Nitrite supplementation reverses vascular endothelial dysfunction and large elastic artery stiffness with aging.

Cardiovascular diseases (CVD) remain the leading cause of death in modern societies (Lloyd-Jones et al.). Advancing age is the major risk factor for CVD (Lakatta 2002; Lakatta & Levy 2003). Most CVD are linked to disorders of arteries (Lloyd-Jones et al.) and it is now recognized that aging increases risk of CVD in large part by causing arterial dysfunction, which then leads to clinical vascular and cardiac diseases (Lakatta & Levy 2003). Many changes to arteries likely contribute to the increased risk of CVD with aging. One of these is the development of vascular endothelial dysfunction, as most commonly indicated by impaired endothelium-dependent dilation (EDD) (Celermajer et al. 1994; Taddei et al. 1995; Lakatta & Levy 2003). Impaired EDD with aging is mediated by excessive superoxide, at least in part as a result of increases in the oxidant enzyme nicotinamide adenine dinucleotide phosphate-oxidase (NADPH oxidase) (Zalba et al. 2000; Bedard & Krause 2007; Donato et al. 2007; Durrant et al. 2009). Increased superoxide reduces bioavailability of nitric oxide (NO) both by reacting with NO to form peroxynitrite and by oxidizing tetrahydrobiopterin (BH4), an essential co-factor for NO production by endothelial nitric oxide synthase (eNOS) (Cosentino et al. 1998; Taddei et al. 2001; Landmesser et al. 2003; Blackwell et al. 2004; Shi et al. 2004). Another key vascular change with aging is the stiffening of large elastic arteries (Lakatta & Levy 2003), possibly linked in part to development of vascular endothelial dysfunction (Fitch et al. 2001; Wilkinson et al. 2004). Large elastic artery stiffness has emerged as a major independent risk factor for age-associated CVD and a key therapeutic target (Lakatta & Levy 2003; Nilsson et al. 2009; Mitchell et al.; Vlachopoulos et al. 2010). Oxidative stress and inflammation are believed to be important mechanisms contributing to these effects of aging on arteries (Brandes et al. 2005; Donato et al. 2007; Csiszar et al. 2008; Donato et al. 2008; Ungvari et al. 2010). Arterial oxidative stress develops with aging as a consequence of excessive production of superoxide by NADPH oxidase or other mechanisms (eNOS uncoupling, mitochondrial dysfunction) (Vasquez-Vivar et al. 1998; Ungvari et al. 2008; Durrant et al. 2009; Rippe et al. 2010) and possibly via selective reductions in antioxidant enzymes, including superoxide dismutase (SOD) (Sun et al. 2004; Rippe et al.). As a result of oxidative stress, injury or other causes, vascular inflammation also develops with aging, as indicated by increases in expression of pro-inflammatory cytokines such as interleukins 1 and 6 (IL-1, IL6), interferon γ (INFγ) and tumor necrosis factor α (TNFα) (Csiszar et al. 2008; Donato et al. 2008; Rippe et al. 2010; Ungvari et al. 2010). Given the above, treatments that improve or reverse vascular endothelial dysfunction and large elastic artery stiffness in middle/older age may prevent much of the increase in CVD risk with aging (Lakatta & Levy 2003; Lonn et al. 2010; Seals 2010). In this context, the nitrite anion (inorganic nitrite) would seem to hold promise. Once considered an inert byproduct of NO metabolism, nitrite now is recognized as a physiologically important storage form of NO (Lundberg & Weitzberg 2008). Nitrite is a cytoprotective molecule considered to have broad therapeutic potential for the prevention and treatment of CVD (Calvert & Lefer; Lundberg & Weitzberg 2008). Sodium nitrite administration protects against ischemia/reperfusion injury (Bryan et al. 2007; Bryan et al. 2008; Gonzalez et al. 2008) and improves vascular endothelial dysfunction in hypercholesterolemic mice (Stokes et al. 2009). However, the efficacy of sodium nitrite for treating arterial aging is entirely unknown. In the present study, we hypothesized that short-term oral sodium nitrite treatment would improve or reverse vascular endothelial dysfunction and large elastic artery stiffness in old mice while having no effects in young mice. As a secondary aim, we sought to gain initial insight into the mechanisms by which nitrite-induced improvements in function in the old mice might be mediated, particularly those related to reduced oxidative stress and inflammation.