Cholesterol: A modulator of the phagocyte NADPH oxidase activity - A cell-free study

Abstract

The NADPH oxidase Nox2, a multi-subunit enzyme complex comprising membrane and cytosolic proteins, catalyzes a very intense production of superoxide ions O2•−, which are transformed into other reactive oxygen species (ROS). In vitro, it has to be activated by addition of amphiphiles like arachidonic acid (AA). It has been shown that the membrane part of phagocyte NADPH oxidase is present in lipid rafts rich in cholesterol. Cholesterol plays a significant role in the development of cardio-vascular diseases that are always accompanied by oxidative stress. Our aim was to investigate the influence of cholesterol on the activation process of NADPH oxidase. Our results clearly show that, in a cell-free system, cholesterol is not an efficient activator of NADPH oxidase like arachidonic acid (AA), however it triggers a basal low superoxide production at concentrations similar to what found in neutrophile. A higher concentration, if present during the assembly process of the enzyme, has an inhibitory role on the production of O2•−. Added cholesterol acts on both cytosolic and membrane components, leading to imperfect assembly and decreasing the affinity of cytosolic subunits to the membrane ones. Added to the cytosolic proteins, it retains their conformations but still allows some conformational change induced by AA addition, indispensable to activation of NADPH oxidase.

Source of Dietary Fat in Pig Diet Affects Adipose Expression of Genes Related to Cancer, Cardiovascular, and Neurodegenerative Diseases

“…This is in accordance with our transcriptomic results since beef tallow is highly hypercholesterolemic [33]. Specifically, it was observed that cholesterol at high concentrations triggers inhibition of the activity of NADPH oxidase in the presence of pro-inflammatory signals [32].…”

“…However, due to the difficulty of reliable quantitative ROS detection, this issue remains not well understood and requires further investigations [31]. Interestingly, recently, it has been shown that cholesterol is a modulator of the phagocyte NADPH oxidase activity [32]. This is in accordance with our transcriptomic results since beef tallow is highly hypercholesterolemic [33].…”

Titanium Dioxide Nanoparticles Increase Superoxide Anion Production by Acting on NADPH Oxidase

“…In this study, we have used a construction called trimera, which consisted of the following domains Nter-p47 phox (amino acids 1-286) linked to the N-ter p67 phox (amino acids 1-212), and the full length Rac1 Q61L [ 44 ]. In a previous paper, we have verified that the rates of production of superoxide anions were similar when the classic cytosolic proteins are replaced by trimera protein to activate the cyt b558 and also that the dependences of the activity in function of the enzyme activator AA concentration were also found comparable with the cytosolic fractions and the trimera [ 45 ]. Thus, the trimera was chosen in order to avoid complications due to some assembly steps and to activate the cyt b558 in a reproducible manner.…”

The physicochemical properties of membranes correlate with the NADPH oxidase activity

“…It has been reported that disruption of lipid rafts resulted in the downregulation of NADPH oxidase subunits in breast cancer cells [27] and in a compromised superoxide production [28], while lipid rafts have also been shown to maintain NADPH oxidase in an inactive state [29]. We could also observe that sterolenriched membranes decrease the rate of superoxide anion produced by recombinant NADPH oxidase [30,31]. This might be explained by different types of lipid raft redox platforms as described in [32].…”