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Association of extremely high levels of high-density lipoprotein cholesterol with endothelial dysfunction in men

Open AccessPublished:June 19, 2019DOI:https://doi.org/10.1016/j.jacl.2019.06.004

      Highlights

      • Flow-mediated vasodilation (FMD) was smaller in the low high-density lipoprotein cholesterol (HDL-C) than in the high HDL-C group.
      • FMD was smaller in the extremely high HDL-C group than in the high HDL-C group.
      • There was no difference in FMD between the low and extremely high HDL-C groups.
      • Extremely high HDL-C was associated with a lower quartile of FMD.

      Background

      It is not clear whether a high level of high-density lipoprotein cholesterol (HDL-C) is associated with lower risk of atherosclerosis. It is likely that HDL-C is a double-edged sword for atherosclerosis.

      Objective

      The purpose of this study was to evaluate the relationship between HDL-C levels and endothelial function in men.

      Methods

      This was a cross-sectional study. We evaluated flow-mediated vasodilation (FMD) and serum levels of HDL-C in 5842 men aged 18 to 92 years who were not receiving lipid-lowering therapy. All participants were divided into four groups by HDL-C level: low HDL-C (<40 mg/dL), moderate HDL-C (40–59 mg/dL), high HDL-C (60–79 md/dL), and extremely high HDL-C (≥80 mg/dL). We were not able to evaluate the amount of alcohol intake because there was limited information on the amount of alcohol drinking in our database.

      Results

      FMD values were significantly smaller in the low group and the extremely high group than in the high group (P = .001 and P = .016, respectively). There was no significant difference in FMD between the low group and the extremely high group. Multiple logistic regression analysis revealed that extremely high HDL-C, but not low HDL-C, was independently associated with the lowest quartile of FMD (odds ratio: 1.39, 95% confidence interval: 1.09–1.77; P = .009).

      Conclusions

      An extremely high level of HDL-C in men (8.1% of this population) was associated with a significant reduction in FMD.

      Keywords

      Introduction

      Several lines of evidence have shown an inverse relationship between high-density lipoprotein cholesterol (HDL-C) levels and cardiovascular disease.
      • Gordon T.
      • Castelli W.P.
      • Hjortland M.C.
      • Kannel W.B.
      • Dawber T.R.
      High density lipoprotein as a protective factor against coronary heart disease. The Framingham Study.
      • Assmann G.
      • Schulte H.
      Relation of high-density lipoprotein cholesterol and triglycerides to incidence of atherosclerotic coronary artery disease (the PROCAM experience). Prospective Cardiovascular Munster study.
      • Gordon D.J.
      • Probstfield J.L.
      • Garrison R.J.
      • et al.
      High-density lipoprotein cholesterol and cardiovascular disease. Four prospective American studies.
      It is well known that HDL has an atheroprotective effect through transportation of excess cholesterol from macrophages in the liver and bile, namely, cholesterol reverse transport.
      • Rader D.J.
      • Alexander E.T.
      • Weibel G.L.
      • Billheimer J.
      • Rothblat G.H.
      The role of reverse cholesterol transport in animals and humans and relationship to atherosclerosis.
      In addition, HDL activates endothelial nitric oxide synthase (eNOS), has antioxidant and anti-inflammatory effects and an antithrombotic effect, and prevents apoptosis of endothelial cells induced by tumor necrosis factor alpha.
      • Yuhanna I.S.
      • Zhu Y.
      • Cox B.E.
      • et al.
      High-density lipoprotein binding to scavenger receptor-BI activates endothelial nitric oxide synthase.
      • Navab M.
      • Hama S.Y.
      • Cooke C.J.
      • et al.
      Normal high density lipoprotein inhibits three steps in the formation of mildly oxidized low density lipoprotein: step 1.
      • Cockerill G.W.
      • Rye K.A.
      • Gamble J.R.
      • Vadas M.A.
      • Barter P.J.
      High-density lipoproteins inhibit cytokine-induced expression of endothelial cell adhesion molecules.
      • Sugano M.
      • Tsuchida K.
      • Makino N.
      High-density lipoproteins protect endothelial cells from tumor necrosis factor-alpha-induced apoptosis.
      However, cohort studies have shown that an extremely high HDL-C group had high mortality and morbidity rates of cardiovascular disease and that there was a U-shaped curve between HDL-C levels and mortality and morbidity rates of cardiovascular disease.
      • Hirata A.
      • Okamura T.
      • Sugiyama D.
      • et al.
      The relationship between very high levels of serum high-density lipoprotein cholesterol and cause-specific mortality in a 20-year follow-up study of Japanese general population.
      • Madsen C.M.
      • Varbo A.
      • Nordestgaard B.G.
      Extreme high high-density lipoprotein cholesterol is paradoxically associated with high mortality in men and women: two prospective cohort studies.
      • Hirata A.
      • Sugiyama D.
      • Watanabe M.
      • et al.
      Association of extremely high levels of high-density lipoprotein cholesterol with cardiovascular mortality in a pooled analysis of 9 cohort studies including 43,407 individuals: The EPOCH-JAPAN study.
      In addition, recent large clinical trials have shown that an increase in HDL-C levels with pharmacologic interventions does not reduce the risk of cardiovascular events. Cholesteryl ester transfer protein (CETP) modulates the transfer of cholesteryl esters from HDL to apolipoprotein B–containing lipoproteins.
      • Tall A.R.
      Plasma cholesteryl ester transfer protein.
      Pharmacologic inhibition of CETP raises HDL-C levels and decreases low-density lipoprotein cholesterol (LDL-C) levels.
      • Barter P.J.
      • Caulfield M.
      • Eriksson M.
      • et al.
      Effects of torcetrapib in patients at high risk for coronary events.
      • Schwartz G.G.
      • Olsson A.G.
      • Abt M.
      • et al.
      Effects of dalcetrapib in patients with a recent acute coronary syndrome.
      • Lincoff A.M.
      • Nicholls S.J.
      • Riesmeyer J.S.
      • et al.
      Evacetrapib and cardiovascular outcomes in high-risk vascular disease.
      • Group H.T.R.C.
      • Bowman L.
      • Hopewell J.C.
      • et al.
      Effects of anacetrapib in patients with atherosclerotic vascular disease.
      Randomized clinical trials using CETP inhibitors such as torcetrapib, dalcetrapib, and evacetrapib have shown an increase in mortality or a lack of efficacy, whereas CETP inhibitors increased HDL-C levels.
      • Barter P.J.
      • Caulfield M.
      • Eriksson M.
      • et al.
      Effects of torcetrapib in patients at high risk for coronary events.
      • Schwartz G.G.
      • Olsson A.G.
      • Abt M.
      • et al.
      Effects of dalcetrapib in patients with a recent acute coronary syndrome.
      • Lincoff A.M.
      • Nicholls S.J.
      • Riesmeyer J.S.
      • et al.
      Evacetrapib and cardiovascular outcomes in high-risk vascular disease.
      On the other hand, the CETP inhibitor anacetrapib reduced the risk of major coronary events during a 4-year treatment period.
      • Group H.T.R.C.
      • Bowman L.
      • Hopewell J.C.
      • et al.
      Effects of anacetrapib in patients with atherosclerotic vascular disease.
      The mechanism of this reduction seems to be largely explained by lowering of non–HDL-C rather than increase in HDL-C.
      • Group H.T.R.C.
      • Bowman L.
      • Hopewell J.C.
      • et al.
      Effects of anacetrapib in patients with atherosclerotic vascular disease.
      Results of recent human genetic studies have shown that genetic conditions leading to increased HDL-C levels may not always be associated with lower risk of atherosclerosis.
      • Vitali C.
      • Khetarpal S.A.
      • Rader D.J.
      HDL cholesterol metabolism and the risk of CHD: new insights from human genetics.
      Moreover, a loss-of-function coding variant in SCARB1 leads to increased risk of coronary heart disease despite elevation in HDL-C levels.
      • Zanoni P.
      • Khetarpal S.A.
      • Larach D.B.
      • et al.
      Rare variant in scavenger receptor BI raises HDL cholesterol and increases risk of coronary heart disease.
      These findings suggest that extremely high HDL-C levels are associated with an increase in the risk of mortality and morbidity of cardiovascular disease. HDL-C may be a double-edged sword for atherosclerosis.
      Endothelial dysfunction is the initial step of atherosclerosis and plays an important role in the development of atherosclerosis.
      • Ross R.
      Atherosclerosis--an inflammatory disease.
      • Higashi Y.
      • Noma K.
      • Yoshizumi M.
      • Kihara Y.
      Endothelial function and oxidative stress in cardiovascular diseases.
      Flow-mediated vasodilation (FMD) of the brachial artery has been used for noninvasive assessment of endothelial function, and FMD has been shown to be significantly associated with cardiovascular risk factors including HDL-C and to be an independent predictor of cardiovascular events.
      • Corretti M.C.
      • Anderson T.J.
      • Benjamin E.J.
      • et al.
      Guidelines for the ultrasound assessment of endothelial-dependent flow-mediated vasodilation of the brachial artery: a report of the International Brachial Artery Reactivity Task Force.
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      • Keyes M.J.
      • et al.
      Clinical correlates and heritability of flow-mediated dilation in the community: the Framingham Heart Study.
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      • et al.
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      • Rossi R.
      Prognostic role of reversible endothelial dysfunction in hypertensive postmenopausal women.
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      • Burke G.L.
      • et al.
      Predictive value of brachial flow-mediated dilation for incident cardiovascular events in a population-based study: the multi-ethnic study of atherosclerosis.
      However, the role of marked elevation of HDL-C in endothelial function remains unclear.
      • Maruhashi T.
      • Soga J.
      • Fujimura N.
      • et al.
      Relationship between flow-mediated vasodilation and cardiovascular risk factors in a large community-based study.
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      • Maruhashi T.
      • Matsumoto T.
      • et al.
      Relationship between serum triglyceride levels and endothelial function in a large community-based study.
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      • Hunter L.M.
      • Watkins M.T.
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      • Vita J.A.
      Risk stratification for postoperative cardiovascular events via noninvasive assessment of endothelial function: a prospective study.
      Therefore, we evaluated the relationship between HDL-C levels and endothelial function assessed by FMD in men not receiving lipid-lowering therapy.

      Materials and methods

      Subjects

      A total of 10,247 Japanese adults (7385 subjects from the FMD-J study and 2862 subjects who underwent a health checkup at Hiroshima University Hospital between August 2007 and August 2016) were enrolled in this study. From this registry, 7682 men aged 18 to 92 years were recruited. Subjects with unclear images of the brachial artery interfaces and subjects without information on HDL-C level were excluded. To eliminate the influence of pharmacologic therapy, subjects who were receiving lipid-lowering medicine (eg, statins, ezetimibe, proprotein convertase subtilisin/kexin type 9 inhibitors, bile acid sequestrants, fibrates, eicosapentaenoic acid, and niacin) were also excluded. Finally, 5842 subjects were enrolled in this study. Hypertension was defined as treatment with oral antihypertensive agents or systolic blood pressure of more than 140 mm Hg or diastolic blood pressure of more than 90 mm Hg measured in a sitting position on at least three different occasions. Diabetes mellitus was defined according to the American Diabetes Association recommendation.
      American Diabetes Association: clinical practice recommendations 1999.
      Dyslipidemia was defined according to the third report of the National Cholesterol Education Program.
      Executive summary of the third report of the national cholesterol education program (NCEP) expert panel on detection, evaluation, and treatment of high blood cholesterol in adults (adult treatment panel III).
      Smokers were defined as those who were current smokers. Coronary heart disease included angina pectoris, myocardial infarction, and unstable angina. Cerebrovascular disease included ischemic stroke, hemorrhagic stroke, and transient ischemic attack. Cardiovascular disease was defined as coronary heart disease and cerebrovascular disease. Framingham risk score was calculated by points of risk factors: age, total cholesterol level, HDL-C level, systolic blood pressure, diabetes mellitus, and smoking status.
      • Wilson P.W.
      • D'Agostino R.B.
      • Levy D.
      • Belanger A.M.
      • Silbershatz H.
      • Kannel W.B.
      Prediction of coronary heart disease using risk factor categories.
      All participants were divided into 4 groups according to the definitions used in a previous study in Japan: low HDL-C (<40 mg/dL), moderate HDL-C (40–59 mg/dL), high HDL-C (60–79 mg/dL), and extremely high HDL-C (≥80 mg/dL).
      • Hirata A.
      • Okamura T.
      • Sugiyama D.
      • et al.
      The relationship between very high levels of serum high-density lipoprotein cholesterol and cause-specific mortality in a 20-year follow-up study of Japanese general population.
      • Moriyama Y.
      • Okamura T.
      • Inazu A.
      • et al.
      A low prevalence of coronary heart disease among subjects with increased high-density lipoprotein cholesterol levels, including those with plasma cholesteryl ester transfer protein deficiency.
      The ethical committees of the participating institutions approved the study protocol. The study was executed in accordance with the Good Clinical Practice guidelines. Informed consent for participation in the study was obtained from all subjects. The protocol was registered in the University Hospital Medical Information Network Clinical Trials Registry (UMIN000012950).

      Study protocol

      Subjects fasted overnight for at least 12 hours and abstained from caffeine, alcohol, smoking, and antioxidant vitamins on the day of the FMD examination. The subjects were kept in the supine position in a quiet, dark, air-conditioned room (constant temperature of 22°C–25°C) throughout the study. A 23-gauge polyethylene catheter was inserted into the deep antecubital vein to obtain blood samples. At 30 minutes of maintaining a supine position, FMD is measured. The observers were blind to the form of examination.

      Measurement of FMD

      We evaluated the vascular response to reactive hyperemia in the brachial artery for assessment of endothelium-dependent FMD. A high-resolution linear artery transducer was coupled to computer-assisted analysis software (UNEXEF18G; UNEX Co, Nagoya, Japan) that used an automated edge detection system for measurement of brachial artery diameter. A blood pressure cuff was placed around the forearm. The brachial artery was scanned longitudinally 5 to 10 cm above the elbow. When the clearest B-mode image of the anterior and posterior intimal interfaces between the lumen and vessel wall was obtained, the transducer was held at the same point throughout the scan by a special probe holder (UNEX Co) to ensure consistency of the image. Depth and gain setting were set to optimize the images of the arterial lumen wall interface. When the tracking gate was placed on the intima, the artery diameter was automatically tracked, and the waveform of diameter changes over the cardiac cycle was displayed in real time using the FMD mode of the tracking system. This allowed the ultrasound images to be optimized at the start of the scan and the transducer position to be adjusted immediately for optimal tracking performance throughout the scan. Pulsed Doppler flow was assessed at baseline and during peak hyperemic flow, which was confirmed to occur within 15 seconds after cuff deflation. Blood flow velocity was calculated from the color Doppler data and was displayed as a waveform in real time. The baseline longitudinal image of the artery was acquired for 30 seconds, and then the blood pressure cuff was inflated to 50 mm Hg above systolic pressure for 5 minutes. The longitudinal image of the artery was recorded continuously until 5 minutes after cuff deflation. Pulsed Doppler velocity signals were obtained for 20 seconds at baseline and for 10 seconds immediately after cuff deflation. Changes in brachial artery diameter were immediately expressed as percentage change relative to the vessel diameter before cuff inflation. FMD was automatically calculated as the percentage change in peak vessel diameter from the baseline value. Percentage of FMD ([peak diameter − baseline diameter]/baseline diameter) was used for analysis. Blood flow volume was calculated by multiplying the Doppler flow velocity (corrected for the angle) by heart rate and vessel cross-sectional area (−r2). Reactive hyperemia was calculated as the maximum percentage increase in flow after cuff deflation compared with baseline flow. The coefficient of variation for FMD was 10.1% in our laboratory. The correlation coefficient between FMD analyzed at the core laboratory and participant institutions was 0.84 (P < .001).

      Statistical analysis

      Results are presented as means ± SD or median (interquartile range) for continuous variables. All reported probability values were 2 sided, and a probability value of <.05 was considered statistically significant. An association between FMD and HDL-C was explored visually using a locally weighted regression smoothing (Locally Weighted Scatterplot Smoothing) plot. The relation between FMD and HDL-C was determined by Pearson's correlation analysis. Comparison of variables among four groups by difference of the HDL-C level was performed using one-way analysis of variance or the Kruskal–Wallis test depending on normality of the data. Normality was assessed by visual inspection of histograms. Tukey's post-hoc test was performed to compare the differences in FMD between groups. Multiple logistic regression analysis was performed to identify independent variables associated with lower quartile of FMD (<3.9%). Age, body mass index, systolic blood pressure, LDL-C, glucose, and presences of current smoking were entered into the multiple logistic regression analysis. As a sensitivity analysis, propensity score analysis was used to generate a set of matched cases (subjects with extremely high HDL-C levels) and controls (subjects with high HDL-C levels). A logistic regression model was used to estimate the propensity of extremely high HDL-C levels based on variables associated with HDL-C, including age, body mass index, systolic blood pressure, total cholesterol, triglycerides, glucose, use of anti-hypertensive drugs (yes or no), current smoking (yes or no), and history of cardiovascular disease (yes or no). With these propensity scores using a caliper width of 0.2 standard deviations of the logit of the propensity score, two well-matched groups based on clinical characteristics were created for comparison of the prevalence of endothelial dysfunction defined as FMD of <3.9%, the division point for the lowest quartile of FMD in all participants. All analyses were conducted using JMP version 13.0 software (SAS Institute, Cary, NC) and Stata version 15 (Stata Corporation, College Station, TX).

      Results

      Baseline characteristics

      Baseline characteristics of all subjects are summarized in Table 1. The age range was 18 to 92 years. Of the 5842 subjects, 2473 (42.3%) had hypertension, 2560 (43.8%) had dyslipidemia, 486 (8.3%) had diabetes mellitus, 259 (4.4%) had previous cardiovascular disease, and 2156 (37.0%) were current smokers. There were significant differences among the four groups in age, body mass index, systolic blood pressure, diastolic blood pressure, heart rate, total cholesterol, triglycerides, HDL-C, LDL-C, glucose, use of antihypertensive drugs and antidiabetic drugs, prevalence of hypertension, dyslipidemia, and diabetes mellitus, and percentage of smokers.
      Table 1Clinical characteristics of the subjects on the basis of HDL-C
      VariablesTotal (n = 5842)Low <40 mg/dL (n = 466)Moderate 40–59 mg/dL (n = 3115)High 60–79 mg/dL (n = 1787)Extremely high ≥80 mg/dL (n = 474)P value for trend
      Age, y50.2 ± 1150.9 ± 1249.8 ± 1150.1 ± 1251.9 ± 11.002
      Body mass index, kg/m223.6 ± 3.225.2 ± 3.524.1 ± 3.222.7 ± 2.922.0 ± 2.7<.001
      Systolic blood pressure, mm Hg128 ± 16128 ± 15129 ± 16127 ± 16129 ± 18.001
      Diastolic blood pressure, mm Hg81 ± 1280 ± 1181 ± 1279 ± 1280 ± 12<.001
      Heart rate, bpm64 ± 1165 ± 1165 ± 1163 ± 1163 ± 12<.001
      Total cholesterol, mg/dL202 ± 33193 ± 37200 ± 33203 ± 32213.7 ± 31<.001
      Triglycerides, mg/dL112 (79, 161)185 (128, 269)125 (90, 175)90 (67, 123)77 (58, 109)<.001
      HDL-C, mg/dL58 ± 1536 ± 350 ± 668 ± 591 ± 11<.001
      LDL-C, mg/dL119 ± 30117 ± 32123 ± 29116 ± 28106 ± 29<.001
      Glucose, mg/dL101 ± 22105 ± 26102 ± 24100 ± 19100 ± 18<.001
      Medications, n (%)
       Antihypertensive therapy1353 (23.2)156 (33.5)746 (24.0)340 (19.0)111 (23.1)<.001
       Antihyperglycemic therapy303 (5.2)49 (10.5)162 (5.2)70 (3.9)22 (4.6)<.001
      Framingham risk score, %9.2 ± 7.812.4 ± 10.010.3 ± 8.37.0 ± 5.97.4 ± 5.8<.001
      Medical history, n (%)
       Hypertension2473 (42.3)219 (47.0)1371 (44.0)668 (37.4)215 (45.4)<.001
       Dyslipidemia2560 (43.8)421 (90.3)1523 (48.9)522 (29.2)94 (19.8)<.001
       Diabetes mellitus486 (8.3)70 (15.0)279 (9.0)108 (6.0)29 (6.1)<.001
       Current smokers2156 (37.0)209 (45.0)1257 (40.5)557 (31.2)133 (28.1)<.001
       Previous cardiovascular disease259 (4.4)30 (6.4)134 (4.3)75 (4.2)20 (4.2).231
      HDL-C, high-density lipoprotein cholesterol; LDL-C, low-density lipoprotein cholesterol.
      Data are presented as mean ± standard deviation or median (interquartile range).

      Relationship between HDL-C and endothelial function

      The scatter plot between FMD and HDL-C with Locally Weighted Scatterplot Smoothing smoothed curve is shown in Figure S1 in the online-only Data Supplement. FMD gradually increased to about 70 to 80 mg/dL of HDL-C levels and decreased in relation to the increase in HDL-C levels after about the 70 to 80 mg/dL of HDL-C. Using a linear regression analysis, there was no significant relationship between FMD and HDL-C (r = 0.03, P = .052). FMD values were 5.6 ± 3.1% in the low HDL-C group, 6.0 ± 3.0% in the moderate HDL-C group, 6.2 ± 3.2% in the high HDL-C group, and 5.7 ± 3.0% in the extremely high HDL-C group. FMD values were significantly smaller in the low HDL-C group and extremely high HDL-C group than in the high HDL-C group (P = .001 and P = .016, respectively; Fig. 1). There was no significant difference in FMD between the low HDL-C group and the extremely high HDL-C group. Multiple logistic regression analysis revealed that extremely high HDL-C was independently associated with the lowest quartile of FMD (odds ratio: 1.39, 95% confidence interval: 1.09–1.77; P = .009) and that low HDL-C and moderate HDL-C were not associated with FMD (Table 2).
      Figure thumbnail gr1
      Figure 1Bar graphs show flow-mediated vasodilation (FMD) in the low levels of high-density lipoprotein cholesterol (HDL-C), moderate levels of HDL-C, high levels of HDL-C, and extremely high levels of HDL-C groups. The error bars indicate the standard deviation.
      Table 2Multivariate analysis of the relationship between FMD and HDL-C
      VariablesModel 1Model 2Model 3Model 4
      OR (95% CI)P valueOR (95% CI)P valueOR (95% CI)P valueOR (95% CI)P value
      HDL-C, mg/dL
       <401.42 (1.13–1.79).0021.39 (1.10–1.77).0071.09 (0.84–1.41).5231.21 (0.94–1.57).135
       40–591.10 (0.96–1.26).1851.13 (0.98–1.54).0720.97 (0.84–1.41).3551.02 (0.87–1.18).837
       60–791 (reference)1 (reference)1 (reference)1 (reference)
       ≥801.39 (1.11–1.75).0051.30 (1.02–1.65).0291.36 (1.07–1.73).0141.39 (1.09–1.77).009
      BMI, body mass index; CI, confidence interval; FMD, flow-mediated vasodilation; HDL-C, high-density lipoprotein cholesterol; OR, odds ratio for lowest quartile of FMD (<3.9%).
      Model 1: unadjusted model.
      Model 2: adjusted for age.
      Model 3: adjusted for age, BMI, the presence of hypertension, dyslipidemia, diabetes, and current smoking.
      Model 4: adjusted for age, BMI, systolic blood pressure, low-density lipoprotein cholesterol, glucose, and current smoking.
      We next evaluated the association between extremely high HDL-C and FMD in the propensity score–matched population. There were significant differences in triglycerides, HDL-C, LDL-C, Framingham risk score, and prevalence of dyslipidemia (Table S1 in the online-only Data Supplement). There were no significant differences in other baseline variables between the two groups. After matching for confounding factors, FMD was significantly lower in subjects with extremely high HDL-C than in subjects with high HDL-C (P < .001; Figure S2 in the online-only Data Supplement).

      Discussion

      In the present study, we demonstrated for the first time that endothelial function was impaired not only in subjects with low HDL-C levels but also in subjects with extremely high HDL-C levels. After adjustment for traditional cardiovascular risk factors, extremely high levels of HDL-C were significantly associated with endothelial dysfunction.
      It is well known that treatment with drugs such as fibrates, CETP inhibitors, and statins increases HDL-C levels, and lipid-lowering therapy, including HDL-C-elevating therapy, per se improves endothelial function. Therefore, in the present study, we carefully excluded subjects receiving lipid-lowering therapy to eliminate the influence of lipid-lowering medication on HDL-C levels.
      Previous studies have shown that there is an inverse association between HDL-C levels and incidence of events of coronary heart disease.
      • Gordon T.
      • Castelli W.P.
      • Hjortland M.C.
      • Kannel W.B.
      • Dawber T.R.
      High density lipoprotein as a protective factor against coronary heart disease. The Framingham Study.
      • Assmann G.
      • Schulte H.
      Relation of high-density lipoprotein cholesterol and triglycerides to incidence of atherosclerotic coronary artery disease (the PROCAM experience). Prospective Cardiovascular Munster study.
      • Gordon D.J.
      • Probstfield J.L.
      • Garrison R.J.
      • et al.
      High-density lipoprotein cholesterol and cardiovascular disease. Four prospective American studies.
      HDL plays an important role in atherosclerosis through cholesterol reverse transport, activation of eNOS, antioxidation, anti-inflammation, and prevention of endothelial cell apoptosis.
      • Rader D.J.
      • Alexander E.T.
      • Weibel G.L.
      • Billheimer J.
      • Rothblat G.H.
      The role of reverse cholesterol transport in animals and humans and relationship to atherosclerosis.
      • Yuhanna I.S.
      • Zhu Y.
      • Cox B.E.
      • et al.
      High-density lipoprotein binding to scavenger receptor-BI activates endothelial nitric oxide synthase.
      • Navab M.
      • Hama S.Y.
      • Cooke C.J.
      • et al.
      Normal high density lipoprotein inhibits three steps in the formation of mildly oxidized low density lipoprotein: step 1.
      • Cockerill G.W.
      • Rye K.A.
      • Gamble J.R.
      • Vadas M.A.
      • Barter P.J.
      High-density lipoproteins inhibit cytokine-induced expression of endothelial cell adhesion molecules.
      • Sugano M.
      • Tsuchida K.
      • Makino N.
      High-density lipoproteins protect endothelial cells from tumor necrosis factor-alpha-induced apoptosis.
      However, some studies showed that high levels of HDL-C are paradoxically associated with high mortality and vascular events.
      • Hirata A.
      • Okamura T.
      • Sugiyama D.
      • et al.
      The relationship between very high levels of serum high-density lipoprotein cholesterol and cause-specific mortality in a 20-year follow-up study of Japanese general population.
      • Madsen C.M.
      • Varbo A.
      • Nordestgaard B.G.
      Extreme high high-density lipoprotein cholesterol is paradoxically associated with high mortality in men and women: two prospective cohort studies.
      • Hirata A.
      • Sugiyama D.
      • Watanabe M.
      • et al.
      Association of extremely high levels of high-density lipoprotein cholesterol with cardiovascular mortality in a pooled analysis of 9 cohort studies including 43,407 individuals: The EPOCH-JAPAN study.
      In clinical trials, elevating HDL-C levels by inhibition of CETP failed to decrease cardiovascular disease.
      • Barter P.J.
      • Caulfield M.
      • Eriksson M.
      • et al.
      Effects of torcetrapib in patients at high risk for coronary events.
      • Schwartz G.G.
      • Olsson A.G.
      • Abt M.
      • et al.
      Effects of dalcetrapib in patients with a recent acute coronary syndrome.
      • Lincoff A.M.
      • Nicholls S.J.
      • Riesmeyer J.S.
      • et al.
      Evacetrapib and cardiovascular outcomes in high-risk vascular disease.
      The first CETP inhibitor, torcetrapib, increased the risk of mortality and morbidity of cardiovascular disease rather than reducing the risk of cardiovascular events, while torcetrapib increased HDL-C levels.
      • Barter P.J.
      • Caulfield M.
      • Eriksson M.
      • et al.
      Effects of torcetrapib in patients at high risk for coronary events.
      In addition, torcetrapib increased systolic blood pressure and impaired endothelial function because of unexpected off-target effects.
      • Simic B.
      • Hermann M.
      • Shaw S.G.
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      Torcetrapib impairs endothelial function in hypertension.
      Dalcetrapib, another CETP inhibitor, increased HDL-C levels without affecting blood pressure or biomarkers of inflammation and oxidative stress but did not reduce the risk of cardiovascular events and did not improve endothelial function after 12 and 36 weeks in a high-risk population.
      • Schwartz G.G.
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      The CETP inhibitor evacetrapib also increased HDL-C levels, but the trial of this CETP inhibitor was terminated early because of a lack of efficacy.
      • Lincoff A.M.
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      Evacetrapib and cardiovascular outcomes in high-risk vascular disease.
      The Randomized Evaluation of the Effects of Anacetrapib through Lipid Modification trial showed a significant reduction of cardiovascular events during a 4-year follow-up period.
      • Group H.T.R.C.
      • Bowman L.
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      Effects of anacetrapib in patients with atherosclerotic vascular disease.
      The reasons for the discrepant results of trials using CETP inhibitors are unclear. Although anacetrapib reduced cardiovascular events in the Randomized Evaluation of the Effects of Anacetrapib through Lipid Modification trial, it has been proposed that a reduction of non–HDL-C rather than an increase in HDL-C is the main mechanism of cardiovascular risk reduction.
      • Group H.T.R.C.
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      Effects of anacetrapib in patients with atherosclerotic vascular disease.
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      The present study showed that FMD was significantly smaller in subjects with extremely high HDL-C as well as subjects with low HDL-C than in subjects with high HDL-C. After adjustment for cardiovascular risk factors, extremely high HDL-C, but not low HDL-C, was significantly associated with endothelial dysfunction.
      Although environmental factors such as obesity, smoking, alcohol, diet, and physical activity have been shown to influence HDL-C levels, it has been estimated that heritability of HDL-C levels is about 40% to 60%.
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      Loss-of-function mutations in the genes LIPG, SCARB1, and CETP cause conditions of extremely high HDL-C.
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      HDL cholesterol metabolism and the risk of CHD: new insights from human genetics.
      The LIPG gene encodes endothelial lipase, which has been shown to mediate HDL catabolism.
      • Voight B.F.
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      • et al.
      Plasma HDL cholesterol and risk of myocardial infarction: a mendelian randomisation study.
      A Mendelian randomization study has shown that a loss-of-function variant in the LIPG gene is not associated with risk of myocardial infarction.
      • Voight B.F.
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      Plasma HDL cholesterol and risk of myocardial infarction: a mendelian randomisation study.
      The SRARB1 gene encodes scavenger receptor class B type 1.
      • Zanoni P.
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      • et al.
      Rare variant in scavenger receptor BI raises HDL cholesterol and increases risk of coronary heart disease.
      Scavenger receptor class B type 1 is an HDL receptor and promotes the uptake of HDL cholesteryl esters into cells, especially hepatocytes.
      • Zanoni P.
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      • Larach D.B.
      • et al.
      Rare variant in scavenger receptor BI raises HDL cholesterol and increases risk of coronary heart disease.
      It has been shown that a loss-of-function variant in SCARB1, P376L, is associated with increased coronary heart disease (odds ratio = 1.79, P = .0018).
      • Zanoni P.
      • Khetarpal S.A.
      • Larach D.B.
      • et al.
      Rare variant in scavenger receptor BI raises HDL cholesterol and increases risk of coronary heart disease.
      CETP deficiency has been reported to be one of the major causes of increased HDL-C levels in Japan.
      • Inazu A.
      • Brown M.L.
      • Hesler C.B.
      • et al.
      Increased high-density lipoprotein levels caused by a common cholesteryl-ester transfer protein gene mutation.
      • Koizumi J.
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      Deficiency of serum cholesteryl-ester transfer activity in patients with familial hyperalphalipoproteinaemia.
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      It is thought that the presence of CETP deficiency contributes to a longevity condition.
      • Moriyama Y.
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      A low prevalence of coronary heart disease among subjects with increased high-density lipoprotein cholesterol levels, including those with plasma cholesteryl ester transfer protein deficiency.
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      A pedigree of homozygous familial hyperalphalipoproteinemia.
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      • et al.
      Genetic cholesteryl ester transfer protein deficiency is extremely frequent in the Omagari area of Japan. Marked hyperalphalipoproteinemia caused by CETP gene mutation is not associated with longevity.
      It has been shown that subjects with CETP deficiency have large and cholesteryl ester–rich HDL-C and polydisperse LDL-C.
      • Yamashita S.
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      • Sakai N.
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      • Tarui S.
      • Hui D.Y.
      Accumulation of apolipoprotein E-rich high density lipoproteins in hyperalphalipoproteinemic human subjects with plasma cholesteryl ester transfer protein deficiency.
      Triglyceride-rich LDL decreases affinity for the LDL receptor.
      • Sakai N.
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      Decreased affinity of low density lipoprotein (LDL) particles for LDL receptors in patients with cholesteryl ester transfer protein deficiency.
      It remains controversial whether cholesteryl ester–rich HDL-C reduces or enhances the ability to promote cholesterol efflux from macrophages.
      • Ishigami M.
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      • et al.
      Large and cholesteryl ester-rich high-density lipoproteins in cholesteryl ester transfer protein (CETP) deficiency can not protect macrophages from cholesterol accumulation induced by acetylated low-density lipoproteins.
      • Matsuura F.
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      • Chen W.
      • Jiang X.C.
      • Tall A.R.
      HDL from CETP-deficient subjects shows enhanced ability to promote cholesterol efflux from macrophages in an apoE- and ABCG1-dependent pathway.
      Gomaraschi et al.
      • Gomaraschi M.
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      • Pozzi S.
      • et al.
      eNOS activation by HDL is impaired in genetic CETP deficiency.
      showed that HDL-C isolated from carriers of CETP deficiency impaired the activation of eNOS. In addition, inhibition of CETP may lead to increases in prooxidant and proinflammatory properties by an increase in dysfunctional HDL-C production. Measurements of markers of oxidative stress and inflammation would enable more specific conclusions concerning the role of HDL-C in endothelial function to be drawn. It is likely that an increase in dysfunctional HDL attenuates the eNOS/NO pathway. When considering the effects of HDL on atherosclerosis including vascular function, attention should be paid to HDL function as well as circulating levels of HDL-C.
      In the present study, FMD was significantly smaller in the low HDL-C group (<40 mg/dL) than in the high HDL-C group (60–79 md/dL). However, after adjustment for cardiovascular risk factors, there was no significant difference in FMD between the low HDL-C group and the high HDL-C group. In the low HDL-C group, the prevalence of hypertension, prevalence of diabetes mellitus, and percentage of current smokers were higher than those in the other groups, suggesting that these confounding factors reflect endothelial function in subjects with low HDL-C. In clinical trials, isolated low levels of HDL-C were not associated with coronary heart disease and mortality in Japan.
      • Hirata A.
      • Sugiyama D.
      • Watanabe M.
      • et al.
      Association of extremely high levels of high-density lipoprotein cholesterol with cardiovascular mortality in a pooled analysis of 9 cohort studies including 43,407 individuals: The EPOCH-JAPAN study.
      • Hirata T.
      • Sugiyama D.
      • Nagasawa S.Y.
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      A pooled analysis of the association of isolated low levels of high-density lipoprotein cholesterol with cardiovascular mortality in Japan.

      Study limitations

      There are a number of limitations in the present study. First, this study was a cross-sectional study. We were able to evaluate association but not causality. Second, we were not able to evaluate the amount of alcohol drinking because there was limited information on the amount of alcohol drinking in our database. It has been shown that there is a positive gradient of HDL-C levels with alcohol consumption.
      • Ernst N.
      • Fisher M.
      • Smith W.
      • et al.
      The association of plasma high-density lipoprotein cholesterol with dietary intake and alcohol consumption. The Lipid Research Clinics Prevalence Study.
      We previously reported that endothelial function is impaired in relation to alcohol intake in men.
      • Oda N.
      • Kajikawa M.
      • Maruhashi T.
      • et al.
      Endothelial function is impaired in relation to alcohol intake even in the case of light alcohol consumption in Asian men; Flow-mediated Dilation Japan (FMD-J) Study.
      The amount of alcohol drinking might have an impact on the relationship between HDL-C and endothelial function. Third, we did not assess CETP mutations. Finally, HDL-C functions other than endothelial function, including anti-inflammation and proinflammation, antioxidation and pro-oxidation, antithrombosis and prothrombosis, eNOS/NO pathway, and cholesterol efflux capacity, were not evaluated. Future studies are needed to confirm the effects of HDL functions, especially cholesterol efflux capacity, on endothelial function.

      Conclusions

      In the present study, an extremely high level of HDL-C in men (8.1% of this population) was associated with a significant reduction in FMD. Subjects with extremely high HDL-C may have dysfunctional HDL that has harmful effects on vascular function.

      Acknowledgments

      The authors would like to thank all patients who participated in this study. In addition, they thank Miki Kumiji, Megumi Wakisaka, Ki-ichiro Kawano, and Satoko Michiyama for their excellent secretarial assistance; FMD-J investigators of Takayuki Hidaka, MD, PhD; Shuji Nakamura, MD, PhD; Junko Soga, MD, PhD; Yuichi Fujii, MD, PhD; Naomi Idei, MD; Noritaka Fujimura, MD, PhD; Shinsuke Mikami, MD, PhD; Yumiko Iwamoto, MD; Akimichi Iwamoto, MD, PhD; Takeshi Matsumoto, MD, PhD; Nozomu Oda, MD, PhD (Department of Cardiovascular Medicine, Hiroshima University Graduate School of Biomedical Sciences, Hiroshima, Japan); Kana Kanai, PhD; Hraruka Morimoto, PhD (Department of Cardiovascular Regeneration and Medicine, Research Institute for Radiation Biology and Medicine, Hiroshima University, Hiroshima, Japan); Tomohisa Sakashita, MD, PhD; Yoshiki Kudo, MD, PhD (Department of Obstetrics and Gynecology, Hiroshima University Graduate School of Biomedical Sciences, Hiroshima, Japan); Taijiro Sueda, MD, PhD (Department of Surgery, Hiroshima University Graduate School of Biomedical Sciences, Hiroshima, Japan); Hirofumi Tomiyama, MD, PhD, FAHA; Akira Yamashina, MD, PhD (Department of Cardiology, Tokyo Medical University, Tokyo, Japan); Bonpei Takase, MD, PhD, FAHA (Division of Biomedical Engineering, National Defense Medical College Research Institute, Tokorozawa, Japan); Takahide Kohro, MD, PhD (Department of Cardiology, Tokyo Medical University, Tokyo, Japan); Toru Suzuki, MD, PhD (Cardiovascular Medicine, University of Leicester, Leicester, UK); Tomoko Ishizu, MD, PhD (Cardiovascular Division, Institute of Clinical Medicine, University of Tsukuba, Ibaraki, Japan); Shinichiro Ueda, MD, PhD (Department of Clinical Pharmacology and Therapeutics, University of the Ryukyu School of Medicine, Okinawa, Japan); Tsutomu Yamazaki, MD, PhD (Clinical Research Support Center, Faculty of Medicine, The University of Tokyo, Tokyo, Japan); Tomoo Furumoto, MD, PhD (Department of Cardiovascular Medicine, Hokkaido University Graduate School of Medicine, Hokkaido, Japan); Kazuomi Kario, MD, PhD (Division of Cardiovascular Medicine, Jichi Medical University School of Medicine, Tochigi, Japan); Teruo Inoue, MD, PhD (Department of Cardiovascular Medicine, Dokkyo Medical University, Mibu, Tochigi, Japan); Shinji Koba, MD, PhD (Department of Medicine, Division of Cardiology, Showa University School of Medicine, Tokyo, Japan); Kentaro Watanabe, MD, PhD (Department of Neurology, Hematology, Metaboism, Endocrinology and Diabetology (DNHMED), Yamagata University School of Medicine, Yamagata, Japan); Yasuhiko Takemoto, MD, PhD (Department of Internal Medicine and Cardiology, Osaka City University Graduate School of Medicine, Osaka, Japan); Takuzo Hano, MD, PhD (Department of Medical Education and Population-based Medicine, Postgraduate School of Medicine, Wakayama Medical University, Wakayama, Japan); Masataka Sata, MD, PhD (Department of Cardiovascular Medicine, Institute of Health Biosciences, The University of Tokushima Graduate School, Tokushima, Japan); Yutaka Ishibashi, MD, PhD (Department of General Medicine, Shimane University Faculty of Medicine, Izumo, Japan); Koichi Node, MD, PhD (Department of Cardiovascular and Renal Medicine, Saga University, Saga, Japan); Koji Maemura, MD, PhD (Department of Cardiovascular Medicine, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki, Japan); Yusuke Ohya, MD, PhD (The Third Department of Internal Medicine, University of the Ryukyus, Okinawa, Japan); Taiji Furukawa, MD, PhD (Department of Internal Medicine, Teikyo University School of Medicine, Tokyo, Japan); Hiroshi Ito, MD, PhD (Department of Cardiovascular Medicine, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Japan); Hisao Ikeda, MD, PhD (Faculty of Fukuoka Medical Technology, Teikyo University, Omuta, Japan).

      Financial disclosure

      Grant-in-Aid for Scientific Research from the Ministry of Education,Science and Culture of Japan (18590815 and 21590898 to Y.H.) and a Grant in Aid of Japanese Arteriosclerosis Prevention Fund (to Y.H.).

      Appendix

      Table S1Clinical characteristics in propensity score matched subjects
      VariablesHigh 60–79 mg/dL (n = 451)Extremely high ≥80 mg/dL (n = 451)P value
      Age, y51.4 ± 1151.5 ± 11.892
      Body mass index, kg/m222.1 ± 2.622.0 ± 2.6.623
      Systolic blood pressure, mmHg129 ± 17129 ± 17.695
      Diastolic blood pressure, mmHg81 ± 1280 ± 12.239
      Heart rate, bpm63 ± 1163 ± 12.956
      Total cholesterol, mg/dL216 ± 32213 ± 30.138
      Triglycerides, mg/dL84 (64, 109)77 (58, 110).021
      HDL-C, mg/dL68 ± 691 ± 11<.001
      LDL-C, mg/dL129 ± 29105 ± 28<.001
      Glucose, mg/dL101 ± 20100 ± 17.500
      Medications, n (%)
       Anti-hypertensive therapy98 (21.7)104 (23.1).632
       Anti-hyperglycemic therapy19 (4.2)18 (4.0).867
      Framingham risk score, %4.0 ± 3.53.1 ± 3.4<.001
      Medical history, n (%)
       Hypertension200 (44.4)203 (45.0).841
       Dyslipidemia165 (36.6)86 (19.1)<.001
       Diabetes mellitus29 (6.4)24 (5.3).479
       Current smokers127 (28.2)131 (29.1).768
       Previous cardiovascular disease12 (2.7)16 (3.6).442
      HDL-C indicates high-density lipoprotein cholesterol; LDL-C, low-density lipoprotein cholesterol.
      Data are presented as mean ± SD or median (interquartile range).
      Figure thumbnail figs1
      Figure S1Scatter plot shows the relationship between flow-mediated vasodilation (FMD) and high-density lipoprotein cholesterol (HDL-C). Red line represents the estimated Lowest smoothed curve.
      Figure thumbnail figs2
      Figure S2Bar graphs show flow-mediated vasodilation (FMD) in high levels of high-density lipoprotein cholesterol (HDL-C) and in extreme high levels of HDL-C in propensity score matched subjects. The error bars indicate the standard deviation.

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