Skeletal muscle overexpression of nicotinamide phosphoribosyl transferase in mice coupled with voluntary exercise augments exercise endurance

Objective: Nicotinamide phosphoribosyl transferase (NAMPT) is the rate-limiting enzyme in the salvage pathway that produces nicotinamide adenine dinucleotide (NAD+), an essential co-substrate regulating a myriad of signaling pathways. We produced a mouse that overexpressed NAMPT in skeletal muscle (NamptTg) and hypothesized that NamptTg mice would have increased oxidative capacity, endurance performance, and mitochondrial gene expression, and would be rescued from metabolic abnormalities that developed with high fat diet (HFD) feeding. Methods: Insulin sensitivity (hyperinsulinemic-euglycemic clamp) was assessed in NamptTg and WT mice fed very high fat diet (VHFD, 60% by kcal) or chow diet (CD). The aerobic capacity (VO2max) and endurance performance of NamptTg and WT mice before and after 7 weeks of voluntary exercise training (running wheel in home cage) or sedentary conditions (no running wheel) were measured. Skeletal muscle mitochondrial gene expression was also measured in exercised and sedentary mice and in mice fed HFD (45% by kcal) or low fat diet (LFD, 10% by kcal). Results: NAMPT enzyme activity in skeletal muscle was 7-fold higher in NamptTg mice versus WT mice. There was a concomitant 1.6-fold elevation of skeletal muscle NAD+. NamptTg mice fed VHFD were partially protected against body weight gain, but not against insulin resistance. Notably, voluntary exercise training elicited a 3-fold higher exercise endurance in NamptTg versus WT mice. Mitochondrial gene expression was higher in NamptTg mice compared to WT mice, especially when fed HFD. Mitochondrial gene expression was higher in exercised NamptTg mice than in sedentary WT mice. Conclusions: Our studies have unveiled a fascinating interaction between elevated NAMPT activity in skeletal muscle and voluntary exercise that was manifest as a striking improvement in exercise endurance

Incomplete Inhibition of Sphingosine 1-Phosphate Lyase Modulates Immune System Function yet Prevents Early Lethality and Non-Lymphoid Lesions

BACKGROUND: S1PL is an aldehyde-lyase that irreversibly cleaves sphingosine 1-phosphate (S1P) in the terminal step of sphingolipid catabolism. Because S1P modulates a wide range of physiological processes, its concentration must be tightly regulated within both intracellular and extracellular environments. METHODOLOGY: In order to better understand the function of S1PL in this regulatory pathway, we assessed the in vivo effects of different levels of S1PL activity using knockout (KO) and humanized mouse models. PRINCIPAL FINDINGS: Our analysis showed that all S1PL-deficient genetic models in this study displayed lymphopenia, with sequestration of mature T cells in the thymus and lymph nodes. In addition to the lymphoid phenotypes, S1PL KO mice (S1PL(-/-)) also developed myeloid cell hyperplasia and significant lesions in the lung, heart, urinary tract, and bone, and had a markedly reduced life span. The humanized knock-in mice harboring one allele (S1PL(H/-)) or two alleles (S1PL(H/H)) of human S1PL expressed less than 10 and 20% of normal S1PL activity, respectively. This partial restoration of S1PL activity was sufficient to fully protect both humanized mouse lines from the lethal non-lymphoid lesions that developed in S1PL(-/-) mice, but failed to restore normal T-cell development and trafficking. Detailed analysis of T-cell compartments indicated that complete absence of S1PL affected both maturation/development and egress of mature T cells from the thymus, whereas low level S1PL activity affected T-cell egress more than differentiation. SIGNIFICANCE: These findings demonstrate that lymphocyte trafficking is particularly sensitive to variations in S1PL activity and suggest that there is a window in which partial inhibition of S1PL could produce therapeutic levels of immunosuppression without causing clinically significant S1P-related lesions in non-lymphoid target organs

Targeted Metabolomic Profiling of Plasma and Survival in Heart Failure Patients

OBJECTIVES: This study sought to derive and validate plasma metabolite associations with survival in heart failure (HF) patients. BACKGROUND: Profiling of plasma metabolites to predict the course of HF appears promising, but validation and incremental value of these profiles are less established. METHODS: Patients (n = 1,032) who met Framingham HF criteria with a history of reduced ejection fraction were randomly divided into derivation and validation cohorts (n = 516 each). Amino acids, organic acids, and acylcarnitines were quantified using mass spectrometry in fasting plasma samples. We derived a prognostic metabolite profile (PMP) in the derivation cohort using Lasso-penalized Cox regression. Validity was assessed by 10-fold cross validation in the derivation cohort and by standard testing in the validation cohort. The PMP was analyzed as both a continuous variable (PMPscore) and dichotomized at the median (PMPcat), in univariate and multivariate models adjusted for clinical risk score and N-terminal pro-B-type natriuretic peptide. RESULTS: Overall, 48% of patients were African American, 35% were women, and the average age was 69 years. After a median follow-up of 34 months, there were 256 deaths (127 and 129 in derivation and validation cohorts, respectively). Optimized modeling defined the 13 metabolite PMPs, which was cross validated as both the PMPscore (hazard ratio [HR]: 3.27; p \u3c 2 × 10 CONCLUSIONS: Plasma metabolite profiles varied across HF subgroups and were associated with survival incremental to conventional predictors. Additional investigation is warranted to define mechanisms and clinical applications

Targeted Metabolomic Profiling of Plasma and Survival in Heart Failure Patients

OBJECTIVES: This study sought to derive and validate plasma metabolite associations with survival in heart failure (HF) patients. BACKGROUND: Profiling of plasma metabolites to predict the course of HF appears promising, but validation and incremental value of these profiles are less established. METHODS: Patients (n = 1,032) who met Framingham HF criteria with a history of reduced ejection fraction were randomly divided into derivation and validation cohorts (n = 516 each). Amino acids, organic acids, and acylcarnitines were quantified using mass spectrometry in fasting plasma samples. We derived a prognostic metabolite profile (PMP) in the derivation cohort using Lasso-penalized Cox regression. Validity was assessed by 10-fold cross validation in the derivation cohort and by standard testing in the validation cohort. The PMP was analyzed as both a continuous variable (PMPscore) and dichotomized at the median (PMPcat), in univariate and multivariate models adjusted for clinical risk score and N-terminal pro-B-type natriuretic peptide. RESULTS: Overall, 48% of patients were African American, 35% were women, and the average age was 69 years. After a median follow-up of 34 months, there were 256 deaths (127 and 129 in derivation and validation cohorts, respectively). Optimized modeling defined the 13 metabolite PMPs, which was cross validated as both the PMPscore (hazard ratio [HR]: 3.27; p \u3c 2 × 10-16) and PMPcat (HR: 3.04; p = 2.93 × 10-8). The validation cohort showed similar results (PMPscore HR: 3.9; p \u3c 2 × 10-16 and PMPcat HR: 3.99; p = 3.47 × 10-9). In adjusted models, PMP remained associated with mortality in the cross-validated derivation cohort (PMPscore HR: 1.63; p = 0.0029; PMPcat HR: 1.47; p = 0.081) and the validation cohort (PMPscore HR: 1.54; p = 0.037; PMPcat HR: 1.69; p = 0.043). CONCLUSIONS: Plasma metabolite profiles varied across HF subgroups and were associated with survival incremental to conventional predictors. Additional investigation is warranted to define mechanisms and clinical applications

Use of Ion Chromatography/Mass Spectrometry for Targeted Metabolite Profiling of Polar Organic Acids

Organic acids (OAs) serve as metabolites that play pivotal roles in a host of different metabolic and regulatory pathways. The polar nature of many OAs poses a challenge to their measurement using widely practiced analytical methods. In this study, a targeted metabolomics method was developed using ion chromatography/triple quadrupole mass spectrometry (IC/MS) to quantitate 28 polar OAs with limits of quantitation ranging from 0.25 to 50 μM. The interday assay precisions ranged from 1% to 19%, with accuracies ranging from 82% to 115%. The IC/MS assay was used to quantitate OAs in quadriceps muscle from sedentary mice compared to fatigued mice subjected to either a low intensity, long duration (LILD) or high intensity, short duration (HISD) forced treadmill regimen. Among the OAs examined, significant differences were detected for hippuric acid, malic acid, fumaric acid, and 2-ketoglutaric acid between the sedentary and fatigued mice. In conclusion, the IC/MS method enabled the separation and quantitative survey of a broad range of polar OAs that are difficult to analyze by chromatographic techniques