Stephen L. Dodd

Hsp70 overexpression inhibits NF-κB and Foxo3a transcriptional activities and prevents skeletal muscle atrophy

Heat shock protein 70 (Hsp70) is a highly conserved and ubiquitous protein that is reported to provide cytoprotection in various cell types and tissues. However, the importance of Hsp70 expression during skeletal muscle atrophy, when Hsp70 levels are significantly decreased, is not known. The current study aimed to determine whether plasmid‐mediated overexpression of Hsp70, in the soleus muscle of rats, was sufficient to regulate specific atrophy signaling pathways and attenuate skeletal muscle disuse atrophy. We found that Hsp70 overexpression prevented disuse muscle fiber atrophy and inhibited the increased promoter activities of atrogin‐1 and MuRF1. Importantly, the transcriptional activities of Foxo3a and NF‐κB, which are implicated in the regulation of atrogin‐1 and MuRF1, were abolished by Hsp70. These data suggest that Hsp70 may regulate key atrophy genes through inhibiting Foxo3a and NF‐κB activities during disuse. Indeed, we show that specific inhibition of Foxo3a prevented the increases in both atrogin‐1 and MuRF1 promoter activities during disuse. However, inhibition of NF‐κB did not affect the activation of either promoter, suggesting its requirement for disuse atrophy is through its regulation of other atrophy genes. We conclude that overexpression of Hsp70 is sufficient to inhibit key atrophy signaling pathways and prevent skeletal muscle atrophy.— Senf, S. M., Dodd, S. L., McClung, J. M., Judge, A. R. Hsp70 overexpression inhibits NF‐κB and Foxo3a transcriptional activities and prevents skeletal muscle atrophy. FASEB J. 22, 3836–3845 (2008)

High intensity training-induced changes in skeletal muscle antioxidant enzyme activity

These experiments tested the hypothesis that high intensity (interval) training is superior to moderate intensity (continuous) exercise training in the upregulation of antioxidant enzyme activity in skeletal muscle. To test this postulate, we examined changes in oxidative and antioxidant enzyme activities in rat skeletal muscle following 12 wk of either interval (6 x approximately 5-min intervals at approximately 80-95% VO2max) or continuous (45 min at approximately 70% VO2max) exercise training. Both continuous and interval training resulted in significantly elevated (P < 0.05) succinate dehydrogenase (SDH) and 3-hydroxyacyl-CoA-dehydrogenase (HADH) activities in the gastrocnemius (G) and soleus (S) muscles compared with controls. SDH and HADH activities in the G and S muscles did not differ between the two exercise groups. Glutathione peroxidase (GPX) activity exceeded controls (P < 0.05) in only the interval trained S muscle. Soleus superoxide dismutase (SOD) activity was higher (P < 0.05) in both exercise groups compared with controls. No differences in SOD activity existed between interval and continuous trained animals. We conclude that when matched for oxygen cost, interval and continuous exercise training result in similar increases in SOD activity. However, high intensity interval exercise is superior to moderate intensity continuous exercise in the promotion of GPX activity in the S.

Incidence of exercise induced hypoxemia in elite endurance athletes at sea level

SummaryRecent evidence suggests that exercise-induced hypoxemia (EIH) may occur in healthy trained endurance athletes. However, at present, no data exist to describe the regularity of EIH in athletes or non-athletes. Therefore, the purpose of the present investigation was to determine the incidence of EIH during exercise in healthy subjects varying in physical fitness. Subjects (N=68) performed an incremental cycle ergometer test to volitional fatigue with percent arterial oxyhemoglobin saturation (%SaO2) measured min-by-min. For the purpose of data analysis subjects were divided into three groups according to their level of physical training: 1) untrained (N=16), 2) moderately trained (N=27), and 3) elite highly trained endurance athletes (N=25). EIH was defined as a %SaO2 of ≤91% during exercise. EIH did not occur in any of the untrained subjects or the moderately trained subjects. However, EIH occurred in 52% of the highly trained endurance athletes tested and was highly reproducible (r=0.95; P<0.05). These findings further confirm the existence of EIH in healthy highly trained endurance athletes and suggests a rather high incidence of EIH in this healthy population. Hence, it is important that the clinician or physiologist performing exercise testing in elite endurance athletes recognize that EIH can and does occur in the elite endurance athlete in the absence of lung disease.

FOXO signaling is required for disuse muscle atrophy and is directly regulated by Hsp70.

The purpose of the current study was to determine whether heat shock protein 70 (Hsp70) directly regulates forkhead box O (FOXO) signaling in skeletal muscle. This aim stems from previous work demonstrating that Hsp70 overexpression inhibits disuse-induced FOXO transactivation and prevents muscle fiber atrophy. However, although FOXO is sufficient to cause muscle wasting, no data currently exist on the requirement of FOXO signaling in the progression of physiological muscle wasting, in vivo. In the current study we show that specific inhibition of FOXO, via expression of a dominant-negative FOXO3a, in rat soleus muscle during disuse prevented >40% of muscle fiber atrophy, demonstrating that FOXO signaling is required for disuse muscle atrophy. Subsequent experiments determined whether Hsp70 directly regulates FOXO3a signaling when independently activated in skeletal muscle, via transfection of FOXO3a. We show that Hsp70 inhibits FOXO3a-dependent transcription in a gene-specific manner. Specifically, Hsp70 inhibited FOXO3a-induced promoter activation of atrogin-1, but not MuRF1. Further studies showed that a FOXO3a DNA-binding mutant can activate MuRF1, but not atrogin-1, suggesting that FOXO3a activates these two genes through differential mechanisms. In summary, FOXO signaling is required for physiological muscle atrophy and is directly inhibited by Hsp70.

The myopathy of peripheral arterial occlusive disease: Part 2. Oxidative stress, neuropathy, and shift in muscle fiber type.

In recent years, an increasing number of studies have demonstrated that a myopathy is present, contributes, and, to a certain extent, determines the pathogenesis of peripheral arterial occlusive disease. These works provide evidence that a state of repetitive cycles of exercise-induced ischemia followed by reperfusion at rest operates in patients with peripheral arterial occlusive disease and mediates a large number of structural and metabolic changes in the muscle, resulting in reduced strength and function. The key players in this process appear to be defective mitochondria that, through multilevel failure in their roles as energy, oxygen radical species, and apoptosis regulators, produce and sustain a progressive decline in muscle performance. In this 2-part review, the currently available evidence that characterizes the nature and mechanisms responsible for this myopathy is highlighted. In part 1, the functional and histomorphological characteristics of the myopathy were reviewed, and the main focus was on the biochemistry and bioenergetics of its mitochondriopathy. In part 2, accumulating evidence that oxidative stress related to ischemia reperfusion is probably the major operating mechanism of peripheral arterial occlusive disease myopathy is reviewed. Important new findings of a possible neuropathy and a shift in muscle fiber type are also reviewed. Learning more about these mechanisms will enhance our understanding of the degree to which they are preventable and treatable.

The Myopathy of Peripheral Arterial Occlusive Disease: Part 1. Functional and Histomorphological Changes and Evidence for Mitochondrial Dysfunction

In recent years, an increasing number of studies have demonstrated that a myopathy is present, contributes, and, to a certain extent, determines the pathogenesis of peripheral arterial occlusive disease (PAD). These works provide evidence that a state of repetitive cycles of exercise-induced ischemia followed by reperfusion at rest operates in PAD patients and mediates a large number of structural and metabolic changes in the muscle, resulting in reduced strength and function. The key players in this process appear to be defective mitochondria that, through multilevel failure in their roles as energy, oxygen radical species, and apoptosis regulators, produce and sustain a progressive decline in muscle performance. In this 2-part review, we highlight the currently available evidence that characterizes the nature and mechanisms responsible for this myopathy. In part 1, the authors review the functional and histomorphological characteristics of the myopathy and focus on the biochemistry and bioenergetics of its mitochondriopathy. In part 2, they then review accumulating evidence that oxidative stress related to ischemia reperfusion is probably the major operating mechanism of PAD myopathy. Important new findings of a possible neuropathy and a shift in muscle fiber type are also reviewed. Learning more about these mechanisms will enhance our understanding of the degree to which they are preventable and treatable.

Calpain activation causes a proteasome‐dependent increase in protein degradation and inhibits the Akt signalling pathway in rat diaphragm muscle

The role of the calpain proteases in skeletal muscle atrophy is poorly understood. One goal of these experiments was to clarify whether calpains act upstream of the ubiquitin–proteasome pathway (UPP). Calpain activation may also inhibit the anabolic signalling of Akt, since a molecular chaperone previously shown to mediate Akt activity, heat shock protein 90 (HSP 90), is a calpain substrate. Thus, an additional objective was to determine whether calpain activation affects the Akt signalling pathway. Ex vivo experiments were conducted using isolated rat diaphragm muscle. Calpain activation increased total protein degradation by 65%. Proteasome inhibition prevented this large rise in proteolysis, demonstrating that the proteasome was necessary for calpain‐activated protein degradation. In addition, calpain activation increased proteasome‐dependent proteolysis by 144%, further supporting the idea of sequential proteolytic pathways. Calpain reduced Akt and mammalian target of rapamycin (mTOR) phosphorylation by 35 and 50%, respectively, and activated glycogen synthase kinase‐3 beta (GSK‐3β) by 40%. Additionally, calpain activation reduced HSP 90β and mTOR protein content by 33 and 50%, respectively. These data suggest that calpains play a dual role in protein metabolism by concomitantly activating proteasome‐dependent proteolysis and inhibiting the Akt pathway of protein synthesis.

Caffeine and exercise performance. An update.

SummaryThree principal cellular mechanisms have been proposed to explain the ergogenic potential of caffeine during exercise: (a) increased myofilament affinity for calcium and/or increased release of calcium from the sarcoplasmic reticulum in skeletal muscle; (b) cellular actions caused by accumulation of cyclic-3′,5′-adenosine monophosphate (cAMP) in various tissues including skeletal muscle and adipocytes; and (c) cellular actions mediated by competitive inhibition of adenosine receptors in the central nervous system and somatic cells. The relative importance of each of the above mechanisms in explaining in vivo physiological effects of caffeine during exercise continues to be debated. However, growing evidence suggests that inhibition of adenosine receptors is one of the most important, if not the most important, mechanism to explain the physiological effects of caffeine at nontoxic plasma concentrations. Numerous animal studies using high caffeine doses have reported increased force development in isolated skeletal muscle in both in vitro and in situ preparations. In contrast, in vivo human studies have not consistently shown caffeine to enhance muscular performance during high intensity, short term exercise. Further, recent evidence supports previous work that shows caffeine does not improve performance during short term incremental exercise. Although controversy exists, the major part of published evidence evaluating performance supports the notion that caffeine is ergogenic during prolonged (>30 min), moderate intensity (≈75 to 80% V̇2max) exercise. The mechanism to explain these findings may be linked to a caffeine-mediated glycogen sparing effect secondary to an increased rate of lipolysis.

Effects of clenbuterol on contractile and biochemical properties of skeletal muscle

We investigated the effects of clenbuterol on the muscle mass, contractile properties, myosin phenotype, and bioenergetic enzyme activity in the gastrocnemius (GS)-plantaris (PL)-soleus (SO) muscle complex. Rats were sham-injected or treated with clenbuterol (2 mg.kg-1, subcutaneously) for 14 d. Clenbuterol increased (P < 0.05) body weight and muscle complex weight. Also, clenbuterol treatment resulted in an increase in total muscle force production and maximal shortening velocity (P < 0.05). No difference (P > 0.05) in relative force production (force.g-1 muscle) existed between experimental groups. However, muscle fatigue increased with clenbuterol treatment. Myosin heavy chain (MHC) composition was not altered in the GS or PL muscles, but shifted toward the fast Type II MHC in the SO. Myosin light chain (MLC) composition was not altered in any of the muscles. Clenbuterol caused a decrease in oxidative and glycolytic enzyme activity in the GS and PL, but not the SO. These data suggest that the clenbuterol-induced increase in muscle mass and maximal force generation is due to hypertrophy of both fast and slow fibers. Furthermore, these findings support the notion that beta-agonists may be beneficial in combating conditions that result in muscle wasting and dysfunction.

Oxygen uptake kinetics in trained athletes differing in \(\dot V_{{\text{O}}_{{\text{2max}}} }\)

SummaryPrevious work has shown that when