Mitochondrial dysfunction in type 2 diabetes leads to oxidative stress, which drives disease progression and diabetes complications. L-carnosine, an endogenous dipeptide, improves metabolic control, wound healing and kidney function in animal models of type 2 diabetes. Coenzyme Q (CoQ), a component of the mitochondrial electron transport chain, possesses similar protective effects on diabetes complications. We aimed to study the effect of carnosine on CoQ, and assess any synergistic effects of carnosine and CoQ on improved mitochondrial function in a mouse model of type 2 diabetes. Carnosine enhanced CoQ gene expression and increased hepatic CoQ biosynthesis in db/db mice, a type 2 diabetes model. Co-administration of Carnosine and CoQ improved mitochondrial function, lowered ROS formation and reduced signs of oxidative stress. Our work suggests that carnosine exerts beneficial effects on hepatic CoQ synthesis and when combined with CoQ, improves mitochondrial function and cellular redox balance in the liver of diabetic mice. (4) Conclusions: L-carnosine has beneficial effects on oxidative stress both alone and in combination with CoQ on hepatic mitochondrial function in an obese type 2 diabetes mouse model.

Hyperglycemia, hyperlipidemia, and adiposity are the main factors that cause inflammation in type 2 diabetes due to excessive ROS production, leading to late complications. To counteract the effects of increased free radical production, we searched for a compound with effective antioxidant properties that can induce coenzyme Q biosynthesis without affecting normal cellular functions. Tocotrienols are members of the vitamin E family, well-known as efficient antioxidants that are more effective than tocopherols. Deh-T3β is a modified form of the naturally occurring tocotrienol-β. The synthesis of this compound involves the sequential modification of geranylgeraniol. In this study, we investigated the effects of this compound in different experimental models of diabetes complications. Deh-T3β was found to possess multifaceted capacities. In addition to enhanced wound healing, deh-T3β improved kidney and liver functions, reduced liver steatosis, and improved heart recovery after ischemia and insulin sensitivity in adipose tissue in a mice model of type 2 diabetes. Deh-T3β exerts these positive effects in several organs of the diabetic mice without reducing the non-fasting blood glucose levels, suggesting that both its antioxidant properties and improvement in mitochondrial function are involved, which are central to reducing diabetes complications.

Diabetes mellitus is characterized by hyperglycemia and capillary hypoxia that causes excessive production of free radicals and impaired antioxidant defense, resulting in oxidative stress and diabetes complications such as impaired wound healing. We have previously shown that modified forms of tocotrienols possess beneficial effects on the biosynthesis of the mevalonate pathway lipids including increase in mitochondrial CoQ. The aim of this study is to investigate the effects of mono-epoxy-tocotrienol-α on in vitro and in vivo wound healing models as well as its effects on mitochondrial function. Gene profiling analysis and gene expression studies on HepG2 cells and human dermal fibroblasts were performed by microarray and qPCR, respectively. In vitro wound healing using human fibroblasts was studied by scratch assay and in vitro angiogenesis using human dermal microvascular endothelial cells was studied by the tube formation assay. In vivo wound healing was performed in the diabetic db/db mouse model. For the study of mitochondrial functions and oxygen consumption rate Seahorse XF-24 was employed. In vitro, significant increase in wound closure and cell migration (p<0.05) both in normal and high glucose and in endothelial tube formation (angiogenesis) (p<0.005) were observed. Microarray profiling analysis showed a 20-fold increase of KIF26A gene expression and 11-fold decrease of lanosterol synthase expression. Expression analysis by qPCR showed significant increase of the growth factors VEGFA and PDGFB. The epoxidated compound induced a significantly higher basal and reserve mitochondrial capacity in both HDF and HepG2 cells. Additionally, in vivo wound healing in db/db mice, demonstrated a small but significant enhancement on wound healing upon local application of the compound compared to treatment with vehicle alone. Mono-epoxy-tocotrienol-α seems to possess beneficial effects on wound healing by increasing the expression of genes involved in cell growth, motility and angiogenes as well as on mitochondrial function.

The possible beneficial effects of coenzyme Q10 (CoQ10) supplementation on disease progression and oxidant status in diabetes remains debated. In the present study, patients with type 1 and type 2 diabetes were treated with oral CoQ10, 100mg twice daily for 12weeks. We assessed total antioxidant capacity, intra- and extracellular levels of the redox regulating protein glutaredoxin 1 (Grx1), CoQ10, oxidized LDL-cholesterol, lipid profile and HbA1c. We have previously shown that extracellular Grx1 is increased in patients with type 2 diabetes compared to healthy subjects. In the present study, CoQ10 treatment significantly decreased serum Grx1 activity as well as total antioxidant capacity independent of type of diabetes, indicating an improvement to a less oxidized extracellular environment. The effect on serum Grx1 activity was more prominent in patients not on statin treatment. Conversely, intracellular Grx1 activity as well as mRNA levels increased independent of statin treatment. There was a significant improvement in oxidized LDL-cholesterol and lipid profile, with a tendency to improved metabolic control (HbA1c). Additionally, we describe for the first time that CoQ10 is a direct substrate for glutathione, and that Grx1 catalyzes this reaction, thus presenting a novel mechanism for CoQ10 reduction which could explain our findings of an increased intracellular Grx1. In conclusion, 12weeks CoQ10 treatment significantly improved the extracellular redox balance and lipid profile, indicating that prolonged treatment may have beneficial effects also on clinical outcome in diabetes.

ObjectiveOur study aimed to explore associations between metabolic control, oxidative stress and coenzyme Q10 (CoQ10) in relation to diabetes complications in a representative population of type 2 diabetes.Research design and methodsA geographic cohort of 156 subjects was recruited. Serum concentrations of CoQ10 and vitamin E were measured by HPLC. ROS was determined by free oxygen radicals testing (FORT). Glutaredoxin (Grx) activity, oxidized LDL cholesterol (oxLDLc), high sensitive CRP (hsCRP), HbA1c, urine albumin, serum creatinine, serum cystatin C, and plasma lipids were assayed with routine laboratory protocols.ResultsSerum CoQ10 was higher than in nondiabetics. HbA1c, fP-glucose, hyperlipidemia, inflammation (hsCRP), and increased BMI were associated with signs of oxidative stress as increased levels of FORT, Grx activity and/or increased levels of oxLDLc Oxidative stress was found to be strongly correlated with prevalence of cardiovascular disease (CVD) and peripheral sensory neuropathy (PSN). In both gender groups there were positive correlations between CoQ10 and oxLDLc, and between BMI and the ratio CoQ10/chol. Grx activity was inversely correlated to oxLDLc and CoQ10. Women with CVD and PSN had higher waist index, oxLDLc, and FORT levels compared to men but lower CoQ10 levels. Men had worse kidney function and lower vitamin E. Multiple regression analysis showed increased levels of CoQ10 to be significantly correlated with increased levels of cholesterol, triglycerides, vitamin E, fB-glucose and BMI.ConclusionsHyperlipidemia, hyperglycemia and inflammation were associated with oxidative stress, which was correlated to the prevalence of diabetes complications. CoQ10 was increased in response to oxidative stress. There were gender differences in the risk factors associated with diabetes complications.

2,3-Oxidosqualene is an intermediate in cholesterol biosynthesis and 2,3:22,23-dioxidosqualene act as the substrate for an alternative pathway that produces 24(S),25-epoxycholesterol which effects cholesterol homeostasis. In light of our previous findings concerning the biological effects of certain epoxidated all-trans-polyisoprenes, the effects of squalene carrying epoxy moieties on the second and third isoprene residues were investigated here. In cultures of HepG2 cells both monoepoxides of squalene and one of their hydrolytic products inhibited cholesterol synthesis and stimulated the synthesis of coenzyme Q (CoQ). Upon prolonged treatment the cholesterol content of these cells and its labeling with [(3)H]mevalonate were reduced, while the amount and labeling of CoQ increased. Injection of the squalene monoepoxides into mice once daily for 6days elevated the level of CoQ in their blood, but did not change the cholesterol level. The same effects were observed upon treatment of apoE-deficient mice and diabetic GK-rats. This treatment increased the hepatic level of CoQ10 in mice, but the amount of CoQ9, which is the major form, was unaffected. The presence of the active compounds in the blood was supported by the finding that cholesterol synthesis in the white blood cells was inhibited. Since the ratio of CoQ9/CoQ10 varies depending on the experimental conditions, the cells were titrated with substrate and inhibitors, leading to the conclusion that the intracellular isopentenyl-PP pool is a regulator of this ratio. Our present findings indicate that oxidosqualenes may be useful for stimulating both the synthesis and level of CoQ both in vitro and in vivo.

Exposure of the general population to polycyclic aromatic hydrocarbons (PAH) is ubiquitous. The aim of this study was to analyze biomarkers associated with the uptake of PAH in 428 non-smoking women from Łodz (Poland), Viterbo (Italy), Belgrade (Serbia) and from the Pančevo area, where the petrochemical complex was destroyed by the air raids in 1999. Urinary excretion of PAH metabolites was lowest in Italian women, intermediary for Serbian and highest in Polish women, who predominantly excreted hydroxy phenanthrenes as metabolites of phenanthrene. Bulky DNA adduct levels were highest in Italian and Polish women. Genotype or PAH ambient air levels could not explain the dissimilarities between the study groups with respect to biomarker patterns, which probably reflected differences in life style-associated factors.

The organ content of the mevalonate pathway lipids was investigated in liver-X-receptor (LXR) α, β and double knock-out mice. An extensive or moderate increase of total cholesterol in the double KO mice was found in all organs elicited by the increase of the esterified form. In LXRα and double KO mice, coenzyme Q (CoQ) was decreased in liver and increased in spleen, thymus and lung, while dolichol was increased in all organs investigated. This effect was confirmed using LXR- agonist GW 3965. Analysis of CoQ distribution in organelles showed that the modifications are present in all cellular compartments and that the increase of the lipid in mitochondria was the result of a net increase of CoQ without changing the number of mitochondria. It appears that LXR influences not only cellular cholesterol homeostasis but also the metabolism of CoQ and dolichol, in an indirect manner.

Skeletal muscle regeneration relies on the pool of satellite cells that in adult muscle reside between sarcolemma and basal lamina surrounding muscle fiber. Once skeletal muscle becomes injured these cells are activated, re-enter cell cycle, fuse into multinucleate myotubes, and finally reconstruct the injured tissue. However, some pathological conditions, such as reoccurring skeletal muscle injuries/regeneration accompanying muscular dystrophies, can lead to the irreversible exhaustion of satellite cells and to the failure in muscle repair. Mobilization of endogenous or implantation of exogenous stem cells are considered as the therapeutic approaches aiming at the improvement of regeneration. In order to be used participate in tissue repair exogenous cells need to be able to undergo myogenic differentiation, to form myotubes, and then myofibres. Among the cells extensively tested as a candidates for “replacement” cells are embryonic stem and induced pluripotent stem cells, and also variety of multipotent stem cells isolated from fetal or adult tissues. The simplest and non-offensive, however, the least efficient means of the induction of stem cells differentiation relies on their exposure to the myogenic environment, i.e. in vitro co-culture with myoblasts, or transplantation into injured skeletal muscle. Other techniques inducing myogenic differentiation base on the treatment of these cells either with various growth factors, or with reagents impacting at the overall gene expression leading to the induction of myogenic program. The most efficient way to obtain myoblasts from stem cells relies, however, on the overexpression of crucial factors regulating myogenesis, e.g. Pax3, Pax7, MyoD. Regardless of the system used detailed molecular, cellular, and functional analyses are required to prove that the stem cells differentiation was efficient and proper, and thus, that these cells can be possibly considered as suitable for the therapy of injured or diseased muscles. L17.2

In addition to its role as a component of the mitochondrial respiratory chain and our only lipid-soluble antioxidant synthesized endogenously, in recent years coenzyme Q (CoQ) has been found to have an increasing number of other important functions required for normal metabolic processes. A number of genetic mutations that reduce CoQ biosynthesis are associated with serious functional disturbances that can be eliminated by dietary administration of this lipid, making CoQ deficiencies the only mitochondrial diseases which can be successfully treated at present. In connection with certain other diseases associated with excessive oxidative stress, the level of CoQ is elevated as a protective response. Aging, certain experimental conditions and several human diseases reduce this level, resulting in serious metabolic disturbances. Since dietary uptake of this lipid is limited, up-regulation of its biosynthetic pathway is of considerable clinical interest. One approach for this purpose is administration of epoxidated all-trans polyisoprenoids, which enhance both CoQ biosynthesis and levels in experimental systems.

Exposure to environmental contaminants such as polycyclic aromatic hydrocarbons (PAHs), life style and nutritional status of a population are important factors that may influence normal serum levels of antioxidants and the insulin-like growth factor system. In this study we examined serum levels of insulin-like growth factor-I (IGF-I), insulin-like growth factor binding protein-1 (IGFBP-1), coenzyme Q10 (CoQ) and vitamin E in healthy female populations (n = 4 x 100) aged 19-59 years from Poland (PL), Sweden (SE), Serbia I (SR I) and Serbia II (SR II). The last group lived in an environmental emergency area affected by the bombings of 1999 in Serbia.The Polish and SR I cohorts exhibited low IGFSD-score levels, (-2 to +/-0), compared to females from SE with IGFSD-score 0. In the SIR II population, the IGFSD range was between -1 and 1. The IGFBP-1 levels of the Polish and SIR I groups were lower than in the Swedish population, while the SR II levels showed a broader distribution, 20-80 mu g/l. The CoQ values in the Swedish and Polish samples were around 1 nmol/ml. In contrast. the SIR I cohorts exhibited higher concentrations, 1.5-3.5 nmol/ml and the SIR II group had extremely low levels, <0.5 nmol/ml. The vitamin E concentrations were similar in the Polish and Swedish populations, 20-40 nmol/ml, while it was twice as high, 40-80 nmol/ml in the SR I and very low in the SIR II group, which is half of the Polish and Swedish cohorts.These results suggest that different lifestyles and environmental factors affect both the IGF system and the antioxidants CoQ10 and vitamin E in female populations in Europe. The females living in the polluted area had different patterns of both the IGF and antioxidant systems. These findings may explain differences in morbidity and mortality in these countries. (C) 2009 Elsevier Ltd. All rights reserved.

Uptake of dietary coenzyme Q (CoQ) into organs is limited but there are some exceptions such as adrenal glands and ovaries. Under deficient conditions an optimal solution could be stimulation of the endogenous synthesis. In rodent exercise, cold exposure and a few substances elevate the CoQ levels to some extent. Investigations of the nuclear receptors PPARα, RXRα and LXRα&β did not answer the question which nuclear receptor regulates CoQ biosynthesis and at present we cannot design a ligand for upregulation of the synthesis. Upon ultraviolet irradiation of CoQ a number of products are formed which influence the synthesis of the mevalonate pathway lipids. Among them epoxidated derivatives were identified. Upon chemical epoxidation of a series of polyisoprenoids it was found that none of the tested poly‐cis polyisoprenols had any effect but some of the all‐trans polyisoprenols stimulated CoQ synthesis and in some cases also inhibited cholesterol biosynthesis. Tocotrienol epoxides were proved to be very efficient, those having one epoxide in the side chain doubled or trebled the CoQ synthesis while those with two epoxides additionally also inhibited cholesterol synthesis by 50–90%. The elevation of CoQ synthesis was elicited by increased mRNA levels for biosynthetic enzymes while the inhibition point in the cholesterol synthesis was localized to oxidosqualene cyclase.

In our search for compounds that up-regulate the biosynthesis of coenzyme Q (CoQ), we discovered that irradiation of CoQ with ultraviolet light results in the formation of a number of compounds that influence the synthesis of mevalonate pathway lipids by HepG2 cells. Among the compounds that potently stimulated CoQ synthesis while inhibiting cholesterol synthesis, derivatives of CoQ containing 1-4 epoxide moieties in their polyisoprenoid side chains were identified. Subsequently, chemical epoxidation of all-trans-polyprenols of different lengths revealed that the shorter farnesol and geranylgeraniol derivatives were without effect, whereas the longer derivatives of solanesol enhanced CoQ and markedly reduced cholesterol biosynthesis. In contrast, none of the modified trans-trans-poly-cis-polyprenols exerted noticeable effects. Tocotrienol epoxides were especially potent in our system; those with one epoxide moiety in the side-chain generally up-regulated CoQ biosynthesis by 200-300%, whereas those with two such moieties also decreased cholesterol synthesis by 50-90%. Prolonged treatment of HepG2 cells with tocotrienol epoxides for 26 days elevated their content of CoQ by 30%. In addition, the levels of mRNA encoding enzymes involved in CoQ biosynthesis were also elevated by the tocotrienol epoxides. The site of inhibition of cholesterol synthesis was shown to be oxidosqualene cyclase. In conclusion, epoxide derivatives of certain all-trans-polyisoprenoids cause pronounced stimulation of CoQ synthesis and, in some cases, simultaneous reduction of cholesterol biosynthesis by HepG2 cells.

Cationic linear poly-cis-isoprenoid prepared from natural plant polyprenol in a mixture with dioleyl phosphatidylethanolamine was found to be an effective lipofection agent for eukaryotic cells. The transfecting activity is related to the poly-cis structure of the polyprenyl chain.

Coenzyme Q (CoQ) deficiency occurs in genetic disorders, during aging and various diseases. Diagnosis requires skin fibroblasts in tissue culture. [3H]Mevalonate incorporation was appropriate to measure the rate of CoQ synthesis in fibroblasts and hepatoblastoma cells. [14C]p-Hydroxybenzoate had limited permeability, but it could be increased with Fugene and cyclodextrin. Inhibition of decaprenyl-4-hydroxybenzoate transferase results in the accumulation of decaprenyl diphosphate, an indicator of enzyme deficiency. Also, analysis of the corresponding mRNAs in this case is useful. In vitro assays to measure trans-prenyltransferase and decaprenyl-4-hydroxybenzoate transferase activities are not available. Neither measurement of methyltransferases is reliable in human cells. In vitro reconstruction of CoQ synthesis, in opposite to cholesterol synthesis, proved to be unsuccessful. Thus, the biochemical characterization of the CoQ biosynthetic system in human cells is restricted to a few reliable analytical procedures.