Musculoskeletal health worsened from carnitine supplementation and not impacted by a novel individualized treadmill training protocol.

INTRODUCTION: Chronic kidney disease (CKD) negatively affects musculoskeletal health, leading to reduced mobility and quality of life. In healthy populations, carnitine supplementation and aerobic exercise have been reported to improve musculoskeletal health. However, there are inconclusive results regarding their effectiveness and safety in CKD. We hypothesized that carnitine supplementation and individualized treadmill exercise would improve musculoskeletal health in CKD. METHODS: We used a spontaneously progressive CKD rat model (Cy/+ rat) (n=11-12/gr): 1) Cy/+ (CKD-Ctrl), 2) CKD-carnitine (CKD-Carn), and 3) CKD-treadmill (CKD-TM). Carnitine (250mg/kg) was injected daily for 10-weeks. Rats in the treadmill group ran 4 days/week on a 5° incline for 10-weeks progressing from 30 min/day for week one to 40 min/day for week two to 50 min/day for the remaining eight weeks. At 32 weeks of age, we assessed overall cardiopulmonary fitness, muscle function, bone histology and architecture, and kidney function. Data was analyzed by one-way ANOVA with Tukey's multiple comparisons tests. RESULTS: Moderate to severe CKD was confirmed by biochemistries for blood urea nitrogen (mean 43±5 mg/dl CKD-Ctrl), phosphorus (mean 8±1 mg/dl CKD-Ctrl), parathyroid hormone (PTH; mean 625±185 pg/ml CKD-Ctrl), and serum creatinine (mean 1.1±0.2 mg/ml CKD-Ctrl). Carnitine worsened phosphorous (mean 11±3 mg/dl CKD-Carn; p<0.0001), PTH (mean 1738±1233 pg/ml CKD-Carn; p<0.0001), creatinine (mean 1±0.3 mg/dl CKD-Carn; p<0.0001), cortical bone thickness (mean 0.5±0.1 mm CKD-Ctrl, 0.4±0.1 mm CKD-Carn; p<0.05). Treadmill running significantly improve maximal aerobic capacity when compared to CKD-Ctrl (mean 14±2 min CKD-TM, 10±2 min CKD-Ctrl; p<0.01). CONCLUSION: Carnitine supplementation worsened CKD progression, mineral metabolism biochemistries and cortical porosity, and did not have an impact on physical function. Individualized treadmill running improved maximal aerobic capacity but did not have an impact on CKD progression or bone properties. Future studies should seek to better understand carnitine doses in conditions of compromised renal function to prevent toxicity which may result from elevated carnitine levels and to optimize exercise prescriptions for musculoskeletal health.

Skeletal muscle atrophy in clinical and preclinical models of chronic kidney disease: A systematic review and meta-analysis.

Patients with chronic kidney disease (CKD) are often regarded as experiencing wasting of muscle mass and declining muscle strength and function, collectively termed sarcopenia. The extent of skeletal muscle wasting in clinical and preclinical CKD populations is unclear. We evaluated skeletal muscle atrophy in preclinical and clinical models of CKD, with multiple sub-analyses for muscle mass assessment methods, CKD severity, sex and across the different preclinical models of CKD. We performed a systematic literature review of clinical and preclinical studies that measured muscle mass/size using the following databases: Ovid Medline, Embase and Scopus. A random effects meta-analysis was utilized to determine standard mean difference (SMD; Hedges' g) between healthy and CKD. Heterogeneity was evaluated using the I2 statistic. Preclinical study quality was assessed via the Systematic Review Centre for Laboratory Animal Experimentation and clinical studies quality was assessed via the Newcastle-Ottawa Scale. This study was registered in PROSPERO (CRD42020180737) prior to initiation of the search. A total of 111 studies were included in this analysis using the following subgroups: 106 studies in the primary CKD analysis, 18 studies that accounted for diabetes and 7 kidney transplant studies. Significant atrophy was demonstrated in 78% of the preclinical studies and 49% of the clinical studies. The random effects model demonstrated a medium overall SMD (SMD = 0.58, 95% CI = 0.52-0.64) when combining clinical and preclinical studies, a medium SMD for the clinical population (SMD = 0.48, 95% CI = 0.42-0.55; all stages) and a large SMD for preclinical CKD (SMD = 0.95, 95% CI = 0.76-1.14). Further sub-analyses were performed based upon assessment methods, disease status and animal model. Muscle atrophy was reported in 49% of the clinical studies, paired with small mean differences. Preclinical studies reported significant atrophy in 78% of studies, with large mean differences. Across multiple clinical sub-analyses such as severity of CKD, dialysis modality and diabetes, a medium mean difference was found. Sub-analyses in both clinical and preclinical studies found a large mean difference for males and medium for females suggesting sex-specific implications. Muscle atrophy differences varied based upon assessment method for clinical and preclinical studies. Limitations in study design prevented conclusions to be made about the extent of muscle loss with disease progression, or the impact of dialysis. Future work would benefit from the use of standardized measurement methods and consistent clinical staging to improve our understanding of atrophy changes in CKD progression, and analysis of biological sex differences.

Skeletal Muscle Complications in Chronic Kidney Disease.

PURPOSE OF REVIEW: To provide an overview of the recent literature investigating the pathophysiology of skeletal muscle changes, interventions for skeletal muscle, and effects of exercise in chronic kidney disease (CKD). RECENT FINDINGS: There are multiple CKD-related changes that negatively impact muscle size and function. However, the variability in the assessment of muscle size, in particular, hinders the ability to truly understand the impact it may have in CKD. Exercise interventions to improve muscle size and function demonstrate inconsistent responses that warrant further investigation to optimize exercise prescription. Despite progress in the field, there are many gaps in the knowledge of the pathophysiology of sarcopenia of CKD. Identifying these gaps will help in the design of interventions that can be tested to target muscle loss and its consequences such as impaired mobility, falls, and poor quality of life in patients with CKD.

Skeletal muscle blood flow during exercise is reduced in a rat model of pulmonary hypertension.

Pulmonary arterial hypertension (PAH) is characterized by exercise intolerance. Muscle blood flow may be reduced during exercise in PAH; however, this has not been directly measured. Therefore, we investigated blood flow during exercise in a rat model of monocrotaline (MCT)-induced pulmonary hypertension (PH). Male Sprague-Dawley rats (∼200 g) were injected with 60 mg/kg MCT (MCT, n = 23) and vehicle control (saline; CON, n = 16). Maximal rate of oxygen consumption (V̇o2max) and voluntary running were measured before PH induction. Right ventricle (RV) morphology and function were assessed via echocardiography and invasive hemodynamic measures. Treadmill running at 50% V̇o2max was performed by a subgroup of rats (MCT, n = 8; CON, n = 7). Injection of fluorescent microspheres determined muscle blood flow via photo spectroscopy. MCT demonstrated a severe phenotype via RV hypertrophy (Fulton index, 0.61 vs. 0.31; P < 0.001), high RV systolic pressure (51.5 vs. 22.4 mmHg; P < 0.001), and lower V̇o2max (53.2 vs. 71.8 mL·min-1·kg-1; P < 0.0001) compared with CON. Two-way ANOVA revealed exercising skeletal muscle blood flow relative to power output was reduced in MCT compared with CON (P < 0.001), and plasma lactate was increased in MCT (10.8 vs. 4.5 mmol/L; P = 0.002). Significant relationships between skeletal blood flow and blood lactate during exercise were observed for individual muscles (r = -0.58 to -0.74; P < 0.05). No differences in capillarization were identified. Skeletal muscle blood flow is significantly reduced in experimental PH. Reduced blood flow during exercise may be, at least in part, consequent to reduced exercise intensity in PH. This adds further evidence of peripheral muscle dysfunction and exercise intolerance in PAH.

Klotho: An Emerging Factor With Ergogenic Potential.

Sarcopenia and impaired cardiorespiratory fitness are commonly observed in older individuals and patients with chronic kidney disease (CKD). Declines in skeletal muscle function and aerobic capacity can progress into impaired physical function and inability to perform activities of daily living. Physical function is highly associated with important clinical outcomes such as hospitalization, functional independence, quality of life, and mortality. While lifestyle modifications such as exercise and dietary interventions have been shown to prevent and reverse declines in physical function, the utility of these treatment strategies is limited by poor widespread adoption and adherence due to a wide variety of both perceived and actual barriers to exercise. Therefore, identifying novel treatment targets to manage physical function decline is critically important. Klotho, a remarkable protein with powerful anti-aging properties has recently been investigated for its role in musculoskeletal health and physical function. Klotho is involved in several key processes that regulate skeletal muscle function, such as muscle regeneration, mitochondrial biogenesis, endothelial function, oxidative stress, and inflammation. This is particularly important for older adults and patients with CKD, which are known states of Klotho deficiency. Emerging data support the existence of Klotho-related benefits to exercise and for potential Klotho-based therapeutic interventions for the treatment of sarcopenia and its progression to physical disability. However, significant gaps in our understanding of Klotho must first be overcome before we can consider its potential ergogenic benefits. These advances will be critical to establish the optimal approach to future Klotho-based interventional trials and to determine if Klotho can regulate physical dysfunction.

Voluntary Wheel Running Has Beneficial Effects in a Rat Model of CKD-Mineral Bone Disorder (CKD-MBD).

BACKGROUND: Reduced bone and muscle health in individuals with CKD contributes to their higher rates of morbidity and mortality. METHODS: We tested the hypothesis that voluntary wheel running would improve musculoskeletal health in a CKD rat model. Rats with spontaneous progressive cystic kidney disease (Cy/+ IU) and normal littermates (NL) were given access to a voluntary running wheel or standard cage conditions for 10 weeks starting at 25 weeks of age when the rats with kidney disease had reached stage 2-3 of CKD. We then measured the effects of wheel running on serum biochemistry, tissue weight, voluntary grip strength, maximal aerobic capacity (VO2max), body composition and bone micro-CT and mechanics. RESULTS: Wheel running improved serum biochemistry with decreased creatinine, phosphorous, and parathyroid hormone in the rats with CKD. It improved muscle strength, increased time-to-fatigue (for VO2max), reduced cortical porosity and improved bone microarchitecture. The CKD rats with voluntary wheel access also had reduced kidney cystic weight and reduced left ventricular mass index. CONCLUSIONS: Voluntary wheel running resulted in multiple beneficial systemic effects in rats with CKD and improved their physical function. Studies examining exercise interventions in patients with CKD are warranted.

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 ( NO3- )/nitrite ( NO2- )/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 NO3- and NO2- in biopsy-sized muscle samples. NO3- and NO2- 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, NO3- and NO2- were measured using HPLC. Mechanical homogenization + ultrasound resulted in the lowest NO3- content (62 ± 20 pmol/mg), with high variability [coefficient of variation (CV) >50%] across samples from the same muscle. The NO2- / NO3- ratio (0.019 ± 0.006) was also elevated, suggestive of NO3- reduction during tissue processing. Bead beating or pulverization yielded lower NO2- and slightly higher NO3- levels, but reproducibility was still poor. Pulverization + 0.5% Triton X-100 provided the highest NO3- content (124 ± 12 pmol/mg) and lowest NO2- / NO3- 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 NO3- and NO2- in <1 mg of muscle. This method should prove highly useful in investigating the role of skeletal muscle in NO3- / NO2- /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.