Hormonal and metabolic responses of older adults to resistance training in normobaric hypoxia.

PURPOSE: In young adults, the hormonal responses to resistance exercise are amplified by normobaric hypoxia. Hormone concentrations and metabolism are typically dysregulated with age, yet the impact of hypoxia on these responses to resistance exercise are uncharacterised. Therefore, this study aimed to characterise the acute and chronic hormonal and metabolic responses of older adults to resistance training in normobaric hypoxia. METHODS: Adults aged 60-75 years completed 8 weeks of resistance training in either normoxia (20.9% O2; n = 10) or normobaric hypoxia (14.4% O2, n = 10) twice weekly at 70% of their predicted 1-repetition maximum. Growth hormone, glucose, lactate, insulin, homeostatic model assessment of insulin resistance (HOMA-IR), cortisol, total testosterone, adrenaline, noradrenaline and dopamine were quantified at pre- and post-training, and in the 60 min following the first training session (untrained state) and the last training session (trained state). RESULTS: Eight weeks of training in hypoxia did not affect the resting levels of the hormones or physiological factors measured. However, hypoxia significantly blunted the acute growth hormone response in the 15 min following the last training session at week eight (43.87% lower in the hypoxic group; p = 0.017). This novel and unexpected finding requires further investigation. All other hormones were unaffected acutely by hypoxia in the 60 min following the first and the last training session. CONCLUSION: Chronic resistance training in normobaric hypoxia supresses the growth hormone response to exercise in older adults. All other hormones and metabolic markers were unaffected both acutely and chronically by hypoxia.

Sustained cardiac programming by short-term juvenile exercise training in male rats.

KEY POINTS: Cardiac hypertrophy following endurance-training is thought to be due to hypertrophy of existing cardiomyocytes. The benefits of endurance exercise on cardiac hypertrophy are generally thought to be short-lived and regress to sedentary levels within a few weeks of stopping endurance training. We have now established that cardiomyocyte hyperplasia also plays a considerable role in cardiac growth in response to just 4 weeks of endurance exercise in juvenile (5-9 weeks of age) rats. The effect of endurance exercise on cardiomyocyte hyperplasia diminishes with age and is lost by adulthood. We have also established that the effect of juvenile exercise on heart mass is sustained into adulthood. ABSTRACT: The aim of this study was to investigate if endurance training during juvenile life 'reprogrammes' the heart and leads to sustained improvements in the structure, function, and morphology of the adult heart. Male Wistar Kyoto rats were exercise trained 5 days week-1 for 4 weeks in either juvenile (5-9 weeks of age), adolescent (11-15 weeks of age) or adult life (20-24 weeks of age). Juvenile exercise training, when compared to 24-week-old sedentary rats, led to sustained increases in left ventricle (LV) mass (+18%; P < 0.05), wall thickness (+11%; P < 0.05), the longitudinal area of binucleated cardiomyocytes (P < 0.05), cardiomyocyte number (+36%; P < 0.05), and doubled the proportion of mononucleated cardiomyocytes (P < 0.05), with a less pronounced effect of exercise during adolescent life. Adult exercise training also increased LV mass (+11%; P < 0.05), wall thickness (+6%; P < 0.05) and the longitudinal area of binucleated cardiomyocytes (P < 0.05), despite no change in cardiomyocyte number or the proportion of mono- and binucleated cardiomyocytes. Resting cardiac function, LV chamber dimensions and fibrosis levels were not altered by juvenile or adult exercise training. At 9 weeks of age, juvenile exercise significantly reduced the expression of microRNA-208b, which is a known regulator of cardiac growth, but this was not sustained to 24 weeks of age. In conclusion, juvenile exercise leads to physiological cardiac hypertrophy that is sustained into adulthood long after exercise training has ceased. Furthermore, this cardiac reprogramming is largely due to a 36% increase in cardiomyocyte number, which results in an additional 20 million cardiomyocytes in adulthood.

Repeated evolution of digital adhesion in geckos: a reply to Harrington and Reeder.

We published a phylogenetic comparative analysis that found geckos had gained and lost adhesive toepads multiple times over their long evolutionary history (Gamble et al., PLoS One, 7, 2012, e39429). This was consistent with decades of morphological studies showing geckos had evolved adhesive toepads on multiple occasions and that the morphology of geckos with ancestrally padless digits can be distinguished from secondarily padless forms. Recently, Harrington & Reeder (J. Evol. Biol., 30, 2017, 313) reanalysed data from Gamble et al. (PLoS One, 7, 2012, e39429) and found little support for the multiple origins hypothesis. Here, we argue that Harrington and Reeder failed to take morphological evidence into account when devising ancestral state reconstruction models and that these biologically unrealistic models led to erroneous conclusions about the evolution of adhesive toepads in geckos.

Striated muscle activator of Rho signalling (STARS) is reduced in ageing human skeletal muscle and targeted by miR-628-5p.

AIM: The striated muscle activator of Rho signalling (STARS) is a muscle-specific actin-binding protein. The STARS signalling pathway is activated by resistance exercise and is anticipated to play a role in signal mechanotransduction. Animal studies have reported a negative regulation of STARS signalling with age, but such regulation has not been investigated in humans. METHODS: Ten young (18-30 years) and 10 older (60-75 years) subjects completed an acute bout of resistance exercise. Gene and protein expression of members of the STARS signalling pathway and miRNA expression of a subset of miRNAs, predicted or known to target members of STARS signalling pathway, were measured in muscle biopsies collected pre-exercise and 2 h post-exercise. RESULTS: For the first time, we report a significant downregulation of the STARS protein in older subjects. However, there was no effect of age on the magnitude of STARS activation in response to an acute bout of exercise. Finally, we established that miR-628-5p, a miRNA regulated by age and exercise, binds to the STARS 3'UTR to directly downregulate its transcription. CONCLUSION: This study describes for the first time the resistance exercise-induced regulation of STARS signalling in skeletal muscle from older humans and identifies a new miRNA involved in the transcriptional control of STARS.

Armored geckos: A histological investigation of osteoderm development in Tarentola (Phyllodactylidae) and Gekko (Gekkonidae) with comments on their regeneration and inferred function.

Osteoderms are bone-rich organs found in the dermis of many scleroglossan lizards sensu lato, but are only known for two genera of gekkotans (geckos): Tarentola and Gekko. Here, we investigate their sequence of appearance, mode of development, structural diversity and ability to regenerate following tail loss. Osteoderms were present in all species of Tarentola sampled (Tarentola annularis, T. mauritanica, T. americana, T. crombei, T. chazaliae) as well as Gekko gecko, but not G. smithii. Gekkotan osteoderms first appear within the integument dorsal to the frontal bone or within the supraocular scales. They then manifest as mineralized structures in other positions across the head. In Tarentola and G. gecko, discontinuous clusters subsequently form dorsal to the pelvis/base of the tail, and then dorsal to the pectoral apparatus. Gekkotan osteoderm formation begins once the dermis is fully formed. Early bone deposition appears to involve populations of fibroblast-like cells, which are gradually replaced by more rounded osteoblasts. In T. annularis and T. mauritanica, an additional skeletal tissue is deposited across the superficial surface of the osteoderm. This tissue is vitreous, avascular, cell-poor, lacks intrinsic collagen, and is herein identified as osteodermine. We also report that following tail loss, both T. annularis and T. mauritanica are capable of regenerating osteoderms, including osteodermine, in the regenerated part of the tail. We propose that osteoderms serve roles in defense against combative prey and intraspecific aggression, along with anti-predation functions.

Effective treatment of alkaline Cr(VI) contaminated leachate using a novel Pd-bionanocatalyst: Impact of electron donor and aqueous geochemistry.

Palladium catalysts offer the potential for the effective treatment of a variety of priority reducible pollutants in natural waters. In this study, microbially synthesized magnetite nanoparticles were functionalized with Pd(0), creating a highly reactive, magnetically recoverable, nano-scale catalyst (Pd-BnM). This was then investigated for the treatment of model Cr(VI) contaminated solutions at a range of pH values, and also alkaline Cr(VI) contaminated leachates from chromite ore processing residue (COPR); a contaminant issue of global concern. The sample of COPR used in this study was obtained from a site in Glasgow, UK, where extensive Cr(VI) contamination has been reported. In initial experiments Pd-BnM was supplied with H2 gas or formate as electron donors, and Cr(VI) removal from model synthetic solutions was quantified at various pH values (2-12). Effective removal was noted at neutral to environmentally relevant alkaline (pH 12) pH values, while the use of formate as an electron donor resulted in loss of performance under acidic conditions (pH 2). Reaction kinetics were then assessed with increasing Pd-BnM loading in both model pH 12 Cr(VI) solutions and the COPR leachate. When formate was used as the electron donor for Pd-BnM, to treat COPR leachate, there was significant inhibition of Cr(VI) removal. In contrast, a promotion of reaction rate, was observed when H2 was employed. Upon sustained reaction with model Cr(VI) solutions, in the presence of excess electron donor (formate or H2), appreciable quantities of Cr(VI) were removed before eventual inactivation of the catalyst. Faster onset of inactivation was reported in the COPR leachates, removing 4% and 64% of Cr(VI) observed from model Cr(VI) solutions, when formate and H2 were used as electron donors, respectively. XAS, TEM-EDX and XPS analysis of the catalysts that had been inactivated in the model solution, showed that the surface had an extensive covering of reduced Cr(III), most likely as a CrOOH phase. COPR reacted catalysts recorded a lower abundance of Cr(III) alongside a high abundance of the leachate components Ca and Si, implicating these elements in the faster onset of inactivation.

Biogenic nano-magnetite and nano-zero valent iron treatment of alkaline Cr(VI) leachate and chromite ore processing residue.

Highly reactive nano-scale biogenic magnetite (BnM), synthesized by the Fe(III)-reducing bacterium Geobacter sulfurreducens, was tested for the potential to remediate alkaline Cr(VI) contaminated waters associated with chromite ore processing residue (COPR). The performance of this biomaterial, targeting aqueous Cr(VI) removal, was compared to a synthetic alternative, nano-scale zero valent iron (nZVI). Samples of highly contaminated alkaline groundwater and COPR solid waste were obtained from a contaminated site in Glasgow, UK. During batch reactivity tests, Cr(VI) removal from groundwater was inhibited by ∼25% (BnM) and ∼50% (nZVI) when compared to the treatment of less chemically complex model pH 12 Cr(VI) solutions. In both the model Cr(VI) solutions and contaminated groundwater experiments the surface of the nanoparticles became passivated, preventing complete coupling of their available electrons to Cr(VI) reduction. To investigate this process, the surfaces of the reacted samples were analyzed by TEM-EDX, XAS and XPS, confirming Cr(VI) reduction to the less soluble Cr(III) on the nanoparticle surface. In groundwater reacted samples the presence of Ca, Si and S was also noted on the surface of the nanoparticles, and is likely responsible for earlier onset of passivation. Treatment of the solid COPR material in contact with water, by addition of increasing weight % of the nanoparticles, resulted in a decrease in aqueous Cr(VI) concentrations to below detection limits, via the addition of ⩾5% w/w BnM or ⩾1% w/w nZVI. XANES analysis of the Cr K edge, showed that the % Cr(VI) in the COPR dropped from 26% to a minimum of 4-7% by the addition of 5% w/w BnM or 2% w/w nZVI, with higher additions unable to reduce the remaining Cr(VI). The treated materials exhibited minimal re-mobilization of soluble Cr(VI) by re-equilibration with atmospheric oxygen, with the bulk of the Cr remaining in the solid fraction. Both nanoparticles exhibited a considerable capacity for the remediation of COPR related Cr(VI) contamination, with the synthetic nZVI demonstrating greater reactivity than the BnM. However, the biosynthesized BnM was also capable of significant Cr(VI) reduction and demonstrated a greater efficiency for the coupling of its electrons towards Cr(VI) reduction than the nZVI.

Androgenic and estrogenic regulation of Atrogin-1, MuRF1 and myostatin expression in different muscle types of male mice.

PURPOSE: The molecular factors targeted by androgens and estrogens on muscle mass are not fully understood. The current study aimed to explore gene and protein expression of Atrogin-1, MuRF1, and myostatin in an androgen deprivation-induced muscle atrophy model. METHODS: We examined the effects of Orx either with or without testosterone (T) or estradiol (E2) administration on Atrogin-1 gene expression, and MuRF1 and myostatin gene and protein expression. Measurements were made in soleus (SOL), extensor digitorum longus (EDL) and levator ani/bulbocavernosus (LA/BC) of male C57BL/6 mice. RESULTS: Thirty days of Orx resulted in a reduction in weight gain and muscle mass. These effects were prevented by T. In LA/BC, Atrogin-1 and MuRF1 mRNA was increased throughout 30 days of Orx, which was fully reversed by T and partially by E2 administration. In EDL and SOL, a less pronounced upregulation of both genes was only detectable at the early stages of Orx. Myostatin mRNA levels were downregulated in LA/BC and upregulated in EDL following Orx. T, but not E2, reversed these effects. No changes in protein levels of MuRF1 and myostatin were found in EDL at any time point following Orx. CONCLUSIONS: The atrophy in SOL and EDL in response to androgen deprivation, and its restoration by T, is accompanied by only minimal changes in atrogenes and myostatin gene expression. The marked differences in muscle atrophy and atrogene and myostatin mRNA between LA/BC and the locomotor muscles suggest that the murine LA/BC is not an optimal model to study Orx-induced muscle atrophy.

Delving into disability in Crohn's disease: dysregulation of molecular pathways may explain skeletal muscle loss in Crohn's disease.

BACKGROUND/AIMS: In Crohn's disease (CD), skeletal muscle mass and function are reduced compared to healthy controls, potentially resulting in disability. Mechanisms contributing to muscle impairment, and thus potential therapeutic targets, are poorly understood. This study aimed to measure and compare skeletal muscle size and molecular targets involved in skeletal muscle growth, in CD subjects and healthy controls. METHODS: CD (n=27) and healthy (n=22) subjects were recruited from the IBD outpatient clinic and via local advertisement respectively. Demographics and clinical data were collected via survey and interview. Quadriceps muscle cross-sectional area was measured using peripheral quantitative CT scanning. Levels of muscle hypertrophy and atrophy signalling targets using quantitative PCR and western blotting were measured in muscle biopsies. RESULTS: Muscle size was 14% lower (p=0.055) and a 54% lower phosphorylated:total (p:t) Akt ratio was measured in the muscle samples (p<0.05), indicating an attenuated muscle hypertrophy pathway in CD compared with controls. In those with CD, a lower p:t Akt ratio (<0.97) was associated with lower serum vitamin D3, lower physical activity indices (49 vs 64 mmol/L, 1.7 vs 2.2×10(6) accelerometer counts respectively, each p<0.05) and a trend towards lower serum ferritin levels (128 vs 322mg/L, p=0.07), compared with CD subjects with normal/high p:t Akt ratios. CONCLUSION: The reduced muscle mass in CD may be explained, in part, by impaired activation of muscle protein synthesis pathways, notably the IGF1-Akt pathway. Normal vitamin D levels and regular exercise may be protective in CD against this trend, though confirmatory longitudinal studies are needed.

Objectively measured muscle fatigue in Crohn's disease: correlation with self-reported fatigue and associated factors for clinical application.

BACKGROUND & AIMS: The association of fatigue with decreased physical performance and underlying mechanisms are poorly understood in Crohn's disease (CD). We aimed to measure and compare self-reported fatigue with skeletal muscle fatigue in CD subjects and healthy controls, and to identify associated factors that may be amenable to change. METHODS: Demographic and clinical data were collected and fatigue assessed using the Fatigue Impact Scale (FIS) in 27 consecutive CD patients and 22 matched healthy controls. Circulating cytokines and growth factors were measured. The rate of quadriceps muscle fatigue was assessed using an isokinetic dynamometer as the decrement of force with 30 contractions performed over a 5-minute period. RESULTS: Compared with healthy controls, CD patients reported greater levels of fatigue (mean global FIS score 45.3 vs 10.5, physical dimension score 12.3 vs 2.7 respectively; each p<0.01) and muscle fatigue (-5.2 vs -1.3 Nm min(-1); p<0.05). The two indices were correlated (r = -0.52 in CD; p<0.01). Patients with CD had lower mean serum IGF-1 levels (16.1 vs 25.4 pmol/L, p<0.01) and higher oxidative stress (TBARS assay 4.3 vs 3.9 µM, p<0.05). On multivariate analysis, low serum vitamin D, IGF-1 and magnesium, and higher IL-6 levels were associated with increased muscle fatigue (all p ≤ 0.05). CONCLUSION: Subjects with CD had more muscle fatigue than matched healthy controls and this correlated well with self-reported fatigue. Of circulating factors that were independently associated with increased muscle fatigue, vitamin D, magnesium and IGF-1 could be targeted in future studies to reduce fatigue and improve physical performance.

Brown adipocyte progenitor population is modified in obese and diabetic skeletal muscle.

Brown adipose tissue mitochondria express the unique thermogenic uncoupling protein-1. Recently, brown adipocyte progenitors have been identified in the CD34+ cell population of human skeletal muscle. The aims of this study were firstly to determine if obesity and diabetes have altered amounts of muscle brown adipocyte progenitors and, secondly, to establish if the latter are correlated with clinical parameters of obesity and diabetes. Body mass index (BMI), plasma glucose, insulin, cholesterol and triglycerides as well as homeostasis model assessment were measured in lean (n=10), obese (n=18) and obese-diabetic (n=15) subjects and muscle biopsies were taken from the rectus abdominus. CD34 being also expressed on endothelial cells, we measured CD31, another endothelial marker, and expressed the brown adipocyte progenitors, as the CD34/CD31 mRNA ratio. The latter was significantly reduced in the obese vs lean subjects suggesting a smaller pool of brown adipocyte progenitors. More strikingly, for lean and obese subjects negative correlations were observed between the CD34/CD31 mRNA ratios and BMI, fasting insulin levels and homeostasis model assessment. These correlations highlight the potential physiological relevance of the muscle CD34/CD31 mRNA ratio.

PPARgamma inhibits NF-kappaB-dependent transcriptional activation in skeletal muscle.

Skeletal muscle pathology associated with a chronic inflammatory disease state (e.g., skeletal muscle atrophy and insulin resistance) is a potential consequence of chronic activation of NF-kappaB. It has been demonstrated that peroxisome proliferator-activated receptors (PPARs) can exert anti-inflammatory effects by interfering with transcriptional regulation of inflammatory responses. The goal of the present study, therefore, was to evaluate whether PPAR activation affects cytokine-induced NF-kappaB activity in skeletal muscle. Using C(2)C(12) myotubes as an in vitro model of myofibers, we demonstrate that PPAR, and specifically PPARgamma, activation potently inhibits inflammatory mediator-induced NF-kappaB transcriptional activity in a time- and dose-dependent manner. Furthermore, PPARgamma activation by rosiglitazone strongly suppresses cytokine-induced transcript levels of the NF-kappaB-dependent genes intracellular adhesion molecule 1 (ICAM-1) and CXCL1 (KC), the murine homolog of IL-8, in myotubes. To verify whether muscular NF-kappaB activity in human subjects is suppressed by PPARgamma activation, we examined the effect of 8 wk of rosiglitazone treatment on muscular gene expression of ICAM-1 and IL-8 in type 2 diabetes mellitus patients. In these subjects, we observed a trend toward decreased basal expression of ICAM-1 mRNA levels. Subsequent analyses in cultured myotubes revealed that the anti-inflammatory effect of PPARgamma activation is not due to decreased RelA translocation to the nucleus or reduced RelA DNA binding. These findings demonstrate that muscle-specific inhibition of NF-kappaB activation may be an interesting therapeutic avenue for treatment of several inflammation-associated skeletal muscle abnormalities.

Thrifty metabolism that favors fat storage after caloric restriction: a role for skeletal muscle phosphatidylinositol-3-kinase activity and AMP-activated protein kinase.

Energy conservation directed at accelerating body fat recovery (or catch-up fat) contributes to obesity relapse after slimming and to excess fat gain during catch-up growth after malnutrition. To investigate the mechanisms underlying such thrifty metabolism for catch-up fat, we tested whether during refeeding after caloric restriction rats exhibiting catch-up fat driven by suppressed thermogenesis have diminished skeletal muscle phosphatidylinositol-3-kinase (PI3K) activity or AMP-activated protein kinase (AMPK) signaling-two pathways required for hormone-induced thermogenesis in ex vivo muscle preparations. The results show that during isocaloric refeeding with a low-fat diet, at time points when body fat, circulating free fatty acids, and intramyocellular lipids in refed animals do not exceed those of controls, muscle insulin receptor substrate 1-associated PI3K activity (basal and in vivo insulin-stimulated) is lower than that in controls. Isocaloric refeeding with a high-fat diet, which exacerbates the suppression of thermogenesis, results in further reductions in muscle PI3K activity and in impaired AMPK phosphorylation (basal and in vivo leptin-stimulated). It is proposed that reduced skeletal muscle PI3K/AMPK signaling and suppressed thermogenesis are interdependent. Defective PI3K or AMPK signaling will reduce the rate of substrate cycling between de novo lipogenesis and lipid oxidation, leading to suppressed thermogenesis, which accelerates body fat recovery and furthermore sensitizes skeletal muscle to dietary fat-induced impairments in PI3K/AMPK signaling.

Slow component of VO(2) during level and uphill treadmill running: relationship to aerobic fitness in endurance runners.

AIM: The aim of this study was to compare the oxygen uptake (VO(2)) slow component (SC) during level and uphill running in endurance runners, and to identify associations between the SC and the following aerobic fitness indicators: peak VO(2), running speed associated with the peak VO(2) (Vpeak), running speed at the lactic threshold and the VO(2) fraction elicited at the lactic threshold. METHODS: Fourteen male endurance-trained runners underwent several 6-min bouts of level (LTR) and 10.5% uphill treadmill running. VO(2) SC was calculated as the difference between mean VO(2) during the 6th and the 3rd minutes. RESULTS: The highest mean values for the SC were 181.9+/-240.2 mL x min(-1) for level running at approximately 94% peak VO(2)2 and 105.4+/-154.6 mL x min(-1) for uphill running at approximately 90% peak VO(2). The SC observed during the last bout of the LTR correlated with peak VO(2) and with Vpeak (-0.71 and -0.76, P<0.05, respectively). CONCLUSION: The results show that for endurance-trained runners the magnitude of the SC is not affected by the treadmill gradient and that within a homogeneous sample of endurance-trained runners the SC does not correlate with indicators of aerobic fitness.

Improved skeletal muscle oxidative enzyme activity and restoration of PGC-1 alpha and PPAR beta/delta gene expression upon rosiglitazone treatment in obese patients with type 2 diabetes mellitus.

OBJECTIVE: To examine whether rosiglitazone alters gene expression of some key genes involved in mitochondrial biogenesis and oxidative capacity in skeletal muscle of type 2 diabetic patients, and whether this is associated with alterations in skeletal muscle oxidative capacity and lipid content. DESIGN: measured in muscle biopsies obtained from diabetic patients, before and after 8 weeks of rosiglitazone treatment, and matched controls. Furthermore, whole-body insulin sensitivity and substrate utilization were assessed. SUBJECTS: Ten obese type 2 diabetic patients and 10 obese normoglycemic controls matched for age and BMI. METHODS: Gene expression and mitochondrial protein content of complexes I-V of the respiratory chain were measured by quantitative polymerase chain reaction and Western blotting, respectively. Histochemical staining was used to quantify lipid accumulation and complex II succinate dehydrogenase (SDH) activity. Insulin sensitivity and substrate utilization were measured during a hyperinsulinemic-euglycemic clamp with indirect calorimetry. RESULTS: Skeletal-muscle mRNA of PGC-1 alpha and PPAR beta/delta--but not of other genes involved in glucose, fat and oxidative metabolism--was significantly lower in diabetic patients (P<0.01). Rosiglitazone significantly increased PGC-1 alpha ( approximately 2.2-fold, P<0.01) and PPAR beta/delta ( approximately 2.6-fold, P<0.01), in parallel with an increase in insulin sensitivity, SDH activity and metabolic flexibility (P<0.01). Surprisingly, none of the measured mitochondrial proteins was reduced in type 2 diabetic patients, nor affected by rosiglitazone treatment. No alterations were seen in muscular fat accumulation upon treatment. CONCLUSION: These results suggest that the insulin-sensitizing effect of rosiglitazone may involve an effect on muscular oxidative capacity, via PGC-1 alpha and PPAR beta/delta, independent of mitochondrial protein content and/or changes in intramyocellular lipid.

Peroxisome proliferator-activated receptor-gamma coactivator-1 and insulin resistance: acute effect of fatty acids.

AIMS/HYPOTHESIS: Peroxisome proliferator-activated receptor (PPAR)-gamma coactivator-1 (PPARGC1), a coactivator regulating the transcription of genes involved in oxidative metabolism, is downregulated in patients with type 2 diabetes and in their first-degree relatives. Whether this downregulation is a cause or effect of early aberrations in the development of insulin resistance, such as disturbances in fat metabolism, is unknown. We examined whether lipid-induced insulin resistance was associated with downregulation of expression of skeletal muscle genes involved in oxidative metabolism and mitochondrial biogenesis in humans. MATERIALS AND METHODS: Nine healthy lean male subjects underwent a 6-h hyperinsulinaemic-euglycaemic clamp with simultaneous infusion of either a lipid emulsion or glycerol as a control. Blood was sampled at regular time points and muscle biopsies were taken before and after every test. Intramuscular triacylglycerol (IMTG) content was determined by Oil Red O staining and gene expression was measured by quantitative PCR. RESULTS: Lipid infusion resulted in a approximately 2.7-fold increase in plasma NEFA levels and a 31+/-6% decrease in insulin sensitivity (p=0.001). The infusion of lipids resulted in a approximately 1.6-fold increase in IMTG (p=0.02), whereas during the clamp with glycerol infusion IMTG tended to decrease to approximately 53% of preinfusion levels (p=0.065). Lipid infusion decreased PPARGC1A, PPARGC1B and PPARA expression to approximately 61, 77 and approximately 52% of basal values respectively, whereas expression of uncoupling protein 3 was upregulated 1.8-fold (all p<0.05). CONCLUSIONS/INTERPRETATION: Acute elevation of plasma NEFA levels, leading to muscular fat accumulation and insulin resistance, downregulates PPARGC1A, PPARGC1B and PPARA expression, suggesting that the decrease in PPARGC1 expression observed in the (pre)diabetic state may be the result, rather than the cause of lipid-induced insulin resistance.

Effect of 2 weeks of endurance training on uncoupling protein 3 content in untrained human subjects.

AIM: The mitochondrial uncoupling protein-3 (UCP3) is able to lower the proton gradient across the inner mitochondrial membrane, thereby uncoupling substrate oxidation from ATP production and dissipating energy as heat. What the effect of endurance training on UCP3 is, is still controversial. Endurance-trained athletes are characterized by lower levels of UCP3, but longitudinal studies in rodents reported no effect of endurance training on muscular UCP3 levels. Here, we examined the effect of a 2-week training programme on skeletal muscle UCP3 protein content in untrained human subjects, and hypothesized that UCP3 will be reduced after the training programme. METHODS: Nine untrained men [age: 23.3 +/- 3.2 years; BMI: 22.6 +/- 2.6 kg m(-2); maximal power output (W(max)): 3.8 +/- 0.6 W kg(-1) body weight] trained for 2 weeks. Before and at least 72 h after the training period, muscle biopsies were taken for determination of UCP3 protein content. RESULTS: UCP3 protein content tended to be lower after the training programme [95 +/- 10 vs. 109 +/- 12 arbitrary units (AU), P = 0.08]. Cytochrome c content tended to increase with 33% in response to endurance training (52 +/- 6 vs. 39 +/- 6 AU, P = 0.08). The ratio UCP3 relative to cytochrome c tended to decrease significantly upon endurance training (2.0 +/- 0.4 vs. 3.2 +/- 0.6 AU, P = 0.01). CONCLUSION: A short-term (2-week) endurance training programme decreased UCP3 protein levels and significantly reduced the ratio of UCP3 to cytochrome c.

Exercise in the fasted state facilitates fibre type-specific intramyocellular lipid breakdown and stimulates glycogen resynthesis in humans.

The effects were compared of exercise in the fasted state and exercise with a high rate of carbohydrate intake on intramyocellular triglyceride (IMTG) and glycogen content of human muscle. Using a randomized crossover study design, nine young healthy volunteers participated in two experimental sessions with an interval of 3 weeks. In each session subjects performed 2 h of constant-load bicycle exercise ( approximately 75% ), followed by 4 h of controlled recovery. On one occasion they exercised after an overnight fast (F), and on the other (CHO) they received carbohydrates before ( approximately 150 g) and during (1 g (kg bw)(-1) h(-1)) exercise. In both conditions, subjects ingested 5 g carbohydrates per kg body weight during recovery. Fibre type-specific relative IMTG content was determined by Oil red O staining in needle biopsies from m. vastus lateralis before, immediately after and 4 h after exercise. During F but not during CHO, the exercise bout decreased IMTG content in type I fibres from 18 +/- 2% to 6 +/- 2% (P = 0.007) area lipid staining. Conversely, during recovery, IMTG in type I fibres decreased from 15 +/- 2% to 10 +/- 2% in CHO, but did not change in F. Neither exercise nor recovery changed IMTG in type IIa fibres in any experimental condition. Exercise-induced net glycogen breakdown was similar in F and CHO. However, compared with CHO (11.0 +/- 7.8 mmol kg(-1) h(-1)), mean rate of postexercise muscle glycogen resynthesis was 3-fold greater in F (32.9 +/- 2.7 mmol kg(-1) h(-1), P = 0.01). Furthermore, oral glucose loading during recovery increased plasma insulin markedly more in F (+46.80 microU ml(-1)) than in CHO (+14.63 microU ml(-1), P = 0.02). We conclude that IMTG breakdown during prolonged submaximal exercise in the fasted state takes place predominantly in type I fibres and that this breakdown is prevented in the CHO-fed state. Furthermore, facilitated glucose-induced insulin secretion may contribute to enhanced muscle glycogen resynthesis following exercise in the fasted state.

Lipotoxicity: the obese and endurance-trained paradox.

The potential lipotoxic effect of intramyocellular triglyceride (IMTG) accumulation has been suggested to be a major component in the development of insulin resistance. Increased levels of IMTGs correlate with insulin resistance in both obese and diabetic patients, but this relationship does not exist in endurance trained (ETr) subjects. This may be, in part, related to differences in the gene expression and activities of key enzymes involved in fatty acid transport and oxidation as well as in the perodixation status of the IMTGs in obese/diabetic patients as compared with ETr subjects. Disruptions in fat and lipid homeostasis in skeletal muscle have been shown to activate protein kinase C (PKC), which acts on several downstream signalling pathways, including the insulin and the IkappaB kinase (IKK)/NFkappaB signalling pathways. Additionally, an increased peroxidation of IMTGs may reduce insulin sensitivity by increasing TNFalpha, which is known to increase the expression of suppressor of cytokine signalling proteins (SOCS). A common characteristic observed when activating both PKC and TNFalpha/SOCS3 is the inhibition of tyrosine phosphorylation of IRS-1 and subsequently an inhibition of its activation of downstream signalling molecules. These may be important players in the development of insulin resistance and understanding their activation and expression in both obese and ETr humans should assist in understanding how and why IMTGs become lipotoxic.