**4. Statins and statin-induced myopathy**

their size (fusion and fission), as well as in the proportions of lipids and proteins, somewhat modified by pathophysiological processes or nutritional and/or pharmacological interven‐ tions. Caveolae are dissimilar to LR as they are deficient in (GPI)-anchored proteins and poor in CHOL but rich in caveolins, the structural proteins assembled to stabilize membrane invaginations [25]. In extreme situations, PM may be subjected to disintegration, protein misfolding, and aggregation, and finally profound dysfunction causes cell death by necrosis. How these nanodomains are segregated within plasma membrane is a matter of debate, although cholesterol molecules establish closeness of PL, GL, GSL, SM, and proteins. Choles‐ terol (alcohol) acts as "glue" with hydroxyl group that combines with the phosphate head of phospholipids, whereas the hydrophobic steroid section works together with phospholipids acyl chains. Growing body of evidence points to physicochemical forces (intermolecular forces including electrostatic interactions, hydrogen bonds, Van der Waals forces, and hydration forces) which determine asymmetric geometry of membranes both laterally and in crosssection. Moreover, integral proteins also influence lipid structure in the membrane. One might bear in mind that according to lipidomics, more than 1000 different lipid forms are to be found in plasma membrane. To meet the requirements of fluidity, membrane components are also subject to considerable qualitative and quantitative seasonal changes adjusted by the cell [26]. It is determined by the environmental conditions (i.e., cold vs. heat) but also by needs of adaptation such as hyperplasia/hypertrophy or resistance to different types of stress (shear stress, oxidative stress including irradiation). In either case, physicochemical properties of membranes have to facilitate cell signaling and motility. In turn, cell signaling and motility are influenced by the glycosylation status of PM proteins, both integral represented by receptors

112 Muscle Cell and Tissue

and peripheral because they are heavily modified on the external leaflet.

**Figure 2.** Schematic illustration of a biomembrane, depicting membrane lipid asymmetry as well as microdomains en‐

riched in particular lipids and those induced by membrane proteins, reprinted from Escriba et al. [12].

Statins, 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase reversible inhibitors, became the most frequently prescribed drugs in modern societies used clinically to improve the lipid profile of hyperlipidemic patients, thereby decreasing the incidence of primary or secondary ischemic cardiac events [27–28]. The primary mechanism of action of statins is to lower CHOL levels by the inhibition of mevalonate formation, the rate limiting step in the cholesterol biosynthesis [29]. Pleiotropic effects of statins which seem to be independent of the

**Figure 4.** *Top*: Simplified model of two muscle sarcomeres in parallel. The sarcomere is composed of the thin (mostly actin) filaments, the thick (mostly myosin) filaments, and the giant filamentous molecule titin. The thin filaments are anchored in the Z-line, where they are cross-linked by α-actinin. The thick filament is centrally located in the sarco‐ mere and constitute the sarcomeric A-band. The myosin heads, or cross-bridges, on the thick filament interact with actin during activation. Titin spans the half-sarcomeric distance from the Z-line to the M-line, thus forming a third sar‐ comeric filament. In the I-band region, titin is extensible and functions as a molecular spring that develops passive ten‐ sion upon stretch. In the A-band, titin is inextensible due to its strong interaction with the thick filament. *Bottom*: Electron microscopy photograph of the ultrastructural organization of sarcomeres in parallel, reprinted from Otten‐ heijm et al. 2008 [134].

inhibiting effect on CHOL formation have also been reported [30–31], although statin-induced release of nitric oxide (NO) and prostaglandins (PGI2) does not explain side effects of statins on skeletal muscle cells (NO stimulates myogenesis, while PGI2 inhibits platelet activation). Even if statins are in general well tolerated and are safe for almost all patients, they were reported to induce different grades of myopathy in a significant part of the population, ranging from mild myalgia to morbid rhabdomyolysis [32–33]. There are risk factors for developing statin-induced myopathy (SIM), the significant component of statin intolerance during statin treatment, such as advanced age, excessive exercise, and multisystem disease as renal or hepatic insufficiencies, diabetes, or hypothyroidism [34]. Impaired metabolism of statins, pharmacokinetic interactions, and genetic effects are all probable causes of statin-induced

**Figure 5.** Dystrophin as a molecular shock absorber. Shown is a hypothetical model for how dystrophin may function to dampen elastic extension during muscle stretch. (I) Relaxed muscle. (II) Muscle stretch imposes forces that uncoil spring-like elements within repeats 1–10 and 18–24. (III) Electrostatic interaction of basic actin-binding repeats 11–17 with acidic actin filaments dampens extension of the spring-like elements. The "nonspecific" electrostatic interaction between the basic spectrin repeats and actin filaments is optimal because it does not require a specific orientation for interaction and would allow sliding between dystrophin and actin. As muscle rapidly shortens during contraction, the electrostatic interaction of the basic actin-binding repeats with acidic actin filaments would also serve to dampen elas‐ tic recoil, reprinted from Ervasti 2007 [23].

inhibiting effect on CHOL formation have also been reported [30–31], although statin-induced release of nitric oxide (NO) and prostaglandins (PGI2) does not explain side effects of statins on skeletal muscle cells (NO stimulates myogenesis, while PGI2 inhibits platelet activation). Even if statins are in general well tolerated and are safe for almost all patients, they were reported to induce different grades of myopathy in a significant part of the population, ranging from mild myalgia to morbid rhabdomyolysis [32–33]. There are risk factors for developing statin-induced myopathy (SIM), the significant component of statin intolerance during statin treatment, such as advanced age, excessive exercise, and multisystem disease as renal or hepatic insufficiencies, diabetes, or hypothyroidism [34]. Impaired metabolism of statins, pharmacokinetic interactions, and genetic effects are all probable causes of statin-induced

heijm et al. 2008 [134].

114 Muscle Cell and Tissue

**Figure 4.** *Top*: Simplified model of two muscle sarcomeres in parallel. The sarcomere is composed of the thin (mostly actin) filaments, the thick (mostly myosin) filaments, and the giant filamentous molecule titin. The thin filaments are anchored in the Z-line, where they are cross-linked by α-actinin. The thick filament is centrally located in the sarco‐ mere and constitute the sarcomeric A-band. The myosin heads, or cross-bridges, on the thick filament interact with actin during activation. Titin spans the half-sarcomeric distance from the Z-line to the M-line, thus forming a third sar‐ comeric filament. In the I-band region, titin is extensible and functions as a molecular spring that develops passive ten‐ sion upon stretch. In the A-band, titin is inextensible due to its strong interaction with the thick filament. *Bottom*: Electron microscopy photograph of the ultrastructural organization of sarcomeres in parallel, reprinted from Otten‐

> myotoxicity (muscle toxicity), although the molecular mechanism has not yet been elucidated in full. The most frightening clinical adverse effect is drug-induced rhabdomyolysis (0.1–0.5% in patients treated with pravastatin) the frequency of which is further increased by coadmi‐

**Figure 6.** A model of the effect of membrane cholesterol depletion on the sarcolemmal distribution of β-DG. The sarco‐ lemmal lipid rafts close to the clear opening of the TT-membrane are enriched in cholesterol, GM1 (ganglioside M1), Cav-3 (caveolin-3), and β-DG (dystroglycan). The schematic diagram illustrates the diminished contact between β-DG and Dys (dystrophin) in the presence of MβCD (methyl β-cyclodextrin); the β-DG/Dys interaction is essential for later‐ al force transmission. SERCA1 (sarcoplasmic reticulum calcium ATPase) and RyR (ryanodine receptor (SR Ca2+ chan‐ nel)) function normally. SL, sarcolemma; SR, sarcoplasmic reticulum; EM, extracellular matrix; LR, lipid raft; Dys, dystrophin; SG, sarcoglycan; CSQ, calsequestrin, adapted from Vega-Moreno et al. 2012 [17].

nistration of fibrates [35]. Reductions in skeletal muscle membrane CHOL were initially thought to account for the range of myopathic reactions. Additionally, the lowering the isoprenoid levels has been suggested to contribute to these pathologies as protein prenylation, and the potential consequences of a generalized insufficiency of this form of protein modifi‐ cation [36] are important for the activity and anchorage of plasma and other membrane proteins (nuclear envelope, dystrophin–glycoprotein complex, cytoskeletal G-proteins, etc.).

There is growing interest to decipher the molecular mechanism of the statin-induced myopa‐ thy, both by scientific community and pharmaceutical companies. One in three people over the age of 45 is taking a statin to reduce heart attack risk, the HMG-CoA antagonist with three orders of magnitude greater affinity to bind and subsequently to inhibit HMG-CoA reductase activity than that of natural substrate (HMG-CoA). Two in five women taking the statin are weaker than before, with one in ten reporting they felt "much worse". As statins became the most frequently prescribed drugs to prevent cardiovascular crisis alongside with the effort to pace physical activity, the issue how to protect from statin-induced myalgia, myositis, and rare cases of rhabdomyolysis is of great concern. In fact, due to muscle toxicity, an estimated 5–

**Figure 7.** Schematic diagram of the sarcomere and costamere protein complexes of striated muscle cells. Major compo‐ nents of the mature sarcomere and costamere are shown, along with the cytoskeletal and motor filament systems, in context with the sarcolemma and organelles of syncytial myocytes. Known chaperone or cochaperone molecules are shown in bold, along with their substrates. Arrows indicate regions where chaperone-mediated protein folding is es‐ sential to incorporate polymeric filament proteins, adapted from Sparrow and Schock 2009 [135], reprinted from Myhre and Pilgrim 2012 [136].

nistration of fibrates [35]. Reductions in skeletal muscle membrane CHOL were initially thought to account for the range of myopathic reactions. Additionally, the lowering the isoprenoid levels has been suggested to contribute to these pathologies as protein prenylation, and the potential consequences of a generalized insufficiency of this form of protein modifi‐ cation [36] are important for the activity and anchorage of plasma and other membrane proteins (nuclear envelope, dystrophin–glycoprotein complex, cytoskeletal G-proteins, etc.).

**Figure 6.** A model of the effect of membrane cholesterol depletion on the sarcolemmal distribution of β-DG. The sarco‐ lemmal lipid rafts close to the clear opening of the TT-membrane are enriched in cholesterol, GM1 (ganglioside M1), Cav-3 (caveolin-3), and β-DG (dystroglycan). The schematic diagram illustrates the diminished contact between β-DG and Dys (dystrophin) in the presence of MβCD (methyl β-cyclodextrin); the β-DG/Dys interaction is essential for later‐ al force transmission. SERCA1 (sarcoplasmic reticulum calcium ATPase) and RyR (ryanodine receptor (SR Ca2+ chan‐ nel)) function normally. SL, sarcolemma; SR, sarcoplasmic reticulum; EM, extracellular matrix; LR, lipid raft; Dys,

**SG**

**CSQ DHPR**

**RyR**

**M**

**M**

**RyR**

**DHPR**

**TT**

116 Muscle Cell and Tissue

**Dys**

**DG**

 **DG**

**Laminin**

**ECM**

**Cav- 3 Cav- 3**

**RS**

dystrophin; SG, sarcoglycan; CSQ, calsequestrin, adapted from Vega-Moreno et al. 2012 [17].

**[Ca2+]**


**ECM**

**RS**

**Dys**

**SG**

**CSQ DHPR**

**RyR [Ca2+]**

**RyR**

**M**

**M**

**DHPR**

**SERCA 1 SERCA 1**

**TT**

**[Ca2+] [Ca2+]**

**Z- disk Z- disk**

**DG**

**Laminin**

There is growing interest to decipher the molecular mechanism of the statin-induced myopa‐ thy, both by scientific community and pharmaceutical companies. One in three people over the age of 45 is taking a statin to reduce heart attack risk, the HMG-CoA antagonist with three orders of magnitude greater affinity to bind and subsequently to inhibit HMG-CoA reductase activity than that of natural substrate (HMG-CoA). Two in five women taking the statin are weaker than before, with one in ten reporting they felt "much worse". As statins became the most frequently prescribed drugs to prevent cardiovascular crisis alongside with the effort to pace physical activity, the issue how to protect from statin-induced myalgia, myositis, and rare cases of rhabdomyolysis is of great concern. In fact, due to muscle toxicity, an estimated 5–

10% of patients discontinue statin use due to myopathic symptoms. Reports of myositis and myopathic symptoms increase with increased statin dose [37], with different classes of statins, or when statins are coupled with other drugs [38], and with exercise [33]. The mechanistic underpinning of statin myopathy are believed to be multifactorial and partially attributed to

**Figure 8.** This figure shows the structure of the costamere and known molecular interactions. Below the membrane bilayer shown is the intracellular space and above it is the extracellular space. In the intracellular space, the costamere is attached to the contractile proteins through dystrophins (for the dystrophin glycoprotein complex, DGC), vinculin, talin, and paxilin (for the integrin complexes; not shown). In the extracellular space, both DGCs and integrin com‐ plexes bind to the components of the basal lamina that is attached to the rest of the extracellular matrix that consists mostly of fibrillar collagens, reprinted from Voermans et al. 2008 [137].

the regulatory effects of statins on apoptosis of muscle cells [39] and proliferation [3]. As skeletal muscle resident satellite cells (RSC) represent physiological reserve of undifferentiated muscle progenitors, it is obvious that activation followed by proliferation and migration are crucial in muscle adaptation to mechanical overload and regeneration from injury [40]. Thus, if statins impair RSC activation these processes could not be initiated. To tackle the problem of reduced CHOL concentration in plasma membrane and associated changes in the function of LR in muscle cells seems to be fundamental. The focal point is LR, where changes (bio‐ chemical and morphological) are presumably attributed to the consequences of disturbed cell signaling. As HMG-CoA reductase activity is ubiquitous, while the side effects of statins are confined to skeletal muscle, it is suggested that muscle tissue is featured by unique response to lower CHOL. RSC are targeted by CHOL depletion and the consequences are cumulative as muscle growth is stopped at the initiation phase (signal transduction). This assumption is supported by the data obtained from sarcolemma examination (single molecule microscopy and molecular studies), which demonstrate the isolation and downregulation of LR and the recruitment of mitochondrial oxidative phosphorylation system during myogenesis [41–42]. CHOL and GSL/SL are also present in the membranes of cellular organelles such as ER and Importance of Plasma Membrane Nanodomains in Skeletal Muscle Regeneration http://dx.doi.org/10.5772/60615 119

**Figure 9.** Schematic representation of a costamere and the focal adhesion complex (FAC). Two laminin receptors, a dystrophin/glycoprotein complex and an integrin receptor complex, are among the sarcolemmal structures that link the contractile apparatus of muscle fibers with the surrounding basal lamina. Components of both receptors, i.e., both dystrophin and the integrin-associated cytoskeletal proteins (talin, vinculin, α-actinin), colocalize in subsarcolemmal complexes which connect through γ-actin and the intermediate-filament proteins desmin and vimentin to the Z-disk of skeletal muscle fibers, adapted from Patel and Lieber 1997; Rybakova et al., 2000 [138–139], adapted from Fluck et al. 2002 [20].

Golgi complex (GC). Their role, however, remains obscure as methods to study effects of CHOL depletion in organelles are limited. The main concern should be placed on the sarcolemma and mitochondria, since these organelles control both muscle growth and development [43, 44].

### **5. Statin-induced myopathy and mitochondria**

the regulatory effects of statins on apoptosis of muscle cells [39] and proliferation [3]. As skeletal muscle resident satellite cells (RSC) represent physiological reserve of undifferentiated muscle progenitors, it is obvious that activation followed by proliferation and migration are crucial in muscle adaptation to mechanical overload and regeneration from injury [40]. Thus, if statins impair RSC activation these processes could not be initiated. To tackle the problem of reduced CHOL concentration in plasma membrane and associated changes in the function of LR in muscle cells seems to be fundamental. The focal point is LR, where changes (bio‐ chemical and morphological) are presumably attributed to the consequences of disturbed cell signaling. As HMG-CoA reductase activity is ubiquitous, while the side effects of statins are confined to skeletal muscle, it is suggested that muscle tissue is featured by unique response to lower CHOL. RSC are targeted by CHOL depletion and the consequences are cumulative as muscle growth is stopped at the initiation phase (signal transduction). This assumption is supported by the data obtained from sarcolemma examination (single molecule microscopy and molecular studies), which demonstrate the isolation and downregulation of LR and the recruitment of mitochondrial oxidative phosphorylation system during myogenesis [41–42]. CHOL and GSL/SL are also present in the membranes of cellular organelles such as ER and

mostly of fibrillar collagens, reprinted from Voermans et al. 2008 [137].

118 Muscle Cell and Tissue

**Figure 8.** This figure shows the structure of the costamere and known molecular interactions. Below the membrane bilayer shown is the intracellular space and above it is the extracellular space. In the intracellular space, the costamere is attached to the contractile proteins through dystrophins (for the dystrophin glycoprotein complex, DGC), vinculin, talin, and paxilin (for the integrin complexes; not shown). In the extracellular space, both DGCs and integrin com‐ plexes bind to the components of the basal lamina that is attached to the rest of the extracellular matrix that consists

> The development of statin-induced rhabdomyolysis is a morbid side effect of HMG-CoA reductase inhibition, occurring in less than 1 in 1000 statin-treated patients [45]. Even though occurrence of myopathy in statin-treated individuals has been estimated to range from 1 to 10%. Studies performed on rats confirmed that atorvastatin treatment reduces exercise capacity manifested by higher fatigability [46]. It is more common in statin users regularly exercised or statin-treated athletes pointing to the high correlation between the muscle contractive activity and muscle-associated complications of statin administration. Among several hypotheses raised to explain the aforementioned causal relationship between statins and physical exercise, growing body of evidence indicate both reactive oxygen species (ROS) and abnormal mitochondrial activity as probable inciting factors implicated in the deleterious effects of statins [47–48]. Actually, the inhibition of HMG-CoA reductase that hampers the

mevalonate pathway in addition to impaired cholesterol synthesis also reduces the synthesis of isoprenoids such as farnesyl pyrophosphate (FPP) and geranylgeranyl pyrophosphate (GGPP), intermediates affecting number of nonsteroid isoprenoids including coenzyme Q10/ ubiquinone [49], and finally it adversely affects selenoprotein synthesis as well as the biosyn‐ thesis of dolichols, which are involved in the process of protein glycosylation (Figure 10). The latter mechanism is rate limiting by dolichyl phosphate, which acts as a donor of oligosac‐ charides in glycoprotein synthesis [50]. The importance of glycoproteins in skeletal muscle growth and regeneration is further emphasized by the examination of N-linked glycoproteome of C2C12 mouse myoblasts and myotubes [51]. It is clear from this study that approximately 128 (117 transmembrane, 4 glycosylphospatidylinositol-anchored, 5 ECM, and 2 membraneassociated) proteins were identified, while a few N-linked glycoproteins (including aquapor‐ in-1 and β-sarcoglycan) were apparently of paramount value when myoblasts differentiate to myotubes. There was an evident decrease of aquaporin-1 and cadherin-2, whereas β-sarco‐ glycan expression level increased as the myoblasts fused and formed multinucleated syncy‐ tium. Ubiquinone is an important electron carrier between the complex I and complex III of respiratory chain in mitochondria. Thus, decrease in nonsteroid isoprenoids such as CoQ10 leads to the inhibition of complex I, incomplete reduction of oxygen at the level of cytochrome *c* oxidase (complex IV), and induces oxidative stress (augmentation of superoxide anion radical, hydrogen peroxide, and finally hydroxyl radical) postulated to cause myotoxicity of statins [48]. Myotoxicity is therefore at least in part the consequence of damage to lipids, proteins, and DNA, although lipids in cellular membranes are most likely targets of ROS (extensive lipid peroxidation). Additionally, limited access of proteins to prenylation impairs important lipid anchorages essential for PM attachment and function of a variety of proteins involved in cell signaling (i.e., heterotrimeric guanine nucleotide-binding protein-coupled receptors – GPCRs, GTP-binding small/G-proteins Rap1, Ras, Rac, and Rho).

#### **6. LR partners control skeletal muscle regeneration**

GPCRs, the most abundant PM receptors, regulate wide range of cellular processes through intracellular heterotrimeric G protein (GTP-binding protein). The latter acts as a signal transducer to control the activity of several catalytic proteins central to the message amplifi‐ cation and its intracellular broadening to effector proteins. Heterotrimeric proteins are composed of three subunits (Gαβγ). Agonist-mediated activation of GPCRs brings about conformational changes which lead to the exchange of guanosine diphosphate (GDP) for GTP on the Gα-subunit which then dissociates from Gβγ dimer. Now, Gα may translocate to the target protein(s), whereas Gβγ dimer inactivates the receptor through phosphorylation (it recruits G protein–coupled receptor kinase – GRK to inactivate the receptor). As GPCRs constitute the largest family of membrane receptors, there are at least 16 types of Gα subunits, 5 of Gβ subunits, and 12 types of Gγ subunits [52]. Gα signaling is stopped when GTP is converted to GDP by the intrinsic GTPase activity of the protein itself. In consequence, Gα-GDP is reassembled with Gβγ dimer and G-protein complex is reestablished. Importantly, target proteins for heterotrimeric G-proteins are membrane bound suggesting that lipid–

mevalonate pathway in addition to impaired cholesterol synthesis also reduces the synthesis of isoprenoids such as farnesyl pyrophosphate (FPP) and geranylgeranyl pyrophosphate (GGPP), intermediates affecting number of nonsteroid isoprenoids including coenzyme Q10/ ubiquinone [49], and finally it adversely affects selenoprotein synthesis as well as the biosyn‐ thesis of dolichols, which are involved in the process of protein glycosylation (Figure 10). The latter mechanism is rate limiting by dolichyl phosphate, which acts as a donor of oligosac‐ charides in glycoprotein synthesis [50]. The importance of glycoproteins in skeletal muscle growth and regeneration is further emphasized by the examination of N-linked glycoproteome of C2C12 mouse myoblasts and myotubes [51]. It is clear from this study that approximately 128 (117 transmembrane, 4 glycosylphospatidylinositol-anchored, 5 ECM, and 2 membraneassociated) proteins were identified, while a few N-linked glycoproteins (including aquapor‐ in-1 and β-sarcoglycan) were apparently of paramount value when myoblasts differentiate to myotubes. There was an evident decrease of aquaporin-1 and cadherin-2, whereas β-sarco‐ glycan expression level increased as the myoblasts fused and formed multinucleated syncy‐ tium. Ubiquinone is an important electron carrier between the complex I and complex III of respiratory chain in mitochondria. Thus, decrease in nonsteroid isoprenoids such as CoQ10 leads to the inhibition of complex I, incomplete reduction of oxygen at the level of cytochrome *c* oxidase (complex IV), and induces oxidative stress (augmentation of superoxide anion radical, hydrogen peroxide, and finally hydroxyl radical) postulated to cause myotoxicity of statins [48]. Myotoxicity is therefore at least in part the consequence of damage to lipids, proteins, and DNA, although lipids in cellular membranes are most likely targets of ROS (extensive lipid peroxidation). Additionally, limited access of proteins to prenylation impairs important lipid anchorages essential for PM attachment and function of a variety of proteins involved in cell signaling (i.e., heterotrimeric guanine nucleotide-binding protein-coupled

120 Muscle Cell and Tissue

receptors – GPCRs, GTP-binding small/G-proteins Rap1, Ras, Rac, and Rho).

GPCRs, the most abundant PM receptors, regulate wide range of cellular processes through intracellular heterotrimeric G protein (GTP-binding protein). The latter acts as a signal transducer to control the activity of several catalytic proteins central to the message amplifi‐ cation and its intracellular broadening to effector proteins. Heterotrimeric proteins are composed of three subunits (Gαβγ). Agonist-mediated activation of GPCRs brings about conformational changes which lead to the exchange of guanosine diphosphate (GDP) for GTP on the Gα-subunit which then dissociates from Gβγ dimer. Now, Gα may translocate to the target protein(s), whereas Gβγ dimer inactivates the receptor through phosphorylation (it recruits G protein–coupled receptor kinase – GRK to inactivate the receptor). As GPCRs constitute the largest family of membrane receptors, there are at least 16 types of Gα subunits, 5 of Gβ subunits, and 12 types of Gγ subunits [52]. Gα signaling is stopped when GTP is converted to GDP by the intrinsic GTPase activity of the protein itself. In consequence, Gα-GDP is reassembled with Gβγ dimer and G-protein complex is reestablished. Importantly, target proteins for heterotrimeric G-proteins are membrane bound suggesting that lipid–

**6. LR partners control skeletal muscle regeneration**

**Figure 10.** The biosynthetic pathway of cholesterol and other cometabolites, reprinted from Vaklavas et al. 2009 [58].

protein and/or lipid–lipid interactions are fundamental for G-protein-mediated signaling. Actually, all known Gγ proteins undergo isoprenylation on cysteine residues (either geranyl‐ geranyl or farnesyl moieties) pointing to increased affinity to hexagonal nonraft phase (e.g., PE) of PM [53–54]. In contrast, Gα subunits are modified by myristoylation (Gα<sup>i</sup> ) and/or reversible palmitoylation (Gα) allowing them to get access to lamellar regions of PM (e.g., lipid rafts). It also explains how Gα subunits migrate to their cognate targets in LR after dissociation form Gβγ assembled to GPCRs. Minetti et al. [55] have showed in elegant study that skeletal muscle hypertrophy and differentiation are greatly influenced by signaling induced by lysophosphatidic acid (LPA) acting on GPCRs which in turn activate a Gα<sup>i</sup> protein. Besides, Gα<sup>i</sup> enhanced muscle regeneration and caused switch to oxidative fibers and can act as a counterbalance to MuRF1 and MAFbx/atrogin-1. To sum up, lipid structures play active role in signal propagation with resulting localization of Gα and Gβγ proteins.

Uncommon myopathic changes resultant from statin therapy offer the opportunity to gain new insight for the function of biochemical pathways downstream to HMG-CoA reductase in skeletal muscles. Irrespective of the type of statin treatment (hydrophilic or hydrophobic), the viability of skeletal muscle cells is considerably reduced, though the effect depends largely on the pharmacokinetic and pharmacodynamic properties of statins [56–57], while the signaling pathway(s) and molecular mechanisms are still not fully understood. Sometimes, signal transduction is dependent on small GTPase proteins that cycle between an inactive guanosine diphosphate (GDP)-bound and active guanosine triphosphate (GTP)-bound state. The posttranslational prenylation of these proteins occurs by the covalent addition of only two types of isoprenoids, FPP and GGPP, to cysteine residues at or near the C-terminus. Upon tyrosine kinase receptor activation, the prenylated (PM protoplasmic/inner leaflet attached) small GTPase protein Ras (MAPK kinase kinase kinase) binds GTP and becomes activated to initiate MAPK cascade ending with the stimulation of muscle cell growth (hyperplasia). In addition, small GTPase protein Rab1 (more than 60 Rab small GTPase isoforms have been identified) is involved in organelle biogenesis and intracellular vesicular trafficking [58]. Overall growth and survival signals depend on the activation of both protein receptor and nonreceptor tyrosine kinases.

Several lines of evidence suggest particular significance of IGF-1/PI3-K/AKT cascade in maintaining muscle cell growth and viability [59–61] likely through the suppression of FOXOdependent activity of atrogin/MAFbx ubiquitin ligase gene required for the development of muscle atrophy [62–63]. Moreover, the statin-induced muscle damage is controlled by PGC-1α, a transcriptional coactivator that induces mitochondrial biogenesis and protects against the development of statin-induced muscle atrophy [64]. In *in vivo* model, simvastatin downregulated PI3-K/AKT signaling and upregulated FOXO transcription factors and downstream gene targets known to be implicated in muscle cachexia [63]. Insulin and IGF-1 are widely known agonists of their cognate receptors (IR and IGF-1R, respectively), although at concentrations higher than physiological cross-reactivity of insulin to IGF-1R and IGF-1 to IR were observed. On the other hand, LR have been shown to be platforms to initiate cellular signal transduction of IGF-1 and insulin-inducing skeletal muscle differentiation and hyper‐ trophy. Notably, the impaired insulin/IGF-1 signaling [65–67] mimics the side effects of statin administration, whereas insulin and/or IGF-1 were reported to overcome statin-induced myopathy [61]. IR and IGF-1R with their intrinsic tyrosine kinase activities transduce the signal by recruiting insulin receptor substrate-1 (IRS-1) with its src-homology 2 domains (SH2) to the receptor phosphotyrosines. IRS-1 activates PI3-K/AKT/mTOR and Ras/Raf/MEK/ERK path‐ ways, however, phosphoinositide 3-kinase (PI3-K) as a lipid kinase converts plasma mem‐ brane phosphatidylinositol-4,5-biphosphate (PIP2) to phosphatidylinositol-3,4,5-triphosphate (PIP3). The latter attracts kinases with pleckstrin homology domain (PH) downstream to PI3- K including phosphoinositide-dependent kinase 1 and 2 (PDK1, PDK2) and AKT. Depletion of CHOL and sphingolipids blocks IGF-1-induced AKT membrane recruitment, whereas LR reconstitution in CHOL- and sphingolipid-deficient cells restored AKT membrane recruitment and phosphorylation [68]. Thus, LR-localized PIP3 is essential for AKT membrane association, while AKT seems to promote formation of PIP3-containing rafts.
