[Show abstract][Hide abstract] ABSTRACT: The significance of nasal resonance and anti-resonance to voice production is a classical issue in vocal pedagogy and voice research. The complex structure of the nasal tract produces a complex frequency response. This complexity must be heavily influenced by the morphology of the paranasal cavities, but their contributions are far from being entirely understood. Detailed analyses of these cavities are difficult because of their limited accessibility. Here we test different methods aiming at documenting the acoustical properties of the paranasal tract. The first set of experiments was performed under in vivo conditions, where the middle meatus was occluded by means of targeted application of a maltodextrin mass under endoscopic control. The efficiency of this occlusion method was verified by measuring the nasal nitric oxide (NO) output during humming. In another experiment the frequency responses to sine sweep excitation of an epoxy mould of a nasal cavity were measured, with and without elimination of paranasal sinuses. The third experiment was conducted in a cadaveric situs, with and without maltodextrin occlusion of the middle meatus and the sphenoidal ostia. The results show that some nasal tract resonances were unaffected by the manipulation of the paranasal cavities. Providing access to a maxillary sinus resulted in marked dips in the response curve while access to the sphenoidal ostium caused only minor effects.
[Show abstract][Hide abstract] ABSTRACT: Renal oxidative stress and nitric oxide (NO) deficiency are key events in hypertension. Stimulation of a nitrate-nitrite-NO pathway with dietary nitrate reduces blood pressure, but the mechanisms or target organ are not clear. We investigated the hypothesis that inorganic nitrate and nitrite attenuate reactivity of renal microcirculation and blood pressure responses to angiotensin II (ANG II) by modulating nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activity and NO bioavailability. Nitrite in the physiological range (10(-7)-10(-5) mol/L) dilated isolated perfused renal afferent arterioles, which were associated with increased NO. Contractions to ANG II (34%) and simultaneous NO synthase inhibition (56%) were attenuated by nitrite (18% and 26%). In a model of oxidative stress (superoxide dismutase-1 knockouts), abnormal ANG II-mediated arteriolar contractions (90%) were normalized by nitrite (44%). Mechanistically, effects of nitrite were abolished by NO scavenger and xanthine oxidase inhibitor, but only partially attenuated by inhibiting soluble guanylyl cyclase. Inhibition of NADPH oxidase with apocynin attenuated ANG II-induced contractility (35%) similar to that of nitrite. In the presence of nitrite, no further effect of apocynin was observed, suggesting NADPH oxidase as a possible target. In preglomerular vascular smooth muscle cells and kidney cortex, nitrite reduced both basal and ANG II-induced NADPH oxidase activity. These effects of nitrite were also abolished by xanthine oxidase inhibition. Moreover, supplementation with dietary nitrate (10(-2) mol/L) reduced renal NADPH oxidase activity and attenuated ANG II-mediated arteriolar contractions and hypertension (99±2-146±2 mm Hg) compared with placebo (100±3-168±3 mm Hg). In conclusion, these novel findings position NADPH oxidase in the renal microvasculature as a prime target for blood pressure-lowering effects of inorganic nitrate and nitrite.
[Show abstract][Hide abstract] ABSTRACT: The maximum power principle dictates that open biological systems tend to self-organize to a level of efficiency that allows maximal power production. Applying this principle to cellular energetics and whole-body physiology would suggest that for every metabolic challenge, an optimal efficiency exists that maximizes power production. On exposure to hypoxia, it would be favorable if metabolic efficiency would rapidly adjust so as to better preserve work performance. We tested this idea in humans by measuring metabolic efficiency and exercise tolerance under normoxic (Fi(O)2=20.9%) and hypoxic (Fi(O)2=16%) conditions, where Fi(O)2 is fraction of inhaled oxygen. The results were compared with respirometric analyses of skeletal muscle mitochondria from the same individuals. We found that among healthy trained subjects (n=14) with a wide range of metabolic efficiency (ME), those with a high ME during normoxic exercise were able to better maintain exercise capacity (Wmax) in hypoxia. On hypoxic exposure, these subjects acutely decreased their efficiency from 19.2 to 17.4%, thereby likely shifting it closer to a degree of efficiency where maximal power production is achieved. In addition, mitochondria from these subjects had a lower intrinsic respiration compared to subjects that showed a large drop in Wmax in hypoxia An acute shift in efficiency was also demonstrated in isolated mitochondria exposed to physiological levels of hypoxia as P/O ratio increased from 0.9 to 1.3 with hypoxic exposure. These findings suggest the existence of a physiological adaptive response by which metabolic efficiency is dynamically optimized to maximize power production.-Schiffer, T. A., Ekblom, B., Lundberg, J. O., Weitzberg, E., Larsen, F. J. Dynamic regulation of metabolic efficiency explains tolerance to acute hypoxia in humans.
FASEB journal : official publication of the Federation of American Societies for Experimental Biology. 06/2014;
[Show abstract][Hide abstract] ABSTRACT: Nitrate, which is an inorganic anion abundant in vegetables, increases the efficiency of isolated human mitochondria. Such an effect might be reflected in changes in the resting metabolic rate (RMR) and formation of reactive oxygen species. The bioactivation of nitrate involves its active accumulation in saliva followed by a sequential reduction to nitrite, nitric oxide, and other reactive nitrogen species.
We studied effects of inorganic nitrate, in amounts that represented a diet rich in vegetables, on the RMR in healthy volunteers.
In a randomized, double-blind, crossover study, we measured the RMR by using indirect calorimetry in 13 healthy volunteers after a 3-d dietary intervention with sodium nitrate (NaNO3) or a placebo (NaCl). The nitrate dose (0.1 mmol ⋅ kg(-1) ⋅ d(-1)) corresponded to the amount in 200-300 g spinach, beetroot, lettuce, or other vegetable that was rich in nitrate. Effects of direct nitrite exposure on cell respiration were studied in cultured human primary myotubes.
The RMR was 4.2% lower after nitrate compared with placebo administration, and the change correlated strongly to the degree of nitrate accumulation in saliva (r(2) = 0.71). The thyroid hormone status, insulin sensitivity, glucose uptake, plasma concentration of isoprostanes, and total antioxidant capacity were unaffected by nitrate. The administration of nitrite to human primary myotubes acutely inhibited respiration.
Dietary inorganic nitrate reduces the RMR. This effect may have implications for the regulation of metabolic function in health and disease.
American Journal of Clinical Nutrition 02/2014; · 6.50 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The discovery of nitric oxide and its role in almost every facet of human biology opened a new avenue for treatment through manipulation of its canonical signaling and by attempts to augment endogenous nitric oxide generation through provision of substrate and co-factors to the endothelial nitric oxide synthase complex. This has been particularly so in the cardiovascular system and it is well recognized that there is reduced biovailable nitric oxide in patients with both cardiovascular risk factors and manifest vascular disease. However, these attempts have failed to deliver the expected benefits of such an approach. Recently, an alternative pathway for nitric oxide synthesis has been elucidated that can produce authentic nitric oxide from the 1 electron reduction of inorganic nitrite. Furthermore, it has long been known that symbiotic, facultative, oral microflora can facilitate the reduction of inorganic nitrate, that is ingested in the average diet in millimolar amounts, to inorganic nitrite itself. Thus, there exists an alternative reductive pathway from nitrate, via nitrite as an intermediate, to nitric oxide that provides a novel pathway that may be amenable to therapeutic manipulation. As such, various research groups have explored the utility of manipulation of this nitrate-nitrite-nitric oxide pathway in situations in which nitric oxide is known to have a prominent role. Animal and early-phase human studies of both inorganic nitrite and nitrate supplementation have shown beneficial effects in blood pressure control, platelet function, vascular health and exercise capacity. This review considers in detail the pathways of inorganic nitrate bioactivation and the evidence of clinical utility to date on the cardiovascular system.
[Show abstract][Hide abstract] ABSTRACT: Aims: Inorganic nitrate and nitrite from endogenous and dietary sources have emerged as alternative substrates for nitric oxide (NO) formation in addition to the classic L-arginine NO synthase (NOS)-dependent pathway. Here we investigated a potential crosstalk between these two pathways in regulation of vascular function. Results: Long-term dietary supplementation with sodium nitrate (0.1 and 1 mmol kg-1 day-1) in rats caused a reversible dose-dependent reduction in phosphorylated eNOS (Ser1177) in aorta and a concomitant increase in phosphorylation at Thr495. Moreover, eNOS-dependent vascular responses were attenuated in vessels harvested from nitrate-treated mice or when nitrite was acutely added to control vessels. The citrulline-to-arginine ratio in plasma, as a measure of eNOS activity, was reduced in nitrate-treated rodents. Telemetry measurements revealed that a low dietary nitrate dose reduced blood pressure, whereas the higher dose was associated with a paradoxical elevation. Finally, plasma cGMP increased in mice treated with a low dietary nitrate dose and decreased with a higher dose. Innovation & Conclusions: These results demonstrate the existence of a crosstalk between the nitrate-nitrite-NO pathway and the NOS-dependent pathway in control of vascular NO homeostasis.
[Show abstract][Hide abstract] ABSTRACT: Antibacterial nitrogen oxides including nitric oxide are formed from nitrite under acidic conditions. In a continuous-flow model of the urinary bladder we used the retention cuff of an all-silicone Foley catheter as a depot for export of nitrogen oxides. The cuff was filled with sodium nitrite (50mM) and an acidic buffer solution (pH 3.6) and the growth of nine common uropathogens in the surrounding artificial urine was measured along with biofilm formation on the catheter surface. In experiments with control catheters (NaCl) bacteria grew readily and biofilm developed within hours in five out of nine strains. In contrast, with test catheters bacterial counts were markedly reduced and biofilm formation by P. aeruginosa, K. pneumoniae and E. cloace was prevented while E. coli and S. aureus was unaffected. We conclude that antibacterial nitrogen oxides generated in the retention cuff of a urinary catheter diffuse into urine and prevent the growth of urinary pathogens and biofilm formation. Although promising, future studies will reveal if this novel approach can be clinically useful for the prevention of catheter-associated urinary tract infections.
Free Radical Biology and Medicine 09/2013; · 5.27 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The theory that red blood cells (RBCs) generate and release nitric oxide (NO)-like bioactivity has gained considerable interest. However, it remains unclear whether it can be produced by endothelial NO synthase (eNOS), which is present in RBCs, and whether NO can escape scavenging by hemoglobin. The aim of this study was to test the hypothesis that arginase reciprocally controls NO formation in RBCs by competition with eNOS for their common substrate arginine and that RBC-derived NO is functionally active following arginase blockade. We show that rodent and human RBCs contain functional arginase 1 and that pharmacological inhibition of arginase increases export of eNOS-derived nitrogen oxides from RBCs under basal conditions. The functional importance was tested in an ex vivo model of myocardial ischemia-reperfusion injury. Inhibitors of arginase significantly improved postischemic functional recovery in rat hearts if administered in whole blood or with RBCs in plasma. By contrast, arginase inhibition did not improve postischemic recovery when administered with buffer solution or plasma alone. The protective effect of arginase inhibition was lost in the presence of a NOS inhibitor. Moreover, hearts from eNOS(-/-) mice were protected when the arginase inhibitor was given with blood from wild-type donors. In contrast, when hearts from wild-type mice were given blood from eNOS(-/-) mice, the arginase inhibitor failed to protect against ischemia-reperfusion. These results strongly support the notion that RBCs contain functional eNOS and release NO-like bioactivity. This process is under tight control by arginase 1 and is of functional importance during ischemia-reperfusion.
Proceedings of the National Academy of Sciences 08/2013; · 9.81 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The circulation of nitrogen in nature is a prerequisite for life on earth. In the nitrogen cycle atmospheric nitrogen is fixated by bacteria into forms that can be utilized by plants and mammals. Nitrate and nitrite are obligate intermediates in this cycle, and for more than half a century these anions have interested nutritional scientists, mostly in relation to cancer, because of their ability to form nitrosamines. However, after the discovery of mammalian endogenous nitric oxide (NO) generation and later that its oxidation products nitrate and nitrite can be recycled back to bioactive NO, a novel field of research has emerged that explores a potentially beneficial role of these anions in physiology, nutrition, and therapeutics. In our diet, vegetables are the major source of nitrate that can fuel a nitrate-nitrite-NO pathway. Herein we discuss the nutritional aspects of this pathway and what is presently known about the implications for human health. Expected final online publication date for the Annual Review of Nutrition Volume 33 is July 17, 2013. Please see http://www.annualreviews.org/catalog/pubdates.aspx for revised estimates.
Annual Review of Nutrition 04/2013; · 9.16 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Liver first-pass metabolism differs considerably among organic nitrates but little information exists on the mechanism of denitration of these compounds in hepatic tissue. The metabolism of nitrooxybutyl-esters of flurbiprofen and ferulic-acid, a class of organic nitrates with potential therapeutic implication in variety of different conditions, was investigated in comparison with glyceryl trinitrate (GTN) in human liver by a multiple approach, using a spontaneous metabolism-independent NO donor (NOC-5) as a reference tool. Nitrooxybutyl-esters were rapidly and quantitatively metabolized to their respective parent compounds and the organic nitrate moiety nitrooxybutyl-alcohol (NOBA). Differently from GTN which was rapidly and completely metabolized to nitrite, NOBA was slowly metabolized to nitrate. In contrast to the spontaneous NO donor NOC-5, NOBA and GTN did not generate detectable NO and failed to suppress the activity of cytochromeP450; an enzyme known to be inhibited by NO. The direct identification of NOBA following liver metabolism targets this compound as the functional organic nitrate metabolite of nitrooxybutyl-esters. Moreover the investigation of the pathways for denitration of NOBA and GTN suggests that organic nitrates are not primarily metabolized to NO in the liver but to different extents of nitrite or nitrate depending from their different chemical structure. It follows that cytochromeP450-dependent metabolism of concomitant drugs is not likely to be affected by oral co-administration of organic nitrates. However the first-pass may differently affect the pharmacological profile of organic nitrates in connection with the different extent of denitration and the distinct bioactive species generated and exported from the liver (nitrate or nitrite).
Journal of Pharmacology and Experimental Therapeutics 04/2013; · 3.89 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Rats with adenine-induced chronic renal failure (A-CRF) develop metabolic and cardiovascular abnormalities resembling those in patients with chronic kidney disease. The aim of this study was to investigate the mechanisms of hypertension in this model and to assess aortic stiffness in vivo. Male Sprague-Dawley rats were equipped with radiotelemetry probes for arterial pressure recordings and received either chow containing adenine or normal control diet. At 7 to 11 weeks after study start, blood pressure responses to high NaCl (4%) diet and different pharmacological interventions were analyzed. Aortic pulse wave velocity was measured under isoflurane anesthesia. Baseline 24 h mean arterial pressure (MAP) was 101±10 and 119±9 mmHg in controls and A-CRF animals, respectively (P<0.01). After 5 days of high NaCl diet MAP had increased by 24±6 mmHg in A-CRF animals vs. 2±1 mmHg in controls (P<0.001). Candesartan (10 mg/kg by gavage) produced a more pronounced reduction of MAP in controls vs. A-CRF animals (-12±3 vs. -5±5 mmHg, P<0.05). Aortic pulse wave velocity was elevated in A-CRF rats (5.10±0.51 vs. 4.58±0.17 m/s, P<0.05). Plasma levels of creatinine were markedly elevated in A-CRF animals (259±46 vs. 31±2 µM, P<0.001) whereas plasma renin activity was suppressed (0.6±0.5 vs. 12.3±7.3 µg/L/h, P<0.001). In conclusion, hypertension in A-CRF animals is characterized by low plasma renin activity and is aggravated by high NaCl diet suggesting a pathogenic role for sodium retention and hypervolemia probably secondary to renal insufficiency. Additionally, aortic stiffness was elevated in A-CRF animals as indicated by increased aortic pulse wave velocity.
[Show abstract][Hide abstract] ABSTRACT: Hydrogen sulfide (H2S), generated through various endogenous enzymatic and non-enzymatic pathways, is emerging as a regulator of physiological and pathological events throughout the body. Bacteria in the gastrointestinal tract also produce significant amounts of H2S that regulates microflora growth and virulence responses. However, the impact of the microbiota on host global H2S bioavailability and metabolism remain unknown. To address this question, we examined H2S bioavailability in its various forms (free, acid labile or bound sulfane sulfur), cystathionine gamma lyase (CSE) activity and cysteine levels in tissues from germ free versus conventionally housed mice. Free H2S levels were significantly reduced in plasma and gastrointestinal tissues of germ free mice. Bound sulfane sulfur levels were decreased by 50-80% in germ free mouse plasma, adipose and lung tissues. Tissue CSE activity was significantly reduced in many organs from germ free mice, whereas tissue cysteine levels were significantly elevated compared to conventional mice. These data reveal that the microbiota profoundly regulates systemic bioavailability and metabolism of H2S.
Free Radical Biology and Medicine 03/2013; · 5.27 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Background
Inorganic nitrate and nitrite have emerged as alternative substrates for nitric oxide (NO) generation in the gastrointestinal tract, and have shown to be protective against drug-induced gastric injury. The aim of this study was to investigate the preventive and therapeutic effects of nitrate and nitrite in a model of experimental colitis.Methods
Colitis was induced in mice by administrating dextran sulfate sodium (DSS) with concurrent administration of nitrite (1 mM) or nitrate (10 mM) in the drinking water for 7 days. A therapeutic approach was also investigated by initiating nitrite treatment 3 days after DSS-induced colitis. Clinical and inflammatory markers were assessed and the colonic mucus thickness was measured in vivo. The effect of nitrite on wound healing was evaluated using colon epithelial cells.ResultsConcurrent administration of DSS and nitrite (1 mM) alleviated inflammation as determined by reduced disease activity index score (DAI) and increased colon length, while nitrate (10 mM) only reduced the DAI-score. Nitrite also displayed therapeutic effects by ameliorating established colonic inflammation with reduced colonic expression of iNOS and improving histopathology. DSS-induced decrease in colonic mucus thickness was completely prevented by nitrite administration. In addition, goblet cell abundance was lower by DSS treatment, but was increased by addition of nitrite. Further studies using colon epithelial cells revealed an NO-dependent improvement in wound healing with nitrite administration.Conclusion
Nitrite exerts both preventive and therapeutic effects in colonic inflammation. The protective effects involve preservation of an intact adherent mucus layer and regulation of epithelial cell restitution.
[Show abstract][Hide abstract] ABSTRACT: OBJECTIVE: Dietary nitrate is reduced to nitrite and nitric oxide ((•)NO) in the gut, producing reactive species able to nitrate proteins and lipids. We investigated intragastric production of (•)NO and nitrating agents in vivo by examining selective nitration of pepsinogen and pepsin. We further address the functional impact of nitration on peptic activity by evaluating the progression of secretagogue-induced ulcers. DESIGN: Pepsinogen nitration was assessed in healthy and diclofenac-induced ulcerated rat stomachs. Both groups were fed nitrite or water by oral gavage. Protein nitration was studied by immunofluorescence and immunoprecipitation. In parallel experiments, pentagastrin was administered to rats and nitrite was then instilled intragastrically. (•)NO levels were measured before and after nitrite administration by chemiluminescence. Macroscopic damage was assessed and nitrated pepsin was examined in the margin of ulcers. RESULTS: Protein nitration was detected physiologically in the stomach of healthy animals. Nitrite had a dual effect on intragastric nitration: overall nitration was decreased under physiological conditions but enhanced by acute inflammation. Pepsin and pepsinogen were also nitrated via a nitrite-dependent pathway. Nitration of both pepsin and its zymogen lead to decreased peptic activity in response to classical substrates (e.g. collagen). Under conditions of acute ulceration, nitrite-dependent pepsin nitration prevented the development of gastric ulcers. CONCLUSION: Dietary nitrite generates nitrating agents in the stomach in vivo, markedly decreasing peptic activity. Under inflammatory and ulcerogenic conditions pepsin nitration attenuates the progression of gastric ulceration. These results suggest that dietary nitrite-dependent nitration of pepsin may have a novel anti-ulcerogenic effect in vivo.
Free Radical Biology and Medicine 12/2012; · 5.27 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Circulating nitrate (NO(3)(-)), derived from dietary sources or endogenous nitric oxide production, is extracted from blood by the salivary glands, accumulates in saliva and is then reduced to nitrite (NO(2)(-)) by the oral microflora. This process has historically been viewed as harmful, since nitrite can promote formation of potentially carcinogenic N-nitrosamines. More recent research, however, suggests that nitrite can also serve as a precursor for systemic generation of vasodilatory nitric oxide, and exogenous administration of nitrate reduces blood pressure in humans. However, whether oral nitrate-reducing bacteria participate in 'setting' blood pressure is unknown. We investigated whether suppression of the oral microflora affects systemic nitrite levels and hence blood pressure in healthy individuals. We measured blood pressure (clinic, home and 24h ambulatory) in 19 healthy volunteers during an initial 7 day control period followed by a 7 day treatment period with a chlorhexidine-based antiseptic mouthwash. Oral nitrate-reducing capacity and nitrite levels were measured after each study period. Antiseptic mouthwash treatment reduced oral nitrite production by 90% (p<0.001) and plasma nitrite levels by 25% (p=0.001) compared to the control period. Systolic and diastolic blood pressure increased by 2-3.5mmHg; increases correlated to decrease in circulating nitrite concentrations (r(2)=0.56, p=0.002). The blood pressure effect appeared within one day of disruption of the oral microflora and was sustained during the 7 day mouthwash intervention. These results suggest that the recycling of endogenous nitrate by oral bacteria plays an important role in determination of plasma nitrite levels and thereby in the physiological control of blood pressure.
Free Radical Biology and Medicine 11/2012; · 5.27 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Inorganic nitrate has been shown to reduce oxygen cost during exercise. Since the nitrate-nitrite-NO pathway is facilitated during hypoxia, we investigated the effects of dietary nitrate on oxygen consumption and cardiovascular responses during apnea. These variables were measured in two randomized, double-blind, placebo-controlled, crossover protocols at rest and ergometer exercise in competitive breath-hold divers. Subjects held their breath for predetermined times along with maximum effort apneas after two separate 3-day periods with supplementation of potassium nitrate/placebo. In contrast to our hypothesis, nitrate supplementation led to lower arterial oxygen saturation (SaO(2), 77±3%) compared to placebo (80±2%) during static apnea, along with lower end-tidal fraction of oxygen (FETO(2)) after 4min of apnea (nitrate 6.9±0.4% vs. placebo 7.6±0.4%). Maximum apnea duration was shorter after nitrate (329±13s) compared to placebo (344±13s). During cycle ergometry nitrate had no effect on SaO(2), FETO(2) or maximum apnea duration. The negative effects of inorganic nitrate during static apnea may be explained by an attenuated diving response.