NADPH oxidases: functions and pathologies in the vasculature

Abstract

Reactive oxygen species are ubiquitous signaling molecules in biological systems. Four members of the NADPH oxidase (Nox) enzyme family are important sources of reactive oxygen species in the vasculature: Nox1, Nox2, Nox4, and Nox5. Signaling cascades triggered by stresses, hormones, vasoactive agents, and cytokines control the expression and activity of these enzymes and of their regulatory subunits, among which p22phox, p47phox, Noxa1, and p67phox are present in blood vessels. Vascular Nox enzymes are also regulated by Rac, ClC-3, Poldip2, and protein disulfide isomerase. Multiple Nox subtypes, simultaneously present in different subcellular compartments, produce specific amounts of superoxide, some of which is rapidly converted to hydrogen peroxide. The identity and location of these reactive oxygen species, and of the enzymes that degrade them, determine their downstream signaling pathways. Nox enzymes participate in a broad array of cellular functions, including differentiation, fibrosis, growth, proliferation, apoptosis, cytoskeletal regulation, migration, and contraction. They are involved in vascular pathologies such as hypertension, restenosis, inflammation, atherosclerosis, and diabetes. As our understanding of the regulation of these oxidases progresses, so will our ability to alter their functions and associated pathologies.

Figure 1. Spatial and molecular organization of vascular Nox enzymes

Nox1, 2 and 5 are represented here in different cellular compartments, but can be located either within cells or at the plasma membrane, thus releasing O₂•− inside vesicles or extracellularly, following activation of receptor (R) by ligand (L). O₂•− may affect cytosolic signaling after crossing membranes via anion channels, reversible protonation, or conversion to H₂O₂. In contrast, Nox4 is always intracellular, and constitutively produces a higher proportion of membrane-permeable H₂O₂ than other oxidases. All oxidases, except Nox5, form a membrane complex with p22phox. Cytosolic activators vary with oxidase subtype: Rac, p47phox and Noxa1 for Nox1 in VSMCs; Rac, p47phox and p67phox for Nox2; Poldip2 for Nox4; Ca⁺⁺ for Nox5. Charge compensation mechanisms are unknown, except for VSMC endosomal ClC-3 that supports Nox1 activity. All vascular cells express multiple Nox subtypes simultaneously.

Figure 2. Nox enzymes in multiple tissues contribute to AngII-induced hypertension

AngII infusion increases Nox activity in tissues involved in blood pressure regulation, including brain, kidney, heart and blood vessels. Brain Nox controls efferent sympathetic stimulation of lymphoid tissue, leading to activation of T lymphocytes and Nox-dependent upregulation of the CCR5 chemokine receptor and the CD44 hyaluronan receptor. Simultaneously, AngII upregulates the adhesion molecule ICAM-1 and the chemokine RANTES in vessels, leading to T cell migration to vascular periadventitia. AngII and Nox in T cells lead to secretion of TNFα, which upregulates vascular Nox., In adventitial fibroblasts, VSMC and endothelial cells, Nox generates O₂•−, which depletes NO• via conversion to peroxynitrate (ONOO⁻), leading to endothelial dysfunction. ROS production is further amplified by ONOO⁻-induced nitric oxide synthase (NOS) uncoupling and mitochondrial dysfunction. Whereas NO• induces vasodilation and growth arrest in healthy vessels, O₂•− and H₂O₂ contribute to vessel contraction and hypertrophy, leading to sustained hypertension. Vascular inflammation produced by mediators such as IL-17 may also induce chronic vascular disease.