Beneficial Effects of Fenofibrate to Improve Endothelial Dysfunction and Raise Adiponectin Levels in Patients With Primary Hypertriglyceridemia

RESEARCH DESIGN AND METHODS

This study was randomized, double blind, placebo controlled, and crossover in design. None had diabetes, were smokers, or had previous angina. We excluded patients with moderate or severe hypertension, nephrotic syndrome, hypothyroidism, coronary artery disease, or peripheral vascular disease. No patient had taken any cholesterol-lowering agent, hormone replacement therapy, or antioxidant vitamin supplements during the preceding 2 months. We administered placebo or micronized fenofibrate 200 mg daily to 46 patients with primary hypertriglyceridemia (>150 mg/dl) for 8 weeks, with the second treatment period initiated upon completion of the first treatment period (without washout phase). A research nurse counted pills at the end of treatment to monitor compliance. Baseline total cholesterol, triglyceride, LDL cholesterol, HDL cholesterol, non-HDL cho-lesterol, apolipoprotein B, and apolipoprotein A-I levels were 236 ± 8, 293 ± 15, 133 ± 8, 44 ± 1, 192 ± 7, 132 ± 4, and 153 ± 3 mg/dl, respectively. The mean age was 57 ± 1 years, and 18 (39%) were men. Mean BMI was 25.78 ± 0.43 kg/m². Ten of 46 patients were mildly hypertensive.

No additional medications, including antihypertensive drugs, aspirin, or nonsteroidal antiinflammatory drugs, were allowed during the study period to avoid other drugs’ effects. Twenty-four of 46 patients had metabolic syndrome according to the definition of National Cholesterol Education Program Adult Treatment Panel III (7). The study was approved by the Gil Hospital Institute Review Board, and all participants gave written informed consent.

Laboratory assays

Blood samples for laboratory assays were obtained at ∼8:00 a.m. following an overnight fast before and at the end of each treatment period. Placebo or fenofibrate was taken at ∼7:00 p.m. following meals. These samples were immediately coded so that investigators performing laboratory assays were blinded to subject identity or study sequence. Assays for lipids, glucose, and plasma adiponectin were performed in duplicate by enzyme-linked immunosorbent assay (R&D Systems, Minneapolis, MN) and assays for high-sensitivity C-reactive protein (hsCRP) levels by latex agglutination (CRP-Latex (II); Denka-Seiken, Tokyo, Japan). Assays for plasma insulin levels were performed in duplicate by immunoradiometric assay (Insulin-Riabead II; Abbott Japan, Tokyo, Japan). All samples from the same patient (batch samples) were measured in blinded pairs on the same enzyme-linked immunosorbent assay kit to minimize run-to-run variability. The interassay and intra-assay coefficients of variation (CVs) were <6%. Quantitative insulin sensitivity check index (QUICKI), a surrogate index of insulin sensitivity, was calculated as follows (insulin is expressed in μU/ml and glucose in mg/dl): QUICKI = 1/[log(insulin) + log(glucose)] (8).

Vascular studies

Imaging studies of the right brachial artery were performed using an ATL HDI 3000 ultrasound machine (Bothell, WA) equipped with a 10-MHz linear-array transducer and lower-arm occlusion technique, based on a previously published technique (9,10). All images were transmitted to a personal computer via ethernet with DICOM (digital imaging and communication in medicine) format and then saved on the hard disk of a personal computer in bitmap format. Arterial diameters were measured with Image Tool for Windows version 2.0 (University of Texas Health Science Center, San Antonio, TX). Measurements were performed by investigators blinded to the subject’s identity and medication status. Measurements of maximum diameter and percent flow-mediated dilation were made in 10 studies selected at random. The inter- and intraobserver variability for repeated measurement of maximum diameter were 0.01 ± 0.06 and 0.008 ± 0.05 mm, respectively. The inter- and intraobserver variability for repeated measurement of percent flow-mediated dilation were 0.12 ± 1.31 and 0.10 ± 1.29%, respectively.

Statistical analysis

Data are expressed as means ± SE or median (range 25–75%). After testing data for normality, we used Student’s paired t or Wilcoxon signed rank test to compare the relative changes in values in response to placebo and fenofibrate treatments, as reported in Tables 1 and 2. We did not incorporate a washout period between placebo and fenofibrate treatments based on our previous study (9). However, to assess the possibility of a carryover effect from the initial treatment periods to the next treatment period, we compared the percent changes of 1) the first treatment placebo and the second treatment placebo with 2) the first treatment fenofibrate and the second treatment fenofibrate relative to baseline values by using Student’s unpaired t or Mann-Whitney rank sum test. No significant differences were found in the above two comparisons. In other words, there were no detectable carryover effects. Pearson or Spearman correlation coefficient analysis was used to assess associations between measured parameters. We calculated that 30 subjects would provide 80% power for detecting an absolute increase of ≥1.5% in flow-mediated dilation of the brachial artery between placebo and fenofibrate, with α = 0.05 based on our previous studies (9–11). The comparison of endothelium-dependent dilation between placebo and fenofibrate was prospectively designated as the primary end point of the study. All other comparisons were considered secondary. A value of P < 0.05 was considered to be statistically significant.

RESULTS

Effects of therapies on lipids and vasomotor function

There were no significant differences in BMI between placebo and fenofibrate, and BMI was unchanged by either therapy. Compared with placebo, fenofibrate significantly changed lipoprotein levels. As expected, fenofibrate decreased total cholesterol, non-HDL cholesterol, apolipoprotein B, and triglycerides and increased HDL cholesterol and apolipoprotein A-I (all P < 0.001; Table 1) and tended to decrease LDL cholesterol (P = 0.069). Fenofibrate significantly improved the percent flow-mediated dilator response to hyperemia by 48 ± 5% (P < 0.001; Fig. 1, Table 1). However, the brachial artery dilator response to nitroglycerin was not significantly changed (P = 0.169). Of interest, there was a siginificant inverse correlation between the flow-mediated dilation following placebo and the percent change in flow-mediated dilation following fenofibrate (r = −0.745, P < 0.0001). Following fenofibrate therapy, improvement in flow-mediated dilation inversely correlated with changes in total cholesterol levels (r = −0.322 and P = 0.029), LDL cholesterol levels (r = −0.302 and P = 0.041), and non-HDL cholesterol levels (r = −0.389, P = 0.008). However, changes in flow-mediated dilation were not significantly correlated with changes in triglyceride levels (r = −0.181 and P = 0.229) and HDL cholesterol levels (r = 0.244, P = 0.102).

Effects of therapies on adiponectin and insulin resistance

There were inverse correlations between BMI and baseline plasma adiponectin levels (r = −0.283, P = 0.057). There were significant correlations between baseline adiponectin levels and baseline triglycerides (r = −0.331, P = 0.025), HDL cholesterol (r = 0.339, P = 0.021), insulin (r = −0.337, P = 0.022), or QUICKI (r = 0.391, P = 0.007).

Compared with placebo, fenofibrate therapy significantly increased plasma levels of adiponectin by 14 ± 5% (P = 0.008; Fig. 1). Fenofibrate therapy did not significantly change fasting insulin and glucose levels. However, compared with placebo, fenofibrate therapy significantly increased QUICKI by 6 ± 2% (P = 0.048). There were correlations between percent changes in adiponectin levels and percent changes in flow-mediated dilation (r = 0.401, P = 0.006), hsCRP (r = −0.443, P = 0.002), QUICKI (r = 0.292, P = 0.049), or insulin (r = −0.273, P = 0.067). The relationship between changes in flow-mediated dilation and changes in adiponectin levels was also investigated in a multivariate setting with other predictors (glucose, HDL cholesterol, triglyc-erides, and BMI). Only changes in adiponectin levels persisted as an independent predictor of changes in flow-mediated dilation (r = 0.504, P = 0.013).

Effects of therapies on hsCRP and fibrinogen

Compared with placebo, fenofibrate significantly lowered plasma levels of hsCRP from 0.80 to 0.70 mg/l (P = 0.001) and fibrinogen levels by 16 ± 3% (P < 0.001).

Effects of therapies in patients with metabolic syndrome

We analyzed 24 patients with metabolic syndrome, as reported in Table 2. Overall, compared with the effects of fenofibrate in 46 hypertriglyceridemic patients, we observed similar results in 24 patients with metabolic syndrome. Compared with placebo, fenofibrate significantly changed lipoprotein and fibrinogen levels and significantly improved the percent flow-mediated dilator response to hyperemia by 47 ± 6% (P < 0.001; Table 2). Compared with placebo, fenofibrate therapy significantly increased plasma levels of adiponectin by 12 ± 6% (P = 0.021). Fenofibrate therapy did not significantly change fasting insulin and glucose levels. However, compared with placebo, fenofibrate therapy significantly increased QUICKI by 6 ± 3% (P = 0.026). Fenofibrate therapy did not significantly reduce hsCRP levels (P = 0.092).

CONCLUSIONS

We observed that fenofibrate therapy significantly improved the percent flow-mediated dilator response to hyperemia, reduced levels of inflammation markers, increased adiponectin levels, and improved insulin sensitivity in hypertriglyceridemic patients.

Several studies (12,13) have examined the effect of fibrates on vasomotor function. In the current study, we observed that fenofibrate significantly improved the percent flow-mediated dilator response to hyperemia consistent with other studies (12,13). Plausible mechanisms underlying improvement of flow-mediated dilation following fenofibrate include changes of lipoproteins. In support of this speculation, we observed significant correlations between the improvement in flow-mediated dilation and the changes in total cholesterol, LDL cholesterol, and non-HDL cholesterol levels. Niacin treatment increases HDL cholesterol levels, improves flow-mediated dilation, and increases endothelial NO synthase expression in patients, suggesting that HDL-mediated increases in endothelial NO synthase expression and activity may contribute to the observed enhancement in vasorelaxation (14). In the current study, we observed that fenofibrate treatment increased HDL cholesterol levels and improved flow-mediated dilation; however, we did not observe a significant correlation between the improvement in flow-mediated dilation and the changes in HDL cholesterol. We also observed that fenofibrate therapy caused a small, but statistically significant, decrease in hsCRP levels, consistent with the findings of others (12). When evaluating the physiological relevance of this finding, it may be important to consider that our subjects were patients with primary hypertriglyceridemia without coronary artery disease or other risk factors who had relatively low pretreatment hsCRP levels (median 0.8 mg/l), and, thus, the reduction of absolute levels of hsCRP by fenofibrate, while statistically significant, is small. Moreover, it is important to note that in our study, multiple independent markers (including triglyceride levels, adiponectin levels, and QUICKI) all changed in a manner that is consistent with improved insulin sensitivity. This reinforces and supports our hypothesis that these factors are linked by pathophysiological mechanisms. Nevertheless, conclusive demonstration of the clinical relevance of these small, but statistically significant, changes will require a prospectively designed large-scale clinical trial that is beyond the scope of the present study.

Fenofibrate therapy resulted in significant elevation of adiponectin levels and increased insulin sensitivity (assessed by QUICKI). QUICKI is a reliable surrogate index for insulin sensitivity that has an especially excellent correlation with the reference standard glucose clamp method in insulin-resistant subjects with type 2 diabetes or obesity (r = ∼0.9 in subjects with these diseases) (8). In addition, test characteristics of QUICKI including CV (CV = 0.05) and discriminant ratio are significantly better than other simple surrogate indexes and comparable with those of the glucose clamp (15). In a number of relevant clinical conditions, including type 2 diabetes, gestational diabetes, and hypertension, QUICKI can appropriately follow changes in insulin sensitivity after various therapeutic interventions when compared directly with glucose clamp results (15,16). Moreover, a large meta-analysis of insulin-resistant subjects demonstrated that QUICKI is among the best surrogate indexes in terms of predictive power for the onset of diabetes (17). Of particular relevance to this study, decreasing values for QUICKI are also a reliable predictor of increased carotid intima-media thickness (18). Since changes in insulin sensitivity were a secondary outcome for this study, our data on improved insulin sensitivity should be regarded as pilot data for a more focused study on insulin sensitivity.

The present study is the first report demonstrating that fenofibrate therapy increases adiponectin levels. Adiponectin is an adipose-derived factor that augments and mimics metabolic actions of insulin. Adiponectin can directly stimulate NO production from endothelium via activation of AMP-activated protein kinase (6). Therefore, increasing adiponectin levels would be predicted to improve both insulin sensitivity and endothelial function by multiple mechanisms. Regulation of metabolic homeostasis and hemodynamic homeostasis may be coupled by vascular actions of insulin to stimulate production of NO (6). Thus, improvements in endothelial function may increase insulin sensitivity, while increased insulin sensitivity may improve endothelial function (3). On the other hand, there may be additional mechanisms for fenofibrate therapy to improve insulin sensitivity that are independent of endothelial function. PPARα activators improve insulin sensitivity and reduce adiposity in rodent models (19). Effects of fenofibrate therapy to increase adiponectin levels may mediate, in part, improved insulin sensitivity supported by significant correlations shown in the present study. Fenofibrate therapy for 2 months’ treatment increased adiponectin levels without a change in body weight. This raises the possibility that drug therapy is directly altering adiponectin levels independent of adiposity. Thus, it is possible that increased adiponectin levels are contributing to improvement in insulin sensitivity rather than simply reflecting a change in adiposity.

Metabolic syndrome is associated with atherosclerotic disease. Obesity is one of the most important contributors to cardiovascular disease. Adipose tissue secretes various bioactive molecules that may directly contribute to the development of obesity-related diseases (20). Dysregulation of adipocyte-derived endocrine factors caused by overnutrition may directly participate in the development of atherosclerosis. Adiponectin may couple regulation of insulin sensitivity with energy metabolism, serve to link obesity with insulin resistance, and possess antiatherogenic properties. In the present study, more than half (57%) of subjects were overweight. Furthermore, compared with the effects of fenofibrate in 46 hypertriglyceridemic patients, we observed similar results in a subgroup of 24 patients with the metabolic syndrome. Thus, our study has implications for the treatment of patients with the metabolic syndrome. Of interest, we observed significant correlations between the degree of changes in adiponectin levels and flow-mediated dilation, insulin levels, QUICKI, or hsCRP levels following fenofibrate therapy. Thus, our findings are consistent with those of Ouchi et al. (21), who demonstrated that plasma hsCRP levels were negatively correlated with plasma adiponectin levels in male coronary artery disease patients. Our multivariate regression analysis demonstrated that only changes in adiponectin levels persisted as an independent predictor of changes in flow-mediated dilation (r = 0.504, P = 0.013).

In conclusion, fenofibrate therapy significantly improved percent flow-mediated dilator response to hyperemia, reduced levels of inflammation markers, increased adiponectin levels, and improved insulin sensitivity in hypertriglyceridemic patients with or without metabolic syndrome. The effects of fenofibrate treatment to improve endothelial function and insulin sensitivity and to suppress inflammatory markers relevant to cardiovascular disease are likely to have important beneficial health consequences in patients with hypertriglyceridemia or the metabolic syndrome.