Dietary nitrate enhances the contractile properties of human skeletal muscle

Dietary nitrate, a source of nitric oxide (NO), improves the contractile properties of human muscle. We present the hypothesis that this is due to nitrosylation of the ryanodine receptor and increased NO signaling via the soluble guanyl cyclase-cyclic guanosine monophosphate-protein kinase G pathway, which together increase the free intracellular Ca concentration along with the Ca sensitivity of the myofilaments themselves

What Is in Your Beet Juice? Nitrate and Nitrite Content of Beet Juice Products Marketed to Athletes

Consumption of beetroot juice (BRJ) supplements has become popular among athletes because beets tend to be rich in nitrate (NO3 − ), which can enhance exercise performance by increasing nitric oxide production. The NO3 − content of beets can vary significantly, however, making it difficult to know how much NO3 − any product actually contains. Samples from 45 different lots of 24 different BRJ products from 21 different companies were therefore analyzed for NO3 − (and nitrite [NO2 − ]) concentration using high-performance liquid chromatography. The NO3 − and NO2 − content (i.e., amount per serving) was then calculated based on either (a) the manufacturer’s recommended serving size (for prepackaged/single dose products) or (b) as used in previous studies, a volume of 500 ml (for BRJ sold in bulk containers). There was moderate-to-large variability in NO3 − content between samples of the same product, with a mean coefficient of variation of 30% ± 26% (range 2–83%). There was even greater variability between products, with a ∼50-fold range in NO3 − content between the lowest and highest. Only five products consistently provided ≥5 mmol of NO3 − /serving, which seems to be the minimal dose required to enhance exercise performance in most individuals. NO2 − contents were generally low (i.e., ≤0.5% compared with NO3 − ), although two products contained 10% and 14%. The results of this study may be useful to athletes and their support staff contemplating which (if any) BRJ product to utilize. These data may also offer insight into variability in the literature with respect to the effects of BRJ on exercise performance

In vivo creatine kinase reaction kinetics at rest and stress in type II diabetic rat heart

The effects of type II diabetes on cardiac creatine kinase (CK) enzyme activity and/or flux are unknown. We therefore measured steady‐state phosphocreatine (PCr) and adenosine triphosphate (ATP) content and forward CK reaction kinetic parameters in Zucker Diabetic Fatty (ZDF) rat hearts, a type II diabetes research model. At baseline the PCr to ATP ratio (PCr/ATP) was significantly lower in diabetic heart when compared with matched controls (1.71 ± 0.21 vs. 2.26 ± 0.24, P < 0.01). Furthermore, the forward CK reaction rate constant (k(f)) was higher in diabetic animals (0.52 ± 0.09 s(−1) vs. 0.35 ± 0.06 s(−1), P < 0.01) and CK flux calculated as a product of PCr concentration ([PCr]) and k(f) was similar between two groups (4.32 ± 1.05 μmol/g/s vs. 4.94 ± 1.23 μmol/g/s, P = 0.20). Dobutamine administration resulted in similar increases in heart rate (~38%) and k(f) (~0.12 s(−1)) in both groups. No significant change in PCr and ATP content was observed with dobutamine. In summary, our data showed reduced PCr/ATP in diabetic myocardium as an indicator of cardiac energy deficit. The forward CK reaction rate constant is elevated at baseline which might reflect a compensatory mechanics to support energy flux through the CK shuttle and maintain constant ATP supply. When hearts were stimulated similar increase in k(f) was observed in both groups thus it seems that CK shuttle does not limit ATP supply for the range of workload studied

Skeletal muscle contractile function in heart failure with reduced ejection fraction - A focus on nitric oxide

Despite advances over the past few decades, heart failure with reduced ejection fraction (HFrEF) remains not only a mortal but a disabling disease. Indeed, the New York Heart Association classification of HFrEF severity is based on how much exercise a patient can perform. Moreover, exercise capacity-both aerobic exercise performance and muscle power-are intimately linked with survival in patients with HFrEF. This review will highlight the pathologic changes in skeletal muscle in HFrEF that are related to impaired exercise performance. Next, it will discuss the key role that impaired nitric oxide (NO) bioavailability plays in HFrEF skeletal muscle pathology. Lastly, it will discuss intriguing new data suggesting that the inorganic nitrate \u27enterosalivary pathway\u27 may be leveraged to increase NO bioavailability via ingestion of inorganic nitrate. This ingestion of inorganic nitrate has several advantages over organic nitrate (e.g., nitroglycerin) and the endogenous nitric oxide synthase pathway. Moreover, inorganic nitrate has been shown to improve exercise performance: both muscle power and aerobic capacity, in some recent small but well-controlled, cross-over studies in patients with HFrEF. Given the critical importance of better exercise performance for the amelioration of disability as well as its links with improved outcomes in patients with HFrEF, further studies of inorganic nitrate as a potential novel treatment is critical

Dietary nitrate's effects on exercise performance in heart failure with reduced ejection fraction (HFrEF)

Heart failure with reduced ejection fraction (HFrEF) is a deadly and disabling disease. A key derangement contributing to impaired exercise performance in HFrEF is decreased nitric oxide (NO) bioavailability. Scientists recently discovered the inorganic nitrate pathway for increasing NO. This has advantages over organic nitrates and NO synthase production of NO. Small studies using beetroot juice as a source of inorganic nitrate demonstrate its power to improve exercise performance in HFrEF. A larger-scale trial is now underway to determine if inorganic nitrate may be a new arrow for physicians' quiver of HFrEF treatments

Dietary nitrate-induced increases in human muscle power: High versus low responders

Maximal neuromuscular power is an important determinant of athletic performance and also quality of life, independence, and perhaps even mortality in patient populations. We have shown that dietary nitrate (NO3- ), a source of nitric oxide (NO), improves muscle power in some, but not all, subjects. The present investigation was designed to identify factors contributing to this interindividual variability. Healthy men (n = 13) and women (n = 7) 22-79 year of age and weighing 52.1-114.9 kg were studied using a randomized, double-blind, placebo-controlled, crossover design. Subjects were tested 2 h after ingesting beetroot juice (BRJ) either containing or devoid of 12.3 ± 0.8 mmol of NO3- . Plasma NO3- and nitrite (NO2- ) were measured as indicators of NO bioavailability and maximal knee extensor speed (Vmax ), power (Pmax ), and fatigability were determined via isokinetic dynamometry. On average, dietary NO3- increased (P < 0.05) Pmax by 4.4 ± 8.1%. Individual changes, however, ranged from -9.6 to +26.8%. This interindividual variability was not significantly correlated with age, body mass (inverse of NO3- dose per kg), body mass index (surrogate for body composition) or placebo trial Vmax or fatigue index (in vivo indicators of muscle fiber type distribution). In contrast, the relative increase in Pmax was significantly correlated (r = 0.60; P < 0.01) with the relative increase in plasma NO2- concentration. In multivariable analysis female sex also tended (P = 0.08) to be associated with a greater increase in Pmax. We conclude that the magnitude of the dietary NO3- -induced increase in muscle power is dependent upon the magnitude of the resulting increase in plasma NO2- and possibly female sex

Measurement of nitrate and nitrite in biopsy-sized muscle samples using HPLC

Studies of rats have indicated that skeletal muscle plays a central role in whole-body nitrate ( NO−3 )/nitrite ( NO−2 )/nitric oxide (NO) metabolism. Extending these results to humans, however, is challenging due to the small size of needle biopsy samples. We therefore developed a method to precisely and accurately quantify NO−3 and NO−2 in biopsy-sized muscle samples. NO−3 and NO−2 were extracted from rat soleus samples using methanol combined with mechanical homogenization + ultrasound, bead beating, pulverization at liquid N2 temperature or pulverization + 0.5% Triton X-100. After centrifugation to remove proteins, NO−3 and NO−2 were measured using HPLC. Mechanical homogenization + ultrasound resulted in the lowest NO−3 content (62 ± 20 pmol/mg), with high variability [coefficient of variation (CV) >50%] across samples from the same muscle. The NO−2 / NO−3 ratio (0.019 ± 0.006) was also elevated, suggestive of NO−3 reduction during tissue processing. Bead beating or pulverization yielded lower NO−2 and slightly higher NO−3 levels, but reproducibility was still poor. Pulverization + 0.5% Triton X-100 provided the highest NO−3 content (124 ± 12 pmol/mg) and lowest NO−2 / NO−3 ratio (0.008 ± 0.001), with the least variability between duplicate samples (CV ~15%). These values are consistent with literature data from larger rat muscle samples analyzed using chemiluminescence. Samples were stable for at least 5 wk at -80°C, provided residual xanthine oxidoreductase activity was blocked using 0.1 mmol/l oxypurinol. We have developed a method capable of measuring NO−3 and NO−2 in <1 mg of muscle. This method should prove highly useful in investigating the role of skeletal muscle in NO−3 / NO−2 /NO metabolism in human health and disease. NEW & NOTEWORTHY Measurement of nitrate and especially nitrite in small, i.e., biopsy-sized, muscle samples is analytically challenging. We have developed a precise, accurate, and convenient method for doing so using an affordable commercial HPLC system

Cardiovascular Functional Changes in Chronic Kidney Disease:Integrative Physiology, Pathophysiology and Applications of Cardiopulmonary Exercise Testing

The development of cardiovascular disease during renal impairment involves striking multi-tiered, multi-dimensional complex alterations encompassing the entire oxygen transport system. Complex interactions between target organ systems involving alterations of the heart, vascular, musculoskeletal and respiratory systems occur in Chronic Kidney Disease (CKD) and collectively contribute to impairment of cardiovascular function. These systemic changes have challenged our diagnostic and therapeutic efforts, particularly given that imaging cardiac structure at rest, rather than ascertainment under the stress of exercise, may not accurately reflect the risk of premature death in CKD. The multi-systemic nature of cardiovascular disease in CKD patients provides strong rationale for an integrated approach to the assessment of cardiovascular alterations in this population. State-of-the-art cardiopulmonary exercise testing (CPET) is a powerful, dynamic technology that enables the global assessment of cardiovascular functional alterations and reflects the integrative exercise response and complex machinery that form the oxygen transport system. CPET provides a wealth of data from a single assessment with mechanistic, physiological and prognostic utility. It is an underutilized technology in the care of patients with kidney disease with the potential to help advance the field of cardio-nephrology. This article reviews the integrative physiology and pathophysiology of cardio-renal impairment, critical new insights derived from CPET technology, and contemporary evidence for potential applications of CPET technology in patients with kidney disease

DIFFERENTIAL EFFECTS OF VARYING DOSES OF DIETARY NITRATE ON MUSCLE FUNCTION AND BLOOD PRESSURE IN OLDER SUBJECTS

We have recently demonstrated that dietary nitrate, a source of nitric oxide via the enterosalivary pathway, can improve muscle contractile function in healthy older men and women. Nitrate ingestion has also been shown to reduce blood pressure in older individuals. However, the optimal dose for eliciting these beneficial effects is unknown. We therefore performed a randomized, double-blind, crossover study to determine the effects of ingesting 3.3 mL/kg of beetroot juice (BRJ) containing 0, 212, or 425 µmol/kg of nitrate in six healthy older (age 69±3 y) subjects. Maximal knee extensor speed (Vmax) and power (Pmax) were measured 2 h after BRJ ingestion using isokinetic dynamometry; blood pressure was monitored periodically throughout each study. Mean arterial pressure (in mmHg) was lower (P<0.05) after the high (80±4) vs. the low (84±3) or placebo (88±2) doses. Vmax (in rad/s), however, was higher (P<0.05) after the low dose (11.7±0.8), but not the high dose (10.8±1.0), compared to the placebo (10.5±1.0). Pmax (in W/kg) also tended to be higher (P=0.11) in the low (3.9±0.5) compared to the placebo (3.7±0.5) or high (3.7±0.5) trials. Five out of six subjects achieved a higher Vmax and Pmax after the low vs. the high dose. We conclude that dietary nitrate has differential effects on muscle function and blood pressure in older individuals. A high dose of nitrate intake further lowers blood pressure but does not enhance muscle contractility as much as a lower dose. Supported by Indiana University Purdue University Indianapolis and by the NIA (R21 AG053606)

Simultaneous Pharmacokinetic Analysis of Nitrate and its Reduced Metabolite, Nitrite, Following Ingestion of Inorganic Nitrate in a Mixed Patient Population

Purpose: The pharmacokinetic properties of plasma NO3- and its reduced metabolite, NO2-, have been separately described, but there has been no reported attempt to simultaneously model their pharmacokinetics following NO3- ingestion. This report describes development of such a model from retrospective analyses of concentrations largely obtained from primary endpoint efficacy trials. Methods: Linear and non-linear mixed effects analyses were used to statistically define concentration dependency on time, dose, as well as patient and study variables, and to integrate NO3- and NO2- concentrations from studies conducted at different times, locations, patient groups, and several studies in which sample range was limited to a few hours. Published pharmacokinetic studies for both substances were used to supplement model development. Results: A population pharmacokinetic model relating NO3- and NO2- concentrations was developed. The model incorporated endogenous levels of the two entities, and determined these were not influenced by exogenous NO3- delivery. Covariate analysis revealed intersubject variability in NO3- exposure was partially described by body weight differences influencing volume of distribution. The model was applied to visualize exposure versus response (muscle contraction performance) in individual patients. Conclusions: Extension of the present first-generation model, to ultimately optimize NO3- dose versus pharmacological effects, is warranted