bims-noxint Biomed News
on NADPH oxidases in tumorigenesis
Issue of 2019‒12‒15
eight papers selected by
Laia Caja Puigsubira
Uppsala University


  1. Oxid Med Cell Longev. 2019 ;2019 3585390
    Youm TH, Woo SH, Kwon ES, Park SS.
      Myoblast fusion is an essential step in skeletal muscle development and regeneration. NADPH oxidase 4 (Nox4) regulates cellular processes such as proliferation, differentiation, and survival by producing reactive oxygen species (ROS). Insulin-like growth factor 1 induces muscle hypertrophy via Nox4, but its function in myoblast fusion remains elusive. Here, we report a ROS-dependent role of Nox4 in myoblast differentiation. Regenerating muscle fibers after injury by cardiotoxin had a lower cross-sectional area in Nox4-knockout (KO) mice than myofibers in wild-type (WT) mice. Diameters and fusion index values of myotubes differentiated from Nox4-KO primary myoblasts were significantly lower than those of myotubes derived from WT myoblasts. However, no difference was observed in the differentiation index and expression of MyoD, myogenin, and myosin heavy chain 3 (MHC) between KO and WT myotubes. The decreased fusion index was also observed during differentiation of primary myoblasts and C2C12 cells with suppressed Nox4 expression. In contrast, in C2C12 cells overexpressing Nox4, the fusion index was increased, whereas the differentiation index and MHC and myogenin protein expression were not affected compared to control. Interestingly, the expression of myomaker (Tmem8c), a fusogenic protein that controls myoblast fusion, was reduced in Nox4-knockdown C2C12 cells. The myomaker expression level was proportional to the cellular ROS level, which was regulated by of Nox4 expression level. These results suggests that Nox4 contributes to myoblast fusion, possibly through the regulation of myomaker expression via ROS production, and that Nox4-dependent ROS may promote skeletal muscle regeneration and growth.
    DOI:  https://doi.org/10.1155/2019/3585390
  2. Cardiovasc Res. 2019 Dec 10. pii: cvz331. [Epub ahead of print]
    Schnelle M, Sawyer I, Anilkumar N, Mohamed BA, Richards DA, Toischer K, Zhang M, Catibog N, Sawyer G, Mongue-Din H, Schröder K, Hasenfuss G, Shah AM.
      AIMS: Chronic pressure or volume overload induce concentric versus eccentric left ventricular (LV) remodelling, respectively. Previous studies suggest that distinct signalling pathways are involved in these responses. NADPH oxidase-4 (Nox4) is a reactive oxygen species (ROS)-generating enzyme that can limit detrimental cardiac remodelling in response to pressure overload. This study aimed to assess its role in volume overload-induced remodelling.METHODS AND RESULTS: We compared the responses to creation of an aortocaval fistula (Shunt) to induce volume overload in Nox4-null mice (Nox4-/-) versus wild-type (WT) littermates. Induction of Shunt resulted in a significant increase in cardiac Nox4 mRNA and protein levels in WT mice as compared to Sham controls. Nox4-/- mice developed less eccentric LV remodelling than WT mice (echocardiographic relative wall thickness: 0.30 vs 0.27, p < 0.05), with less LV hypertrophy at organ level (increase in LV weight/tibia length ratio of 25% vs 43%, p < 0.01) and cellular level (cardiomyocyte cross-sectional area: 323 µm2 vs 379 μm2, p < 0.01). LV ejection fraction, foetal gene expression, interstitial fibrosis, myocardial capillary density and levels of myocyte apoptosis after Shunt were similar in the two genotypes. Myocardial phospho-Akt levels were increased after induction of Shunt in WT mice whereas levels decreased in Nox4-/- mice (+29% vs -21%, p < 0.05), associated with a higher level of phosphorylation of the S6 ribosomal protein (S6) and the eIF4E-binding protein 1 (4E-BP1) in WT compared to Nox4-/- mice. We identified that Akt activation in cardiac cells is augmented by Nox4 via a Src kinase-dependent inactivation of protein phosphatase 2A (PP2A).
    CONCLUSION: Endogenous Nox4 is required for the full development of eccentric cardiac hypertrophy and remodelling during chronic volume overload. Nox4-dependent activation of Akt and its downstream targets S6 and 4E-BP1 may be involved in this effect.
    TRANSLATIONAL PERSPECTIVE: Cardiac volume overload resulting, for example, from aortic or mitral regurgitation contributes to the development of heart failure. Its underlying pathophysiology differs from that of cardiac pressure overload. Our study identifies the reactive oxygen species generating enzyme NADPH oxidase-4 (Nox4) as an important regulator of volume overload-induced cardiac remodelling by promoting eccentric LV hypertrophy, an adaptive response to the increased volume. As Nox inhibition is currently being developed as a potential therapeutic approach for several human diseases (e.g. lung fibrosis), our findings highlight the importance of assessing the potential impact on cardiac function in patients with co-existent valvular regurgitation.
    DOI:  https://doi.org/10.1093/cvr/cvz331
  3. Antioxid Redox Signal. 2019 Dec 11.
    Chocry M, Leloup L.
      SIGNIFICANCE: The oxidative stress, resulting from an imbalance in the production and scavenging of reactive oxygen species (ROS), is known to be involved in the development and progression of several pathologies. The excess of ROS production is often due to an overactivation of NADPH oxidases (NOX) and for this reason these enzymes became promising therapeutic targets. However, even if NOX are now well characterized, the development of new therapies is limited by the lack of highly isoform-specific inhibitors. Recent Advances: In the last decade, several groups and laboratories have screened thousands of molecules to identify new specific inhibitors with low off-target effects. These works have led to the characterization of several new potent NOX inhibitors; however, their specificity varies a lot depending on the molecules.CRITICAL ISSUES: Here we are reviewing more than 25 known NOX inhibitors, focusing mainly on the newly identified ones such as APX-115, NOS31, Phox-I1 and 2, GLX7013114 and GSK2795039. To have a better overall view of these molecules, the inhibitors were classified according to their specificity, from pan-NOX inhibitors to highly isoform-specific ones. We are also presenting the use of these compounds in vitro and in vivo.
    FUTURE DIRECTIONS: Several of these new molecules are potent and very specific inhibitors that could be good candidates for the development of new drugs. Even if the results are very promising, most of these compounds were only validated in vitro or in mice models and further investigations will be required before using them as potential therapies.
    DOI:  https://doi.org/10.1089/ars.2019.7915
  4. Biochem Pharmacol. 2019 Dec 04. pii: S0006-2952(19)30444-7. [Epub ahead of print] 113745
    Yu J, Shi L, Lin W, Lu B, Zhao Y.
      Uncoupling protein 2 (UCP2) is a mitochondrial anion carrier which plays a key role in energy homeostasis. UCP2 is deregulated in several human cancers and has been suggested to regulate cancer metabolism. However, the role of UCP2 in gallbladder cancer has not been defined. Using clinical samples, we found highly expressed UCP2 in gallbladder cancer tissues, and higher expression levels of UCP2 correlated with worse clinical characteristics. To study whether UCP2 promotes gallbladder cancer growth, UCP2 stable knockdown cells were generated, and cell proliferation was suppressed in these knockdown cells. Further studies demonstrated that glycolysis was inhibited and IKKβ, as well as the downstream signaling molecules NF-κB/FAK/β-catenin, were downregulated in UCP2 knockdown cells. More importantly, gallbladder cancer cells became sensitive to gemcitabine treatments when UCP2 was inhibited. UCP2 knockdown suppressed the activation of the NF-κB/β-catenin axis and promoted the increases in mitochondrial ROS in gallbladder cancer cells exposed to gemcitabine treatments. The UCP2 inhibitor genipin suppressed xenograft tumor growth and sensitized grafted tumors to gemcitabine treatments. These results suggest targeting UCP2 as a novel therapeutic strategy for the treatment of gallbladder cancer.
    Keywords:  NF-κB; ROS; UCP2; chemotherapy resistance; gallbladder cancer; glycolysis; β-catenin
    DOI:  https://doi.org/10.1016/j.bcp.2019.113745
  5. Arch Biochem Biophys. 2019 Dec 05. pii: S0003-9861(19)30782-9. [Epub ahead of print] 108220
    França KC, Martinez PA, Prado ML, Lo SM, Borges BE, Zanata SM, San Martin A, Nakao LS.
      Quiescent and contractile VSMC can switch to proliferative and migratory phenotype in response to growth factors and cytokines, an effect underscored by Nox family NADPH oxidases, particularly Nox1. We previously showed that quiescin/sulfhydryl oxidase 1 (QSOX1) has a role in neointima formation in balloon-injured rat carotid. Here, we investigated the intracellular redox mechanisms underlying these effects in primary VSMC. Our results show that exogenous incubation with wild type QSOX1b (wt QSOX), or with secreted QSOX1, but not with the inactive C452S QSOX 1b (C452S QSOX) or secreted inactive C455S QSOX1, induces VSMC migration and chemotaxis. PEG-catalase (PEG-CAT) prevented, while PEG-superoxide dismutase (PEG-SOD) increased migration induced by wt QSOX. Moreover, wt QSOX-induced migration was abrogated in NOX1-null VSMC. In contrast, both wt QSOX and C452S QSOX, and both secreted QSOX1 and C455S QSOX1, induce cell proliferation. Such effect was unaltered by PEG-CAT, while being inhibited by PEG-SOD. However, QSOX1-induced proliferation was not significantly affected in NOX1-null VSMC, compared with WT VSMC. These results indicate that hydrogen peroxide and superoxide mediate, respectively, migration and proliferation. However, Nox1 was required only for QSOX1-induced migration. In parallel, QSOX1-induced proliferation was independent of its redox activity, although mediated by intracellular superoxide.
    Keywords:  Cell migration; Hydrogen peroxide; Nox1; Quiescin/sulfhydryl oxidase 1b; Smooth muscle cells; Superoxide
    DOI:  https://doi.org/10.1016/j.abb.2019.108220
  6. J Immunol Res. 2019 ;2019 1278301
    Lewis CV, Vinh A, Diep H, Samuel CS, Drummond GR, Kemp-Harper BK.
      Aims: To date, the ROS-generating capacities of macrophages in different activation states have not been thoroughly compared. This study is aimed at determining the nature and levels of ROS generated following stimulation with common activators of M1 and M2 macrophages and investigating the potential for this to impact fibrosis.Results: Human primary and THP-1 macrophages were treated with IFN-γ+LPS or IL-4-activating stimuli, and mRNA expression of established M1 (CXCL11, CCR7, IL-1β) and M2 (MRC-1, CCL18, CCL22) markers was used to confirm activation. Superoxide generation was assessed by L-012-enhanced chemiluminescence and was increased in both M(IFN-γ+LPS) and M(IL-4) macrophages, as compared to unpolarised macrophages (MΦ). This signal was attenuated with NOX2 siRNA. Increased expression of the p47phox and p67phox subunits of the NOX2 oxidase complex was evident in M(IFN-γ+LPS) and M(IL-4) macrophages, respectively. Amplex Red and DCF fluorescence assays detected increased hydrogen peroxide generation following stimulation with IL-4, but not IFN-γ+LPS. Coculture with human aortic adventitial fibroblasts revealed that M(IL-4), but not M(IFN-γ+LPS), enhanced fibroblast collagen 1 protein expression. Macrophage pretreatment with the hydrogen peroxide scavenger, PEG-catalase, attenuated this effect.
    Conclusion: We show that superoxide generation is not only enhanced with stimuli associated with M1 macrophage activation but also with the M2 stimulus IL-4. Macrophages activated with IL-4 also exhibited enhanced hydrogen peroxide generation which in turn increased aortic fibroblast collagen production. Thus, M2 macrophage-derived ROS is identified as a potentially important contributor to aortic fibrosis.
    DOI:  https://doi.org/10.1155/2019/1278301
  7. J Clin Immunol. 2019 Dec 08.
    Sacco KA, Smith MJ, Bahna SL, Buchbinder D, Burkhardt J, Cooper MA, Hartog NL, Kobrynski L, Patel KP, Abraham RS.
      PURPOSE: Chronic granulomatous disease (CGD) is an innate immune deficiency, primarily affecting the phagocytic compartment, and presenting with a diverse phenotypic spectrum ranging from severe childhood infections to monogenic inflammatory bowel disease. Dihydrorhodamine (DHR) flow cytometry is the standard diagnostic test for CGD, and correlates with NADPH oxidase activity. While there may be genotype correlation with the DHR flow pattern in some patients, in several others, there is no correlation. In such patients, assessment by flow cytometric evaluation of NADPH oxidase-specific (NOX) proteins provides a convenient and rapid means of genetic triage, though immunoblotting has long been used for this purpose.METHODS AND RESULTS: We describe the clinical utility of the NOX flow cytometry assay through assessment of X-linked and autosomal recessive CGD patients and their first-degree relatives. The assessment of specific NOX proteins was correlated with overall NADPH oxidase function (DHR flow), clinical phenotype and genotype. NOX-specific protein assessment is a valuable adjunct to DHR assessment and genotyping to classify and characterize CGD patients.
    CONCLUSIONS: The atypical clinical presentation of some CGD patients can make genotype-phenotype correlation with DHR flow data challenging. Genetic testing, while useful for confirmation of diagnosis, can take several weeks, and in some patients does not provide a conclusive answer. However, NADPH-oxidase-specific protein flow assessment offers a rapid alternative to identification of the underlying genetic defect in cellular subsets, and can be utilized as a reflex test to an abnormal DHR flow. Further, it can provide insight into correlation between oxidative burst relative to protein expression in granulocytes and monocytes.
    Keywords:  CYBA; CYBB; Chronic granulomatous disease; NADPH oxidase; NCF1; flow cytometry; gp91phox; p22phox; p47phox; primary immunodeficiencies
    DOI:  https://doi.org/10.1007/s10875-019-00712-6
  8. Nutrients. 2019 Dec 09. pii: E3006. [Epub ahead of print]11(12):
    Lai S, Molfino A, Testorio M, Perrotta AM, Currado A, Pintus G, Pietrucci D, Unida V, La Rocca D, Biocca S, Desideri A.
      INTRODUCTION: The gut microbiota has coevolved with humans for a mutually beneficial coexistence and plays an important role in health and disease. A dysbiotic gut microbiome may contribute to progression to chronic kidney disease (CKD) and CKD-related complications such as cardiovascular disease. Microbiota modulation through the administration of prebiotics may represent an important therapeutic target.AIM: We sought to evaluate the effects of a low-protein diet (LPD) (0.6 g/kg/day) with or without the intake of the prebiotic inulin (19 g/day) on microbiota and clinical parameters in CKD patients.
    MATERIALS AND METHODS: We performed a longitudinal, prospective, controlled, and interventional study on 16 patients: 9 patients treated with LPD (0.6 g/kg/day) and inulin (19 g/day) and 7 patients (control group) treated only with LPD (0.6 g/kg/day). Clinical evaluations were performed and fecal samples were collected for a subsequent evaluation of the intestinal microbiota in all patients. These tests were carried out before the initiation of LPD, with or without inulin, at baseline (T0) and at 6 months (T2). The microbiota of 16 healthy control (HC) subjects was also analyzed in order to identify potential dysbiosis between patients and healthy subjects.
    RESULTS: Gut microbiota of CKD patients was different from that of healthy controls. The LPD was able to significantly increase the frequencies of Akkermansiaceae and Bacteroidaceae and decrease the frequencies of Christensenellaceae, Clostridiaceae, Lactobacillaceae, and Pasteurellaceae. Only Bifidobacteriaceae were increased when the LPD was accompanied by oral inulin intake. We showed a significant reduction of serum uric acid (SUA) and C-reactive protein (CRP) in patients treated with LPD and inulin (p = 0.018 and p = 0.003, respectively), an improvement in SF-36 (physical role functioning and general health perceptions; p = 0.03 and p = 0.01, respectively), and a significant increase of serum bicarbonate both in patients treated with LPD (p = 0.026) or with LPD and inulin (p = 0.01). Moreover, in patients treated with LPD and inulin, we observed a significant reduction in circulating tumor necrosis factor alpha (TNF-α) (p = 0.041) and plasma nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX2) (p = 0.027) levels. We did not find a significant difference in the circulating levels of Interleukin (IL)-1β (p = 0.529) and IL-6 (p = 0.828) in the two groups.
    CONCLUSIONS: LPD, associated or not with inulin, modified gut microbiota and modulated inflammatory and metabolic parameters in patients with CKD. Our results suggest that interventions attempting to modulate the gut microbiome may represent novel strategies to improve clinical outcomes in CKD patients and may provide useful therapeutic effects.
    Keywords:  chronic kidney disease; endothelial dysfunction; inulin; low-protein diet; microbiota; prebiotic therapy
    DOI:  https://doi.org/10.3390/nu11123006