**4. Skeletal muscle in metabolic health and disease**

Skeletal muscle represents a large mass of tissue, and its primary function is to use energy, though quite inefficiently, to enable us the 3D life, voluntary positioning and moving our bodies in a surrounding space. This makes an active muscle to be the most effective energy burner. In addition to obvious metabolic consequences, regular exercise activates central reward mechanisms and makes us happy (Figure 7.) (Sher 1998; Boecker et al. 2008).

Fig. 7. Correlation of opidergic ligand 6-O-(2-[18F]fluoroethyl)-6-O-desmethyldiprenorphine binding in right orbitofrontal cortex (OFC) with the visual analog mood scale (for euphoria) in runners (**A**) and effect of exercise on the individual's mood expressed in the visual analog mood scale (**B**) (Boecker et al. 2008).

Adipose Tissue and Skeletal Muscle Plasticity in Obesity and Metabolic Disease 153

The recent identification of skeletal muscle as an endocrine organ that produces and releases biologically active substances, "myokines", expands our knowledge on how the endocrine, immune and nervous systems contribute to the maintenance of homeostasis, especially when energy demands are increased (Pedersen & Febbraio 2008). To date, only a few muscle cell-secreted proteins with auto-, para-, or endocrine functions have been identified. It could be hypothesized that skeletal muscle releases a large number of biologically active substances which participate to cell-to-cell and organ-to-organ cross-talk. It also has to be noted that specific biological functions of known myokines are very incompletely

Certain myokines, such as calprotectin (Mortensen et al. 2008), IL15 (Nielsen & Pedersen 2007) and IL6 (Jonsdottir et al. 2000), are acutely induced by muscle contraction but might not necessarily be increased in response to muscle training (Pedersen & Febbraio 2008; Haugen et al. 2010). Exercise training involves multiple adaptations including increased preexercise skeletal muscle glycogen content (Kirwan et al. 1990), enhanced activity of key enzymes involved in -oxidation (Schantz 1986), increased sensitivity of adipose tissue to epinephrine-induced lipolysis (Crampes et al. 1986), and increased muscle capacity to oxidize fat (Holloszy & Booth 1976; Phillips et al. 1996). It could therefore be hypothesized that secretory activity of muscle subjected to inadequate physical activity would be qualitatively and quantitatively distinct from that of the trained athlete, and that it could simply be regulated by e.g. the glycogen level (Keller et al. 2001; Steensberg et al. 2001), reactive oxygen species production (Kosmidou et al. 2002; Steensberg et al. 2007) or by modulating biological availability of various forms of lipids (Peter et al. 2009), such as found

Previous reports indicate that calprotectin, IL6 and IL15 might contribute to homeostatic control of glucose and lipid metabolism (Van Hall et al. 2003; Febbraio et al. 2004). In addition, fibroblast growth factor-21 (FGF-21), the potent metabolic regulator, shown to improve glucose metabolism and insulin sensitivity in animal models, had recently been found to be expressed and secreted *in vitro* from murine muscle cells and *in vivo* from human muscle in response to insulin stimulation (hyperinsulinemic-euglycemic clamp) (Hojman et al. 2009). Follistatin-like 1 (Fstl-1) is another myokine whose functional significance in physiological and pathological processes is incompletely understood. Preliminary evidence indicates that Fstl-1 promotes endothelial cell function and stimulates revascularization in response to ischemic insult through its ability to activate Akt-eNOS signaling in muscle (Ouchi et al. 2008). Interleukin-8 is a CXC family chemokine increased in human muscle in response to concentric exercise (Akerstrom et al. 2005), which has also been shown to have angiogenic actions associated with activation of CXCR2 receptors in the human microvascular endothelial cells (Bek et al. 2002; Frydelund-Larsen et al. 2007). Recent report by Drevon's laboratory describes interleukin-7 as a novel myokine affecting myogenesis *in vitro* in human primary muscle cells. Interleukin-7 is up-regulated by exercise training in male individuals undergoing a strength training program (Haugen et al. 2010).

**4.2 Mitochondrial biogenesis in skeletal muscle – Energetic remodeling of muscle** 

Mitochondria are energy power plants of the cells, believed to have evolved over billions of years from invading prokaryotic oxygen utilizing "quite energizing" eubacterium to early eucaryotic cells, giving the life on earth new energy spark (Lanza & Nair 2010). Their

**phenotype in obesity, insulin resistance and exercise** 

**4.1 Exercise and skeletal muscle endocrinology** 

understood.

in obesity.

More importantly, inadequate physical activity, associated with defects in mitochondrial function and changes in ultrastructure as well as muscle endocrine properties, largely contributes to the imbalance between energy intake and energy expenditure and is tightly associated with many chronic metabolic and cardiovascular diseases (Bluher & Zimmer 2010; Pedersen 2011). Physical activity is a key factor to bring individuals living in a modern society with plenty of palatable food choices to energy balance. The mechanisms that tie muscle activity to health are unclear. Generation of "exercise pill" targeting organ systems involved in facultative thermogenesis had been envisaged (Himms-Hagen 2004). And results of studies aimed at identifying the endocrine properties of exercising muscle are encouraging our thinking in this respect. In our recent study we observed on the sample of 71 individuals with a broad range of BMI that overweight and obesity is associated with decreased physical activity. This might be not so surprising. But low physical activity level was also associated with decreased insulin sensitivity, increased fat cell size and expanded visceral adiposity all independent on BMI (Fig. 8.). In addition, the basic metabolic rate was positively and respiratory quotient negatively associated with the duration of the daily physical activity representing thus a direct link between physical activity and major determinants of energy homeostasis (Ukropcova et al*.,* unpublished observations).

Fig. 8. Free-living ambulatory activity (number of steps in 24h) correlates with insulin sensitivity, modulates adipocyte diameter as well as visceral adiposity (Ukropcova et al., unpublished observations).

This complements the previous observations by others indicating that inactivity initiates unique cellular processes that are qualitatively different from the exercise responses. Inactivity physiology studies are beginning to raise a new concern with potentially major clinical and public health significance. Sedentary lifestyle threatens our society. The average nonexercising person may become even more metabolically unfit in the coming years if they sit too much and thereby lower the normally high volume of intermittent nonexercise physical activity in everyday life (Fig. 8) (Hamilton et al. 2007; Levine et al. 2008). Dynamic interrelations of skeletal muscle and adipose tissue during exercise are necessary to support muscle performance. This requires precise spacio-temporal management of the adipose tissue metabolic flux. The transcriptional coactivator PGC1 has recently been shown to regulate several exercise-associated aspects of muscle function. There is mounting evidence suggesting that this transcription factor controls muscle plasticity, suppresses a broad inflammatory response and integrates many beneficial effects of exercise on metabolic health (Handschin & Spiegelman 2008).

## **4.1 Exercise and skeletal muscle endocrinology**

152 Dyslipidemia - From Prevention to Treatment

More importantly, inadequate physical activity, associated with defects in mitochondrial function and changes in ultrastructure as well as muscle endocrine properties, largely contributes to the imbalance between energy intake and energy expenditure and is tightly associated with many chronic metabolic and cardiovascular diseases (Bluher & Zimmer 2010; Pedersen 2011). Physical activity is a key factor to bring individuals living in a modern society with plenty of palatable food choices to energy balance. The mechanisms that tie muscle activity to health are unclear. Generation of "exercise pill" targeting organ systems involved in facultative thermogenesis had been envisaged (Himms-Hagen 2004). And results of studies aimed at identifying the endocrine properties of exercising muscle are encouraging our thinking in this respect. In our recent study we observed on the sample of 71 individuals with a broad range of BMI that overweight and obesity is associated with decreased physical activity. This might be not so surprising. But low physical activity level was also associated with decreased insulin sensitivity, increased fat cell size and expanded visceral adiposity all independent on BMI (Fig. 8.). In addition, the basic metabolic rate was positively and respiratory quotient negatively associated with the duration of the daily physical activity representing thus a direct link between physical activity and major determinants of energy homeostasis (Ukropcova et al*.,* unpublished

Fig. 8. Free-living ambulatory activity (number of steps in 24h) correlates with insulin sensitivity, modulates adipocyte diameter as well as visceral adiposity (Ukropcova et al.,

This complements the previous observations by others indicating that inactivity initiates unique cellular processes that are qualitatively different from the exercise responses. Inactivity physiology studies are beginning to raise a new concern with potentially major clinical and public health significance. Sedentary lifestyle threatens our society. The average nonexercising person may become even more metabolically unfit in the coming years if they sit too much and thereby lower the normally high volume of intermittent nonexercise physical activity in everyday life (Fig. 8) (Hamilton et al. 2007; Levine et al. 2008). Dynamic interrelations of skeletal muscle and adipose tissue during exercise are necessary to support muscle performance. This requires precise spacio-temporal management of the adipose tissue metabolic flux. The transcriptional coactivator PGC1 has recently been shown to regulate several exercise-associated aspects of muscle function. There is mounting evidence suggesting that this transcription factor controls muscle plasticity, suppresses a broad inflammatory response and integrates many beneficial effects of exercise on metabolic

observations).

unpublished observations).

health (Handschin & Spiegelman 2008).

The recent identification of skeletal muscle as an endocrine organ that produces and releases biologically active substances, "myokines", expands our knowledge on how the endocrine, immune and nervous systems contribute to the maintenance of homeostasis, especially when energy demands are increased (Pedersen & Febbraio 2008). To date, only a few muscle cell-secreted proteins with auto-, para-, or endocrine functions have been identified. It could be hypothesized that skeletal muscle releases a large number of biologically active substances which participate to cell-to-cell and organ-to-organ cross-talk. It also has to be noted that specific biological functions of known myokines are very incompletely understood.

Certain myokines, such as calprotectin (Mortensen et al. 2008), IL15 (Nielsen & Pedersen 2007) and IL6 (Jonsdottir et al. 2000), are acutely induced by muscle contraction but might not necessarily be increased in response to muscle training (Pedersen & Febbraio 2008; Haugen et al. 2010). Exercise training involves multiple adaptations including increased preexercise skeletal muscle glycogen content (Kirwan et al. 1990), enhanced activity of key enzymes involved in -oxidation (Schantz 1986), increased sensitivity of adipose tissue to epinephrine-induced lipolysis (Crampes et al. 1986), and increased muscle capacity to oxidize fat (Holloszy & Booth 1976; Phillips et al. 1996). It could therefore be hypothesized that secretory activity of muscle subjected to inadequate physical activity would be qualitatively and quantitatively distinct from that of the trained athlete, and that it could simply be regulated by e.g. the glycogen level (Keller et al. 2001; Steensberg et al. 2001), reactive oxygen species production (Kosmidou et al. 2002; Steensberg et al. 2007) or by modulating biological availability of various forms of lipids (Peter et al. 2009), such as found in obesity.

Previous reports indicate that calprotectin, IL6 and IL15 might contribute to homeostatic control of glucose and lipid metabolism (Van Hall et al. 2003; Febbraio et al. 2004). In addition, fibroblast growth factor-21 (FGF-21), the potent metabolic regulator, shown to improve glucose metabolism and insulin sensitivity in animal models, had recently been found to be expressed and secreted *in vitro* from murine muscle cells and *in vivo* from human muscle in response to insulin stimulation (hyperinsulinemic-euglycemic clamp) (Hojman et al. 2009). Follistatin-like 1 (Fstl-1) is another myokine whose functional significance in physiological and pathological processes is incompletely understood. Preliminary evidence indicates that Fstl-1 promotes endothelial cell function and stimulates revascularization in response to ischemic insult through its ability to activate Akt-eNOS signaling in muscle (Ouchi et al. 2008). Interleukin-8 is a CXC family chemokine increased in human muscle in response to concentric exercise (Akerstrom et al. 2005), which has also been shown to have angiogenic actions associated with activation of CXCR2 receptors in the human microvascular endothelial cells (Bek et al. 2002; Frydelund-Larsen et al. 2007). Recent report by Drevon's laboratory describes interleukin-7 as a novel myokine affecting myogenesis *in vitro* in human primary muscle cells. Interleukin-7 is up-regulated by exercise training in male individuals undergoing a strength training program (Haugen et al. 2010).

#### **4.2 Mitochondrial biogenesis in skeletal muscle – Energetic remodeling of muscle phenotype in obesity, insulin resistance and exercise**

Mitochondria are energy power plants of the cells, believed to have evolved over billions of years from invading prokaryotic oxygen utilizing "quite energizing" eubacterium to early eucaryotic cells, giving the life on earth new energy spark (Lanza & Nair 2010). Their

Adipose Tissue and Skeletal Muscle Plasticity in Obesity and Metabolic Disease 155

prolong lifespan, is also inducing mitochondrial biogenesis in human skeletal muscle (Civitarese et al. 2007). Many scientists are on a quest, pursuing the vision of exercise mimicking pill, capable of induction of mitochondrial biogenesis *in vivo*. "Exercise in a pill" (another option would be a pill mimicking caloric restriction) is by many considered a putatively great tool to combat obesity and civilization diseases. However, healthy lifestyle intervention, with sufficient physical activity and matching caloric intake still proves to be the most natural and effective way how to stay fit, healthy and with increased chances to

Our organism can be viewed as a very complex society of tissues that need to communicate with one another in order to maintain metabolic health. Tissue cross-talk plays the central role in the regulation of food intake, energy expenditure, oxidative capacity, adaptation to changes in physical activity, nutritional status etc. As mentioned above, adipose tissue (as well as many other tissues in our body) is (are) a (the) source of many biologically active substances with autocrine, paracrine and endocrine activities, exerting effects over many

Adiponectin is the most studied adipocytokine which is in relatively high quantities secreted from adipose tissue into the bloodstream. Adiponectin has very positive effects on our metabolic health as it activates glucose and fatty acid metabolism and improves insulin sensitivity. Adiponectin levels are inversely correlated with body fat mass and positively with insulin sensitivity (Hara et al. 2005) and it also displays anti-atherogenic and antiinflammatory effects (Antoniades et al. 2009). This hormone was first characterized in mice as a transcript overexpressed in preadipocytes (precursors of fat cells) differentiating into adipocytes. The human homologue was identified as the most abundant transcript in adipose tissue. Contrary to expectations and despite being produced in adipose tissue, adiponectin was found to be decreased in obesity. The gene was localized to chromosome 3p27, a region highlighted as affecting genetic susceptibility to T2D and obesity. Supplementation by differing forms of adiponectin was able to improve insulin control, blood glucose and triglyceride levels in mouse models. The question remains what are the

The molecular mechanisms leading to mitochondrial dysfunction in obesity and T2D remain largely unknown. Bergeron et al (Bergeron et al. 2001) demonstrated that activation of cAMP-activated protein kinase (AMPK) increases both mitochondrial biogenesis and oxidative capacity in skeletal muscle of rodents. In animal models of T2D, the activation of AMPK by adiponectin increases muscle and hepatic fat oxidation and improves insulin sensitivity (Yamauchi et al. 2001). Studies in obese and diabetic rhesus monkey demonstrate that plasma adiponectin level declines in the early phases of obesity and in parallel to the progressive development of insulin resistance (Hotta et al. 2001). Furthermore, circulating plasma adiponectin levels and the expression of both adiponectin receptors are reduced in subjects with a family history of diabetes (Civitarese et al. 2004), while prospective studies in Pima Indians show that high concentrations of adiponectin is protective against the development of T2D (Lindsay et al. 2002). Collectively, these data define a pathway in skeletal muscle by which adiponectin contributes to energy homeostasis by modulating mitochondrial number and function (Civitarese et al. 2006). Early defects in the secretion of adiponectin or in adiponectin signaling might contribute to the lower mitochondrial content

live up to be a hundred.

**4.2.3 Adipose tissue and skeletal muscle interplay** 

different neighboring as well as distant tissues and organs.

mechanisms underlying positive effects of adiponectin on metabolism?

structure and function is orchestrated by a strict coordination of nuclear and mitochondrial genome. Of ~1.000 mitochondrial proteins, only 13 are encoded by the mitochondrial genome, remaining proteins are translated from nuclear genome and transported across the inner mitochondrial membrane (Lanza & Nair 2010). Mitochondria cover majority of energetic needs of cells by coupling substrate oxidation with ATP formation, the process known as oxidative phosphorylation. This process also generates reactive oxygen species (ROS). It has been estimated that 0.2 – 2% of oxygen taken up by the cell is converted into ROS (Harper et al. 2004). Mechanisms for detoxifying the ROS are quite well developed in a eukaryotic cell which is another reason for their long lasting partnership with "dangerous" mitochondria. Sustained excessive production may accumulate amount of ROS exceeding the antioxidant capacity of the specific cell, eventually leading to cell damage and death (Harman 1956). During recent years, mitochondria , though not only those found in skeletal muscle, were put on the spot as organelles involved in aging and associated chronic civilization diseases such as Alzheimer's disease (Reddy 2009), some forms of cancer, obesity and type 2 diabetes (Johannsen & Ravussin 2009).
