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Original Research Communication |
1 From the Neuroendocrine Unit (CH, JR, GM, NA, and SG) and the Program in Nutritional Metabolism (CH, GM, NA, and SG), Massachusetts General Hospital, Boston; the Clinical Research Center, Massachusetts Institute of Technology, Cambridge (JB); and Harvard Medical School, Boston (CH, GM, and SG).
2 Supported in part by NIH grants K23-DK02844, R01-DK59535, M01-RR300088, and P30 DK40561. 3 Reprints not available. Address correspondence to C Hadigan, Massachusetts General Hospital, Neuroendocrine Unit, BUL 457B, 55 Fruit Street, Boston, MA 02114. E-mail: chadigan{at}partners.org.
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ABSTRACT |
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 TOP ABSTRACT INTRODUCTIONSUBJECTS AND METHODSRESULTSDISCUSSIONREFERENCES |
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Objective: We determined the effects of acute inhibition of lipolysis on insulin sensitivity in HIV-infected patients with fat redistribution who were receiving a protease inhibitor.
Design: Seven HIV-infected men [age: 45 ± 2 y; body mass index (in kg/m2): 28.8 ± 1.9] with a fasting insulin concentration 
104 pmol/L (15 µIU/mL), combined visceral adiposity and peripheral lipoatrophy, and receiving a protease inhibitor were studied. Tolbutamide-modified frequently sampled intravenous-glucose-tolerance tests (FSIGTTs) were performed after randomized double-blind administration of acipimox (500 mg at -90 and 0 min), a potent inhibitor of lipolysis, and placebo. The subjects completed 2 FSIGTTs separated by 3–7 d.
Results: At baseline, fasting insulin and fatty acid concentrations were 27.6 ± 5.0 µIU/mL and 0.83 ± 0.08 mmol/L (normal range: 0.1–0.6 mmol/L), respectively. Fatty acid concentrations were significantly reduced after acipimox compared with placebo (fatty acid area under the curve: acipimox = 73 ± 8 compared with placebo = 122 ± 12 mmol · 270 min/L, P = 0.002). Acipimox treatment resulted in a significant increase in the insulin sensitivity index (acipimox = 1.63 ± 0.5 compared with placebo = 0.88 ± 0.3 x 10-4 · min-1 · µIU/mL, P = 0.015).
Conclusions: Acute inhibition of lipolysis and reduction in fatty acid concentrations are associated with improved insulin sensitivity in patients with HIV lipodystrophy and hyperinsulinemia. Further studies are needed to determine whether long-term antilipolytic strategies to reduce fatty acid concentrations may be useful in treating the metabolic disturbances associated with HIV lipodystrophy.
Key Words: Lipodystrophy • insulin resistance • fatty acids • acipimox • HIV • men
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONSUBJECTS AND METHODSRESULTSDISCUSSIONREFERENCES |
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A potential mechanism by which increased visceral adiposity and subcutaneous fat loss may result in insulin resistance is through increased concentrations of circulating fatty acids, which in turn may contribute to hepatic insulin resistance and decreased insulin signaling through insulin receptor substrate 1 (6). For example, among non-HIV-infected patients with congenital lipodystrophy and in animal models of lipoatrophy (A-ZIP/F-1 mice), fatty acid concentrations are elevated and are thought to contribute to insulin resistance (7, 8). Increased fatty acid concentrations were shown in HIV-infected patients with fat redistribution and insulin resistance (9, 10). Furthermore, we recently showed that fatty acid concentrations are a strong predictor of insulin response to standard oral glucose challenge in HIV-infected patients with lipodystrophy, independent of the effects of age, body mass index (BMI), sex, and body composition (10). To test the hypothesis that increased circulating fatty acids contribute to insulin resistance in patients with HIV infection and lipodystrophy, we determined the effect of acute inhibition of lipolysis and fatty acid reduction on insulin sensitivity in HIV-infected men with fat redistribution and hyperinsulinemia. Our data, showing significant improvement in insulin sensitivity after an acute lipolytic blockade, suggest the importance of long-term studies of therapies to decrease lipolysis and improve insulin resistance in this population.
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SUBJECTS AND METHODS |
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TOPABSTRACTINTRODUCTIONSUBJECTS AND METHODSRESULTSDISCUSSIONREFERENCES |
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3 mo before the study, hyperinsulinemia (fasting insulin 15 µIU/mL), increased waist-to-hip ratio (> 0.95), and evidence of fat redistribution. Fat redistribution was scored by a single investigator (CH) on the basis of evidence of fat accumulation in the trunk, breast, or neck and loss of fat in the face or extremities on physical examination. Subjects were excluded if they had a history of significant kidney or liver disease; < 110 g hemoglobin/L; a history of diabetes mellitus or > 7.0 mmol fasting glucose/L (126 mg/dL); concurrent therapy with insulin, antidiabetic agents, glucocorticoids, testosterone, growth hormone, estrogen, or anabolic steroids; concurrent substance abuse; or an opportunistic infection within the past 6 wk. All subjects gave written informed consent, and the study was approved by the Human Research Committee of the Massachusetts General Hospital.
Protocol
After eligibility was established, the subjects returned for 2 test days separated by 3–7 d. Each day the subjects reported to the Clinical Research Center after a 12-h overnight fast and completed a frequently sampled intravenous-glucose-tolerance test (FSIGTT; 11). One day the subjects received acipimox (500 mg orally; Olbetam Pharmacia, Milan, Italy) at -90 min and again at 0 min during the FSIGTT; on the other day the subjects received an identical placebo at the same time points. Two doses were administered 90 min apart to provide maximal lipolytic blockade for the duration of the FSIGTT. The dosing interval was based on the pharmacologic half-life of acipimox. The order of acipimox and placebo was randomized, and both the investigators and subjects were blinded to the order of drug assignment. Randomization was performed by the Massachusetts General Hospital pharmacy.
The tolbutamide-modified FSIGTT as developed by Bergman et al (11) was used. The FSIGTT provides a precise measure of insulin sensitivity and was shown to correlate well with euglycemic insulin clamp studies in populations with insulin resistance (12, 13). A bolus of 25% glucose solution (0.5 g/kg, with a maximal dose of 35 g) was administered over 1–3 min at the start of the study. Intravenous tolbutamide was given 20 min after glucose administration at the following doses: subjects with a BMI (in kg/m2) < 30 received 300 mg, and subjects with a BMI 30 received 500 mg. Venous blood samples were obtained at -15, -10, -5, -1, 2, 3, 4, 5, 6, 8, 10, 12, 14, 16, 19, 22, 24, 25, 27, 30, 40, 50, 60, 70, 90, 100, 120, 140, and 180 min for the determination of plasma glucose and insulin concentrations. Insulin sensitivity and acute insulin response to glucose were calculated from these values with the nonlinear mathematical model of glucose disappearance (11). Fatty acid concentrations were determined at -90 min and every 30 min thereafter for the remainder of the testing period.
Cross-sectional abdominal computed tomography scans were performed as described by Borkan et al (14) to assess the distribution of subcutaneous and visceral adipose tissue. A lateral scout image was obtained to identify the level of the L4 pedicle, which served as the landmark for the 1-cm single-slice image. The subjects also underwent total-body dual-energy X-ray absorptiometry with the Hologic QDR-4500A scanner (Hologic Inc, Bedford, MA). The DXA scan was used to determine total-body fat and lean mass as well as regional body fat measurements, such as percentage trunk fat (trunk fat/total trunk mass) and percentage extremity fat (extremity fat/total extremity mass).
Biochemical and immunologic assays
Concentrations of glucose, cholesterol, triacylglycerol, HDL, and LDL were determined by methods previously reported (2). Nonesterified fatty acid concentrations were measured by using an in vitro enzymatic colorimetric assay kit (Wako Chemicals USA, Inc, Richmond, VA). The intraassay CV for fatty acids ranged from 1.1% to 2.7%. The published normal range for fatty acids is 0.1–0.6 mmol/L. Insulin concentrations were measured in serum by radioimmunoassay (Diagnostic Product Corp, Los Angeles). The intraassay and interassay CVs ranged from 4.7% to 7.7% and from 5.5% to 9.2%, respectively. The CD4+ count was determined by flow cytometry (Becton Dickinson Immunocytochemistry Systems, San Jose, CA), and the HIV viral load was determined by ultrasensitive assay (Amplicor HIV-1 Monitor Assay; Roche Molecular Systems, Branchburg, NJ), with limits of detection of 50–75 000 copies/mL.
Statistical analyses
Univariate clinical characteristics are presented as means ± SEMs. The primary outcome variables were insulin sensitivity index (SI) and fatty acid area under the curve during an FSIGTT. The SI was calculated by using MINIMAL MODEL IDENTIFICATION (version 3.0, developed by Richard Bergman, 1994). The effect of acipimox on these outcome variables was determined by using paired t tests, comparing results after acipimox to placebo for each subject. Confirmatory analyses were performed with a nonparametric Wilcoxon rank-sum test. Simple linear regression was used to assess the relation between continuous variables (eg, SI and fatty acid area under the curve). All statistical analyses were performed with SAS JMP (version 3.2.2; SAS Institute, Inc, Cary, NC), and statistical significance was defined as a two-tailed 
value of P < 0.05.
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RESULTS |
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TOPABSTRACTINTRODUCTIONSUBJECTS AND METHODSRESULTSDISCUSSIONREFERENCES |
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. Each subject was receiving a protease inhibitor and and at least one nucleoside reverse transcriptase inhibitor; only one patient was receiving a nonnucleoside reverse transcriptase inhibitor. The subjects showed significant central adiposity (waist-to-hip ratio: 0.99 ± 0.01) as well as extremity fat loss, which was rated as moderate to severe in all cases. The basal fasting insulin concentration was 27.6 ± 5.0 µIU/mL, and the baseline fatty acid concentration was increased above the normal range for the assay (0.83 ± 0.08 mmol/L; normal range: 0.1–0.6 mmol/L).
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). The fatty acid area under the curve between -90 min and 180 min was 122 ± 12 mmol · 270 min/L after placebo but 73 ± 8 mmol · 270 min/L after acipimox (P = 0.002). Insulin sensitivity was significantly increased after administration of acipimox compared with placebo (Figure 2). The SI was 0.88 ± 0.3 x 10-4 · min-1 · µIU/mL with placebo and increased significantly after fatty acid suppression with acipimox (1.63 ± 0.5 x 10-4 · min-1 · µIU/mL, P = 0.015). Results were similar with nonparametric testing. The mean change in the SI after administration of acipimox was 0.74 ± 0.2 x 10-4 · min-1 · µIU/mL, and all but one subject showed an increase in the SI with acipimox compared with placebo. Acute insulin response to glucose decreased, but not significantly, after administration of acipimox (acute insulin response to glucose with placebo = 975 ± 178 and with acipimox = 853 ± 101 µIU/mL · min). The fatty acid area under the curve after acipimox administration was inversely correlated with insulin sensitivity (r = -0.75, P = 0.05).
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30 min after receiving acipimox, but this resolved spontaneously 1 h. Another subject reported developing a transient rash 3 d after receiving acipimox that resolved spontaneously in 1 d and was not believed to be related to the medication. |
DISCUSSION |
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TOPABSTRACTINTRODUCTIONSUBJECTS AND METHODSRESULTSDISCUSSIONREFERENCES |
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Our data suggest that HIV-infected patients with significant visceral adiposity and peripheral subcutaneous fat loss, selected on the basis of fasting hyperinsulinemia, show increased fatty acid concentrations. To test the hypothesis that increased lipolysis and fatty acids contribute to insulin resistance among HIV-infected patients, we investigated the effects of lipolytic blockade in this population. Treatment with acipimox, a potent nicotinic acid analogue, resulted in decreased fatty acid concentrations in our study population. Acipimox is widely used as a lipid-lowering therapy for the treatment of hypercholesterolemia and hypertriglyceridemia, and the presumed mechanism of action is through reduction in release of fatty acids and decreased VLDL cholesterol.
The subjects in the present study, all of whom showed evidence of fat redistribution and who were receiving a protease inhibitor, were highly insulin resistant, with a reduced basal SI (SI = 0.88). With the use of FSIGTT, Kosmiski et al (18) previously showed a similarly low SI (SI = 1.00) in 14 protease inhibitor–treated HIV-infected patients with lipodystrophy. In the present study, acute inhibition of lipolysis resulted in an almost 2-fold increase in insulin sensitivity in HIV-infected patients with fat redistribution and hyperinsulinemia. Importantly, fatty acid concentrations after acipimox administration were inversely associated with insulin sensitivity during FSIGTT, suggesting that the significant increase in insulin sensitivity after acipimox is indeed an effect of inhibition of lipolysis. However, despite a significant increase, the SI after acipimox remained low (1.63 ± 0.05). The persistent low insulin sensitivity may be caused by several factors in this population. All the subjects were required to be receiving a protease inhibitor–containing regimen for 3 mo before testing. Recent data suggest that protease inhibitors acutely inhibit glucose transport at physiologic concentrations (15, 19) and therefore may contribute to the low insulin sensitivity in our subjects after acipimox. The potential effect of acipimox in the absence of a protease inhibitor is not known and will be important to investigate in future studies. In addition, the subjects received acute, 1-d dosing of acipimox. Chronic inhibition of lipolysis may result in a greater improvement in insulin sensitivity. Finally, increased intramyocellular lipid content was shown in HIV-infected patients with lipodystrophy (20, 21) and may contribute to insulin resistance in these patients.
In non-HIV-infected subjects with obesity, the administration of acipimox reduces fasting fatty acid concentrations and improves hyperinsulinemia (22, 23). Acute inhibition of lipolysis was also shown to improve insulin sensitivity in obese and lean patients with or without type II diabetes mellitus (24). In comparison, subjects in the present study were not simply obese but had a pattern of mixed lipodystrophy with increased visceral fat and decreased subcutaneous fat. Our data, derived by using acipimox as an acute physiologic probe, suggest that increased fatty acids may contribute to severe insulin resistance in nondiabetic HIV-infected patients with fasting hyperinsulinemia and fat redistribution. Despite the significant improvement in insulin sensitivity, the SI did not normalize, suggesting that additional factors also mediate insulin resistance in patients with HIV infection and fat redistribution.
Increased central adiposity and reduced subcutaneous fat may play an important role in the development of insulin resistance in patients with HIV lipodystrophy. Increased visceral adiposity may contribute to insulin resistance through the production of increased fatty acids, which may lead to increased hepatic insulin resistance and hepatic gluconeogenesis. Among obese non-HIV-infected subjects, increased endogenous concentrations of fatty acids were shown to correlate significantly with and contribute to hyperinsulinemia (25, 26). Further, Boden et al (27) showed that administration of fatty acids results in both decreased glucose uptake in peripheral muscle and inhibition of the usual suppressive effect of insulin on hepatic glucose production. Griffin et al (6) hypothesized that increased plasma fatty acid concentrations induce skeletal muscle insulin resistance by inhibition of insulin signaling through insulin receptor substrate 1. Increased fatty acid concentrations are also seen in patients with congenital lipoatrophy in combination with severe insulin resistance (7). Further, Gavrilova et al (8) showed that fatty acid concentrations and insulin resistance improved with implantation of subcutaneous fat depots in the A-ZIP/F-1 fatless mouse.
The present study shows the effect of lipolytic blockade in HIV-infected patients with fat redistribution and helps to elucidate the role of fatty acids in insulin resistance in protease inhibitor–treated patients with central adiposity. These data suggest a potential approach to breaking the metabolic cycle in these patients in whom fat redistribution results in insulin resistance and leads to worsening glucose homeostasis in part through increased fatty acid concentrations. Although it remains uncertain whether long-term administration of a lipolytic blocking agent would be beneficial to the larger population of HIV-infected patients with lipodystrophy, data from our acute study with acipimox suggest a potential role for adipogenic, antilipolytic agents in the long-term treatment of patients with HIV lipodystrophy. For example, the thiazolidinediones activate peroxisome proliferator activated receptor g and stimulate adipogenesis, resulting in reduced lipolysis (7). Our data provide a rationale for the investigation of the thiazolidinediones to improve fat redistribution and insulin resistance in HIV-infected patients, in whom acute reduction in lipolysis significantly improves insulin sensitivity.
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ACKNOWLEDGMENTS |
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REFERENCES |
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release from abdominal subcutaneous adipose tissue in HIV-infected subjects with lipodystrophy. Abstracts of the 3rd International Workshop on Adverse Drug Reactions and Lipodystrophy in HIV. 23-26 October 2001. Athens, Greece. Antivir Ther 2001;6 (suppl):16 (abstr).
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) and acipimox (
; 500 mg/dose). The fatty acid area under the curve was significantly different after acipimox (73 ± 8 mmol · 270 min/L) than after placebo (122 ± 12 mmol · 270 min/L), P = 0.002 (paired t test). n = 7.