bims-noxint 2020-11-22 papers

bims-noxint

Biomed News

on NADPH oxidases in tumorigenesis

Issue of 2020–11–22
six papers selected by
Laia Caja Puigsubira, Uppsala University

Neuronal NADPH oxidase 2 regulates growth cone guidance downstream of slit2/robo2.
Nox4 Facilitates TGFβ1-Induced Fibrotic Response in Human Tenon's Fibroblasts and Promotes Wound Collagen Accumulation in Murine Model of Glaucoma Filtration Surgery.
Nox2 Upregulation and p38α MAPK Activation in Right Ventricular Hypertrophy of Rats Exposed to Long-Term Chronic Intermittent Hypobaric Hypoxia.
Endothelium-specific deletion of Nox4 delays retinal vascular development and mitigates pathological angiogenesis.
2,5-hexanedione induces NLRP3 inflammasome activation and neurotoxicity through NADPH oxidase-dependent pathway.
Interplay between Nox2 Activity and Platelet Activation in Patients with Sepsis and Septic Shock: A Prospective Study.

Dev Neurobiol. 2020 Nov 15.

Neuronal NADPH oxidase 2 regulates growth cone guidance downstream of slit2/robo2.

Aslihan Terzi, Haley Roeder, Cory J Weaver, Daniel M Suter.

  NADPH oxidases (Nox) are membrane-bound multi-subunit protein complexes producing reactive oxygen species (ROS) that regulate many cellular processes. Emerging evidence suggests that Nox-derived ROS also control neuronal development and axonal outgrowth. However, whether Nox act downstream of receptors for axonal growth and guidance cues is presently unknown. To answer this question, we cultured retinal ganglion cells (RGCs) derived from zebrafish embryos and exposed these neurons to netrin-1, slit2, and brain-derived neurotrophic factor (BDNF). To test the role of Nox in cue-mediated growth and guidance, we either pharmacologically inhibited Nox or investigated neurons from mutant fish that are deficient in Nox2. We found that slit2-mediated growth cone collapse, and axonal retraction was eliminated by Nox inhibition. Though we did not see an effect of either BDNF or netrin-1 on growth rates, growth in the presence of netrin-1 was reduced by Nox inhibition. Furthermore, attractive and repulsive growth cone turning in response to gradients of BDNF, netrin-1, and slit2, respectively, were eliminated when Nox was inhibited in vitro. ROS biosensor imaging showed that slit2 treatment increased growth cone hydrogen peroxide levels via mechanisms involving Nox2 activation. We also investigated the possible relationship between Nox2 and slit2/Robo2 signaling in vivo. astray/nox2 double heterozygote larvae exhibited decreased area of tectal innervation as compared to individual heterozygotes, suggesting both Nox2 and Robo2 are required for establishment of retinotectal connections. Our results provide evidence that Nox2 acts downstream of slit2/robo2 by mediating growth and guidance of developing zebrafish RGC neurons.

Keywords:  NADPH oxidase; Reactive oxygen species; Robo2; axon growth and guidance; hydrogen peroxide; optic chiasm; optic tectum; retinal ganglion cell; slit2

DOI:  https://doi.org/10.1002/dneu.22791
Antioxidants (Basel). 2020 Nov 13. pii: E1126. [Epub ahead of print]9(11):

Nox4 Facilitates TGFβ1-Induced Fibrotic Response in Human Tenon's Fibroblasts and Promotes Wound Collagen Accumulation in Murine Model of Glaucoma Filtration Surgery.

Manisha H Shah, Elsa C Chan, Nicole J Van Bergen, Surinder S Pandav, Sze Ng, Jonathan G Crowston, Hitesh M Peshavariya.

  Collagen accumulation in sub-conjunctival tissue at the surgical wound is one of the major complications associated with glaucoma filtration surgery (GFS). This process often leads to unwanted fibrotic scar formation at the lesion site and dysfunction of tissues. Previously, we demonstrated that NADPH oxidase 4 (Nox4) is implicated in transforming growth factor-beta (TGFβ)-induced collagen production in ocular fibroblasts and scarring responses in a mouse model of corneal injury. Here, we propose that Nox4 is an important facilitator of TGFβ-induced responses. We tested this hypothesis in human Tenon's fibroblasts (HTF) and also assessed a role of Nox4 in an experimental mouse model of GFS. TGFβ1 induced Nox4 mRNA expression but downregulated Nox5 in HTF. Targeting Nox4 gene expression with an adenovirus carrying a Nox4 small interfering RNA (siRNA) (Ad-Nox4i) or removal of hydrogen peroxide (H2O2) with EUK-134 (25 μM) in HTFs significantly reduced TGFβ1-induced Nox4 expression, H2O2 production, and collagen synthesis (p < 0.05, n = 3-6). SIS3 (5 μM) that prevents Smad3 phosphorylation is found to suppress TGFβ1-induced collagen production in HTFs. Furthermore, Ad-Nox4i and EUK-134 both abolished TGFβ1-stimulated proliferation of HTFs. We also compared collagen deposition at the wound arising from GFS between wildtype (WT) and Nox4 knockout (KO) mice. Both collagen deposition and fibrovascularization at the wound were significantly decreased in Nox4 KO mice at 14 days after GFS. Our results provide comprehensive evidence that Nox4 is an important mediator for TGFβ1-induced responses in HTFs and collagen deposition in surgical wound following GFS in mice. As such, pharmacological inhibition of Nox4 would be a viable therapeutic strategy for the control of scarring after glaucoma surgery.

Keywords:  NADPH Oxidase; collagen; glaucoma filtration surgery; reactive oxygen species; transforming growth factor-beta

DOI:  https://doi.org/10.3390/antiox9111126
Int J Mol Sci. 2020 Nov 13. pii: E8576. [Epub ahead of print]21(22):

Nox2 Upregulation and p38α MAPK Activation in Right Ventricular Hypertrophy of Rats Exposed to Long-Term Chronic Intermittent Hypobaric Hypoxia.

Eduardo Pena, Patricia Siques, Julio Brito, Silvia M Arribas, Rainer Böger, Juliane Hannemann, Fabiola León-Velarde, M Carmen González, M Rosario López, Ángel Luis López de Pablo.

  One of the consequences of high altitude (hypobaric hypoxia) exposure is the development of right ventricular hypertrophy (RVH). One particular type of exposure is long-term chronic intermittent hypobaric hypoxia (CIH); the molecular alterations in RVH in this particular condition are less known. Studies show an important role of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase complex-induced oxidative stress and protein kinase activation in different models of cardiac hypertrophy. The aim was to determine the oxidative level, NADPH oxidase expression and MAPK activation in rats with RVH induced by CIH. Male Wistar rats were randomly subjected to CIH (2 days hypoxia/2 days normoxia; n = 10) and normoxia (NX; n = 10) for 30 days. Hypoxia was simulated with a hypobaric chamber. Measurements in the RV included the following: hypertrophy, Nox2, Nox4, p22phox, LOX-1 and HIF-1α expression, lipid peroxidation and H2O2 concentration, and p38α and Akt activation. All CIH rats developed RVH and showed an upregulation of LOX-1, Nox2 and p22phox and an increase in lipid peroxidation, HIF-1α stabilization and p38α activation. Rats with long-term CIH-induced RVH clearly showed Nox2, p22phox and LOX-1 upregulation and increased lipid peroxidation, HIF-1α stabilization and p38α activation. Therefore, these molecules may be considered new targets in CIH-induced RVH.

Keywords:  cardiac hypertrophy; high altitude; kinases and NADPH oxidase; oxidative stress

DOI:  https://doi.org/10.3390/ijms21228576
Angiogenesis. 2020 Nov 17.

Endothelium-specific deletion of Nox4 delays retinal vascular development and mitigates pathological angiogenesis.

Xixiang Tang, Joshua J Wang, Jinli Wang, Hanna E Abboud, Yanming Chen, Sarah X Zhang.

  NADPH oxidase 4 (Nox4) is a major isoform of NADPH oxidases playing an important role in many biological processes. Previously we have shown that Nox4 is highly expressed in retinal blood vessels and is upregulated in oxygen-induced retinopathy (OIR). However, the exact role of endothelial Nox4 in retinal angiogenesis remains elusive. Herein, using endothelial cell (EC)-specific Nox4 knockout (Nox4EC-KO) mice, we investigated the impact of endothelial Nox4 deletion on retinal vascular development and pathological angiogenesis during OIR. Our results show that deletion of Nox4 in ECs led to retarded retinal vasculature development with fewer, blunted-end tip cells and sparser, dysmorphic filopodia at vascular front, and reduced density of vascular network in superficial, deep, and intermediate layers in postnatal day 7 (P7), P12, and P17 retinas, respectively. In OIR, loss of endothelial Nox4 had no effect on hyperoxia-induced retinal vaso-obliteration at P9 but significantly reduced aberrant retinal neovascularization at P17 and decreased the deep layer capillary density at P25. Ex vivo study confirmed that lack of Nox4 in ECs impaired vascular sprouting. Mechanistically, loss of Nox4 significantly reduced expression of VEGF, p-VEGFR2, integrin αV, angiopoietin-2, and p-ERK1/2, attenuating EC migration and proliferation. Taken together, our results indicate that endothelial Nox4 is important for retinal vascular development and contributes to pathological angiogenesis, likely through regulation of VEGF/VEGFR2 and angiopoietin-2/integrin αV/ERK pathways. In addition, our study suggests that endothelial Nox4 appears to be essential for intraretinal revascularization after hypoxia. These findings call for caution on targeting endothelial Nox4 in ischemic/hypoxic retinal diseases.

Keywords:  Angiogenesis; Development; Endothelial cells; Nox4; Retina; Vascularization

DOI:  https://doi.org/10.1007/s10456-020-09757-3
Free Radic Biol Med. 2020 Nov 16. pii: S0891-5849(20)31611-7. [Epub ahead of print]

2,5-hexanedione induces NLRP3 inflammasome activation and neurotoxicity through NADPH oxidase-dependent pathway.

Ruixue Huang, Liyan Hou, Xingyue Zhai, Zhengzheng Ruan, Wei Sun, Dongdong Zhang, Xiulan Zhao, Qingshan Wang.

  Chronic exposure to n-hexane causes sensorimotor neuropathy, which is mediated by 2,5-hexanedione (HD), a toxic metabolite of n-hexane. Activation of the nucleotide-binding and oligomerization domain, leucine-rich repeat, and pyrin domain-containing 3 (NLRP3) inflammasome is involved in multiple neurodegenerative diseases. However, whether the NLRP3 inflammasome contributes to HD-induced neurotoxicity remains unclear. In this study, the effects of HD on NLRP3 inflammasome activation and the underlying mechanisms were determined by using HD-treated rat and cell culture models. Increased NLRP3 expression, caspase-1 activation and interleukin-1β production were observed in both the brain and spinal cord of HD-treated rats. Double-immunofluorescence staining showed that ASC speck formation and caspase-1 expression were mainly localized in microglia. HD-induced activation of the NLRP3 inflammasome was further mirrored in BV2 microglial cells and was associated with NADPH oxidase activation. Interestingly, inhibition of NADPH oxidase by apocynin or specific siRNAs significantly mitigated HD-induced NLRP3 inflammasome activation. Furthermore, apocynin suppressed activation of the MAPK and NF-κB signaling pathways. Blocking activation of p38-MAPK and NF-κB significantly reduced HD-induced capase-1 activation and interleukin-1β maturation, indicating a critical role of NADPH oxidase and downstream MAPK and NF-κB pathways in regulating activation of NLRP3 inflammasome, in HD-treated microglia. Finally, we found that inhibition of microglial NLRP3 inflammasome and NADPH oxidase activation abrogated HD-induced microglial activation and neurodegeneration in both SHSY5Y neuronal cells and primary cortical neuron-glia cultures. Altogether, our findings suggest that NADPH oxidase-dependent activation of microglial NLRP3 inflammasome contributes to HD-induced neurotoxicity, providing novel insight into the mechanisms of this solvent-induced neuropathy.

Keywords:  Hexane; Microglia; NADPH oxidase; NLRP3 inflammasome

DOI:  https://doi.org/10.1016/j.freeradbiomed.2020.11.013
Oxid Med Cell Longev. 2020 ;2020 4165358

Interplay between Nox2 Activity and Platelet Activation in Patients with Sepsis and Septic Shock: A Prospective Study.

Giusy Tiseo, Elena Cavarretta, Arianna Forniti, Cristina Nocella, Sebastiano Sciarretta, Ornella Spagnolello, Enrico Baldini, Mariangela Peruzzi, Giuliano Bertazzoni, Francesco Menichetti, Antonino G M Marullo, Fabio Miraldi, Andrea Morelli, Giacomo Frati, Roberto Carnevale, Falcone Marco.

Background: Although preclinical studies highlighted the potential role of NADPH oxidase (NOX) in sepsis, only few studies evaluated the oxidative stress in patients with sepsis and septic shock. The objective of the study is to appraise the oxidative stress status and platelet function in patients with sepsis and septic shock compared to healthy controls.
Methods and Results: Patients with sepsis or septic shock admitted to the hospital Policlinico Umberto I (Sapienza University, Rome) underwent a blood sample collection within 1 hour from admission. Platelet aggregation, serum thromboxane B2 (TxB2), soluble NOX2-derived peptides (sNox2-dp), and hydrogen peroxide breakdown activity (HBA) were measured and compared to those of healthy volunteers. Overall, 33 patients were enrolled; of these, 20 (60.6%) had sepsis and 13 (39.4%) septic shock. Compared to healthy controls (n = 10, age 67.8 ± 3.2, male 50%), patients with sepsis and septic shock had higher platelet aggregation (49% (IQR 45-55), 60% (55.75-67.25), and 73% (IQR 69-80), respectively, p < 0.001), higher serum TxB2 (77.5 (56.5-86.25), 122.5 (114-131.5), and 210 (195-230) pmol/L, respectively, p < 0.001), higher sNox2-dp (10 (7.75-12), 19.5 (17.25-21), and 33 (29.5-39) pg/mL, respectively, p < 0.001), and lower HBA (75% (67.25-81.5), 50% (45-54.75), and 27% (21.5-32.5), respectively, p < 0.001). Although not statistically significant, a trend in higher levels of serum TxB2 and sNox2-dp in patients who died was observed.
Conclusions: Patients with septic shock exhibit higher Nox2 activity and platelet activation than patients with sepsis. These insights joined to better knowledge of these mechanisms could guide the identification of future prognostic biomarkers and new therapeutic strategies in the scenario of septic shock.

DOI: https://doi.org/10.1155/2020/4165358