Tumor necrosis factor-alpha-induced nuclear factor-kappaB activation in human cardiomyocytes is mediated by NADPH oxidase

An elevated level of tumor necrosis factor (TNF)-α is implicated in several cardiovascular diseases including heart failure. Numerous reports have demonstrated that TNF- alfa activates nuclear factor (NF)-kappaB, resulting in the upregulation of several genes that regulate inflammation, proliferation, and apoptosis of cardiomyocytes. Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, a major source of reactive oxygen species (ROS), is also activated by TNF- alfa and plays a crucial role in redox-sensitive signaling pathways. The present study investigated whether NADPH oxidase mediates TNF-alfa-induced NF-kappaB activation and NF-kappaB-mediated gene expression. Human cardiomyocytes were treated with recombinant TNF- alfa with or without pretreatment with diphenyleneiodonium (DPI) and apocynin, inhibitors of NADPH oxidase. TNF-α-induced ROS production was measured using 5-(and-6)-chloromethyl-2', 7'-dichlorodihydrofluorescein diacetate assay. TNF- alfa-induced NF-kappaB activation was also examined using immunoblot; NF-kappaB binding to its binding motif was determined using a Cignal reporter luciferase assay and an electrophoretic mobility shift assay. TNF- alfa -induced upregulation of interleukin (IL)-1β and vascular cell adhesion molecule (VCAM)-1 was investigated using real-time PCR and immunoblot. TNF- alfa -induced ROS production in cardiomyocytes was mediated by NADPH oxidase. Phosphorylation of IKK- alfa /β and p65, degradation of IkappaBalfa, binding of NF-kappaB to its binding motif, and upregulation of IL-1β and VCAM-1 induced by TNF- alfa were significantly attenuated by treatment with DPI and apocynin. Collectively, these findings demonstrate that NADPH oxidase plays a role in regulation of TNF- alfa -induced NF-kappaB activation and upregulation of proinflammatory cytokines, IL-1β and VCAM-1, in human cardiomyocytes

The senescence-accelerated mouse prone-8 (SAM-P8) oxidative stress is associated with upregulation of renal NADPH oxidase system

Herein, we investigate whether the NADPH oxidase might be playing a key role in the degree of oxidative stress in the senescence-accelerated mouse prone-8 (SAM-P8). To this end, the activity and expression of the NADPH oxidase, the ratio of glutathione and glutathione disulfides (GSH/GSSG), and the levels of malonyl dialdehyde (MDA) and nitrotyrosine (NT) were determined in renal tissue from SAM-P8 mice at the age of 1 and 6 months. The senescence-accelerated-resistant mouse (SAM-R1) was used as control. At the age of 1 month, NADPH oxidase activity and Nox2 protein expression were higher in SAM-P8 than in SAM-R1 mice. However, we found no differences in the GSH/GSSG ratio, MDA, NT, and Nox4 levels between both groups of animals. At the age of 6 months, SAM-R1 mice in comparison to SAM-P8 mice showed an increase in NADPH oxidase activity, which is associated with higher levels of NT and increased Nox4 and Nox2 expression levels. Furthermore, we found oxidative stress hallmarks including depletion in GSH/GSSG ratio and increase in MDA levels in the kidney of SAM-P8 mice. Finally, NADPH oxidase activity positively correlated with Nox2 expression in all the animals (r = 0.382, P < 0.05). Taken together, our data allow us to suggest that an increase in NADPH oxidase activity might be an early hallmark to predict future oxidative stress in renal tissue during the aging process that takes place in SAM-P8 mice (AU)

Estallido respiratorio de los fagocitos / Phagocyte respiratory burst

El estallido respiratorio de los fagocitos, producido por la NADPH-oxidasa, complejo enzimático que cataliza la formación de radical superóxido, constituye una de las fuentes endógenas más importantes de especies reactivas del oxígeno en el organismo. Este sistema comprende un complejo flavocitocromo b558 unido a membrana, y factores citosolicos p47phox, p67phox, p40phox y la pequeña GTPasa Rac2, que se trastocan a la membrana plasmática donde experimentan un proceso de ensamblaje que conforma el sistema enzimático activo. El conocimiento de las interacciones proteina-proteína que permiten el ensamblaje y el mecanismo de acción enzimático, ha permitido detectar los cambios que transcurren en el estado activo. La importancia de la NADPH oxidasa se muestra en la enfermedad granulomatosa crónica, patología transmitida por herencia, en la cual un componente de la NADPH oxidasa está ausente o defectivo. Tales individuos padecen infecciones recurrentes crónicas y severas debido a la incapacidad de sus neutrófilos para destruir microbios

The phagocyte respiratory burst produced by the NADPH oxidase, enzyme complex which catalyzes the production of superoxide radical, is one of the main endogenous sources of reactive oxygen species in the body. NADPH oxidase consists of a membrane-bound flavocytochrome b558 complex, and cytosolic factors p47phox, p67phox, p40phox and the small GTPase Rac, which translocate to the membrane to assemble the active complex following cell activation. A great deal of current research involves understanding the protein protein interactions involved in the assembly and enzyme mechanism and how these change with the activation state. The importance of the NADPH oxidase is illustrated by the inherited condition Chronic Granulomatous Disease in which a component of the respiratory burst oxidase is absent or defective. Affected individuals suffer from recurrent, chronic and severe infections due to the inability of their neutrophils to kill microbes