Macrophage NADPH oxidase activation, impaired cholesterol fluxes, and increased cholesterol biosynthesis in diabetic mice: A stimulatory role for d-glucose

Efficient mechanisms of H+ ion extrusion are crucial for normal NADPH oxidase function. However, whether the NADPH oxidase—in analogy with mitochondrial cytochromes—has an inherent H+ channel activity remains uncertain: electrophysiological studies did not find altered H+ currents in cells from patients with chronic granulomatous disease (CGD), challenging earlier reports in intact cells. In this study, we describe the presence of two different types of H+ currents in human eosinophils. The “classical” H+ current had properties similar to previously described H+ conductances and was present in CGD cells. In contrast, the “novel” type of H+ current had not been described previously and displayed unique properties: (a) it was absent in cells from gp91- or p47-deficient CGD patients; (b) it was only observed under experimental conditions that allowed NADPH oxidase activation; (c) because of its low threshold of voltage activation, it allowed proton influx and cytosolic acidification; (d) it activated faster and deactivated with slower and distinct kinetics than the classical H+ currents; and (e) it was ∼20-fold more sensitive to Zn2+ and was blocked by the histidine-reactive agent, diethylpyrocarbonate (DEPC). In summary, our results demonstrate that the NADPH oxidase or a closely associated protein provides a novel type of H+ conductance during phagocyte activation. The unique properties of this conductance suggest that its physiological function is not restricted to H+ extrusion and repolarization, but might include depolarization, pH-dependent signal termination, and determination of the phagosomal pH set point.

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Olfactomedin 4 contributes to hydrogen peroxide-induced NADPH oxidase activation and apoptosis in mouse neutrophils

AJP Cell Physiology ◽

10.1152/ajpcell.00336.2017 ◽

2018 ◽

Vol 315 (4) ◽

pp. C494-C501 ◽

Cited By ~ 3

Author(s):

Wenli Liu ◽

Yueqin Liu ◽

Hongzhen Li ◽

Griffin P. Rodgers

Keyword(s):

Hydrogen Peroxide ◽

Nadph Oxidase ◽

Molecular Mechanisms ◽

Oxidase Inhibitor ◽

Superoxide Production ◽

Wild Type ◽

Escherichia Coli Bacteria ◽

Induced Apoptosis ◽

Nadph Oxidase Activation ◽

Deficient Mice

Neutrophils increase production of reactive oxygen species, including superoxide, hydrogen peroxide (H2O2), and hydroxyl radical, to destroy invading microorganisms under pathological conditions. Conversely, oxidative stress conditions, such as the presence of H2O2, induce neutrophil apoptosis, which helps to remove neutrophils after inflammation. However, the detailed molecular mechanisms that are involved in the latter process have not been elucidated. In this study, we investigated the potential role of olfactomedin 4 (Olfm4) in H2O2-induced superoxide production and apoptosis in mouse neutrophils. We have demonstrated that Olfm4 is not required for maximal-dosage PMA- and Escherichia coli bacteria-induced superoxide production, but Olfm4 contributes to suboptimal-dosage PMA- and H2O2-induced superoxide production. Using an NADPH oxidase inhibitor and gp91phox-deficient mouse neutrophils, we found that NAPDH oxidase was required for PMA-stimulated superoxide production and that Olfm4 mediated H2O2-induced superoxide production through NADPH oxidase, in mouse neutrophils. We have shown that neutrophils from Olfm4-deficient mice exhibited reduced H2O2-induced apoptosis compared with neutrophils from wild-type mice. We also demonstrated that neutrophils from Olfm4-deficient mice exhibited reduced H2O2-stimulated mitochondrial damage and membrane permeability, and as well as reduced caspase-3 and caspase-9 activity, compared with neutrophils from wild-type mice. Moreover, the cytoplasmic translocation of the proapoptotic mitochondrial proteins Omi/HtrA2 and Smac/DIABLO in response to H2O2was reduced in neutrophils from Olfm4-deficient mice compared with neutrophils from wild-type mice. Our study demonstrates that Olfm4 contributes to H2O2-induced NADPH oxidase activation and apoptosis in mouse neutrophils. Olfactomedin 4 might prove to be a potential target for future studies on inflammatory neutrophil biology and for inflammatory disease treatment.

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P2X7 receptor is critical in α-synuclein–mediated microglial NADPH oxidase activation

Neurobiology of Aging ◽

10.1016/j.neurobiolaging.2015.03.015 ◽

2015 ◽

Vol 36 (7) ◽

pp. 2304-2318 ◽

Cited By ~ 51

Author(s):

Tianfang Jiang ◽

Jake Hoekstra ◽

Xin Heng ◽

Wenyan Kang ◽

Jianqing Ding ◽

...

Keyword(s):

Nadph Oxidase ◽

P2x7 Receptor ◽

Nadph Oxidase Activation

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Protective Role of Endogenous Kallistatin in Vascular Injury and Senescence by Inhibiting Oxidative Stress and Inflammation

Oxidative Medicine and Cellular Longevity ◽

10.1155/2018/4138560 ◽

2018 ◽

Vol 2018 ◽

pp. 1-8 ◽

Cited By ~ 10

Author(s):

Julie Chao ◽

Youming Guo ◽

Lee Chao

Keyword(s):

Oxidative Stress ◽

Endothelial Cells ◽

Nadph Oxidase ◽

Vascular Injury ◽

Oxidase Activity ◽

Sirtuin 1 ◽

Mouse Lung ◽

Heparin Binding ◽

Diabetic Mice ◽

Nadph Oxidase Activity

Kallistatin was identified in human plasma as a tissue kallikrein-binding protein and a serine proteinase inhibitor. Kallistatin exerts pleiotropic effects on angiogenesis, oxidative stress, inflammation, apoptosis, fibrosis, and tumor growth. Kallistatin levels are markedly reduced in patients with coronary artery disease, sepsis, diabetic retinopathy, inflammatory bowel disease, pneumonia, and cancer. Moreover, plasma kallistatin levels are positively associated with leukocyte telomere length in young African Americans, indicating the involvement of kallistatin in aging. In addition, kallistatin treatment promotes vascular repair by increasing the migration and function of endothelial progenitor cells (EPCs). Kallistatin via its heparin-binding site antagonizes TNF-α-induced senescence and superoxide formation, while kallistatin’s active site is essential for inhibiting miR-34a synthesis, thus elevating sirtuin 1 (SIRT1)/eNOS synthesis in EPCs. Kallistatin inhibits oxidative stress-induced cellular senescence by upregulating Let-7g synthesis, leading to modulate Let-7g-mediated miR-34a-SIRT1-eNOS signaling pathway in human endothelial cells. Exogenous kallistatin administration attenuates vascular injury and senescence in association with increased SIRT1 and eNOS levels and reduced miR-34a synthesis and NADPH oxidase activity, as well as TNF-α and ICAM-1 expression in the aortas of streptozotocin- (STZ-) induced diabetic mice. Conversely, endothelial-specific depletion of kallistatin aggravates vascular senescence, oxidative stress, and inflammation, with further reduction of Let-7g, SIRT1, and eNOS and elevation of miR-34a in mouse lung endothelial cells. Furthermore, systemic depletion of kallistatin exacerbates aortic injury, senescence, NADPH oxidase activity, and inflammatory gene expression in STZ-induced diabetic mice. These findings indicate that endogenous kallistatin displays a novel role in protection against vascular injury and senescence by inhibiting oxidative stress and inflammation.

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Function of wild-type or mutant Rac2 and Rap1a GTPases in differentiated HL60 cell NADPH oxidase activation

Blood ◽

10.1182/blood.v85.3.804.bloodjournal853804 ◽

1995 ◽

Vol 85 (3) ◽

pp. 804-811 ◽

Cited By ~ 68

Author(s):

TG Gabig ◽

CD Crean ◽

PL Mantel ◽

R Rosli

Keyword(s):

Plasma Membrane ◽

Nadph Oxidase ◽

Cell Line ◽

Cell Lines ◽

Wild Type ◽

Free System ◽

Hl60 Cells ◽

Cell Free System ◽

Nadph Oxidase Activation ◽

O2 Production

Studies of neutrophil nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activation in a cell-free system showed that the low molecular-weight guanosine triphosphatase (GTPase) Rac was required, and that Rap1a may participate in activation of the catalytic complex. Full-length posttranslationally modified Rac2 was active, whereas only the 1–166 truncated form of Rap1a was functional in the cell-free system, and thus, clarification of the function of Rap1a and Rac2 in intact human phagocytes is needed to provide further insight into their roles as signal transducers from plasma membrane receptors. In the present studies, oligonucleotide-directed mutagenesis was used to introduce a series of mutations into human rap1a or rac2 in the mammalian expression vector pSR alpha neo. HL60 cells transfected with wild-type or mutated rac2 or rap1a cDNA constructs and control HL60 cells transfected with the pSR alpha neo vector containing no inserted cDNA were selected in G418-containing media, then subclones were isolated. Compared with the parent HL60 cells, each of the stable transfected cell lines differentiated similarly into neutrophil-like cells and expressed comparable levels of NADPH oxidase components p47- phox, p67-phox and gp91-phox. The differentiated vector control cell line produced O2. in response to receptor stimulation at rates that were not significantly different from parent HL60 cells. O2-. production by differentiated cell lines expressing mutated N17 Rap1a or N17 Rac2 dominant-negative proteins was inhibited, whereas O2-. production by the subline overexpressing wild-type Rap1a was increased by fourfold. O2-. production by the differentiated cell line expressing GTPase-defective V12 Rap1a was also significantly inhibited, a finding that is consistent with a requirement for cycling between guanosine diphosphate- and GTP-bound forms of Rap1a for continuous NADPH oxidase activation in intact neutrophils. A model is proposed in which Rac2 mediates assembly of the p47 and p67 oxidase components on the cytosolic face of the plasma membrane via cytoskeletal reorganization, whereas Rap1a functions downstream as the final activation switch involving direct physical interaction with the transmembrane flavocytochrome component of the NADPH oxidase.

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Calpain activation mediates microgravity-induced myocardial abnormalities in mice via p38 and ERK1/2 MAPK pathways

Journal of Biological Chemistry ◽

10.1074/jbc.ra119.011890 ◽

2020 ◽

Vol 295 (49) ◽

pp. 16840-16851

Author(s):

Liwen Liang ◽

Huili Li ◽

Ting Cao ◽

Lina Qu ◽

Lulu Zhang ◽

...

Keyword(s):

Nadph Oxidase ◽

Simulated Microgravity ◽

Heart Weight ◽

Calpain Inhibitor ◽

Protective Effects ◽

Tail Suspension ◽

Calpain Activation ◽

Microgravity Conditions ◽

Underlying Mechanisms ◽

Nadph Oxidase Activation

The human cardiovascular system has adapted to function optimally in Earth's 1G gravity, and microgravity conditions cause myocardial abnormalities, including atrophy and dysfunction. However, the underlying mechanisms linking microgravity and cardiac anomalies are incompletely understood. In this study, we investigated whether and how calpain activation promotes myocardial abnormalities under simulated microgravity conditions. Simulated microgravity was induced by tail suspension in mice with cardiomyocyte-specific deletion of Capns1, which disrupts activity and stability of calpain-1 and calpain-2, and their WT littermates. Tail suspension time-dependently reduced cardiomyocyte size, heart weight, and myocardial function in WT mice, and these changes were accompanied by calpain activation, NADPH oxidase activation, and oxidative stress in heart tissues. The effects of tail suspension were attenuated by deletion of Capns1. Notably, the protective effects of Capns1 deletion were associated with the prevention of phosphorylation of Ser-345 on p47phox and attenuation of ERK1/2 and p38 activation in hearts of tail-suspended mice. Using a rotary cell culture system, we simulated microgravity in cultured neonatal mouse cardiomyocytes and observed decreased total protein/DNA ratio and induced calpain activation, phosphorylation of Ser-345 on p47phox, and activation of ERK1/2 and p38, all of which were prevented by calpain inhibitor-III. Furthermore, inhibition of ERK1/2 or p38 attenuated phosphorylation of Ser-345 on p47phox in cardiomyocytes under simulated microgravity. This study demonstrates for the first time that calpain promotes NADPH oxidase activation and myocardial abnormalities under microgravity by facilitating p47phox phosphorylation via ERK1/2 and p38 pathways. Thus, calpain inhibition may be an effective therapeutic approach to reduce microgravity-induced myocardial abnormalities.

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