Biochemical rationale and experimental data on the antiaging properties of CoQ(10) at skin level

Coenzyme Q(10) (CoQ(10) ) is a key component of the mitochondrial respiratory chain and, therefore, is essential for the bioenergetics of oxidative phosphorylation. It is also endowed with antioxidant properties, and recent studies pointed out its capability of affecting the expression of different genes. In this review, we analyze the data on the mechanisms by which CoQ(10) interacts with skin aging processes. The effect of CoQ(10) in preserving mitochondrial function cooperates in maintaining a proper energy level, which serves to prevent the aging skin from switching to anaerobic energy production mechanisms. Furthermore, the antioxidant capacity of CoQ(10) contributes to a positive effect against UV-mediated oxidative stress. Some of these effects have been assessed also in vivo, by the sensitive technique of ultraweak photoemission. Finally, CoQ(10) has been shown to influence, through a gene induction mechanism, the synthesis of some key proteins of the skin and to decrease the expression of some metalloproteinase such as collagenase. These mechanisms may also contribute to preserve collagen content of the skin.     

Effect of mitochondrial dysfunction and oxidative stress on endogenous levels of coenzyme Q(10) in human cells

Little is known about the regulation of endogenous CoQ(10) levels in response to mitochondrial dysfunction or oxidative stress although exogenous CoQ(10) has been extensively used in humans. In this study, we first demonstrated that acute treatment of antimycin A, an inhibitor of mitochondrial complex III, and the absence of mitochondrial DNA suppressed CoQ(10) levels in human 143B cells. Because these two conditions also enhanced formation of reactive oxygen species (ROS), we further investigated whether oxidative stress or mitochondrial dysfunction primarily contributed to the decrease of CoQ(10) levels. Results showed that H(2)O(2) augmented CoQ(10) levels, but carbonyl cyanide-p-trifluoromethoxyphenylhydrazone (FCCP), a chemical uncoupler, decreased CoQ(10) levels in 143B cells. However, H(2)O(2) and FCCP both increased mRNA levels of multiple COQ genes for biosynthesis of CoQ(10) . Our findings suggest that ROS induced CoQ(10) biosynthesis, whereas mitochondrial energy deficiency caused secondary suppression of CoQ(10) levels possibly due to impaired import of COQ proteins into mitochondria.     

Cytoprotective and anticancer properties of coenzyme Q versus capsaicin

Coenzyme Q (CoQ) is an essential component of the mitochondrial electron transport chain and serves as an electron donor and acceptor in mitochondrial energy-linked respiration. CoQ1 was shown to prevent ROS formation and cell death in complex 1 inhibited cells. Low concentrations of capsaicin like CoQ1 inhibited ROS formation but CoQ1 was more effective at restoring the mitochondrial membrane potential collapse caused by complex 1 inhibitors such as rotenone. At low concentrations, capsaicin acts as a CoQ mimic by protecting against rotenone induced ROS formation and mitochondrial membrane potential collapse. Lipid peroxidation in isolated rat hepatocytes induced by cumene hydroperoxide and chloroacetaldehyde was also prevented. At higher concentrations, capsaicin and CoQ1 became cytotoxic. Hep G2 cells were more susceptible than hepatocytes. The cytotoxic mechanism for both capsaicin and CoQ1 was shown to involve a collapse of the mitochondrial membrane potential, however, only capsaicin caused ROS formation. The capsaicin side chain was required for capsaicin induced cytotoxicity. The anticancer properties of CoQ1 and capsaicin should prove useful for inducing tumor cell apoptosis.     

Coenzyme Q10, a cutaneous antioxidant and energizer.

The processes of aging and photoaging are associated with an increase in cellular oxidation. This may be in part due to a decline in the levels of the endogenous cellular antioxidant coenzyme Q10 (ubiquinone, CoQ10). Therefore, we have investigated whether topical application of CoQ10 has the beneficial effect of preventing photoaging. We were able to demonstrate that CoQ10 penetrated into the viable layers of the epidermis and reduce the level of oxidation measured by weak photon emission. Furthermore, a reduction in wrinkle depth following CoQ10 application was also shown. CoQ10 was determined to be effective against UVA mediated oxidative stress in human keratinocytes in terms of thiol depletion, activation of specific phosphotyrosine kinases and prevention of oxidative DNA damage. CoQ10 was also able to significantly suppress the expression of collagenase in human dermal fibroblasts following UVA irradiation. These results indicate that CoQ10 has the efficacy to prevent many of the detrimental effects of photoaging.     

Effects of inhibiting CoQ10 biosynthesis with 4-nitrobenzoate in human fibroblasts

Coenzyme Q(10) (CoQ(10)) is a potent lipophilic antioxidant in cell membranes and a carrier of electrons in the mitochondrial respiratory chain. We previously characterized the effects of varying severities of CoQ(10) deficiency on ROS production and mitochondrial bioenergetics in cells harboring genetic defects of CoQ(10) biosynthesis. We observed a unimodal distribution of ROS production with CoQ(10) deficiency: cells with <20% of CoQ(10) and 50-70% of CoQ(10) did not generate excess ROS while cells with 30-45% of CoQ(10) showed increased ROS production and lipid peroxidation. Because our previous studies were limited to a small number of mutant cell lines with heterogeneous molecular defects, here, we treated 5 control and 2 mildly CoQ(10) deficient fibroblasts with varying doses of 4-nitrobenzoate (4-NB), an analog of 4-hydroxybenzoate (4-HB) and inhibitor of 4-para-hydroxybenzoate:polyprenyl transferase (COQ2) to induce a range of CoQ(10) deficiencies. Our results support the concept that the degree of CoQ(10) deficiency in cells dictates the extent of ATP synthesis defects and ROS production and that 40-50% residual CoQ(10) produces maximal oxidative stress and cell death.     

Effect of UV-C mediated oxidative stress in leukemia cell lines and its relation to ubiquinone content

UV-C radiation is able to impair cellular functions by directly damaging DNA, and by inducing an increased formation of reactive oxygen species that leads to a condition of oxidative stress. In this study we evaluated different responses to UV insult of two leukemia cell lines, HL-60 and Raji, and the relationship with their CoQ10 content. DNA damage was monitored by means of the alkaline single cell gel electrophoresis (Comet assay); intracellular levels of ROS, mitochondrial depolarization and cell viability was measured by flow cytometry. Raji cells appeared more resistant to the UV insult; moreover, they did not show any increase in ROS content and the extent of mitochondrial depolarisation was much lower than in HL 60 cell line. Raji cells also contained significantly higher levels of CoQ10 and their ability to incorporate and to reduce exogenous CoQ10 added to the culture medium was remarkably elevated compared with HL 60.     

Characterization of cellular uptake and distribution of coenzyme Q10 and vitamin E in PC12 cells

Coenzyme Q (CoQ) is a well-known electron transporter in the mitochondrial respiratory chain. Furthermore, ubiquinol (UQH(2))--a reduced form of ubiquinone (UQ)--has been shown to act as a radical-scavenging antioxidant. Some studies have reported the beneficial effect of CoQ addition to cultured cells; however, the cellular uptake and distribution of CoQ have not been elucidated. In the present study, we used rat pheochromocytoma PC12 cells to investigate and compare the cellular uptake and distribution of CoQ(10) and alpha-tocopherol (alphaT). UQ(10) or UQ(10)H(2) treatment resulted in an increase in the cellular content of both CoQ(10) in a time- and concentration-dependent manner. A subcellular fractionation study revealed that the added UQ(10) as well as UQ(10)H(2) mainly localized in the mitochondrial fraction, which is similar to the localization of endogenous CoQ but different from that of alphaT. The cellular distribution of alphaT directly corresponded to the lipid distribution, while the CoQ distribution did not show any relationship with the lipid distribution, particularly in the mitochondrial and microsomal fractions. These results indicate that the cellular distribution of CoQ is completely different from that of alphaT; moreover, a certain system which accumulates CoQ preferentially in mitochondria may be suggested.     

Mitochondrial production of oxygen radical species and the role of Coenzyme Q as an antioxidant

The mitochondrial respiratory chain is a powerful source of reactive oxygen species (ROS), which is considered as the pathogenic agent of many diseases and of aging. We have investigated the role of complex I in superoxide radical production and found by the combined use of specific inhibitors of complex I that the one-electron donor to oxygen in the complex is a redox center located prior to the sites where three different types of Coenzyme Q (CoQ) competitors bind, to be identified with an Fe-S cluster, most probably N2, or possibly an ubisemiquinone intermediate insensitive to all the above inhibitors. Short-chain Coenzyme Q analogs enhance superoxide formation, presumably by mediating electron transfer from N2 to oxygen. The clinically used CoQ analog, idebenone, is particularly effective, raising doubts on its safety as a drug. Cells counteract oxidative stress by antioxidants. CoQ is the only lipophilic antioxidant to be biosynthesized. Exogenous CoQ, however, protects cells from oxidative stress by conversion into its reduced antioxidant form by cellular reductases. The plasma membrane oxidoreductase and DT-diaphorase are two such systems, likewise, they are overexpressed under oxidative stress conditions.     

Coenzyme Q cytoprotective mechanisms for mitochondrial complex I cytopathies involves NAD(P)H: quinone oxidoreductase 1(NQO1)

The commonest mitochondrial diseases are probably those impairing the function of complex I of the respiratory electron transport chain. Such complex I impairment may contribute to various neurodegenerative disorders e.g. Parkinson's disease. In the following, using hepatocytes as a model cell, we have shown for the first time that the cytotoxicity caused by complex I inhibition by rotenone but not that caused by complex III inhibition by antimycin can be prevented by coenzyme Q (CoQ1) or menadione. Furthermore, complex I inhibitor cytotoxicity was associated with the collapse of the mitochondrial membrane potential and reactive oxygen species (ROS) formation. ROS scavengers or inhibitors of the mitochondrial permeability transition prevented cytotoxicity. The CoQ1 cytoprotective mechanism required CoQ1 reduction by DT-diaphorase (NQO1). Furthermore, the mitochondrial membrane potential and ATP levels were restored at low CoQ1 concentrations (5 microM). This suggests that the CoQ1H2 formed by NQO1 reduced complex III and acted as an electron bypass of the rotenone block. However cytoprotection still occurred at higher CoQ1 concentrations (>10 microM), which were less effective at restoring ATP levels but readily restored the cellular cytosolic redox potential (i.e. lactate: pyruvate ratio) and prevented ROS formation. This suggests that CoQ1 or menadione cytoprotection also involves the NQO1 catalysed reoxidation of NADH that accumulates as a result of complex I inhibition. The CoQ1H2 formed would then also act as a ROS scavenger.     

Coenzyme Q10 Protects Astrocytes from ROS-Induced Damage through Inhibition of Mitochondria-Mediated Cell Death Pathway

Coenzyme Q10 (CoQ10) acts by scavenging reactive oxygen species to protect neuronal cells against oxidative stress in neurodegenerative diseases. The present study was designed to examine whether CoQ10 was capable of protecting astrocytes from reactive oxygen species (ROS) mediated damage. For this purpose, ultraviolet B (UVB) irradiation was used as a tool to induce ROS stress to cultured astrocytes. The cells were treated with 10 and 25 μg/ml of CoQ10 for 3 or 24 h prior to the cells being exposed to UVB irradiation and maintained for 24 h post UVB exposure. Cell viability was assessed by MTT conversion assay. Mitochondrial respiration was assessed by respirometer. While superoxide production and mitochondrial membrane potential were measured using fluorescent probes, levels of cytochrome C (cyto-c), cleaved caspase-9, and caspase-8 were detected using Western blotting and/or immunocytochemistry. The results showed that UVB irradiation decreased cell viability and this damaging effect was associated with superoxide accumulation, mitochondrial membrane potential hyperpolarization, mitochondrial respiration suppression, cyto-c release, and the activation of both caspase-9 and -8. Treatment with CoQ10 at two different concentrations started 24 h before UVB exposure significantly increased the cell viability. The protective effect of CoQ10 was associated with reduction in superoxide, normalization of mitochondrial membrane potential, improvement of mitochondrial respiration, inhibition of cyto-c release, suppression of caspase-9. Furthermore, CoQ10 enhanced mitochondrial biogenesis. It is concluded that CoQ10 may protect astrocytes through suppression of oxidative stress, prevention of mitochondrial dysfunction, blockade of mitochondria-mediated cell death pathway, and enhancement of mitochondrial biogenesis.     

Coenzyme Q Cytoprotective Mechanisms for Mitochondrial Complex I Cytopathies Involves NAD(P)H: Quinone Oxidoreductase 1(NQO1)

The commonest mitochondrial diseases are probably those impairing the function of complex I of the respiratory electron transport chain. Such complex I impairment may contribute to various neurodegenerative disorders e.g. Parkinson's disease. In the following, using hepatocytes as a model cell, we have shown for the first time that the cytotoxicity caused by complex I inhibition by rotenone but not that caused by complex III inhibition by antimycin can be prevented by coenzyme Q (CoQ 1 ) or menadione. Furthermore, complex I inhibitor cytotoxicity was associated with the collapse of the mitochondrial membrane potential and reactive oxygen species (ROS) formation. ROS scavengers or inhibitors of the mitochondrial permeability transition prevented cytotoxicity. The CoQ 1 cytoprotective mechanism required CoQ 1 reduction by DT-diaphorase (NQO 1 ). Furthermore, the mitochondrial membrane potential and ATP levels were restored at low CoQ 1 concentrations (5 &#119 M). This suggests that the CoQ 1 H 2 formed by NQO 1 reduced complex III and acted as an electron bypass of the rotenone block. However cytoprotection still occurred at higher CoQ 1 concentrations (>10 &#119 M), which were less effective at restoring ATP levels but readily restored the cellular cytosolic redox potential (i.e. lactate: pyruvate ratio) and prevented ROS formation. This suggests that CoQ 1 or menadione cytoprotection also involves the NQO 1 catalysed reoxidation of NADH that accumulates as a result of complex I inhibition. The CoQ 1 H 2 formed would then also act as a ROS scavenger.     

How to clean the dirtiest place in the cell: cationic antioxidants as intramitochondrial ROS scavengers

Membrane-penetrating triphenyl alkyl phosphonium cations have been suggested for many years in our group as having the ability to measure mitochondrial potential were recently used by Murphy as vehicles to specifically target CoQ to mitochondria. As was shown in our group, the phosphonium derivative of CoQ (MitoQ) easily penetrates a planar bilayer phospholipid membrane as a cation, generating 60 mV electric potential (Deltapsi) per a 10-fold MitoQ gradient. This means that MitoQ should be unequally distributed across the inner mitochondrial membrane, the intramitochondrial [MitoQ] = extramitochondrial [MitoQ] x 10(3) at 180 mV Deltapsi. In line with such a calculation, Murphy and his colleagues reported that antioxidant efficiency of MitoQ added to mitochondria or cells appears to be very much higher than of CoQ. It was found that H2O2-induced apoptosis (Murphy) and the H2O2-mediated bystander killing of the cultivated cells (our group) are completely arrested by pretreatement of the cells with 10(-10) - 10(-8) M MitoQ. These effects indicate that MitoQ and similar compounds may be promising in treatment of heart attack, stroke and other diseases accompanied by massive apoptosis in the injured tissue. The very fact that: (i) MitoQ is not only accumulated by mitochondria but also can be regenerated in its reduced form by mitochondrial respiratory chain, (ii) it is the mitochondrial interior that produces a large portion of reactive oxygen species (ROS) in our body, and (iii) the most sensitive ROS targets are localized in the mitochondrial matrix suggest the MitoQ-like compounds are promising tools of molecular therapy of aerobic cells. In line with this suggestion, we found that addition of MitoQ strongly improves structural and biochemical parameters of cultivated cells. As to cationic tetrapeptides, recently advertised as mitochondrially-targeted Deltapsi-independent antioxidants, their effect is most probably mediated by an opioid activity inherent in some of these substances.     

The effects of dietary coenzyme Q on Drosophila life span

Reactive oxygen species (ROS), generated as by-products of aerobic metabolism, cause damage to proteins and cellular membranes, and are thus thought to influence senescence. Caenorhabditis elegans fed on diets lacking in ubiquinone coenzyme Q (CoQ), a coenzyme in the oxidative phosphorylation pathway, show increased longevity, possibly because of reduced ROS generation. We test the role of dietary CoQ in determining Drosophila melanogaster longevity by measuring survival and cytochrome c-oxidase activity (a proxy for aerobic metabolic performance) in flies fed wild-type yeast, CoQ-less yeast, or respiratory control (RC) yeast replete with CoQ but independently deficient in mitochondrial respiration. We find no evidence that dietary manipulation of CoQ in D. melanogaster increases life span or decreases age-dependent decline in cytochrome c oxidase activity. Instead, we find evidence that flies fed a diet of respiratory-deficient yeast (CoQ-less or RC) tend to have decreased longevity and increased rates of decline in cytochrome c-oxidase activity [corrected]     

A Novel Endogenous Indole Protects Rodent Mitochondria and Extends Rotifer Lifespan

Aging is a multi-factorial process, however, it is generally accepted that reactive oxygen species (ROS) are significant contributors. Mitochondria are important players in the aging process because they produce most of the cellular ROS. Despite the strength of the free-radical hypothesis, the use of free radical scavengers to delay aging has generated mixed results in vertebrate models, and clinical evidence of efficacy is lacking. This is in part due to the production of pro-oxidant metabolites by many antioxidants while scavenging ROS, which counteract their potentially beneficial effects. As such, a more effective approach is to enhance mitochondrial metabolism by reducing electron leakage with attendant reduction of ROS generation. Here, we report on the actions of a novel endogenous indole derivative, indolepropionamide (IPAM), which is similar in structure to melatonin. Our results suggest that IPAM binds to the rate-limiting component of oxidative phosphorylation in complex I of the respiratory chain and acts as a stabilizer of energy metabolism, thereby reducing ROS production. IPAM reversed the age-dependent decline of mitochondrial energetic capacity and increased rotifer lifespan, and it may, in fact, constitute a novel endogenous anti-aging substance of physiological importance.     

Mitochondrial bioenergetics in aging

Mitochondria are strongly involved in the production of reactive oxygen species, considered as the pathogenic agent of many diseases and of aging. The mitochondrial theory of aging considers somatic mutations of mitochondrial DNA induced by oxygen radicals as the primary cause of energy decline; experimentally, complex I appears to be mostly affected and to become strongly rate limiting for electron transfer. Mitochondrial bioenergetics is also deranged in human platelets upon aging, as shown by the decreased Pasteur effect (enhancement of lactate production by respiratory chain inhibition). Cells counteract oxidative stress by antioxidants; among lipophilic antioxidants, coenzyme Q is the only one of endogenous biosynthesis. Exogenous coenzyme Q, however, protects cells from oxidative stress by conversion into its reduced antioxidant form by cellular reductases.     

Mitochondrial effectors of cellular senescence: beyond the free radical theory of aging

Cellular senescence is a process that results from a variety of stresses, leading to a state of irreversible growth arrest. Senescent cells accumulate during aging and have been implicated in promoting a variety of age‐related diseases. Mitochondrial stress is an effective inducer of cellular senescence, but the mechanisms by which mitochondria regulate permanent cell growth arrest are largely unexplored. Here, we review some of the mitochondrial signaling pathways that participate in establishing cellular senescence. We discuss the role of mitochondrial reactive oxygen species (ROS), mitochondrial dynamics (fission and fusion), the electron transport chain (ETC), bioenergetic balance, redox state, metabolic signature, and calcium homeostasis in controlling cellular growth arrest. We emphasize that multiple mitochondrial signaling pathways, besides mitochondrial ROS, can induce cellular senescence. Together, these pathways provide a broader perspective for studying the contribution of mitochondrial stress to aging, linking mitochondrial dysfunction and aging through the process of cellular senescence.     

Ubiquinol and a coenzyme Q reducing system protect platelet mitochondrial function of transfusional buffy coats from oxidative stress

The conditions under which Coenzyme Q (CoQ) may protect platelet mitochondrial function of transfusional buffy coats from aging and from induced oxidative stress were investigated. The Pasteur effect, i.e. the enhancement of lactate production after inhibition of mitochondrial respiratory chain, was exploited as a marker of mitochondrial function as it allows to calculate the ratio of mitochondrial ATP to glycolytic ATP. Reduced CoQ 10 improves platelet mitochondrial function of transfusional buffy coats and protects the cells from induced oxidative stress. Oxidized CoQ is usually less effective, despite the presence, shown for the first time in this study, of quinone reductase activities in the platelet plasma membranes. The addition of a CoQ reducing system to platelets is effective in enhancing the protection of platelet mitochondrial function from the oxidative stress. The results support on one hand a possibility of protection of mitochondrial function in aging by exogenous CoQ intake, on the other a possible application in protection of transfusional buffy coats from storage conditions and oxidative deterioration.