Atherosclerosis 216 (2011) Contents lists available at ScienceDirect. Atherosclerosis

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1 Atherosclerosis 216 (2011) Contents lists available at ScienceDirect Atherosclerosis journal homepage: Reversal of mitochondrial dysfunction by coenzyme Q10 supplement improves endothelial function in patients with ischaemic left ventricular systolic dysfunction: A randomized controlled trial Yuk-Ling Dai a, Ting-Hin Luk a, Kai-Hang Yiu a, Mei Wang a, Pandora M.C. Yip a, Stephen W.L. Lee a, Sheung-Wai Li c, Sidney Tam d, Bonnie Fong d, Chu-Pak Lau a, Chung-Wah Siu a,b, Hung-Fat Tse a,b, a Cardiology Division, Department of Medicine, The University of Hong Kong, Hong Kong b Research Centre of Heart, Brain, Hormone and Healthy Aging, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong c Department of Medicine, Tung Wah Hospital, Hong Kong d Department of Clinical Biochemistry, Queen Mary Hospital, Hong Kong, Hong Kong SAR, China article info abstract Article history: Received 25 November 2010 Received in revised form 3 February 2011 Accepted 4 February 2011 Available online 17 February 2011 Keywords: Mitochondrial function Co-enzyme Q10 Heart failure Aims: Coronary artery disease (CAD) is associated with endothelial dysfunction and mitochondrial dysfunction (MD). The aim of this study was to investigate whether co-enzyme Q10 (CoQ) supplementation, which is an obligatory coenzyme in the mitochondrial respiratory transport chain, can reverse MD and improve endothelial function in patients with ischaemic left ventricular systolic dysfunction (LVSD). Methods and results: We performed a randomized, double-blind, placebo-controlled trial to determine the effects of CoQ supplement (300 mg/day, n = 28) vs. placebo (controls, n = 28) for 8 weeks on brachial flow-mediated dilation (FMD) in patients with ischaemic LVSD(left ventricular ejection fraction <45%). Mitochondrial function was determined by plasma lactate/pyruvate ratio (LP ratio). After 8 weeks, CoQtreated patients had significant increases in plasma CoQ concentration (treatment effect 2.20 g/ml, P < 0.001) and FMD (treatment effect 1.51%, P = 0.03); and decrease in LP ratio (treatment effect 2.46, P = 0.03) compared with controls. However, CoQ treatment did not alter nitroglycerin-mediated dilation, blood pressure, blood levels of fasting glucose, haemoglobin A1c, lipid profile, high-sensitivity C-reactive protein and oxidative stress as determined by serum superoxide dismutase and 8-isoprostane (all P > 0.05). Furthermore, the reduction in LP ratio significantly correlated with improvement in FMD (r = 0.29, P = 0.047). Conclusion: In patients with ischaemic LVSD, 8 weeks supplement of CoQ improved mitochondrial function and FMD; and the improvement of FMD correlated with the change in mitochondrial function, suggesting that CoQ improved endothelial function via reversal of mitochondrial dysfunction in patients with ischaemic LVSD Elsevier Ireland Ltd. All rights reserved. 1. Introduction Co-enzyme Q10 is a potent antioxidant and an obligatory coenzyme for mitochondrial enzyme complexes in oxidative phosphorylation for the production of adenosine triphosphate (ATP). It has been shown to improve brachial artery flow-mediated dilation (FMD) in patients with coronary artery disease [1], heart failure [2] and type 2 diabetes mellitus [3]. Brachial FMD is an established marker of endothelial dysfunction, which is recognized as a precursor of atherosclerosis and an independent predictor of Corresponding author at: Cardiology Division, Department of Medicine, The University of Hong Kong, Rm 1928, Block K, Queen Mary Hospital, Hong Kong. Tel.: ; fax: address: hftse@hkucc.hku.hk (H.-F. Tse). cardiovascular events [4]. However, the precise mechanism that mediates the vasoprotective effects of co-enzyme Q10 on endothelial function remains unclear. Previous clinical studies have demonstrated the important role of co-enzyme Q10 on mitochondrial bioenergetics in patients with cardiovascular diseases [5]. Co-enzyme Q10 supplementation in patients with cardiac diseases increased myocardial tissue mitochondrial co-enzyme Q10 levels and improved mitochondrial function and in vitro contractility of myocardial tissue [6]. Furthermore, mitochondrial dysfunction has also been implicated in the pathogenesis of atherosclerosis [7]. Cardiovascular risk factors such as aging, hyperglycemia [8] and dyslipidemia [9] induce mitochondrial dysfunction and increase production of reactive oxygen species, which can lead to endothelial dysfunction [7,10]. Our recent cross-sectional study has demonstrated the association /$ see front matter 2011 Elsevier Ireland Ltd. All rights reserved. doi: /j.atherosclerosis

2 396 Y.-L. Dai et al. / Atherosclerosis 216 (2011) between endothelial dysfunction and mitochondrial dysfunction in patients with coronary artery disease [11]. Nevertheless, it remains unclear whether reversal of mitochondrial dysfunction in patients with cardiovascular diseases can improve endothelial function. The aim of this study was to investigate whether co-enzyme Q10 supplementation can reverse mitochondrial dysfunction and improve endothelial function in patients with ischaemic left ventricular systolic dysfunction. 2. Methods 2.1. Study population A total of 56 consecutive patients with ischaemic left ventricular systolic dysfunction were recruited from the Queen Mary Hospital cardiac specialist clinic during the period November 2007 to April All patients had stable CAD, evidenced by >50% stenosis in at least 1 of the major coronary arteries as documented by coronary angiogram and left ventricular ejection fraction <45% as documented by echocardiogram. All patients received stable medications for at least 1-year before enrollment and medications were not changed during the study. Patients with recent acute coronary syndrome or percutaneous coronary intervention in the past 3 months, significant valvular heart disease, significant renal or hepatic dysfunction, or those who were on habitual antioxidant supplementation (Vitamin C, E, A, or co-enzyme Q10) were excluded from the study. The research protocol was approved by the local institutional review board, and all subjects provided written informed consent. This study complied with the Declaration of Helsinki and was registered at number HKCTR Study design and materials A randomized, double-blind, placebo controlled trial was conducted and the study design is shown in Fig. 1. Initially, 70 eligible patients as determined from medical records were interviewed by phone for recruitment into the study. Among them, 62 patients verbally consented to participate in the study. During the initial assessment, 6 patients were deemed ineligible for the exclusion criteria. The remaining 56 enrolled patients were randomized to receive either oral co-enzyme Q10 supplementation (100 mg 3 times per day, i.e., 300 mg per day, Q-absorb, Jarrow Formulas, Los Angeles, CA, USA) or matched placebo for a period of 8 weeks according to a computer generated sequence. Specifically, an independent nurse received the identity list of recruited patients from the investigators and was requested to assign each patient into one of the intervention groups by a randomly generated number (0 or 1) from computer. Block randomizations of n = 20 were used throughout the drug dispensing procedure: 28 patients were allocated into the intervention group and 28 into the placebo group. The medications were indistinguishable in packing and were dispensed by the nurse independent of the investigators, such that the allocation of patients to the intervention or placebo group was concealed to the patients, to the personnel conducting the clinical tests, and to the laboratory people involved in the analysis. The codes were kept by the nurse who dispensed the capsules and were broken when the trial was over and patients had completed the tests. At baseline and the final follow-up at week 8, all patients underwent vascular ultrasound measurement and blood samples were taken to measure fasting glucose, glycosylated haemoglobin A1c (HbA1c), lipid profile and high-sensitivity C-reactive protein (hs-crp). Body anthropometry and systolic and diastolic blood pressures were also documented. For all the medications, including co-enzyme Q10 and metformin, the last dose was taken the evening before the final assessment at week Demographic, dietary and laboratory evaluations Baseline demographic data and cardiovascular medications were documented in all subjects. Cardiovascular risk factors, including tobacco smoking, diabetes mellitus, hypercholesterolemia, hypertension, and family history of cardiovascular diseases diagnosed in first-degree relatives before 55 years of age were assessed. The body height and weight, blood pressure and body mass index (BMI) of all subjects were measured as previously described [12]. Hypertension was defined as either systolic or diastolic blood pressure 140/90 mmhg at 2 different clinical visits or the use of medications. Diabetes mellitus was defined as fasting serum glucose of 7.0 mmol/l or on medications. Hypercholesterolemia was defined as a fasting total serum cholesterol level of 4.9 mmol/l or on medications. Smoking status was recorded as either smoker (past or current) or non-smoker. Fasting blood samples were obtained from all subjects to determine serum creatinine, glucose and lipid levels, including total cholesterol, low-density lipoprotein cholesterol, highdensity lipoprotein cholesterol, triglyceride, apolipoprotein A-I, apolipoprotein B and lipoprotein (a). Plasma hs-crp level was measured by using a Hitachi 747 analyzer (Boehringer Mannheim, Mannheim, Germany), and a particle-enhanced immunoturbidimetric assay (Roche Diagnostics, Mannheim, Germany). Furthermore, venous blood samples of lactate and pyruvate were collected by non-tourniquet sampling and immediately added to fluoride oxalate medium and a titrated volume of chilled perchlorate, respectively. The plasma lactate/pyruvate ratio is a good indicator of impaired oxidative phosphorylation in the mitochondria, as excessive hydrolysis of ATP produces more protons that drive the equilibrium reaction from pyruvate to lactate; leading to a raised lactate/pyruvate ratio. An increased lactate/pyruvate ratio has been widely used clinically as a non-invasive reflection of mitochondrial dysfunction [11,13 15]. After thorough mixing, the samples were then centrifuged at 3000 rpm for 20 min and the supernatants snap-frozen at 80 C until assay. Oxidative stress was measured as superoxide dismutase (SOD), an important antioxidant enzyme and 8-isoprostane, a marker of lipid peroxidation [16]. Lactate was measured by an enzyme-coupled colorimetric assay automated on Hitachi-912 analyzer (Roche Diagnostics, GmbH, Mannheim, Germany). Pyruvate was measured by reverse phase high performance liquid chromatography (Waters Corp, Milford, Massachusetts, USA) using a C18 column with isocratic elution. To determine the total plasma concentration of co-enzyme Q10, high performance liquid chromatography was performed with Alltima (Grace Davison, Illinois) reverse phase C18 column (5 m, 150 mm 4.6 mm). Mobile phase was constituted by methanol:ethanol 3:7 (v/v). Flow rate was 1.5 ml/min and UV detection was carried out at 275 nm, as previously described [17,18] Brachial artery FMD Vascular ultrasound was performed with a high-resolution ultrasound system (Agilent Sonos 5500; Philips, Andover, MA) with the use of a 7.5-MHz linear array transducer by a single experienced operator without knowledge of the subjects [11,16]. All of the scanned images were stored digitally and analyzed offline by the same operator, who was blinded to the identity of studied subjects. All subjects were studied in the fasting state, and all vasoactive medications were withheld for 12 h before the study. As previously described [19], longitudinal scans of the brachial artery were obtained at rest, and then brachial FMD was induced by inflation of a pneumatic tourniquet placed on the forearm to a pressure of 250 mmhg for 5 min. The diameter of the brachial artery was allowed to return to baseline. Finally, the diameter of the brachial

3 Y.-L. Dai et al. / Atherosclerosis 216 (2011) Assessed for eligibility Interviewed by phone (n=70) Enrollment (n=56) Excluded (n=14): Refused to participate (n=8) Excluded during initial assessment (n=6) Randomized Coenzyme Q supplementation ti Control Group Lost to follow-up (n=0) Lost to follow-up (n=1) Analyzed Analyzed Fig. 1. Study flowchart. artery was obtained 5 min after administration of 400 g sublingual nitroglycerin spray. FMD was calculated as the percentage change in diameter from baseline to the diameter at 60 s of reactive hyperemia. Nitroglycerin-mediated dilatation (NMD) was defined as the percentage change in the brachial artery diameter by 4 min after nitrate administration. An investigation of 25 patients revealed an intra-observer variability correlation coefficient of 0.90 (P < 0.001) Statistical analysis The primary endpoint of this study was pre-defined as the change in FMD from baseline. Secondary endpoints were changes in endothelium-independent NMD, systolic and diastolic blood pressure, plasma co-enzyme Q10 level, lactate/pyruvate ratio, glycemic control, lipid profile, hs-crp and markers of oxidative stress such as serum superoxide dismutase and 8-isoprostane. Based on the data from our cross-sectional study on a similar patient population [11], we calculated the sample size using a standard deviation of 2.5% for the change of FMD at 8 weeks from baseline. For 1:1 randomization into co-enzyme Q10 and control group we would need 25 patients in each arm, assuming an 80% power to detect an absolute change of 2% in FMD between the treatment and control group with a 5% maximum false-positive rate. Assuming a 5% dropout rate, at least 54 patients would be required. Continuous variables were expressed as mean ± standard deviation (SD). Categorical data were presented as frequencies and percentages. Statistical comparisons were performed with Student s t-test for continuous variables or Pearson s Chi-squared test for categorical variables. Paired t-test was used to compare the co-enzyme Q10 concentration before and after 8 weeks in both intervention and control groups. Bivariate correlations were studied using Pearson s correlation coefficient. All analyses were performed according to intention-to-treat principle, with the missing measurements of patients having defaulted the followup replaced by the baseline values. The analysis of covariance (ANCOVA) was used to adjust for the baseline values in comparing the means of outcome variables, i.e., the follow-up measurement was taken as the dependent variable with group allocation (coenzyme Q10 vs. control) as independent variable correcting for the baseline value. Absolute changes and 95% confidence intervals (CI) of outcome variables were calculated using univariate and multivariate linear regression to determine the effect of increased co-enzyme Q10 levels after treatment. All statistical analyses were performed using the SPSS program (version 15.0). A two-sided P value of <0.05 was considered statistically significant. 3. Results 3.1. Baseline characteristics The baseline characteristics of the study population are shown in Table 1. The overall mean age was 68.9 ± 9.6 years, and 93% were men. Among these patients, 54% had hypertension, 61% had diabetes mellitus, 50% had hypercholesterolemia and 70% had prior history of myocardial infarction. They were treated optimally with medications; with 80% of patients treated with beta-blockers, 90% with angiotensin converting enzyme inhibitors or angiotensin receptor blockers, 95% with statins and 96% with aspirin. There were no significant differences in the baseline characteristics between patients randomized to co-enzyme Q10 treatment or placebo (Table 1, all P > 0.05). Of the 56 patients randomized in the trial, 55 patients completed the final follow-up after 8 weeks of intervention, and 1 patient in the placebo group denied follow-up. The capsules were well tolerated by patients and no adverse effects were reported. Co-enzyme Q10 supplementation resulted in a threefold increase in plasma co-enzyme Q10 level from baseline in the treatment group (from 1.08 to 3.24 g/ml, P < 0.001) while no significant changes were observed in the control group (from 0.95 to 0.98 g/ml, P = 0.42).

4 398 Y.-L. Dai et al. / Atherosclerosis 216 (2011) Table 1 Baseline clinical characteristics of study population. Control (n = 28) Co-enzyme Q10 (n = 28) P value Age, years 70.1 ± ± Males, n (%) 25 (89) 27(96) 0.30 Smoking (past/current), n (%) 17(61) 19(70) 0.45 Systolic BP, mmhg 136 ± ± Diastolic BP, mmhg 81 ± 9 81 ± BMI, kg/m ± ± Hypercholesterolemia, n (%) 11(39) 17(61) 0.11 Hypertension, n (%) 17(61) 13(46) 0.28 Diabetes mellitus, n (%) 10(36) 12(43) 0.58 Fasting blood glucose, mmol/l 6.25 ± ± HbA1c, % 6.82 ± ± Total cholesterol, mmol/l 4.10 ± ± HDL cholesterol, mmol/l 1.21 ± ± LDL cholesterol, mmol/l 2.26 ± ± Triglyceride, mmol/l 1.37 ± ± Apo A1, g/l 1.26 ± ± Apo B, g/l 0.72 ± ± Lp(a), mg/l 228 ± ± hs-crp, mg/l 1.48 ± ± Superoxide dismutase (U/ml) ± ± Isoprostane (pg/ml) 508 ± ± History of MI, n (%) 19(68) 20(71) 0.77 History of revascularization, n (%) 21(75) 22(79) 0.75 PCI, n (%) 18(69) 18(67) 0.84 CABG, n (%) 4(19) 6(33) 0.31 Aspirin, n (%) 27(96) 27(96) 0.99 Statins, n (%) 26(93) 27(96) 0.55 Beta-blockers, n (%) 23(82) 22(79) 0.74 Calcium channel blockers, n (%) 4(14) 4(14) 0.99 ACEI/ARB, n (%) 27(96) 25(89) 0.30 Nitrates, n (%) 11(41) 10(36) 0.70 Co-enzyme Q10 concentration, g/ml 0.95 ± ± Lactate/pyruvate ratio 17.4 ± ± FMD, % 3.68 ± ± NMD, % ± ± Left ventricular ejection fraction, % ± ± Abbreviations: ACEI, angiotensin converting enzyme inhibitor; ARB, angiotensin receptor blocker; Apo, Apolipoprotein; BMI, body mass index; BP, blood pressure; CABG, coronary artery bypass grafting; FMD, flow-mediated dilatation; hs-crp, high-sensitivity C-reactive protein; HbA1c, glycosylated haemoglobin A1c; HDL, high-density lipoprotein; LDL, low-density lipoprotein; Lp, lipoprotein; MI, myocardial infarction; NMD, nitroglycerin-mediated dilatation; PCI, percutaneous coronary intervention Effect of co-enzyme Q10 supplementation on vascular function At 8 weeks, co-enzyme Q10 supplementation significantly improved brachial FMD (treatment effect 1.51%, 95% CI , P = 0.03, Table 2) without any difference in endotheliumindependent NMD (Table 2, P > 0.05). The absolute increase in plasma co-enzyme Q10 level significantly correlated with an improvement in FMD (r = 0.35, P = 0.01, Fig. 2). After adjusting for baseline differences in FMD, for every unit ( g/ml) increase in plasma co-enzyme Q10, there was an absolute 0.57% increase in FMD compared with baseline (95% CI , P = 0.007). In addition, the absolute treatment effect of co-enzyme Q10 on brachial FMD was inversely related to baseline FMD (r = 0.51, P < 0.001), suggesting that the beneficial effect of co-enzyme Q10 supplementation was more pronounced in patients with more severe endothelial dysfunction Effect of co-enzyme Q10 supplementation on plasma lactate/pyruvate ratio At baseline, the increase in plasma lactate/pyruvate ratio significantly correlated with impaired FMD in the overall population (r = 0.41, P < 0.001), suggesting that impaired mitochondrial function was associated with endothelial dysfunction. At 8 weeks, co-enzyme Q10 supplementation significantly lowered plasma lactate/pyruvate ratio (treatment effect 2.46, 95% CI 4.71 to 0.20, P = 0.03, Table 2). The absolute treatment effect of co-enzyme Q10 on lactate/pyruvate ratio was inversely related to baseline lactate/pyruvate ratio (r = 0.53, P < 0.001), suggesting that the beneficial effect of co-enzyme Q10 was also more pronounced in patients with more severe mitochondrial dysfunction. In addition, the reduction in plasma lactate/pyruvate ratio significantly correlated with an improvement in FMD (r = 0.29, P = 0.047, Fig. 3), indicating that the improvement in endothelial function was associated with improvement in mitochondrial function. Although co-enzyme Q10 supplementation lowered lactate/pyruvate ratio, the increase in plasma co-enzyme Q10 levels did not correlate with changes in lactate/pyruvate ratio (r = 0.08, P = 0.57) Subgroup analyses Subgroup analyses were performed to determine the treatment effects of co-enzyme Q10 therapy in different patient subgroups. The therapeutic effect of co-enzyme Q10 on FMD was observed in patients with history of prior myocardial infarction (treatment effect 1.73%, 95% CI , P = 0.045, n = 39), but not in those without (treatment effect 0.86%, 95% CI 1.90 to 3.62, P = 0.52, n = 17). Furthermore, the treatment effect of co-enzyme Q10 on FMD was more prominent in the subset of patients with mitochondrial dysfunction, defined as baseline lactate/pyruvate ratio above median value of 16 (treatment effect 2.03%, 95% CI , P = 0.04, n = 28) compared with those without (treatment effect 1.26%, 95%CI 1.07 to 3.59, P = 0.28, n = 28). There were no differences in treatment effect in diabetics vs. non-diabetics (P > 0.05) or smokers vs. non-smokers (P > 0.05).

5 Y.-L. Dai et al. / Atherosclerosis 216 (2011) Table 2 Treatment effect of 8-week co-enzyme Q10 supplementation. Control (n = 28) Co-enzyme Q10 (n = 28) Treatment effect (ANCOVA) P value * Change (SD) Change (SD) Difference (95% CI) Co-enzyme Q10, g/ml 0.03 (0.19) 2.23 (1.88) (1.45, 2.95) <0.001 Lactate/pyruvate ratio 0.33 (6.71) 0.87 (3.25) 2.46 ( 4.71, 0.20) 0.03 Brachial FMD, % 0.65 (2.79) 1.62 (4.35) 1.51 (0.14, 2.88) 0.03 Brachial NMD, % 1.39 (4.66) 1.25 (5.94) 0.42 ( 2.24, 3.07) 0.75 Mean SBP, mmhg 7 (12) 3 (12) 4.26 ( 1.74, 10.26) 0.16 Mean DBP, mmhg 4 (9) 0 (8) 3.50 ( 0.53, 7.54) 0.09 BMI, kg/m (0.75) 0.05 (0.50) 0.16 ( 0.18, 0.50) 0.36 Total cholesterol, mmol/l 0.06 (0.64) 0.02 (0.73) 0.04 ( 0.37, 0.30) 0.82 HDL cholesterol, mmol/l 0.03 (0.13) 0.02 (0.18) 0.02 ( 0.10, 0.06) 0.64 LDL cholesterol, mmol/l 0.09 (0.56) 0.04 (0.56) 0.08 ( 0.38, 0.22) 0.58 Triglyceride, mmol/l 0.12 (0.50) 0.10 (0.52) 0.01 ( 0.29, 0.27) 0.93 ApoA1, g/l 0.05 (0.11) 0.05 (0.14) 0.02 ( 0.08, 0.05) 0.57 ApoB, g/l 0.03 (0.15) 0.01 (0.16) 0.02 ( 0.10, 0.06) 0.61 Lp(a), mg/l 16 (65) 27 (98) 6 ( 35, 48) 0.76 Fasting blood glucose, mmol/l 0.47 (2.01) 0.35 (1.26) 0.12 ( 1.00, 0.76) 0.78 HbA1c, % 0.06 (0.97) 0.13 (0.48) 0.12 (0.45, 0.20) 0.45 hs-crp, mg/l 0.24 (2.43) 1.21 (9.30) 1.15 ( 2.62, 4.92) Isoprostane, pg/ml 57 (352) 175 (391) 104 ( 227, 18) 0.09 Superoxide dismutase, U/ml (0.031) (0.031) ( 0.008, 0.021) 0.38 Abbreviations as in Table 1. * Treatment effects expressed as mean differences (95% CI) of outcome variables at follow-up using analysis of covariance (ANCOVA), with the follow-up measurement taken as the dependent variable and group allocation (Co-enzyme Q10 vs. control) as independent variable correcting for the baseline value. The changes within treatment group were expressed as mean (SD) Effect of co-enzyme Q10 supplementation on other outcome variables Nevertheless, co-enzyme Q10 supplementation for 8 weeks did not have significant effects on NMD, systolic and diastolic blood pressures, and serum levels of fasting lipid profiles, blood glucose, HbA1c and hs-crp or oxidative stress as determined by serum superoxide dismutase and 8-isoprostane (ANCOVA: all P > 0.05, Table 2). 4. Discussion The results of this study demonstrate that 8 weeks of co-enzyme Q10 supplementation at a dosage of 300 mg/day improved vascular endothelial function and mitochondrial function in patients with ischaemic left ventricular systolic dysfunction. Co-enzyme Q10 supplementation in this study resulted in a threefold increase in plasma co-enzyme Q10 level from baseline in the treatment group, which was comparable to other studies in similar patient groups [20]. Furthermore, the reduction in plasma lactate/pyruvate ratio with co-enzyme Q10 supplementation significantly correlated with the increase in FMD, suggesting that the improvement in endothelial function was mediated through improvement in mitochondrial function after co-enzyme Q10 supplementation. The effect of co-enzyme Q10 supplementation has been widely studied in patients with cardiovascular risk factors. Systematic review and meta-analysis have shown that co-enzyme Q10 has a well documented role as an adjunctive treatment in chronic heart failure, leading to improvement in both clinical and haemodynamic parameters on echocardiography [21,22]. Co-enzyme Q10 has been shown to improve brachial artery FMD in patients with coronary artery disease [1], heart failure [2], and diabetes mellitus [3]. Consistent with these findings, this randomized controlled study also confirmed that co-enzyme Q10 supplementation improved endothelial function in patients with ischaemic heart failure. Furthermore, the improvement in FMD correlated with the increase in plasma co-enzyme Q10 concentration, suggesting a possible dose response relationship as reported previously [1]. The absolute increase in FMD of 1.51% after co-enzyme Q10 supplementation is consistent with previous studies [3,23] and is of clinically significance, as an absolute improvement in FMD of 1% may already translate into a 10 25% reduction in residual cardiovascular risk for these patients [24,25]. The precise mechanism of the vasoprotective effects of coenzyme Q10 is unknown. It has been postulated that co-enzyme Q10 mediates its potential beneficial effects through its bioenergetic properties as an obligatory coenzyme, or through its potent antioxidant properties [5,7]. Tiano et al. [1] showed that coenzyme Q10 supplementation in patients with coronary artery Fig. 2. Relationship between change in brachial flow mediated dilatation (FMD) and change in serum co-enzyme Q10 (CoQ) levels at 8-week follow-up. Fig. 3. Relationship between change in brachial flow mediated dilatation (FMD) and change in serum lactate/pyruvate (LP) ratio at 8-week follow-up.

6 400 Y.-L. Dai et al. / Atherosclerosis 216 (2011) disease resulted in significant improvement in peak oxygen consumption and oxygen pulse, hence providing indirect evidence that co-enzyme Q10 enhances mitochondrial bioenergetics and improves aerobic capacity. In this study, we measured the plasma lactate/pyruvate ratio as a reflection of mitochondrial dysfunction. The plasma lactate/pyruvate ratio is a good indicator of impaired oxidative phosphorylation as it reflects the cytosolic redox state of reduced nicotinamide-adenine dinucleotide (NADH) to oxidized nicotinamide-adenine dinucleotide (NAD+), which cannot be measured directly. In the presence of mitochondrial dysfunction, there is excessive hydrolysis of ATP, producing more protons which drives the equilibrium reaction from pyruvate to lactate and causes the lactate/pyruvate ratio to increase. As a result, elevated lactate/pyruvate ratio has been used as an indirect indicator of mitochondrial dysfunction [11,13]. In the present study, 8 weeks of co-enzyme Q10 supplementation led to a significant reduction in plasma lactate/pyruvate ratio compared with controls, hence providing further evidence to indicate that co-enzyme Q10 supplementation reversed mitochondrial dysfunction in patients with ischaemic left ventricular systolic dysfunction. Furthermore, increased lactate/pyruvate ratio at baseline correlated with impaired FMD, suggesting that mitochondrial dysfunction exerted a detrimental effect on endothelial function. More importantly, the magnitude of reduction in plasma lactate/pyruvate ratio after co-enzyme Q10 supplementation significantly correlated with the changes in FMD. These findings provide new insight into how co-enzyme Q10 improves endothelial function via correction of mitochondrial function. Apart from its important role in mitochondrial bioenergetics, co-enzyme Q10 is also a potent antioxidant [26,27]. Nevertheless, there are conflicting data on the effect of co-enzyme Q10 supplementation on oxidative stress. Tiano et al. [1] showed an improvement in endothelium-bound extracellular superoxide dismutase (ecsod) activity in subjects with coronary artery disease and the increase in ecsod activity significantly correlated with the improvement in FMD. On the other hand, other studies failed to show any significant alteration in markers of oxidative stress such as plasma F2-isoprostane [3,23], urinary 24-h urinary 20-hydroxyeicosatetraenoic acid levels [3] and plasma soluble intercellular adhesion molecule [28] after co-enzyme Q10 supplementation. In this study, co-enzyme Q10 supplementation did not alter hs-crp, serum superoxide dismutase or 8-isoprostane; suggesting that the beneficial effects of co-enzyme Q10 on endothelial function are independent of changes in oxidative stress and systemic inflammation. Moreover, co-enzyme Q10 has also been shown to have beneficial effects on systolic and diastolic pressure, lipid profile, and glycemic control [29 31] which might also contribute to the improvement in endothelial function. Our results showed that coenzyme Q10 supplementation did not have any significant effect on blood pressure, lipid profile, HbA1c and fasting blood glucose in patients with ischaemic left ventricular systolic dysfunction who had received optimal medical therapy. Nevertheless, the beneficial effect of co-enzyme Q10 appears to be more pronounced in patients with a lower FMD and higher lactate/pyruvate ratio at baseline, and in those with a history of prior myocardial infarction. These findings suggest that co-enzyme Q10 supplementation reverses endothelial dysfunction in these groups of patients with significant cardiovascular risks on top of their optimal medical therapy without any adverse effect Study limitations First, blood lactate/pyruvate ratio was used to predict mitochondrial dysfunction in this study. However, this method is only an indirect assessment for mitochondrial dysfunction, and the validation of mitochondrial dysfunction requires muscle biopsy to determine ubiquinone status and cytochrome oxidase activity. Due to the invasive nature of muscle biopsy, direct histological assessment to confirm mitochondrial dysfunction was not performed in this study. Similarly, only plasma co-enzyme Q10 concentration rather than tissue level was measured for the same reason. Although prior study has shown that oral co-enzyme Q10 supplementation can increase mitochondrial and tissue co-enzyme Q10 concentration [6], the relationship between the plasma and tissue levels of co-enzyme Q10 remains unclear. These might account for the lack of correlation between plasma co-enzyme Q10 levels and the changes in lactate/pyruvate ratio in this study. Second, several subgroup analyses have been performed, including patients with or without prior history of myocardial infarction in this study. However, these subgroup analyses are confounded by a number of factors, such as co-existing cardiovascular risk factors and medications. Therefore, further studies are required to confirm the potential difference in the treatment effects of co-enzyme Q10 in these different subgroups of patients. 5. Conclusions In this randomized controlled study, 8 weeks of co-enzyme Q10 supplementation at 300 mg daily significantly improved endothelial function in patients with ischaemic left ventricular systolic dysfunction without any adverse effect. Furthermore, a possible dose response relationship between the improvement in FMD and the increase in plasma co-enzyme Q10 concentration was observed. More importantly, co-enzyme Q10 supplementation significantly reduced the plasma lactate/pyruvate ratio, which correlated with the improvement in FMD. These findings provide further insight into the potential mechanism by which co-enzyme Q10 improves endothelial function through ameliorating mitochondrial dysfunction. Nevertheless, whether co-enzyme Q10 supplementation in patients with ischaemic heart failure can improve clinical outcome merits further studies. Conflict of interest None declared. Acknowledgments This work was supported by the CRCG Small Project Funding of University of Hong Kong (Project No ); and Sun Chieh Yeh Heart Foundation. Jarrow Formulas, Los Angeles, CA, USA kindly supplied the co-enzyme Q10 and matching placebo capsules used in this study. References [1] Tiano L, Belardinelli R, Carnevali P, et al. Effect of coenzyme Q10 administration on endothelial function and extracellular superoxide dismutase in patients with ischaemic heart disease: a double-blind, randomized controlled study. Eur Heart J 2007;28: [2] Belardinelli R, Mucaj A, Lacalaprice F, et al. Coenzyme Q10 and exercise training in chronic heart failure. Eur Heart J 2006;27: [3] Hamilton SJ, Chew GT, Watts GF. Coenzyme Q10 improves endothelial dysfunction in statin-treated type 2 diabetic patients. Diab Care 2009;32: [4] Chan SY, Mancini GB, Kuramoto L, et al. The prognostic importance of endothelial dysfunction and carotid atheroma burden in patients with coronary artery disease. 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