NAD(P)H Oxidase–Derived Superoxide Mediates Hypercholesterolemia-Induced Leukocyte–Endothelial Cell Adhesion

Abstract

Experimental animals placed on a high-cholesterol diet for 2 or more weeks exhibit an inflammatory response in postcapillary venules. The aims of this study were to determine (1) whether superoxide mediates the hypercholesterolemia-induced inflammatory response and (2) whether leukocyte and/or vessel wall NAD(P)H oxidase contributes to this response. Intravital videomicroscopy was used to quantify leukocyte-endothelial cell adhesion in cremasteric postcapillary venules of wild-type (WT) mice, CuZn-superoxide dismutase transgenic (SOD TgN) mice, and mice heterozygous (p47(phox)+/-) or homozygous (p47(phox)-/-) for NAD(P)H oxidase placed on either a normal diet or high-cholesterol diet (HCD) for 2 weeks. The number of adherent and emigrated leukocytes in postcapillary venules of WT HCD mice was significantly higher than that detected in venules of their normal-diet counterparts. However, the HCD-induced recruitment of adherent and emigrated leukocytes was not observed in SOD TgN mice. Whereas hypercholesterolemic p47(phox)+/- and WT mice exhibited similar inflammatory responses, p47(phox)-/- mice did not. Bone marrow chimeras were developed to selectively delete p47(phox) from either the vessel wall or circulating leukocytes. Whereas WT marrow transplanted into WT mice produced a normal inflammatory response of venules to HCD, chimeric mice with p47(phox) deficiency in either the vessel wall or leukocytes exhibited an attenuated inflammatory response to HCD that was comparable with that observed in p47(phox)-/- HCD mice. Our findings indicate that enhanced superoxide production is a critical event that initiates the leukocyte-endothelial cell adhesion in postcapillary venules of HCD mice. NAD(P)H oxidase appears to be an important source of this superoxide.

Platelet-associated NAD(P)H oxidase contributes to the thrombogenic phenotype induced by hypercholesterolemia

“…Here we provide the first direct evidence that NAD(P)H oxidase mediates the arteriolar dysfunction that results from a moderate rise in blood cholesterol concentration. Previously we demonstrated that NAD(P)H oxidase-derived superoxide generated from blood cells and the vessel wall mediates leukocyte recruitment in postcapillary venules of hypercholesterolemic mice [11]. The present study extends these observations to demonstrate that platelet-associated NAD(P) H oxidase is a key factor that contributes to the prothrombogenic response that accompanies hypercholesterolemia.…”

“…Hypercholesterolemia elicits an inflammatory response in the microvasculature of many organs, including skeletal muscle, intestine, brain, and mesentery [3,4,11,12,14]. This inflammatory response is characterized by endothelial dysfunction/activation, and manifests as impaired endothelium-dependent vasodilation in arterioles [3,4], and leukocyte and platelet recruitment in postcapillary venules [4,14,22].…”

“…The left carotid artery was cannulated for systemic arterial pressure measurement. The cremaster muscle was prepared for intravital microscopy as described previously [11]. An upright multi-purpose microscope system (Zeiss, Thornwood, NY) with a 40X water immersion objective lens (Achroplan 40X/0.75 W) was used to observe the cremasteric microcirculation.…”

“…NAD(P)H oxidase-derived superoxide also appears to contribute to the recruitment of both leukocytes and platelets that is elicited in postcapillary venules by hypercholesterolemia [11,12]. The hypercholesterolemia-induced leukocyte adhesion involves superoxide generated from NAD(P)H oxidase that is expressed in the vessel wall as well as circulating blood cells.…”

Modulation of the Inflammatory Response in Cardiovascular Disease

“…37,38 Similarly, mice that are genetically deficient in critical protein subunits (eg, p47 phox ) of NAD(P)H oxidase or transgenic mice that overexpress SOD exhibit attenuated leukocyte adhesion responses in models of oxidative stress. 39 Studies demonstrating that treatment of otherwise normal animals with inhibitors of NOS 40 and endothelial NOS-knockout mice 41 exhibit increased leukocyte adhesion support the view that a critical determinant of whether the vasculature assumes a proinflammatory or an antiinflammatory (and therefore, a prothrombogenic or an antithrombogenic) phenotype is the balance between ROS and NO. Although NO and superoxide per se are often ascribed anti-inflammatory and proinflammatory (and antithrombogenic and prothrombogenic) roles, respectively, the products of their chemical interaction (RNOS) can yield either phenotype, depending on whether there is net oxidation or nitrosation of specific molecular targets that regulate the inflammatory response (Figure 1).…”