Nitric oxide in cardiovascular and renal disease : role of organic nitrates, inorganic nitrate and red blood cells

RATIONALE: Cardiovascular and renal disorders are major health problems, which are often co-existing. Mechanistically, these conditions can be attributed to endothelial dysfunction, a process coupled with reduced nitric oxide (NO) bioavailability. In the vasculature, NO is predominantly formed by endothelial NO-synthase (eNOS), which uses L-arginine and oxygen as substrates. Organic nitrates like glyceryl trinitrate (GTN) are clinically used as an exogenous source of NO, with potent vasodilator actions. Despite their long history of use, the undesired side-effects induced by organic nitrates, such as hypotension, headache and development of tolerance, limit their clinical use and demand a need to develop new nitrate preparations. In addition to the classical NOS system, the nitrate-nitrite-NO pathway can serve as additional source of NO generation. Inorganic nitrate is abundant in certain foods, e.g. leafy green vegetables and beetroot. Stimulating this NOS-independent system, via the diet, has been linked with favorable cardiovascular, metabolic and renal effects in several disease models. Moreover, the recent discovery of existing NOS in red blood cells (RBCs) has started a debate regarding the role and interaction between RBCs and endothelial NOSs in regulation of vascular function. AIM: This thesis I) characterizes the therapeutic value of restoring NO bioavailability in cardiovascular and renal disease models by using novel organic mononitrate(s) and inorganic nitrate, and II) investigates the potential role and interaction between NOS in RBCs and the endothelium in regulation or modulation of vascular function. METHODS & RESULTS: Combination of in vivo disease models, ex vivo vessel reactivity as well as in vitro studies were used. Study I-II: A novel organic nitrate 1,3-bis(hexyloxy)propan- 2-yl nitrate (NDHP) was synthesized and functionally characterized. It was found that NDHPderived NO formation was enzymatically mediated by xanthine oxidoreductase (XOR). Moreover, NDHP treatment in contrast to GTN was not subject to tolerance in isolated small arteries and NDHP attenuated angiotensin II-induced hypertension and endothelial dysfunction in rats. Study III: Inorganic nitrate supplementation, increased NO bioactivity, dampened oxidative stress and inflammation and ameliorated mitochondrial abnormalities, which were associated with protection against development of renal ischemia-reperfusion (IR) injuries in mice. This novel approach may have therapeutic value by reducing the risk of acute and chronic kidney disease as well as cardiovascular co-morbidities. Study IV: Using ex vivo co-incubation of isolated RBCs and aortas from eNOS-deficient and control mice, it was shown that RBCs lacking NOS induced oxidative stress and endothelial dysfunction in healthy vessels. This pathological vascular phenotype could be prevented by inhibition of arginase activity and by scavenging of NADPH oxidase-derived reactive oxygen species. CONCLUSION: Restoring NO bioactivity, by using novel organic nitrates or inorganic nitrate, is coupled with favorable effects in models of cardiorenal disease. In addition, arginase and oxidative stress is involved in the interaction between NOS in RBCs and the endothelium. Future studies are needed to further characterize underlying mechanisms and to investigate the potential therapeutic value