Michael De Lisio

Resistance exercise‐induced increases in putative anabolic hormones do not enhance muscle protein synthesis or intracellular signalling in young men

We aimed to determine whether exercise‐induced elevations in systemic concentration of testosterone, growth hormone (GH) and insulin‐like growth factor‐1 (IGF‐1) enhanced post‐exercise myofibrillar protein synthesis (MPS) and phosphorylation of signalling proteins important in regulating mRNA translation. Eight young men (20 ± 1.1 years, BMI = 26 ± 3.5 kg m−2) completed two exercise protocols designed to maintain basal hormone concentrations (low hormone, LH) or elicit increases in endogenous hormones (high hormone, HH). In the LH protocol, participants performed a bout of unilateral resistance exercise with the elbow flexors. The HH protocol consisted of the same elbow flexor exercise with the contralateral arm followed immediately by high‐volume leg resistance exercise. Participants consumed 25 g of protein after arm exercise to maximize MPS. Muscle biopsies and blood samples were taken as appropriate. There were no changes in serum testosterone, GH or IGF‐1 after the LH protocol, whereas there were marked elevations after HH (testosterone, P < 0.001; GH, P < 0.001; IGF‐1, P < 0.05). Exercise stimulated a rise in MPS in the biceps brachii (rest = 0.040 ± 0.007, LH = 0.071 ± 0.008, HH = 0.064 ± 0.014% h−1; P < 0.05) with no effect of elevated hormones (P= 0.72). Phosphorylation of the 70 kDa S6 protein kinase (p70S6K) also increased post‐exercise (P < 0.05) with no differences between conditions. We conclude that the transient increases in endogenous purportedly anabolic hormones do not enhance fed‐state anabolic signalling or MPS following resistance exercise. Local mechanisms are likely to be of predominant importance for the post‐exercise increase in MPS.

Association of interleukin-6 signalling with the muscle stem cell response following muscle-lengthening contractions in humans.

Background The regulation of muscle stem cells in humans in response to muscle injury remains largely undefined. Recently, interleukin-6 (IL-6) has been implicated in muscle stem cell (satellite cell)-mediated muscle hypertrophy in animals; however, the role of IL-6 in the satellite cell (SC) response following muscle-lengthening contractions in humans has not been studied. Methodology/Principal Findings Eight subjects (age 22±1 y; 79±8 kg) performed 300 maximal unilateral lengthening contractions (3.14 rad.s−1) of the knee extensors. Blood and muscle samples were collected before and at 4, 24, 72, and 120 hours post intervention. IL-6, IL-6 receptor (IL-6Rα), cyclin D1, suppressor of cytokine signling-3 (SOCS3) mRNA were measured using quantitative RT-PCR and serum IL-6 protein was measured using an ELISA kit. JAK2 and STAT3 phosphorylated and total protein was measured using western blotting techniques. Immunohistochemical analysis of muscle cross-sections was performed for the quantification of SCs (Pax7+ cells) as well as the expression of phosphorylated STAT3, IL-6, IL-6Rα, and PCNA across all time-points. The SC response, as defined by an amplification of Pax7+ cells, was rapid, increasing by 24 h and peaking 72 h following the intervention. Muscle IL-6 mRNA increased following the intervention, which correlated strongly (R2 = 0.89, p<0.002) with an increase in serum IL-6 concentration. SC IL-6Rα protein was expressed on the fiber, but was also localized to the SC, and IL-6+ SC increased rapidly following muscle-lengthening contractions and returned to basal levels by 72 h post-intervention, demonstrating an acute temporal expression of IL-6 with SC. Phosphorylated STAT3 was evident in SCs 4 h after lengthening contraction, and the downstream genes, cyclin D1 and SOCS3 were significantly elevated 24 hours after the intervention. Conclusions/Significance The increased expression of STAT3 responsive genes and expression of IL-6 within SCs demonstrate that IL-6/STAT3 signaling occurred in SCs, correlating with an increase in SC proliferation, evidenced by increased Pax7+/PCNA+ cell number in the early stages of the time-course. Collectively, these data illustrate that IL-6 is an important signaling molecule associated with the SC response to acute muscle-lengthening contractions in humans.

Resistance training, sarcopenia, and the mitochondrial theory of aging

Skeletal muscle aging is associated with a significant loss of muscle mass, strength, function, and quality of life. In addition, the healthcare cost of aging and age-related disease is growing, and will continue to grow as a larger proportion of our population reaches retirement age and beyond. The mitochondrial theory of aging has been identified as a leading explanation of the aging process and describes a path leading to cellular senescence that includes electron transport chain deficiency, reactive oxygen species production, and the accumulation of mitochondrial DNA deletions and mutations. It is also quite clear that regular resistance exercise is a potent and effective countermeasure for skeletal muscle aging. In this review, we discuss age-related sarcopenia, the mitochondrial theory of aging, and how resistance exercise may directly affect key components of the mitochondrial theory. It is clear from the data discussed that regular resistance training can effectively disturb processes that contribute to the progression of aging as it pertains to the mitochondrial theory.

IL-6 induced STAT3 signalling is associated with the proliferation of human muscle satellite cells following acute muscle damage.

Background Although the satellite cell (SC) is a key regulator of muscle growth during development and muscle adaptation following exercise, the regulation of human muscle SC function remains largely unexplored. STAT3 signalling mediated via interleukin-6 (IL-6) has recently come to the forefront as a potential regulator of SC proliferation. The early response of the SC population in human muscle to muscle-lengthening contractions (MLC) as mediated by STAT3 has not been studied. Methodology/Principal Findings Twelve male subjects (21±2 y; 83±12 kg) performed 300 maximal MLC of the quadriceps femoris at 180°•s−1 over a 55° range of motion with muscle samples (vastus lateralis) and blood samples (antecubital vein) taken prior to exercise (PRE), 1 hour (T1), 3 hours (T3) and 24 hours (T24) post-exercise. Cytoplasmic and nuclear fractions of muscle biopsies were purified and analyzed for total and phosphorylated STAT3 (p-STAT3) by western blot. p-STAT3 was detected in cytoplasmic fractions across the time course peaking at T24 (p<0.01 vs. PRE). Nuclear total and p-STAT3 were not detected at appreciable levels. However, immunohistochemical analysis revealed a progressive increase in the proportion of SCs expressing p-STAT3 with ∼60% of all SCs positive for p-STAT3 at T24 (p<0.001 vs. PRE). Additionally, cMyc, a STAT3 downstream gene, was significantly up-regulated in SCs at T24 versus PRE (p<0.05). Whole muscle mRNA analysis revealed induction of the STAT3 target genes IL-6, SOCS3, cMyc (peaking at T3, p<0.05), IL-6Rα and GP130 (peaking at T24, p<0.05). In addition, Myf5 mRNA was up-regulated at T24 (p<0.05) with no appreciable change in MRF4 mRNA. Conclusions/Significant Findings We demonstrate that IL-6 induction of STAT3 signaling occurred exclusively in the nuclei of SCs in response to MLC. An increase in the number of cMyc+ SCs indicated that human SCs were induced to proliferate under the control of STAT3 signaling.

Biological Effects and Adaptive Response from Single and Repeated Computed Tomography Scans in Reticulocytes and Bone Marrow of C57BL/6 Mice

This study investigated the biological effects and adaptive responses induced by single and repeated in vivo computed tomography (CT) scans. We postulated that, through the induction of low-level oxidative stress, repeated low-dose CT scans (20 mGy, 2 days/week, 10 weeks) could protect mice (C57BL/6) from acute effects of high-dose radiation (1 Gy, 2 Gy). The micronucleated reticulocyte (MN-RET) count increased linearly after exposure to single CT scans of doses ranging from 20 to 80 mGy (P = 0.033). Ten weeks of repeated CT scans (total dose 400 mGy) produced a slight reduction in spontaneous MN-RET levels relative to levels in sham CT-scanned mice (P = 0.04). Decreases of nearly 10% in γ-H2AX fluorescence levels were observed in the repeated CT-scanned mice after an in vitro challenge dose of 1 Gy (P = 0.017) and 2 Gy (P = 0.026). Spontaneous apoptosis levels (caspase 3 and 7 activation) were also significantly lower in the repeated CT-scanned mice than the sham CT-scanned mice (P < 0.01). In contrast, mice receiving only a single CT scan showed a 19% elevation in apoptosis (P < 0.02) and a 10% increase in γ-H2AX fluorescence levels after a 2-Gy challenge (P < 0.05) relative to sham CT controls. Overall, repeated CT scans seemed to confer resistance to larger doses in mice, whereas mice exposed to single CT scans exhibited transient genotoxicity, enhanced apoptosis, and characteristics of radiation sensitization.

Endurance exercise training promotes medullary hematopoiesis

Endurance exercise is a poorly defined yet powerful mediator of hematopoiesis. The purpose of this study was to directly investigate the effects of endurance exercise training on hematopoiesis and to identify potential mechanisms responsible for any observed changes. Four‐week‐old male C57Bl/6 mice were trained on a treadmill at progressive speeds over a 10‐wk period. Tissues were harvested 2 d following the final training session. Flow cytometry, the cobblestone area‐forming cell assay, and the methycellulose colony‐forming unit assay were used to assess medullary and mobilized hematopoietic stem and progenitor cells. Quantitative real‐time PCR and Western blots were used to measure hematopoietic cytokine production. Histochemistry was also used to assess adaptations to exercise in the bone marrow niche. Depending on the cell type, endurance training increased medullary and mobilized hematopoietic stem and progenitor cell content from 50 to 800%. Training also reduced marrow cavity fat by 78%. Skeletal muscle hematopoietic cytokine expression was also increased at least 60% by training. Sedentary mice served as controls for the above experiments. In conclusion, endurance exercise training greatly promotes hematopoiesis and does so through improvements in medullary niche architecture as well as increased skeletal muscle hematopoietic cytokine production.—Baker, J. M., De Lisio, M., Parise, G. Endurance exercise training promotes medullary hematopoiesis. FASEB J. 25, 4348–4357 (2011). www.fasebj.org

Regulation of Muscle Satellite Cell Activation and Chemotaxis by Angiotensin II

The role of angiotensin II (Ang II) in skeletal muscle is poorly understood. We report that pharmacological inhibition of Ang II signaling or ablation of the AT1a receptor significantly impaired skeletal muscle growth following myotrauma, in vivo, likely due to impaired satellite cell activation and chemotaxis. In vitro experiments demonstrated that Ang II treatment activated quiescent myoblasts as evidenced by the upregulation of myogenic regulatory factors, increased number of β-gal+, Myf5-LacZ myoblasts and the acquisition of cellular motility. Furthermore, exogenous treatment with Ang II significantly increased the chemotactic capacity of C2C12 and primary cells while AT1a−/− myoblasts demonstrated a severe impairment in basal migration and were not responsive to Ang II treatment. Additionally, Ang II interacted with myoblasts in a paracrine-mediated fashion as 4 h of cyclic mechanical stimulation resulted in Ang II-induced migration of cocultured myoblasts. Ang II-induced chemotaxis appeared to be regulated by multiple mechanisms including reorganization of the actin cytoskeleton and augmentation of MMP2 activity. Collectively, these results highlight a novel role for Ang II and ACE inhibitors in the regulation of skeletal muscle growth and satellite cell function.

Exercise-induced mitochondrial p53 repairs mtDNA mutations in mutator mice

BackgroundHuman genetic disorders and transgenic mouse models have shown that mitochondrial DNA (mtDNA) mutations and telomere dysfunction instigate the aging process. Epidemiologically, exercise is associated with greater life expectancy and reduced risk of chronic diseases. While the beneficial effects of exercise are well established, the molecular mechanisms instigating these observations remain unclear.ResultsEndurance exercise reduces mtDNA mutation burden, alleviates multisystem pathology, and increases lifespan of the mutator mice, with proofreading deficient mitochondrial polymerase gamma (POLG1). We report evidence for a POLG1-independent mtDNA repair pathway mediated by exercise, a surprising notion as POLG1 is canonically considered to be the sole mtDNA repair enzyme. Here, we show that the tumor suppressor protein p53 translocates to mitochondria and facilitates mtDNA mutation repair and mitochondrial biogenesis in response to endurance exercise. Indeed, in mutator mice with muscle-specific deletion of p53, exercise failed to prevent mtDNA mutations, induce mitochondrial biogenesis, preserve mitochondrial morphology, reverse sarcopenia, or mitigate premature mortality.ConclusionsOur data establish a new role for p53 in exercise-mediated maintenance of the mtDNA genome and present mitochondrially targeted p53 as a novel therapeutic modality for diseases of mitochondrial etiology.

Skeletal muscle myoblasts possess a stretch-responsive local angiotensin signalling system.

A paucity of information exists regarding the presence of local renin—angiotensin systems (RASs) in skeletal muscle and associated muscle stem cells. Skeletal muscle and muscle stem cells were isolated from C57BL/6 mice and examined for the presence of a local RAS using quantitative reverse transcriptase polymerase chain reaction (qRT-PCR), immunohistochemistry (IHC), Western blotting and liquid chromatography-mass spectrometry (LC-MS). Furthermore, the effect of mechanical stimulation on RAS member gene expression was analysed. Whole skeletal muscle, primary myoblasts and C2C12 derived myoblasts and myotubes differentially expressed members of the RAS including angiotensinogen, angiotensin-converting enzyme (ACE), angiotensin II (Ang II) type 1 (AT1) and type 2 (AT2). Renin transcripts were never detected, however, mRNA for the ‘renin-like’ enzyme cathepsin D was observed and Ang I and Ang II were identified in cell culture supernatants from proliferating myoblasts. AT1 appeared to co-localise with polymerised actin filaments in proliferating myoblasts and was primarily found in the nucleus of terminally differentiated myotubes. Furthermore, mechanical stretch of proliferating and differentiating C2C12 cells differentially induced mRNA expression of angiotensinogen, AT 1 and AT2. Proliferating and differentiated muscle stem cells possess a local stress-responsive RAS in vitro. The precise function of a local RAS in myoblasts remains unknown. However, evidence presented here suggests that Ang II may be a regulator of skeletal muscle myoblasts.

Defining a role for non-satellite stem cells in the regulation of muscle repair following exercise

Skeletal muscle repair is essential for effective remodeling, tissue maintenance, and initiation of beneficial adaptations post-eccentric exercise. A series of well characterized events, such as recruitment of immune cells and activation of satellite cells, constitute the basis for muscle regeneration. However, details regarding the fine-tuned regulation of this process in response to different types of injury are open for investigation. Muscle-resident non-myogenic, non-satellite stem cells expressing conventional mesenchymal stem cell (MSC) markers, have the potential to significantly contribute to regeneration given the role for bone marrow-derived MSCs in whole body tissue repair in response to injury and disease. The purpose of this mini-review is to highlight a regulatory role for Pnon-satellite stem cells in the process of skeletal muscle healing post-eccentric exercise. The non-myogenic, non-satellite stem cell fraction will be defined, its role in tissue repair will be briefly reviewed, and recent studies demonstrating a contribution to eccentric exercise-induced regeneration will be presented.