**3. Statins adverse effects**

**2. Statins pleiotropic effects**

390 Mitochondrial Diseases

thione peroxidase activities [105].

a rat colitis model [108].

**2.2. Statins and cancer**

**2.1. Antioxidant responses triggered by statins**

Statins are among the most commonly prescribed medicines worldwide. They are safe and well-tolerated and seem to present a range of cholesterol-independent protective actions called pleiotropic effects. Indeed, several studies claim that statins act as antioxidants [19, 96], anti-inflammatory agents [97], and can increase stability of the atherosclerotic plaque [98],

Extensive literature reports have indicated that antioxidant effects can be attributed to statins. It has been postulated that statins decrease systemic or local oxidative stress and this appears to confer additional vascular protection. The first possible mechanism for this protective effect could be secondary to statins' main target effect, which is to decrease the concentration of the oxidizable substrate, LDLc. This decrease may lead to a reduction in oxidized-LDL, which

Another antioxidant mechanism frequently attributed to statins is the upregulation of cellular antioxidant defenses. For instance, atorvastatin treatment decreased the expression of essential NAD(P)H oxidase subunits and upregulated catalase expression in cultured rat vascular smooth muscle cells and in the vasculature of spontaneous hypertensive rats (SHR) [104]. Simvastatin treatment restored endothelial function in SHR by increasing superoxide dismutase and gluta-

Other studies have demonstrated a protective effect by statins against oxidative damage of biomolecules. In whole blood leukocytes of non-treated dyslipidemic diabetic type 2 patients, simvastatin treatment [19] protected against DNA oxidative damage. Similarly, rosuvastatin inhibited lipid peroxidation and attenuated the oxidative damage to DNA in treated rat liver [106]. Rosuvastatin-treated HL-60 cells exhibited a glutathione-dependent protective mechanism against DNA oxidation [107]. In addition, simvastatin or fluvastatin administration prevented lipid peroxidation, superoxide generation, cytokine production, and neutrophil accumulation in

With respect to statins' effects on specific mitochondrial redox homeostasis, literature reports are more controversial. It was shown that atorvastatin and simvastatin reduced oxidative stress triggered by Ca2+ and prevented MPT and cytochrome c release in rat liver mitochondria [96]. On the other hand, results from our group and others suggest that statins, when administered to mitochondria, muscle biopsies, or *in vivo* exert pro-oxidant activities (this will be discussed in more detail in the next section) [47, 49, 109]. Thus, our hypothesis for the alleged statin antioxidant effects is based on the mitohormesis concept [37, 38]: mild mitochondrial oxidative stress caused by statins may function as a signal that leads to a cellular adaptive response such as increasing the expression and activity of cellular antioxidant systems in order to overcome this stress.

Statins have been proposed as adjuvant in cancer therapy since the 1990s and, until then, several mechanisms have been proposed for this specific function depending on the type of cancer and

constitutes a very early step involved in atherosclerosis development [101–103].

improve endothelial function [99], and induce cancer cell death [100].

After decades of statins' use, some side effects have been consistently described in a minority of patients, particularly regarding muscle function. Adverse effects other than muscle symptoms such as headache, digestive problems, liver enzymes abnormalities, and neurological dysfunction may occur in some patients [127, 128]. The side effects are often the decisive factor for the noncompliance to statins treatment [129, 130] and its discontinuation usually makes the side effect symptoms disappear [131].

The precise mechanisms involved in statins toxicity and the reasons why only a few subjects are affected remain unclear. Several groups, including ours, have proposed that mitochondria are the main players in statin-induced toxicity.

#### **3.1. Mitochondrial dysfunction caused by statins treatment**

Mitochondrial redox imbalance is associated with aging, degenerative disorders, and druginduced toxicity [26, 132]. Several reports concerning statin *in vitro* effects on isolated tissues or mitochondria from experimental models demonstrated that statins promote inhibition of mitochondrial respiration, mitochondrial oxidative stress, and cell death [47, 49, 109, 133]. It has been previously shown that lipophilic (cerivastatin, fluvastatin, atorvastatin, and simvastatin) and hydrophilic (pravastatin) statins-induced mitochondrial membrane potential decrease in rat skeletal muscle cell line (L6) [133]. The four lipophilic statins also induced mitochondrial swelling, cytochrome c release, and DNA fragmentation in these L6 cells. Mitochondrial β-oxidation enzymes activities were strongly impaired by all lipophilic statins, but in the case of pravastatin, it occurred only at high concentrations. In isolated rat skeletal muscle mitochondria, glutamatesupported state 3 respiration and respiratory control ratios were decreased by all lipophilic statins, but not by pravastatin [133]. According to the authors, this mitochondrial dysfunction caused by lipophilic statins in skeletal muscle might partially explain the muscle symptoms presented by some patients. Abdoli and coworkers demonstrated in isolated rat liver mitochondria that atorvastatin, simvastatin, and lovastatin increased ROS formation followed by lipid peroxidation, inner mitochondrial membrane depolarization, and a decreased GSH/GSSG ratio [47].

**3.3. Statins effects on respiratory chain complexes**

of patients undergoing statin treatment [147].

in the lactone form binds to Q<sup>o</sup>

**4. Muscle sensitivity to statins**

[49, 148].

It is well known that enzymes containing 4Fe-4S clusters are particularly vulnerable to damage by superoxide or peroxynitrite radicals [140–145]. Complexes I and II present six and one of these 4Fe-4S clusters, respectively, thus showing a high superoxide-sensitivity. Some studies have demonstrated that superoxide generation inhibits respiration at complex I and II levels as a result of 4Fe-4S clusters damage. These alterations diminish resistance to Ca2+-induced MPT and induce necrotic cell death [65, 145]. As mentioned before, our group demonstrated that mitochondrial dysfunction caused by simvastatin incubation in permeabilized skeletal muscle was L-carnitine and CoQ10 sensitive [49]. L-carnitine did not protect against CoQ10 depletion, indicating that both CoQ10 and L-carnitine are protecting mitochondrial respiration due to its ROS scavenging properties. Since L-carnitine also binds Fe2+ [146], it is feasible that this antioxidant molecule interacts with 4Fe-4S clusters in complexes I and II of the respiratory chain, protecting these sites against superoxide attack. Simvastatin lowered the ADP-stimulated respiration supported by substrates of complexes I and II in primary human skeletal myotubes and increased susceptibility to MPT, mitochondrial oxidative stress, and apoptosis [48]. These results are in agreement with a decrease in complex I activity in muscle

Mitochondrial Oxidative Stress and Calcium-Dependent Permeability Transition are Key Players…

http://dx.doi.org/10.5772/intechopen.71610

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Another study performed in myoblasts culture (C2C12) incubated with several statins (atorvastatin, cerivastatin, fluvastatin, lovastatin, pitavastatin, pravastatin, rosuvastatin, and simvastatin) showed that the respiratory capacity is reduced not only at the levels of respiratory chain complexes I and II but also in complex III [148]. In this case, it was suggested that statins

activity of muscle from patients presenting myopathies induced by statins was also reduced [148]. On the other hand, statins do not seem to affect the complex IV-supported respiration

It is well known that about 10% of patients undergoing statin treatment develop mild myopathic symptoms such as weakness, muscle pain, exercise intolerance, and other symptoms that are usually with normal or minimally elevated creatine kinase (CK) serum levels [149, 150]. Moreover, myositis, defined as muscle symptoms associated with increased CK, is usually present [151, 152]. Rhabdomyolysis, the most severe adverse effect of statins, is a very rare condition affecting 1.6/100,000 patients-years. It may result in acute renal failure and disseminated intravascular coagulation, leading to death. This condition is frequently related to drug interactions and occurs with CK levels 10-fold higher than the normal limit and elevated levels of creatinine [153, 154]. Increased intracellular lipid stores, cytochrome oxidase-negative myofibers, ragged red fibers, and subsarcolemmal accumulation of mitochondria were found in patients with muscle symptoms during statin therapy [155, 156]. Schick and colleagues also observed reduced mitochondrial DNA levels in patients treated with simvastatin [157]. Muscle-associated statin toxicity seems to be more severe with increasing lipophilicity, whereas more hydrophilic statins exert only mild or no toxicity [133, 153]. The myotoxic effect is attributed to their ability to penetrate and accumulate in cell membranes and alter their structural conformation [158–160].

site of complex III, inhibiting its activity. Similarly, complex III

More recently, mitochondrial redox imbalance [67, 134] was observed in a genetic human familial hypercholesterolemia mouse model, the LDL receptor knockout mouse (*LDLr−/−)* [135]. Mitochondria isolated from several tissues of these mice (liver, heart, and brain) and intact spleen mononuclear cells presented higher ROS production and higher susceptibility to MPT. In addition, these mitochondria showed lower capacity to sustain reduced NADPH [67, 134], which is the most important reducing power involved in reconstituting mitochondrial antioxidant systems [132]. As a consequence, H<sup>2</sup> O2 accumulates and PTP opens [67, 134]. Since cholesterol synthesis consumes a large amount of NADPH, we have proposed that the increased steroidogenesis observed in these mice would be partially responsible for the lower mitochondrial content of NADPH and Krebs cycle intermediates observed in their liver mitochondria [67, 134]. Therefore, we hypothesized that inhibition of cholesterol synthesis by statins treatment could prevent the decrease in NADPH oxidation in *LDLr−/−* mice mitochondria. Unexpectedly, liver mitochondria from wild type and *LDLr−/−* mice treated with lovastatin presented a higher susceptibility to PTP opening, and *in vitro* experiments revealed a drug dose- and class-dependence of this effect [109]. Statin induced PTP opening was shown to be Ca2+-dependent and associated with oxidation of protein thiol groups. Thus, statins induced a direct oxidative damage in mitochondrial proteins [109].

#### **3.2. Ca2+ and statins toxicity**

It has been proposed by our group and others that statins impair cellular Ca2+ homeostasis, leading to mitochondrial dysfunction. Increased cytosolic Ca2+ levels were observed after simvastatin treatment of myoblasts culture [136], rat skeletal muscle [137], and human skeletal muscle fibers, and this was followed by mitochondrial Ca2+ accumulation [138]. Indeed, Hattori and coworkers [139] proposed that statins induced Ca2+ release from the endoplasmic reticulum to the cytosol in human CD19+ primary lymphocytes. As a consequence of high Ca2+ levels in the cytosol, Ca2+ enters the mitochondria and induces MPT as demonstrated by our group in PC3 cells after simvastatin treatment [21, 22].
