potential target for efficacious therapy following ischemic stroke.
    Keywords:  chlorpromazine and promethazine (C+P); middle cerebral artery occlusion (MCAO); neuroprotection; reperfusion