Contraction-induced changes in TNFalpha and Akt-mediated signalling are associated with increased myofibrillar protein in rat skeletal muscle.

Resistance training results in skeletal muscle hypertrophy, but the molecular signalling mechanisms responsible for this altered phenotype are incompletely understood. We used a resistance training (RT) protocol consisting of three sessions [day 1 (d1), day 3 (d3), day 5 (d5)] separated by 48 h recovery (squat exercise, 4 sets x 10 repetitions, 3 min recovery) to determine early signalling responses to RT in rodent skeletal muscle. Six animals per group were killed 3 h after each resistance training session and 24 and 48 h after the last training session (d5). There was a robust increase in TNFalpha protein expression, and IKK(Ser180/181) and p38MAPK(Thr180/Tyr182) phosphorylation on d1 (P < 0.05), which abated with subsequent RT, returning to control levels by d5 for TNFalpha and IKK(Ser180/181). There was a trend for a decrease in MuRF-1 protein expression, 48 h following d5 of training (P = 0.08). Notably, muscle myofibrillar protein concentration was elevated compared to control 24 and 48 h following RT (P < 0.05). Akt(Ser473) and mTOR(Ser2448) phosphorylation were unchanged throughout RT. Phosphorylation of p70S6k(Thr389) increased 3 h post-exercise on d1, d3 and d5 (P < 0.05), whilst phosphorylation of S6(Ser235/236) increased on d1 and d3 (P < 0.05). Our results show a rapid attenuation of inflammatory signalling with repeated bouts of resistance exercise, concomitant with summation in translation initiation signalling in skeletal muscle. Indeed, the cumulative effect of these signalling events was associated with myofibrillar protein accretion, which likely contributes to the early adaptations in response to resistance training overload in the skeletal muscle.

Activation of atypical protein kinase Czeta toward TC10 is regulated by high-fat diet and aerobic exercise in skeletal muscle.

We determined whether sustained aerobic exercise reverses high-fat diet-induced impairments in the c-Cbl associated protein (CAP)/Casitas b-lineage lymphoma (c-Cbl) signaling cascade in rodent skeletal muscle. Sprague-Dawley rats were placed into either control (n = 16) or high-fat-fed (n = 32) diet groups for 4 weeks. During a subsequent 4-week experimental period, 16 high-fat-fed rats remained sedentary, 16 high-fat-fed rats completed 4 weeks of exercise training, and control animals were sedentary and remained on the control diet. After the intervention period, animals were subjected to hind limb perfusions in the presence (n = 8 per group) or absence (n = 8 per group) of insulin. In the plasma membrane fractions, neither high-fat feeding nor exercise training altered adaptor protein with PH and SH2 domains, (APS), c-Cbl, or TC10 protein concentrations. In contrast, CAP protein concentration and insulin-stimulated plasma membrane c-Cbl tyrosine phosphorylation were reduced by high-fat feeding; but exercise training reversed these impairments. Of note was that insulin-stimulated atypical protein kinase Czeta kinase activity toward TC10 was reduced by high-fat feeding but normalized by exercise training. We conclude that sustained (4 weeks) exercise training can reverse high-fat diet-induced impairments on the CAP/c-Cbl pathway in high-fat-fed rodent skeletal muscle. We also provide the first evidence that the CAP/c-Cbl insulin signaling cascade in skeletal muscle may directly interact with components of the classic (phosphoinositide 3-kinase dependent) insulin signaling cascade.

Effect of high-frequency resistance exercise on adaptive responses in skeletal muscle.

PURPOSE: Regulation of skeletal muscle mass is highly dependent on contractile loading. The purpose of this study was to examine changes in growth factor and inflammatory pathways following high-frequency resistance training. METHODS: Using a novel design in which male Sprague-Dawley rats undertook a "stacked" resistance training protocol designed to generate a summation of transient exercise-induced signaling responses (four bouts of three sets x 10 repetitions of squat exercise, separated by 3 h of recovery), we determined the effects of high training frequency on signaling pathways and transcriptional activity regulating muscle mass. RESULTS: The stacked training regimen resulted in acute suppression of insulin-like growth factor 1 mRNA abundance (P < 0.05) and Akt phosphorylation (P < 0.05), an effect that persisted 48 h after the final training bout. Conversely, stacked training elicited a coordinated increase in the expression of tumor necrosis factor alpha, inhibitor kappa B kinase alpha/beta activity (P < 0.05), and p38 mitogen-activated protein kinase phosphorylation (P < 0.05) at 3 h after each training bout. In addition, the stacked series of resistance exercise bouts induced an increase in p70 S6 kinase phosphorylation 3 h after bouts x3 and x4, independent of the phosphorylation state of Akt. CONCLUSIONS: Our results indicate that high resistance training frequency extends the transient activation of inflammatory signaling cascades, concomitant with persistent suppression of key mediators of anabolic responses. We provide novel insights into the effects of the timing of exercise-induced overload and recovery on signal transduction pathways and transcriptional activity regulating skeletal muscle mass in vivo.

Exercise reverses high-fat diet-induced impairments on compartmentalization and activation of components of the insulin-signaling cascade in skeletal muscle.

The aims of this investigation were 1) to determine whether endurance exercise training could reverse impairments in insulin-stimulated compartmentalization and/or activation of aPKCzeta/lambda and Akt2 in skeletal muscle from high-fat-fed rodents and 2) to assess whether the PPARgamma agonist rosiglitazone could reverse impairments in skeletal muscle insulin signaling typically observed after high-fat feeding. Sprague-Dawley rats were placed on chow (NORCON, n = 16) or high-fat (n = 64) diets for 4 wk. During a subsequent 4-wk experimental period, high-fat-fed rats were allocated (n = 16/group) to either sedentary control (HFC), exercise training (HFX), rosiglitazone treatment (HFRSG), or a combination of both exercise training and rosiglitazone (HFRX). Following the 4-wk experimental period, animals underwent hindlimb perfusions. Insulin-stimulated plasma membrane-associated aPKCzeta and -lambda protein concentration, aPKCzeta/lambda activity, GLUT4 protein concentration, cytosolic Akt2, and aPKCzeta/lambda activities were reduced (P < 0.05) in HFC compared with NORCON. Cytosolic Akt2, aPKCzeta, and aPKClambda protein concentrations were not affected in HFC compared with NORCON. Exercise training reversed the deleterious effects of the high-fat diet such that insulin-stimulated compartmentalization and activation of components of the insulin-signaling cascade in HFX were normalized to NORCON. High-fat diet-induced impairments to skeletal muscle glucose metabolism were not reversed by rosiglitazone administration, nor did rosiglitazone augment the effect of exercise. Our findings indicate that chronic exercise training, but not rosiglitazone, reverses high-fat diet induced impairments in compartmentalization and activation of components of the insulin-signaling cascade in skeletal muscle.

Tissue-specific effects of rosiglitazone and exercise in the treatment of lipid-induced insulin resistance.

Both pharmacological intervention (i.e., thiazolidinediones [TZDs]) and lifestyle modification (i.e., exercise training) are clinically effective treatments for improving whole-body insulin sensitivity. However, the mechanism(s) by which these therapies reverse lipid-induced insulin resistance in skeletal muscle is unclear. We determined the effects of 4 weeks of rosiglitazone treatment and exercise training and their combined actions (rosiglitazone treatment and exercise training) on lipid and glucose metabolism in high-fat-fed rats. High-fat feeding resulted in decreased muscle insulin sensitivity, which was associated with increased rates of palmitate uptake and the accumulation of the fatty acid metabolites ceramide and diacylglycerol. Impairments in lipid metabolism were accompanied by defects in the Akt/AS160 signaling pathway. Exercise training, but not rosiglitazone treatment, reversed these impairments, resulting in improved insulin-stimulated glucose transport and increased rates of fatty acid oxidation in skeletal muscle. The improvements to glucose and lipid metabolism observed with exercise training were associated with increased AMP-activated protein kinase alpha1 activity; increased expression of Akt1, peroxisome proliferator-activated receptor gamma coactivator 1, and GLUT4; and a decrease in AS160 expression. In contrast, rosiglitazone treatment exacerbated lipid accumulation and decreased insulin-stimulated glucose transport in skeletal muscle. However, rosiglitazone, but not exercise training, increased adipose tissue GLUT4 and acetyl CoA carboxylase expression. Both exercise training and rosiglitazone decreased liver triacylglycerol content. Although both interventions can improve whole-body insulin sensitivity, our results show that they produce divergent effects on protein expression and triglyceride storage in different tissues. Accordingly, exercise training and rosiglitazone may act as complementary therapies for the treatment of insulin resistance.

High-fat feeding effects on components of the CAP/Cbl signaling cascade in Sprague-Dawley rat skeletal muscle.

The aim of this investigation was to determine whether the CAP (Cbl-associated protein)/Cbl signaling cascade is present and responsive to insulin in skeletal muscle and if high-fat feeding impairs insulin-stimulated activation of this signaling cascade. Sprague-Dawley rats were assigned to either control (n = 16) or high fat-fed (n = 16) dietary groups. After a 12-week dietary period, animals were subjected to hind limb perfusions in the presence (n = 8 per group) or absence (n = 8 per group) of insulin. High-fat feeding reduced rates of insulin-stimulated skeletal muscle phosphatidylinositol 3-kinase activity and 3-O-methylglucose transport. In plasma membrane fractions, neither the high-fat diet nor insulin altered the insulin receptor beta subunit (IR-beta), APS (adaptor protein containing PH and SH2 domains), c-Cbl, or TC10 protein concentration, but high-fat feeding did decrease CAP protein concentration. APS, c-Cbl, CAP, and TC10 messenger RNA were present in the skeletal muscle and reflected the protein concentration of experimental groups. Despite insulin-stimulated plasma membrane IR-beta tyrosine phosphorylation being unaffected by high-fat feeding, c-Cbl tyrosine phosphorylation, the kinase activity of IR-beta toward APS, and glucose transporter 4 protein concentration were all significantly reduced in insulin-stimulated plasma membrane prepared from the skeletal muscle of high fat-fed animals. These findings suggest that the CAP/Cbl signaling cascade is present in skeletal muscle, activated by insulin, and impaired by high-fat feeding.

Insulin-stimulated plasma membrane association and activation of Akt2, aPKC zeta and aPKC lambda in high fat fed rodent skeletal muscle.

Several recent reports using cell lines have suggested that both Akt and atypical protein kinase C (aPKC) zeta/lambda are translocated to the plasma membrane (PM) in response to insulin. However, it has yet to be determined in skeletal muscle whether: (1) insulin increases PM-associated Akt2, aPKC zeta and/or lambda protein concentration, (2) the activity of these kinases is altered by insulin at the PM, and (3) high fat feeding alters the insulin-stimulated PM concentration and/or activity of Akt2 and aPKC zeta/lambda. Sprague-Dawley rats were randomly assigned to either normal (n=16) or high fat (n=16) dietary groups. Following a 12 week dietary period, animals were subjected to hind limb perfusions in the presence (n=8 per group) or absence (n=8 per group) of insulin. In normal skeletal muscle, total PI3-kinase, Akt2 and aPKC zeta/lambda activities were increased by insulin. PM-associated aPKC zeta and lambda, and aPKC zeta/lambda activity, but not Akt2 or Akt2 activity, were increased by insulin in normal muscle. High fat feeding did not alter total skeletal muscle Akt2, aPKC zeta or aPKC lambda protein concentration. Insulin-stimulated total PI3-kinase, Akt2 and aPKC zeta/lambda activities were reduced in the high fat fed animals. Insulin-stimulated PM aPKC zeta, aPKC lambda, aPKC zeta/lambda activity and GLUT4 protein concentration were also reduced in high fat fed animals. These findings suggest that in skeletal muscle, insulin stimulates translocation of aPKC zeta and lambda, but not Akt2, to the PM. In addition, high fat feeding impairs insulin-stimulated activation of total aPKC zeta/lambda and Akt2, as well as PM association and activation of aPKC zeta and lambda.