Arkan Abadi

miRNA in the Regulation of Skeletal Muscle Adaptation to Acute Endurance Exercise in C57Bl/6J Male Mice

MicroRNAs (miRNAs) are evolutionarily conserved small non-coding RNA species involved in post-transcriptional gene regulation. In vitro studies have identified a small number of skeletal muscle-specific miRNAs which play a crucial role in myoblast proliferation and differentiation. In skeletal muscle, an acute bout of endurance exercise results in the up-regulation of transcriptional networks that regulate mitochondrial biogenesis, glucose and fatty acid metabolism, and skeletal muscle remodelling. The purpose of this study was to assess the expressional profile of targeted miRNA species following an acute bout of endurance exercise and to determine relationships with previously established endurance exercise responsive transcriptional networks. C57Bl/6J wild-type male mice (N = 7/group) were randomly assigned to either sedentary or forced-endurance exercise (treadmill run @ 15 m/min for 90 min) group. The endurance exercise group was sacrificed three hours following a single bout of exercise. The expression of miR- 181, 1, 133, 23, and 107, all of which have been predicted to regulate transcription factors and co-activators involved in the adaptive response to exercise, was measured in quadriceps femoris muscle. Endurance exercise significantly increased the expression of miR-181, miR-1, and miR-107 by 37%, 40%, and 56%, respectively, and reduced miR-23 expression by 84% (P≤0.05 for all), with no change in miR-133. Importantly, decreased expression of miRNA-23, a putative negative regulator of PGC-1α was consistent with increased expression of PGC-1α mRNA and protein along with several downstream targets of PGC-1α including ALAS, CS, and cytochrome c mRNA. PDK4 protein content remains unaltered despite an increase in its putative negative regulator, miR-107, and PDK4 mRNA expression. mRNA expression of miRNA processing machinery (Drosha, Dicer, and DGCR8) remained unchanged. We conclude that miRNA-mediated post-transcriptional regulation is potentially involved in the complex regulatory networks that govern skeletal muscle adaptation to endurance exercise in C57Bl/6J male mice.

Limb Immobilization Induces a Coordinate Down-Regulation of Mitochondrial and Other Metabolic Pathways in Men and Women

Advancements in animal models and cell culture techniques have been invaluable in the elucidation of the molecular mechanisms that regulate muscle atrophy. However, few studies have examined muscle atrophy in humans using modern experimental techniques. The purpose of this study was to examine changes in global gene transcription during immobilization-induced muscle atrophy in humans and then explore the effects of the most prominent transcriptional alterations on protein expression and function. Healthy men and women (N = 24) were subjected to two weeks of unilateral limb immobilization, with muscle biopsies obtained before, after 48 hours (48 H) and 14 days (14 D) of immobilization. Muscle cross sectional area (∼5%) and strength (10–20%) were significantly reduced in men and women (∼5% and 10–20%, respectively) after 14 D of immobilization. Micro-array analyses of total RNA extracted from biopsy samples at 48 H and 14 D uncovered 575 and 3,128 probes, respectively, which were significantly altered during immobilization. As a group, genes involved in mitochondrial bioenergetics and carbohydrate metabolism were predominant features at both 48 H and 14 D, with genes involved in protein synthesis and degradation significantly down-regulated and up-regulated, respectively, at 14 D of muscle atrophy. There was also a significant decrease in the protein content of mitochondrial cytochrome c oxidase, and the enzyme activity of cytochrome c oxidase and citrate synthase after 14 D of immobilization. Furthermore, protein ubiquitination was significantly increased at 48 H but not 14 D of immobilization. These results suggest that transcriptional and post-transcriptional suppression of mitochondrial processes is sustained throughout 14 D of immobilization, while protein ubiquitination plays an early but transient role in muscle atrophy following short-term immobilization in humans.

Little change in markers of protein breakdown and oxidative stress in humans in immobilization- induced skeletal muscle atrophy

A number of studies in rodents suggest that disuse atrophy results from a large increase in proteolysis affected by, or accompanying, increased oxidative stress. Little information is available, however, about the effects of immobilization on markers of muscle protein breakdown and oxidative stress in humans. Therefore, the purpose of this investigation was to measure markers of breakdown or oxidative stress in subjects who underwent 14 days of knee-brace-mediated immobilization. Vastus lateralis samples taken from 21 young subjects before, and 2 days and 14 days after, single leg immobilization were measured for ubiquitin-protein conjugates, caspase 3/7 activity, the 14-kDa caspase-3 cleaved actin fragment, 4-hydroxy-2-nonenal (4-HNE) adducts, and protein carbonyls. Quadriceps cross-sectional area decreased by 5.7% +/- 1.1% (p < 0.0001) following immobilization. Ubiquitin-protein conjugates were elevated at 2 days of immobilization (12%, p < 0.05) but were not different from baseline at 14 days. Levels of the 14-kDa actin fragment and caspase 3/7 activity did not change over the immobilization period. The oxidative stress markers, 4-HNE adducts and protein carbonyls, did not change at any time point. These static measures of breakdown and oxidative modification suggest that a small increase in protein ubiquitination occurs early (2 days), but elevations in ubiquitinated or oxidatively modified proteins are not sustained during the later phase (14 days) of uncomplicated disuse atrophy in humans, suggesting that these pathways are not playing a major role in simple disuse-induced atrophic loss of protein mass.

Supplementation with α-Lipoic Acid, CoQ10, and Vitamin E Augments Running Performance and Mitochondrial Function in Female Mice

Antioxidant supplements are widely consumed by the general public; however, their effects of on exercise performance are controversial. The aim of this study was to examine the effects of an antioxidant cocktail (α-lipoic acid, vitamin E and coenzyme Q10) on exercise performance, muscle function and training adaptations in mice. C57Bl/J6 mice were placed on antioxidant supplement or placebo-control diets (n = 36/group) and divided into trained (8 wks treadmill running) (n = 12/group) and untrained groups (n = 24/group). Antioxidant supplementation had no effect on the running performance of trained mice nor did it affect training adaptations; however, untrained female mice that received antioxidants performed significantly better than placebo-control mice (p ≤ 0.05). Furthermore, antioxidant-supplemented females (untrained) showed elevated respiratory capacity in freshly excised muscle fibers (quadriceps femoris) (p ≤ 0.05), reduced oxidative damage to muscle proteins (p ≤ 0.05), and increased expression of mitochondrial proteins (p ≤ 0.05) compared to placebo-controls. These changes were attributed to increased expression of proliferator-activated receptor gamma coactivator 1α (PGC-1α) (p ≤ 0.05) via activation of AMP-activated protein kinase (AMPK) (p ≤ 0.05) by antioxidant supplementation. Overall, these results indicate that this antioxidant supplement exerts gender specific effects; augmenting performance and mitochondrial function in untrained females, but does not attenuate training adaptations.

Elevated Mitochondrial Oxidative Stress Impairs Metabolic Adaptations to Exercise in Skeletal Muscle

Mitochondrial oxidative stress is a complex phenomenon that is inherently tied to energy provision and is implicated in many metabolic disorders. Exercise training increases mitochondrial oxidative capacity in skeletal muscle yet it remains unclear if oxidative stress plays a role in regulating these adaptations. We demonstrate that the chronic elevation in mitochondrial oxidative stress present in Sod2 +/- mice impairs the functional and biochemical mitochondrial adaptations to exercise. Following exercise training Sod2 +/- mice fail to increase maximal work capacity, mitochondrial enzyme activity and mtDNA copy number, despite a normal augmentation of mitochondrial proteins. Additionally, exercised Sod2 +/- mice cannot compensate for their higher amount of basal mitochondrial oxidative damage and exhibit poor electron transport chain complex assembly that accounts for their compromised adaptation. Overall, these results demonstrate that chronic skeletal muscle mitochondrial oxidative stress does not impact exercise induced mitochondrial biogenesis, but impairs the resulting mitochondrial protein function and can limit metabolic plasticity.

Assessing the effects of 35 European-derived BMI-associated SNPs in Mexican children

The prevalence of obesity in Mexico has increased at an alarming rate in both adults and children. This study was undertaken to test in Mexican children the effects of single nucleotide polymorphisms (SNP) that have been associated with body mass index (BMI) and obesity in Europeans.

Evaluating the transferability of 15 European-derived fasting plasma glucose SNPs in Mexican children and adolescents

Genome wide association studies (GWAS) have identified single-nucleotide polymorphisms (SNPs) that are associated with fasting plasma glucose (FPG) in adult European populations. The contribution of these SNPs to FPG in non-Europeans and children is unclear. We studied the association of 15 GWAS SNPs and a genotype score (GS) with FPG and 7 metabolic traits in 1,421 Mexican children and adolescents from Mexico City. Genotyping of the 15 SNPs was performed using TaqMan Open Array. We used multivariate linear regression models adjusted for age, sex, body mass index standard deviation score, and recruitment center. We identified significant associations between 3 SNPs (G6PC2 (rs560887), GCKR (rs1260326), MTNR1B (rs10830963)), the GS and FPG level. The FPG risk alleles of 11 out of the 15 SNPs (73.3%) displayed significant or non-significant beta values for FPG directionally consistent with those reported in adult European GWAS. The risk allele frequencies for 11 of 15 (73.3%) SNPs differed significantly in Mexican children and adolescents compared to European adults from the 1000G Project, but no significant enrichment in FPG risk alleles was observed in the Mexican population. Our data support a partial transferability of European GWAS FPG association signals in children and adolescents from the admixed Mexican population.

Fine-mapping of 98 obesity loci in Mexican children

Background/objectivesMexico has one of the highest prevalence of childhood obesity in the world. Genome-wide association studies (GWAS) for obesity have identified multiple single-nucleotide polymorphisms (SNPs) in populations of European, East Asian, and African descent. The contribution of these loci to obesity in Mexican children is unclear. We assessed the transferability of 98 obesity loci in Mexican children and fine-mapped the association signals.Subjects/methodsThe study included 405 and 390 Mexican children with normal weight and obesity. Participants were genotyped with a genome-wide dense SNP array designed for Latino populations, allowing for the analysis of GWAS index SNPs as well as fine-mapping SNPs, totaling 750 SNPs covering 98 loci. Two genetic risk scores (GRS) were constructed: a “discovery GRS” and a “best-associated GRS”, representing the number of effect alleles at the GWAS index SNPs and at the best-associated SNPs after fine-mapping for each subject.ResultsSeventeen obesity loci were significantly associated with obesity, and five had fine-mapping SNPs significantly better associated with obesity than their corresponding GWAS index SNPs in Mexican children. Six obesity-associated SNPs significantly departed from additive to dominant (N = 5) or recessive (N = 1) models, and a significant interaction was found between rs274609 (TNNI3K) and rs1010553 (ITIH4) on childhood obesity risk. The best-associated GRS was significantly more associated with childhood obesity (OR = 1.21 per additional risk allele [95%CI:1.17–1.25], P = 4.8 × 10−25) than the discovery GRS (OR = 1.05 per additional risk allele [95%CI:1.02–1.08], P = 8.0 × 10−4), and was also associated with waist-to-hip ratio, fasting glucose, fasting insulin and triglyceride levels, the association being mediated by obesity. An overall depletion of obesity risk alleles was observed in Mexican children with normal weight when compared to GWAS discovery populations.ConclusionsOur study indicates a partial transferability of GWAS obesity loci in Mexican children, and supports the pertinence of post-GWAS fine-mapping experiments in the admixed Mexican population.

Penetrance of polygenic obesity susceptibility loci across the body mass index distribution: an update on scaling effects.

A growing number of single nucleotide polymorphisms (SNPs) have been associated with body mass index (BMI) and obesity, but whether the effect of these obesity susceptibility loci is uniform across the BMI distribution remains unclear. We studied the effects of 37 BMI/obesity-associated SNPs in 75,230 adults of European ancestry along BMI percentiles using conditional quantile regression (CQR) and meta-regression (MR) models. The effects of 9 SNPs (24%) increased significantly across the sample BMI distribution including, FTO (rs1421085, p=8.69×10−15), PCSK1 (rs6235, p=7.11×10−06), TCF7L2 (rs7903146, p=9.60×10−06), MC4R (rs11873305, p=5.08×10−05), FANCL (rs12617233, p=5.30×10−05), GIPR (rs11672660, p=1.64×−04), MAP2K5 (rs997295, p=3.25×10−04), FTO (rs6499653, p=6.23×10−04) and NT5C2 (rs3824755, p=7.90×10−04). We showed that such increases stem from unadjusted gene interactions that enhanced the effects of SNPs in persons with high BMI. When 125 height-associated were analyzed for comparison, only one (<1%), IGF1 (rs6219, p=1.80×10−04), showed effects that varied significantly across height percentiles. Cumulative gene scores of these SNPs (GS-BMI and GS-Height, respectively) showed that only GS-BMI had effects that increased significantly across the sample distribution (BMI: p=7.03×10−37, Height: p=0.499). Overall, these findings underscore the importance of gene-gene and gene-environment interactions in shaping the genetic architecture of BMI and advance a method to detect such interactions using only the sample outcome distribution.