Anna E. Thalacker-Mercer

Differential DNA methylation with age displays both common and dynamic features across human tissues that are influenced by CpG landscape

BackgroundDNA methylation is an epigenetic modification that changes with age in human tissues, although the mechanisms and specificity of this process are still poorly understood. We compared CpG methylation changes with age across 283 human blood, brain, kidney, and skeletal muscle samples using methylation arrays to identify tissue-specific age effects.ResultsWe found age-associated CpGs (ageCGs) that are both tissue-specific and common across tissues. Tissue-specific ageCGs are frequently located outside CpG islands with decreased methylation, and common ageCGs show the opposite trend. AgeCGs are significantly associated with poorly expressed genes, but those with decreasing methylation are linked with higher tissue-specific expression levels compared with increasing methylation. Therefore, tissue-specific gene expression may protect against common age-dependent methylation. Distinguished from other tissues, skeletal muscle ageCGs are more associated with expression, enriched near genes related to myofiber contraction, and closer to muscle-specific CTCF binding sites. Kidney-specific ageCGs are more increasingly methylated compared to other tissues as measured by affiliation with kidney-specific expressed genes. Underlying chromatin features also mark common and tissue-specific age effects reflective of poised and active chromatin states, respectively. In contrast with decreasingly methylated ageCGs, increasingly methylated ageCGs are also generally further from CTCF binding sites and enriched within lamina associated domains.ConclusionsOur data identified common and tissue-specific DNA methylation changes with age that are reflective of CpG landscape and suggests both common and unique alterations within human tissues. Our findings also indicate that a simple epigenetic drift model is insufficient to explain all age-related changes in DNA methylation.

Simvastatin impairs ADP-stimulated respiration and increases mitochondrial oxidative stress in primary human skeletal myotubes

Statins, the widely prescribed cholesterol-lowering drugs for the treatment of cardiovascular disease, cause adverse skeletal muscle side effects ranging from fatigue to fatal rhabdomyolysis. The purpose of this study was to determine the effects of simvastatin on mitochondrial respiration, oxidative stress, and cell death in differentiated primary human skeletal muscle cells (i.e., myotubes). Simvastatin induced a dose-dependent decrease in viability of proliferating and differentiating primary human muscle precursor cells, and a similar dose-dependent effect was noted in differentiated myoblasts and myotubes. Additionally, there were decreases in myotube number and size following 48 h of simvastatin treatment (5 μM). In permeabilized myotubes, maximal ADP-stimulated oxygen consumption, supported by palmitoylcarnitine+malate (PCM, complex I and II substrates) and glutamate+malate (GM, complex I substrates), was 32-37% lower (P<0.05) in simvastatin-treated (5 μM) vs control myotubes, providing evidence of impaired respiration at complex I. Mitochondrial superoxide and hydrogen peroxide generation were significantly greater in the simvastatin-treated human skeletal myotube cultures compared to control. In addition, simvastatin markedly increased protein levels of Bax (proapoptotic, +53%) and Bcl-2 (antiapoptotic, +100%, P<0.05), mitochondrial PTP opening (+44%, P<0.05), and TUNEL-positive nuclei in human skeletal myotubes, demonstrating up-regulation of mitochondrial-mediated myonuclear apoptotic mechanisms. These data demonstrate that simvastatin induces myotube atrophy and cell loss associated with impaired ADP-stimulated maximal mitochondrial respiratory capacity, mitochondrial oxidative stress, and apoptosis in primary human skeletal myotubes, suggesting that mitochondrial dysfunction may underlie human statin-induced myopathy.

Differential genomic responses in old vs. young humans despite similar levels of modest muscle damage after resistance loading

Across numerous model systems, aging skeletal muscle demonstrates an impaired regenerative response when exposed to the same stimulus as young muscle. To better understand the impact of aging in a human model, we compared changes to the skeletal muscle transcriptome induced by unaccustomed high-intensity resistance loading (RL) sufficient to cause moderate muscle damage in young (37 yr) vs. older (73 yr) adults. Serum creatine kinase was elevated 46% 24 h after RL in all subjects with no age differences, indicating similar degrees of myofiber membrane wounding by age. Despite this similarity, from genomic microarrays 318 unique transcripts were differentially expressed after RL in old vs. only 87 in young subjects. Follow-up pathways analysis and functional annotation revealed among old subjects upregulation of transcripts related to stress and cellular compromise, inflammation and immune responses, necrosis, and protein degradation and changes in expression (up- and downregulation) of transcripts related to skeletal and muscular development, cell growth and proliferation, protein synthesis, fibrosis and connective tissue function, myoblast-myotube fusion and cell-cell adhesion, and structural integrity. Overall the transcript-level changes indicative of undue inflammatory and stress responses in these older adults were not mirrored in young subjects. Follow-up immunoblotting revealed higher protein expression among old subjects for NF-kappaB, heat shock protein (HSP)70, and IL-6 signaling [total and phosphorylated signal transducer and activator of transcription (STAT)3 at Tyr705]. Together, these novel findings suggest that young and old adults are equally susceptible to RL-mediated damage, yet the muscles of older adults are much more sensitive to this modest degree of damage-launching a robust transcriptome-level response that may begin to reveal key differences in the regenerative capacity of skeletal muscle with advancing age.

Production and supply of high‐quality food protein for human consumption: sustainability, challenges, and innovations

The Food and Agriculture Organization of the United Nations estimates that 843 million people worldwide are hungry and a greater number suffer from nutrient deficiencies. Approximately one billion people have inadequate protein intake. The challenge of preventing hunger and malnutrition will become even greater as the global population grows from the current 7.2 billion people to 9.6 billion by 2050. With increases in income, population, and demand for more nutrient‐dense foods, global meat production is projected to increase by 206 million tons per year during the next 35 years. These changes in population and dietary practices have led to a tremendous rise in the demand for food protein, especially animal‐source protein. Consuming the required amounts of protein is fundamental to human growth and health. Protein needs can be met through intakes of animal and plant‐source foods. Increased consumption of food proteins is associated with increased greenhouse gas emissions and overutilization of water. Consequently, concerns exist regarding impacts of agricultural production, processing and distribution of food protein on the environment, ecosystem, and sustainability. To address these challenging issues, the New York Academy of Sciences organized the conference “Frontiers in Agricultural Sustainability: Studying the Protein Supply Chain to Improve Dietary Quality” to explore sustainable innovations in food science and programming aimed at producing the required quality and quantity of protein through improved supply chains worldwide. This report provides an extensive discussion of these issues and summaries of the presentations from the conference.

Heightened muscle inflammation susceptibility may impair regenerative capacity in aging humans

The regenerative response of skeletal muscle to mechanically induced damage is impaired with age. Previous work in our laboratory suggests this may result from higher proinflammatory signaling in aging muscle at rest and/or a greater inflammatory response to damage. We, therefore, assessed skeletal muscle proinflammatory signaling at rest and 24 h after unaccustomed, loaded knee extension contractions that induced modest muscle damage (72% increase in serum creatine kinase) in a cohort of 87 adults across three age groups (AGE40, AGE61, and AGE76). Vastus lateralis muscle gene expression and protein cell signaling of the IL-6 and TNF-α pathways were determined by quantitative PCR and immunoblot analysis. For in vitro studies, cell signaling and fusion capacities were compared among primary myoblasts from young (AGE28) and old (AGE64) donors treated with TNF-α. Muscle expression was higher (1.5- to 2.1-fold) in AGE76 and AGE61 relative to AGE40 for several genes involved in IL-6, TNF-α, and TNF-like weak inducer of apoptosis signaling. Indexes of activation for the proinflammatory transcription factors signal transducer and activator of transcription-3 and NF-κB were highest in AGE76. Resistance loading reduced gene expression of IL-6 receptor, muscle RING finger 1, and atrogin-1, and increased TNF-like weak inducer of apoptosis receptor expression. Donor myoblasts from AGE64 showed impaired differentiation and fusion in standard media and greater NF-κB activation in response to TNF-α treatment (compared with AGE28). We show for the first time that human aging is associated with muscle inflammation susceptibility (i.e., higher basal state of proinflammatory signaling) that is present in both tissue and isolated myogenic cells and likely contributes to the impaired regenerative capacity of skeletal muscle in the older population.

Does habitual dietary intake influence myofiber hypertrophy in response to resistance training? A cluster analysis

Although resistance exercise training (RT) is a common intervention to stimulate muscle protein synthesis and increase skeletal muscle mass, the optimal daily protein and total energy intakes sufficient to support RT-mediated muscle growth are as yet unclear. Further, the efficacy of RT varies widely among adults of all ages and whether this is attributable to interindividual differences in nutrition is not known. To determine if self-selected daily intake of macronutrients and specific components of dietary protein and fat are predictive of the magnitude of RT-mediated muscle growth, detailed 4-day dietary records were analyzed on 60 subjects previously clustered (K-means cluster analysis) as non-, modest, and extreme responders (non, n = 16; mod, n = 29; xtr, n = 15), based on the magnitudes of change in vastus lateralis myofiber cross-sectional area following a 16-week, 3-day-per-week, high-intensity RT. Despite the marked contrast between 60% myofiber hypertrophy in xtr and zero growth in non, we found no differences among response clusters in daily intakes of energy (mean +/- SEM: non 102 +/- 8; mod 111 +/- 6; xtr 109 +/- 5 kJ.kg-1.day-1), protein (non 0.97 +/- 0.08; mod 1.07 +/- 0.07; xtr 1.05 +/- 0.06 g.kg-1.day-1), carbohydrate (non 3.02 +/- 0.24; mod 3.18 +/- 0.20; xtr 3.14 +/- 0.17 g.kg-1.day-1), and fat (non 0.95 +/- 0.09; mod 1.05 +/- 0.08; xtr 1.03 +/- 0.08 g.kg-1.day-1), which generally met or exceeded dietary recommendations. There were no cluster differences in intakes of branched chain amino acids known to stimulate muscle protein synthesis. Using the novel K-means clustering approach, we conclude from this preliminary study that protein and energy intakes were sufficient to facilitate modest and extreme muscle growth during RT and intrinsic or extrinsic factors other than nutrient ingestion apparently impaired the anabolic response in nonresponders.

Dietary protein intake affects albumin fractional synthesis rate in younger and older adults equally

Inclusion of dietary protein in meals and beverages affects the hepatic synthesis of the protein albumin. Besides dietary protein, several factors can influence albumin metabolism and affect plasma albumin. The role of aging in albumin synthesis is unclear. Recent research documents that albumin synthesis rate is influenced comparably in younger and older adults by dietary protein ingestion and changes in dietary protein quantity. This emphasizes the importance for all adults to consume an adequate amount of dietary protein.

Randomized, four-arm, dose-response clinical trial to optimize resistance exercise training for older adults with age-related muscle atrophy

Purpose: The myriad consequences of age‐related muscle atrophy include reduced muscular strength, power, and mobility; increased risk of falls, disability, and metabolic disease; and compromised immune function. At its root, aging muscle atrophy results from a loss of myofibers and atrophy of the remaining type II myofibers. The purpose of this trial (NCT02442479) was to titrate the dose of resistance training (RT) in older adults in an effort to maximize muscle regrowth and gains in muscle function. Methods: A randomized, four‐arm efficacy trial in which four, distinct exercise prescriptions varying in intensity, frequency, and contraction mode/rate were evaluated: (1) high‐resistance concentric‐eccentric training (H) 3 d/week (HHH); (2) H training 2 d/week (HH); (3) 3 d/week mixed model consisting of H training 2 d/week separated by 1 bout of low‐resistance, high‐velocity, concentric only (L) training (HLH); and (4) 2 d/week mixed model consisting of H training 1 d/week and L training 1 d/week (HL). Sixty‐four randomized subjects (65.5 ± 3.6 y) completed the trial. All participants completed the same 4 weeks of pre‐training consisting of 3 d/week followed by 30 weeks of randomized RT. Results: The HLH prescription maximized gains in thigh muscle mass (TMM, primary outcome) and total body lean mass. HLH also showed the greatest gains in knee extension maximum isometric strength, and reduced cardiorespiratory demand during steady‐state walking. HHH was the only prescription that led to increased muscle expression of pro‐inflammatory cytokine receptors and this was associated with a lesser gain in TMM and total body lean mass compared to HLH. The HL prescription induced minimal muscle regrowth and generally lesser gains in muscle performance vs. the other prescriptions. Major conclusions: The HLH prescription offers distinct advantages over the other doses, while the HL program is subpar. Although limited by a relatively small sample size, we conclude from this randomized dose‐response trial that older adults benefit greatly from 2 d/week high‐intensity RT, and may further benefit from inserting an additional weekly bout of low‐load, explosive RT. Trial registration: ClinicalTrials.gov NCT02442479 HighlightsHLH maximized gains in thigh muscle mass and total body lean mass.HLH induced the greatest gains in knee extension maximum isometric strength.HLH reduced cardiorespiratory demand during steady‐state walking.HHH led to increased muscle expression of pro‐inflammatory cytokine receptors.The HL prescription induced minimal muscle regrowth and lesser gains in performance.

The importance of dietary protein for muscle health in inactive, hospitalized older adults

Dietary protein and amino acids are necessary for overall human health. Insufficient protein intake induces a negative protein balance with adverse outcomes such as muscle atrophy and functional decline—outcomes that are worsened in older adults. Furthermore, during inactivity, such as bed rest/hospitalization, skeletal muscle protein synthesis is reduced, protein balance is negative, and older adults lose significant amounts of muscle. Dietary protein and amino acid supplementation (∼30 g protein and ∼3 g leucine) stimulate skeletal muscle protein anabolism in healthy, community‐dwelling older adults and may be considered as possible nutritional interventions to improve the muscle protein balance and potentially support skeletal muscle maintenance in hospitalized older adults. The following is a timely review of metabolic and dietary challenges faced by hospitalized older adults and potential dietary protein and amino acids solutions for maintaining skeletal muscle health during hospitalization‐induced inactivity in this population.

The metabolic fate of isotopically labeled trimethylamine-N-oxide (TMAO) in humans ☆ ☆☆ ★ ★★

Trimethylamine-N-oxide (TMAO) is associated with chronic disease risk. However, little is known about the metabolic fate of dietary TMAO. This study sought to quantitatively elucidate the metabolic fate of orally consumed TMAO in humans. As part of a crossover feeding study, healthy young men (n=40) consumed 50-mg deuterium-labeled methyl d9-TMAO (d9-TMAO), and enrichments of TMAO and its derivatives were measured in blood for 6 h, urine and stool, as well as skeletal muscle in a subset of men (n=6). Plasma d9-TMAO was detected as early as 15 min, increased until 1 h and remained elevated through the 6-h period. TMAO exhibited an estimated turnover time of 5.3 h, and ~96% of the dose was eliminated in urine by 24 h, mainly as d9-TMAO. No d9-TMAO was detected in feces. Notably, d9-TMAO and d9-trimethylamine were detected in skeletal muscle (n=6) at 6 h, and the enrichment ratio of d9-TMAO to d9-trimethylamine was influenced by a genetic variant in flavin-containing monooxygenase isoform 3 (FMO3 G472A). These results suggest that the absorption of orally consumed TMAO is near complete and does not require processing by gut microbes. TMAO exhibits fast turnover in the circulation with the majority being eliminated in urine within 24 h. A small portion of the dose, however, is taken up by extrahepatic tissue in a manner that appears to be under the influence of FMO3 G472A polymorphism. This trial was registered at clinicaltrials.gov as NCT02558673.