Quantitative Metabolomics by 1H-NMR and LC-MS/MS Confirms Altered Metabolic Pathways in Diabetes

Insulin is as a major postprandial hormone with profound effects on carbohydrate, fat, and protein metabolism. In the absence of exogenous insulin, patients with type 1 diabetes exhibit a variety of metabolic abnormalities including hyperglycemia, glycosurea, accelerated ketogenesis, and muscle wasting due to increased proteolysis. We analyzed plasma from type 1 diabetic (T1D) humans during insulin treatment (I+) and acute insulin deprivation (I-) and non-diabetic participants (ND) by 1H nuclear magnetic resonance spectroscopy and liquid chromatography-tandem mass spectrometry. The aim was to determine if this combination of analytical methods could provide information on metabolic pathways known to be altered by insulin deficiency. Multivariate statistics differentiated proton spectra from I- and I+ based on several derived plasma metabolites that were elevated during insulin deprivation (lactate, acetate, allantoin, ketones). Mass spectrometry revealed significant perturbations in levels of plasma amino acids and amino acid metabolites during insulin deprivation. Further analysis of metabolite levels measured by the two analytical techniques indicates several known metabolic pathways that are perturbed in T1D (I-) (protein synthesis and breakdown, gluconeogenesis, ketogenesis, amino acid oxidation, mitochondrial bioenergetics, and oxidative stress). This work demonstrates the promise of combining multiple analytical methods with advanced statistical methods in quantitative metabolomics research, which we have applied to the clinical situation of acute insulin deprivation in T1D to reflect the numerous metabolic pathways known to be affected by insulin deficiency

Poor glycaemic control is associated with reduced exercise performance and oxygen economy during cardio-pulmonary exercise testing in people with type 1 diabetes

BackgroundTo explore the impact of glycaemic control (HbA1c) on functional capacity during cardio-pulmonary exercise testing in people with type 1 diabetes.MethodsSixty-four individuals with type 1 diabetes (age: 34 ± 8 years; 13 females, HbA1c: 7.8 ± 1% (62 ± 13 mmol/mol), duration of diabetes: 17 ± 9 years) performed a cardio-pulmonary cycle ergometer exercise test until volitional exhaustion. Stepwise linear regression was used to explore relationships between HbA1c and cardio-respiratory data with p ≤ 0.05. Furthermore, participants were divided into quartiles based on HbA1c levels and cardio-respiratory data were analysed by one-way ANOVA. Multiple regression analysis was performed to explore the relationships between changes in time to exhaustion and cardio-respiratory data. Data were adjusted for confounder.ResultsHbA1c was related to time to exhaustion and oxygen consumption at the power output elicited at the sub-maximal threshold of the heart rate turn point (r = 0.47, R2 = 0.22, p = 0.03). Significant differences were found at time to exhaustion between QI vs. QIV and at oxygen consumption at the power output elicited at the heart rate turn point between QI vs. QII and QI vs. QIV (p < 0.05). Changes in oxygen uptake, power output and in oxygen consumption at the power output elicited at the heart rate turn point and at maximum power output explained 55% of the variance in time to exhaustion (r = 0.74, R2 = 0.55, p < 0.01).ConclusionsPoor glycaemic control is related to less economical use of oxygen at sub-maximal work rates and an earlier time to exhaustion during cardio-pulmonary exercise testing. However, exercise training could have the same potential to counteract the influence of poor glycaemic control on functional capacity

Function-Based Discovery of Significant Transcriptional Temporal Patterns in Insulin Stimulated Muscle Cells

Background: Insulin action on protein synthesis (translation of transcripts) and post-translational modifications, especially of those involving the reversible modifications such as phosphorylation of various signaling proteins, are extensively studied but insulin effect on transcription of genes, especially of transcriptional temporal patterns remains to be fully defined. Methodology/Principal Findings: To identify significant transcriptional temporal patterns we utilized primary differentiated rat skeletal muscle myotubes which were treated with insulin and samples were collected every 20 min for 8 hours. Pooled samples at every hour were analyzed by gene array approach to measure transcript levels. The patterns of transcript levels were analyzed based on a novel method that integrates selection, clustering, and functional annotation to find the main temporal patterns associated to functional groups of differentially expressed genes. 326 genes were found to be differentially expressed in response to in vitro insulin administration in skeletal muscle myotubes. Approximately 20 % of the genes that were differentially expressed were identified as belonging to the insulin signaling pathway. Characteristic transcriptional temporal patterns include: (a) a slow and gradual decrease in gene expression, (b) a gradual increase in gene expression reaching a peak at about 5 hours and then reaching a plateau or an initial decrease and other different variable pattern of increase in gene expression over time. Conclusion/Significance: The new method allows identifying characteristic dynamic responses to insulin stimulus, commo

Palmitic acid-rich oils with and without interesterification lower postprandial lipemia and increase atherogenic lipoproteins compared to a MUFA-rich oil: a randomized controlled trial

Background: Interesterified (IE) fats are widely used in place of trans fats; however, little is known about their metabolism. Objective: To test the impact of a commonly consumed IE versus a non-IE equivalent fat on in vivo postprandial and in vitro lipid metabolism, compared with a reference oil (rapeseed oil; RO). Design: A double-blinded, 3-phase crossover, randomized controlled trial was performed in healthy adults (n=20) aged 45-75 years. Postprandial plasma triacylglycerol (TG) and lipoprotein responses (including stable isotope tracing) to a test meal (50g fat) were evaluated over 8 hours. The test fats were IE 80:20 palm stearin/palm kernel fat, an identical non-IE fat, and RO (control). In vitro, mechanisms of digestion were explored using a dynamic gastric model (DGM). Results: Plasma TG 8h incremental area under the curves were lower following non-IE versus RO (-1.7 mmol/L.h (95% confidence interval -3.3, -0.0)), but there were no differences between IE and RO nor IE and non-IE. Low density lipoprotein (LDL) particles were smaller following IE and non-IE versus RO (P=0.005). Extra, extra large (XXL)-, extra large (XL)- and large (L)- VLDL particle concentrations were higher following IE and non-IE versus RO at 6-8 h (P <0.05). No differences in the appearance of [13C]palmitic acid in plasma TG was observed between IE and non-IE fats. DGM revealed differences in phase separation of the IE and non-IE meals and delayed release of saturated fatty acids (SFA) versus RO. Conclusions: Interesterification did not modify fat digestion, postprandial lipemia or lipid metabolism measured by stable isotope and DGM analysis. Despite the lower lipemia following the SFA rich fats, increased pro-atherogenic large TG-rich lipoprotein remnant and small LDL particles following the SFA rich fats relative to RO adds a new postprandial dimension to the mechanistic evidence linking SFA to cardiovascular disease risk

Further Support to the Uncoupling-to-Survive Theory: The Genetic Variation of Human UCP Genes Is Associated with Longevity

In humans Uncoupling Proteins (UCPs) are a group of five mitochondrial inner membrane transporters with variable tissue expression, which seem to function as regulators of energy homeostasis and antioxidants. In particular, these proteins uncouple respiration from ATP production, allowing stored energy to be released as heat. Data from experimental models have previously suggested that UCPs may play an important role on aging rate and lifespan. We analyzed the genetic variability of human UCPs in cohorts of subjects ranging between 64 and 105 years of age (for a total of 598 subjects), to determine whether specific UCP variability affects human longevity. Indeed, we found that the genetic variability of UCP2, UCP3 and UCP4 do affect the individual's chances of surviving up to a very old age. This confirms the importance of energy storage, energy use and modulation of ROS production in the aging process. In addition, given the different localization of these UCPs (UCP2 is expressed in various tissues including brain, hearth and adipose tissue, while UCP3 is expressed in muscles and Brown Adipose Tissue and UCP4 is expressed in neuronal cells), our results may suggest that the uncoupling process plays an important role in modulating aging especially in muscular and nervous tissues, which are indeed very responsive to metabolic alterations and are very important in estimating health status and survival in the elderly

Sex differences in the effects of 12 weeks sprint interval training on body fat mass and the rates of fatty acid oxidation and VO 2 max during exercise

Abstract Background The purpose of this study was to examine whether very short duration, very high intensity sprint interval training (SIT) leads to loss of body fat mass in association with improvements to VO2max and fatty acid oxidation, and to assess the extent of sex dimorphism in these physiological responses. Methods A total of 24 men and 17 women (mean (SEM) age: 39 (±2) years; body mass index 24.6 (0.6)) completed measurements of the maximal rate of oxygen uptake (VO2max) and fatty acid oxidation (FATmax). Body fat and lean mass were measured by dual emission x-ray absorptiometry, and fasting blood lipid, glucose and insulin profiles were assessed before and after training. SIT consisted of 4×20 s sprints on a cycle ergometer at approximately 175% VO2max, three times per week for 12 weeks. Results Fat mass decreased by 1.0 kg, although men lost statistically significantly more fat than women both when expressed in Kg and as % body fat. VO2max increased by around 9%, but women improved VO2max significantly more than men. FATmax improved by around 13%, but fasting plasma glucose, insulin, total triglyceride, total cholesterol and high-density lipoprotein (HDL) did not change after training, while low-density lipoprotein decreased by 8% (p=0.028) and the HDL:Total Cholesterol ratio improved by 6%. There were no sex differences in these metabolic responses to training. Conclusions These results show lower body fat %, and higher rates of fatty acid oxidation and VO2max after 12 weeks of training for just 4 min per week. Notably, women improved VO2max more than men, while men lost more fat than women

Contribution of Heritability and Epigenetic Factors to Skeletal Muscle Mass Variation in United Kingdom Twins

Context: Skeletal muscle mass (SMM) is one of the major components of human body composition, with deviations from normal values often leading to sarcopenia.Objective: Our major aim was to conduct a genome-wide DNA methylation study in an attempt to identify potential genomic regions associated with SMM.Design: This was a mixed cross-sectional and longitudinal study.Setting: Community-based study.Participants: A total of 1550 middle-aged UK twin (monozygotic and dizygotic) twins, 297 of which were repeatedly measured participated in the study.Main Outcome Measure: Appendicular lean mass assessed using DXA technology, and MeDIP-seq DNA methylation profiling genome-wide were obtained from each individual.Results: Heritability estimate of SMM, with simultaneous adjustment for covariates obtained using variance decomposition analysis was h2=0.809±0.050. After quality control and analysis of longitudinal stability, the DNA methylation data comprised of 723,029 genomic sites, with positive correlations between repeated measurements (Rrepeated =0.114–0.905). Correlations between MZ and DZ twins were 0.51 and 0.38 at a genome-wide average, respectively and clearly increased with Rrepeated. Testing for DNA methylation association with SMM in 50 discordant MZ twins revealed 36,081 nominally significant results, of which the top-ranked 134 signals (P&lt;0.01 and Rrepeated&gt;0.40) were subjected to replication in the sample of 1,196 individuals. Seven SMM-methylation association signals replicated at a false discovery rate &lt;0.1, and these were located in or near genes DNAH12, CAND1, CYP4F29P, ZFP64 which have previously been highlighted in muscle-related studies. Adjusting for age, smoking and blood cell heterogeneity did not alter significance of these associations.Conclusion: This epigenome-wide study, testing longitudinally stable methylation sites discovered and replicated a number of associations between DNA methylation at CpG loci and skeletal muscle mass. Four replicated signals were related to genes with potential muscle functions, suggesting that the methylome of whole blood may be informative of SMM variation<br/

Metabolic and molecular changes associated with the increased skeletal muscle insulin action 24–48 h after exercise in young and old humans

The molecular and metabolic mechanisms underlying the increase in insulin sensitivity (i.e. increased insulin-stimulated skeletal muscle glucose uptake, phosphorylation and storage as glycogen) observed from 12 to 48 h following a single bout of exercise inhumans remain unresolved. Moreover, whether these mechanisms differ with age is unclear. It is well established that a single bout of exercise increases the translocation of the glucose transporter, GLUT4, to the plasma membrane. Previous research using unilateral limb muscle contraction models in combination with hyperinsulinaemia has demonstrated that the increase in insulin sensitivity and glycogen synthesis 24 h after exercise is also associated with an increase in hexokinase II (HKII) mRNA and protein content, suggesting an increase in the capacity of the muscle to phosphorylate glucose and divert it towards glycogen synthesis. Interestingly, this response is altered in older individuals for up to 48 h post exercise and is associated with molecular changes in skeletal muscle tissue that are indicative of reduced lipid oxidation, increased lipogenesis, increased inflammation and a relative inflexibility of changes in intramyocellular lipid (IMCL) content. Reduced insulin sensitivity (insulin resistance) is generally related to IMCL content, particularly in the subsarcolemmal (SSL) region, and both are associated with increasing age. Recent research has demonstrated that ageing per se appears to cause an exacerbated lipolytic response to exercise that may result in SSL IMCL accumulation. Further research is required to determine if increased IMCL content affects HKII expression in the days after exercise in older individuals, and the effect of this on skeletal muscle insulin action