Mats I. Nilsson

Simulated resistance training during hindlimb unloading abolishes disuse bone loss and maintains muscle strength.

This study was designed to determine the effectiveness of simulated resistance training (SRT) without weight bearing in attenuating bone and muscle loss during 28 day hindlimb unloading (HU) in mature male rats. An ambulatory control group (CC) and four groups of HU rats were used: HU, HU + anesthesia (ANHU), HU + eccentric muscle contractions (HU + ECC), and HU + isometric and eccentric muscle contractions (HU + ISO/ECC). Animals in the two SRT groups were trained once every other day at 100% daily peak isometric torque (P0). HU resulted in significantly lower plantarflexor muscle mass (−33% versus CC) and reduced isometric strength (−10%), which reductions were partially attenuated in both training groups. Significantly reduced total and cancellous volumetric bone mineral density (vBMD) and total bone mineral content (BMC) at the proximal tibia metaphysis (PTM) also was evidenced in HU and ANHU groups compared with both SRT groups (p < .05). Training resulted in greater increases in cortical bone mass and area compared with all other groups (p < .05). Fourfold higher material properties of PTM cancellous bone were demonstrated in SRT animals versus HU or CC animals. A significant reduction in midshaft periosteal bone formation rate (BFR) in the HU group (−99% versus CC) was completely abolished in HU + ECC (+656% versus CC). These results demonstrate that high‐intensity muscle contractions, independent of weight‐bearing forces, can effectively mitigate losses in muscle strength and provide a potent stimulus to bone during prolonged disuse.

Abnormal protein turnover and anabolic resistance to exercise in sarcopenic obesity

Obesity may impair protein synthesis rates and cause anabolic resistance to growth factors, hormones, and exercise, ultimately affecting skeletal muscle mass and function. To better understand muscle wasting and anabolic resistance with obesity, we assessed protein 24‐h fractional synthesis rates (24‐h FSRs) in selected hind‐limb muscles of sedentary and resistance‐exercised lean and obese Zucker rats. Despite atrophied hind‐limb muscles (–28% vs. lean rats), 24‐h FSRs of mixed proteins were significantly higher in quadriceps (+18%) and red or white gastrocnemius (+22 or +38%, respectively) of obese animals when compared to lean littermates. Basal synthesis rates of myofibrillar (+8%) and mitochondrial proteins (–1%) in quadriceps were not different between phenotypes, while manufacture of cytosolic proteins (+12%) was moderately elevated in obese cohorts. Western blot analyses revealed a robust activation of p70S6k (+178%) and a lower expression of the endogenous mTOR inhibitor DEPTOR (–28%) in obese rats, collectively suggesting that there is an obesity‐induced increase in net protein turnover favoring degradation. Lastly, the protein synthetic response to exercise of mixed (–7%), myofibrillar (+6%), and cytosolic (+7%) quadriceps subfractions was blunted compared to the lean phenotype (+34, +40, and +17%, respectively), indicating a muscle‐ and subfraction‐specific desensitization to the anabolic stimulus of exercise in obese animals.—Nilsson, M. I., Dobson, J. P., Greene, N. P., Wiggs, M. P., Shimkus, K. L., Wudeck, E. V., Davis, A. R., Laureano, M. L., Fluckey, J. D., Abnormal protein turnover and anabolic resistance to exercise in sarcopenic obesity. FASEB J. 27, 3905–3916 (2013). www.fasebj.org

Insulin resistance syndrome blunts the mitochondrial anabolic response following resistance exercise

Metabolic risk factors associated with insulin resistance syndrome may attenuate augmentations in skeletal muscle protein anabolism following contractile activity. The purpose of this study was to investigate whether or not the anabolic response, as defined by an increase in cumulative fractional protein synthesis rates (24-h FSR) following resistance exercise (RE), is blunted in skeletal muscle of a well-established rodent model of insulin resistance syndrome. Four-month-old lean (Fa/?) and obese (fa/fa) Zucker rats engaged in four lower body RE sessions over 8 days, with the last bout occurring 16 h prior to muscle harvest. A priming dose of deuterium oxide ((2)H(2)O) and (2)H(2)O-enriched drinking water were administered 24 h prior to euthanization for assessment of cumulative FSR. Fractional synthesis rates of mixed (-5%), mitochondrial (-1%), and cytosolic (+15%), but not myofibrillar, proteins (-16%, P = 0.012) were normal or elevated in gastrocnemius muscle of unexercised obese rats. No statistical differences were found in the anabolic response of cytosolic and myofibrillar subfractions between phenotypes, but obese rats were not able to augment 24-h FSR of mitochondria to the same extent as lean rats following RE (+14% vs. +28%, respectively). We conclude that the mature obese Zucker rat exhibits a mild, myofibrillar-specific suppression in basal FSR and a blunted mitochondrial response to contractile activity in mixed gastrocnemius muscle. These findings underscore the importance of assessing synthesis rates of specific myocellular subfractions to fully elucidate perturbations in basal protein turnover rates and differential adaptations to exercise stimuli in metabolic disease.

Simulated resistance training, but not alendronate, increases cortical bone formation and suppresses sclerostin during disuse.

Mechanical loading modulates the osteocyte-derived protein sclerostin, a potent inhibitor of bone formation. We hypothesized that simulated resistance training (SRT), combined with alendronate (ALEN) treatment, during hindlimb unloading (HU) would most effectively mitigate disuse-induced decrements in cortical bone geometry and formation rate (BFR). Sixty male, Sprague-Dawley rats (6-mo-old) were randomly assigned to either cage control (CC), HU, HU plus either ALEN (HU+ALEN), or SRT (HU+SRT), or combined ALEN and SRT (HU+SRT/ALEN) for 28 days. Computed tomography scans on days -1 and 28 were taken at the middiaphyseal tibia. HU+SRT and HU+SRT/ALEN rats were subjected to muscle contractions once every 3 days during HU (4 sets of 5 repetitions; 1,000 ms isometric + 1,000 ms eccentric). The HU+ALEN and HU+SRT/ALEN rats received 10 μg/kg ALEN 3 times/wk. Compared with the CC animals, HU suppressed the normal slow growth-induced increases of cortical bone mineral content, cortical bone area, and polar cross-sectional moment of inertia; however, SRT during HU restored cortical bone growth. HU suppressed middiaphyseal tibia periosteal BFR by 56% vs. CC (P < 0.05). However, SRT during HU restored BFR at both periosteal (to 2.6-fold higher than CC) and endocortical (14-fold higher than CC) surfaces (P < 0.01). ALEN attenuated the SRT-induced BFR gains during HU. The proportion of sclerostin-positive osteocytes in cortical bone was significantly higher (+121% vs. CC) in the HU group; SRT during HU effectively suppressed the higher proportion of sclerostin-positive osteocytes. In conclusion, a minimum number of high-intensity muscle contractions, performed during disuse, restores cortical BFR and suppress unloading-induced increases in sclerostin-positive osteocytes.

Cancellous bone formation response to simulated resistance training during disuse is blunted by concurrent alendronate treatment

The purpose of this study was to assess the effectiveness of simulated resistance training (SRT) exercise combined with alendronate (ALEN) in mitigating or preventing disuse‐associated losses in cancellous bone microarchitecture and formation. Sixty male Sprague‐Dawley rats (6 months old) were randomly assigned to either cage control (CC), hind limb unloading (HU), HU plus either ALEN (HU + ALEN), SRT (HU + SRT), or a combination of ALEN and SRT (HU + SRT/ALEN) for 28 days. HU + SRT and HU + SRT/ALEN rats were anesthetized and subjected to muscle contractions once every 3 days during HU (four sets of five repetitions, 1000 ms isometric + 1000 ms eccentric). Additionally, HU + ALEN and HU + SRT/ALEN rats received 10 µg/kg of body weight of ALEN three times per week. HU reduced cancellous bone‐formation rate (BFR) by 80%, with no effect of ALEN treatment (−85% versus CC). SRT during HU significantly increased cancellous BFR by 123% versus CC, whereas HU + SRT/ALEN inhibited the anabolic effect of SRT (−70% versus HU + SRT). SRT increased bone volume and trabecular thickness by 19% and 9%, respectively, compared with CC. Additionally, osteoid surface (OS/BS) was significantly greater in HU + SRT rats versus CC (+32%). Adding ALEN to SRT during HU reduced Oc.S/BS (−75%), Ob.S/BS (−72%), OS/BS (−61%), and serum TRACP5b (−36%) versus CC. SRT and ALEN each independently suppressed a nearly twofold increase in adipocyte number evidenced with HU and inhibited increases in osteocyte apoptosis. These results demonstrate the anabolic effect of a low volume of high‐intensity muscle contractions during disuse and suggest that both bone resorption and bone formation are suppressed when SRT is combined with bisphosphonate treatment.

microRNA‐16 Is Downregulated During Insulin Resistance and Controls Skeletal Muscle Protein Accretion

Insulin resistant diabetes, currently at epidemic levels in developed countries, begins in the skeletal muscle and is linked to altered protein turnover. microRNAs downregulate targeted mRNA translation decreasing the amount of translated protein, thereby regulating many cellular processes. Regulation of miRNAs and their function in skeletal muscle insulin resistance is largely unexplored. The purpose of this study was to identify the effects of insulin resistance on contents of skeletal muscle miRNAs with potential functions in protein turnover. We examined miRs ‐1, ‐16, ‐23, ‐27, ‐133a, ‐133b, and ‐206 in muscles of Zucker rats. miR‐1 was 5‐ to 10‐fold greater in obesity, whereas miRs‐16 and ‐133b were repressed ∼50% in obese compared to lean rats, with no other alterations in miRNA contents. miR‐16 correlated to protein synthesis in lean, but not obese rats. miR‐16 reduction by lipid overload was verified in‐vivo by diet‐induced obesity and in‐vitro using a diacylglycerol analog. A role for miR‐16 in protein turnover of skeletal myocytes was established using transient overexpression and anti‐miR inhibition. miR‐16 overexpression resulted in lower protein synthesis (puromycin incorporation, ∼25–50%), mTOR (∼25%), and p70S6K1 (∼40%) in starved and insulin stimulated myoblasts. Conversely, anti‐miR‐16 increased basal protein synthesis (puromycin incorporation, ∼75%), mTOR (∼100%), and p70S6K1 (∼100%). Autophagy was enhanced by miR‐16 overexpression (∼50% less BCL‐2, ∼100% greater LC3II/I, ∼50% less p62) and impaired with miR‐16 inhibition (∼45% greater BCL‐2, ∼25% less total LC3, ∼50% greater p62). This study demonstrates reduced miR‐16 during insulin resistance and establishes miR‐16 control of protein accretion in skeletal muscle. J. Cell. Biochem. 117: 1775–1787, 2016.

Impaired exercise-induced mitochondrial biogenesis in the obese Zucker rat, despite PGC-1α induction, is due to compromised mitochondrial translation elongation.

Previously, we demonstrated that high-volume resistance exercise stimulates mitochondrial protein synthesis (a measure of mitochondrial biogenesis) in lean but not obese Zucker rats. Here, we examined factors involved in regulating mitochondrial biogenesis in the same animals. PGC-1α was 45% higher following exercise in obese but not lean animals compared with sedentary counterparts. Interestingly, exercised animals demonstrated greater PPARδ protein in both lean (47%) and obese (>200%) animals. AMPK phosphorylation (300%) and CPT-I protein (30%) were elevated by exercise in lean animals only, indicating improved substrate availability/flux. These findings suggest that, despite PGC-1α induction, obese animals were resistant to exercise-induced synthesis of new mitochondrial and oxidative protein. Previously, we reported that most anabolic processes are upregulated in these same obese animals regardless of exercise, so the purpose of this study was to assess specific factors associated with the mitochondrial genome as possible culprits for impaired mitochondrial biogenesis. Exercise resulted in higher mRNA contents of mitochondrial transcription factor A (∼50% in each phenotype) and mitochondrial translation initiation factor 2 (31 and 47% in lean and obese, respectively). However, mitochondrial translation elongation factor-Tu mRNA was higher following exercise in lean animals only (40%), suggesting aberrant regulation of mitochondrial translation elongation as a possible culprit in impaired mitochondrial biogenesis following exercise with obesity.