MicroRNAs (miRNA), small noncoding RNA molecules, may regulate protein synthesis, while resistance exercise training (RT) is an efficient strategy for stimulating muscle protein synthesis in vivo. However, RT increases muscle mass, with a very wide range of effectiveness in humans. We therefore determined the expression level of 21 abundant miRNAs to determine whether variation in these miRNAs was able to explain the variation in RT-induced gains in muscle mass. Vastus lateralis biopsies were obtained from the top and bottom ∼20% of responders from 56 young men who undertook a 5 day/wk RT program for 12 wk. Training-induced muscle mass gain was determined by dual-energy X-ray absorptiometry, and fiber size was evaluated by histochemistry. The expression level of each miRNA was quantified using TaqMan-based quantitative PCR, with the analysis carried out in a blinded manner. Gene ontology and target gene profiling were used to predict the potential biological implications. Of the 21 mature miRNAs examined, 17 were stable during RT in both groups. However, miR-378, miR-29a, miR-26a, and miR-451 were differentially expressed between low and high responders. miR-378, miR-29a, and miR-26a were downregulated in low responders and unchanged in high responders, while miR-451 was upregulated only in low responders. Interestingly, the training-induced change in miR-378 abundance was positively correlated with muscle mass gains in vivo. Gene ontology analysis of the target gene list of miR-378, miR-29a, miR-26a, and miR-451, from the weighted cumulative context ranking methodology, indicated that miRNA changes in the low responders may be compensatory, reflecting a failure to “activate” growth and remodeling genes. We report, for the first time, that RT-induced hypertrophy in human skeletal muscle is associated with selected changes in miRNA abundance. Our analysis indicates that miRNAs may play a role in the phenotypic change and pronounced intergroup variation in the RT response.

Diagnostics of the human ageing process may help predict future healthcare needs or guide preventative measures for tackling diseases of older age. We take a transcriptomics approach to build the first reproducible multi-tissue RNA expression signature by gene-chip profiling tissue from sedentary normal subjects who reached 65 years of age in good health. One hundred and fifty probe-sets form an accurate classifier of young versus older muscle tissue and this healthy ageing RNA classifier performed consistently in independent cohorts of human muscle, skin and brain tissue (n = 594, AUC = 0.83–0.96) and thus represents a biomarker for biological age. Using the Uppsala Longitudinal Study of Adult Men birth-cohort (n = 108) we demonstrate that the RNA classifier is insensitive to confounding lifestyle biomarkers, while greater gene score at age 70 years is independently associated with better renal function at age 82 years and longevity. The gene score is ‘up-regulated’ in healthy human hippocampus with age, and when applied to blood RNA profiles from two large independent age-matched dementia case–control data sets (n = 717) the healthy controls have significantly greater gene scores than those with cognitive impairment. Alone, or when combined with our previously described prototype Alzheimer disease (AD) RNA ‘disease signature’, the healthy ageing RNA classifier is diagnostic for AD. We identify a novel and statistically robust multi-tissue RNA signature of human healthy ageing that can act as a diagnostic of future health, using only a peripheral blood sample. This RNA signature has great potential to assist research aimed at finding treatments for and/or management of AD and other ageing-related conditions.

Alternatively activated macrophages (AAMΦ) are found in abundance during chronic Th2 inflammatory responses to metazoan parasites. Important roles for these macrophages are being defined, particularly in the context of Th2-mediated pathology and fibrosis. However, a full understanding of the requirements for alternative activation, particularly at the innate level, is lacking. We present evidence that alternative activation by the Th2 cytokines IL-4 and IL-13 is an innate and rapid response to tissue injury that takes place even in the absence of an infectious agent. This early response does not require CD4⁺ Th2 cells because it occurred in RAG-deficient mice. However, class II-restricted CD4⁺ T cell help is essential to maintain AAMΦ in response to infection, because AAMΦ were absent in RAG-deficient and MHC class II-deficient, but not B cell-deficient mice after chronic exposure to the nematode parasite, Brugia malayi. The absence of AAMΦ was associated with increased neutrophilia and reduced eosinophilia, suggesting that AAMΦ are involved in the clearance of neutrophils as well as the recruitment of eosinophils. Consistent with this hypothesis, AAMΦ show enhanced phagocytosis of apoptotic neutrophils, but not latex beads. Our data demonstrate that alternative activation by type 2 cytokines is an innate response to injury that can occur in the absence of an adaptive response. However, analogous to classical activation by microbial pathogens, Th2 cells are required for maintenance and full activation during the ongoing response to metazoan parasites.

The molecular pathways that are activated and contribute to physiological remodeling of skeletal muscle in response to endurance exercise have not been fully characterized. We previously reported that ∼800 gene transcripts are regulated following 6 wk of supervised endurance training in young sedentary males, referred to as the training-responsive transcriptome (TRT) (Timmons JA et al. J Appl Physiol 108: 1487–1496, 2010). Here we utilized this database together with data on biological variation in muscle adaptation to aerobic endurance training in both humans and a novel out-bred rodent model to study the potential regulatory molecules that coordinate this complex network of genes. We identified three DNA sequences representing RUNX1, SOX9, and PAX3 transcription factor binding sites as overrepresented in the TRT. In turn, miRNA profiling indicated that several miRNAs targeting RUNX1, SOX9, and PAX3 were downregulated by endurance training. The TRT was then examined by contrasting subjects who demonstrated the least vs. the greatest improvement in aerobic capacity (low vs. high responders), and at least 100 of the 800 TRT genes were differentially regulated, thus suggesting regulation of these genes may be important for improving aerobic capacity. In high responders, proangiogenic and tissue developmental networks emerged as key candidates for coordinating tissue aerobic adaptation. Beyond RNA-level validation there were several DNA variants that associated with maximal aerobic capacity (V̇o_2max) trainability in the HERITAGE Family Study but these did not pass conservative Bonferroni adjustment. In addition, in a rat model selected across 10 generations for high aerobic training responsiveness, we found that both the TRT and a homologous subset of the human high responder genes were regulated to a greater degree in high responder rodent skeletal muscle. This analysis provides a comprehensive map of the transcriptomic features important for aerobic exercise-induced improvements in maximal oxygen consumption.

Skeletal muscle insulin resistance (IR) is considered a critical component of type II diabetes, yet to date IR has evaded characterization at the global gene expression level in humans. MicroRNAs (miRNAs) are considered fine-scale rheostats of protein-coding gene product abundance. The relative importance and mode of action of miRNAs in human complex diseases remains to be fully elucidated. We produce a global map of coding and non-coding RNAs in human muscle IR with the aim of identifying novel disease biomarkers. We profiled >47,000 mRNA sequences and >500 human miRNAs using gene-chips and 118 subjects (n = 71 patients versus n = 47 controls). A tissue-specific gene-ranking system was developed to stratify thousands of miRNA target-genes, removing false positives, yielding a weighted inhibitor score, which integrated the net impact of both up- and down-regulated miRNAs. Both informatic and protein detection validation was used to verify the predictions of in vivo changes. The muscle mRNA transcriptome is invariant with respect to insulin or glucose homeostasis. In contrast, a third of miRNAs detected in muscle were altered in disease (n = 62), many changing prior to the onset of clinical diabetes. The novel ranking metric identified six canonical pathways with proven links to metabolic disease while the control data demonstrated no enrichment. The Benjamini-Hochberg adjusted Gene Ontology profile of the highest ranked targets was metabolic (P < 7.4 × 10-8), post-translational modification (P < 9.7 × 10-5) and developmental (P < 1.3 × 10-6) processes. Protein profiling of six development-related genes validated the predictions. Brain-derived neurotrophic factor protein was detectable only in muscle satellite cells and was increased in diabetes patients compared with controls, consistent with the observation that global miRNA changes were opposite from those found during myogenic differentiation. We provide evidence that IR in humans may be related to coordinated changes in multiple microRNAs, which act to target relevant signaling pathways. It would appear that miRNAs can produce marked changes in target protein abundance in vivo by working in a combinatorial manner. Thus, miRNA detection represents a new molecular biomarker strategy for insulin resistance, where micrograms of patient material is needed to monitor efficacy during drug or life-style interventions.

Ym1 and Fizz1 are secreted proteins that have been identified in a variety of Th2-mediated inflammatory settings. We originally found Ym1 and Fizz1 as highly expressed macrophage genes in aBrugia malayiinfection model. Here, we show that their expression is a generalized feature of nematode infection and that they are induced at the site of infection with both the tissue nematodeLitomosoides sigmodontisand the gastrointestinal nematodeNippostrongylus brasiliensis. At the sites of infection withN. brasiliensis, we also observed induction of other chitinase and Fizz family members (ChaFFs): acidic mammalian chitinase (AMCase) and Fizz2. The high expression of both Ym1 and AMCase in the lungs of infected mice suggests that abundant chitinase production is an important feature of Th2 immune responses in the lung. In addition to expression of ChaFFs in the tissues, Ym1 and Fizz1 expression was observed in the lymph nodes. Expression both in vitro and in vivo was restricted to antigen-presenting cells, with the highest expression in B cells and macrophages. ChaFFs may therefore be important effector or wound-repair molecules at the site of nematode infection, with potential regulatory roles for Ym1 and Fizz1 in the draining lymph nodes.