Differential Effects of Metformin and Exercise on Muscle Adiposity and ...

0021-972X/04/$15.00/0 Printed in U.S.A.

The Journal of Clinical Endocrinology & Metabolism 89(5):2171–2178 Copyright © 2004 by The Endocrine Society doi: 10.1210/jc.2003-031858

Differential Effects of Metformin and Exercise on Muscle Adiposity and Metabolic Indices in Human Immunodeficiency Virus-Infected Patients SUSAN D. DRISCOLL, GARY E. MEININGER, KARIN LJUNGQUIST, COLLEEN HADIGAN, MARTIN TORRIANI, ANNE KLIBANSKI, WALTER R. FRONTERA, AND STEVEN GRINSPOON Program in Nutritional Metabolism (S.D.D., G.E.M., K.L., C.H., S.G.), Neuroendocrine Unit (S.D.D., G.E.M., K.L., C.H., A.K., S.G.), and Division of Musculoskeletal Radiology (M.T.), Massachusetts General Hospital, and Department of Physical Medicine and Rehabilitation (W.R.F.), Spaulding Rehabilitation Hospital and Harvard Medical School, Boston, Massachusetts 02114 The HIV-lipodystrophy syndrome is associated with fat redistribution and metabolic abnormalities, including insulin resistance (IR). The mechanisms and treatment strategies for IR in HIV-lipodystrophy are unclear, but data suggest that intramuscular lipids contribute to IR in this population. We previously showed that metformin and exercise improve hyperinsulinemia more than metformin alone in HIV-lipodystrophy. Now we investigate the effects of these treatment strategies on thigh muscle adiposity measured by computed tomography and additional body composition measures. Twenty-five HIV-infected patients on stable antiretroviral therapy with hyperinsulinemia and fat redistribution participated in a prospective, randomized, 3-month study of metformin alone or metformin and resistance training three times a week. Thigh muscle adiposity decreased significantly more

H

IV-LIPODYSTROPHY IS A heterogeneous syndrome of fat redistribution, characterized by varying degrees of visceral abdominal fat accumulation, loss of sc fat, and metabolic abnormalities, including insulin resistance (1– 8). Increased abdominal fat and loss of sc fat correlate strongly with insulin resistance in HIV- and non-HIVinfected patients (9 –15). In addition to fat in the abdomen and sc compartments, intramuscular fat is an important determinant of insulin resistance in non-HIV-infected patients (16 –22). Recent studies have shown that higher intramyocellular lipid concentrations, measured by magnetic resonance spectroscopy, are strongly associated with insulin resistance in HIV-infected patients (23, 24). In addition, our group recently demonstrated lower muscle attenuation by computed tomography (CT) in patients with HIV-lipodystrophy, indicative of higher muscle adiposity (25). Muscle attenuation was found to be a strong independent predictor of hyperinsulinemia in this population, controlling for body mass index Abbreviations: BMI, Body mass index; CT, computed tomography; DEXA, dual-energy x-ray absorptiometry; FFA, free fatty acid; OGTT, oral glucose tolerance testing; PI, phosphatidyl inositol; WHR, waistto-hip ratio. JCEM is published monthly by The Endocrine Society (http://www. endo-society.org), the foremost professional society serving the endocrine community.

as shown by increased muscle attenuation [2.0 (range, 0.5–5.0) vs. –1.0 (–3.5– 0), P ⴝ 0.04] and sc leg fat tended to decrease more [–3.3 (–7.5– 4.3) vs. 0.8 (–2.1–9.5), P ⴝ 0.06] in the combined treatment group in comparison with metformin alone. In multivariate analysis, change in thigh muscle adiposity remained a significant predictor of change in insulin (P ⴝ 0.04), controlling for changes in other body composition measurements. These data suggest that muscle adiposity, in addition to other fat depots, is an important determinant of hyperinsulinemia and that exercise has complex effects on regional fat depots in HIV-infected patients. Reduction in muscle adiposity may be an important mechanism by which exercise improves hyperinsulinemia in this population. (J Clin Endocrinol Metab 89: 2171–2178, 2004)

(BMI) and abdominal visceral and abdominal sc fat in multivariate regression modeling (25). Additional metabolic indices, such as adiponectin and free fatty acids (FFAs), have also been associated with insulin resistance in HIV-infected patients. Reduced adipose tissue expression and circulating concentrations of adiponectin have been shown in patients with HIV-lipodystrophy in association with insulin resistance, decreased extremity fat, and increased trunk fat (26 –29). Increased plasma FFA concentrations are seen in HIV-infected patients with fat redistribution and also correlate with insulin resistance in this population (30, 31). In this study, we sought to determine the effects of metformin and exercise training on specific and detailed measures of body composition and to assess whether a change in muscle adiposity contributes to the beneficial effects of exercise training in HIV-infected patients with fat redistribution and insulin resistance. We previously showed that exercise training and metformin was more beneficial than metformin alone on measures of insulin (32) and have now analyzed the effects of these treatments on muscle attenuation and sc leg fat and more detailed metabolic indices, including adiponectin and FFA. Our data demonstrate that exercise in combination with metformin improves muscle attenuation more than metformin alone and suggest that a reduction in muscle adiposity is an important mechanistic factor and treatment target for insulin resistance in HIV-lipodystrophy.

2171

2172

J Clin Endocrinol Metab, May 2004, 89(5):2171–2178

Subjects and Methods Subjects From October 2000 to August 2002, 84 subjects were recruited through community advertisements and primary care provider referral. Eligible subjects were seen at the Clinical Research Centers of the Massachusetts General Hospital and the Massachusetts Institute of Technology. As previously described (32), eligible subjects were between 18 and 60 yr of age, HIV positive, on a stable antiretroviral regimen for more than 3 months, with fasting insulin greater than or equal to 15 ␮IU/ml (104 pmol/liter) or 2-h insulin greater than or equal to 75 ␮IU/ml (521 pmol/liter), and evidence of fat redistribution [truncal obesity with a waist-to-hip ratio (WHR) ⬎ 0.90 in men and ⬎0.85 in women and a lipodystrophy score ⬎ 2]. Lipodystrophy scores were calculated based on evaluation of the face, neck/shoulders, arms, abdomen, and hips/ legs with graded values between 0 and 2 for fat loss or accumulation, as previously described (32). Fasting and 2-h hyperinsulinemia were defined in advance based on the 90th percentile for healthy subjects ages 26 –50 yr old in the Framingham Offspring Study (Meigs, J., personal communication). Subjects were excluded from the study if they had been hospitalized because of new opportunistic infections within the 6 wk before enrollment. Subjects with a history of unstable angina, aortic stenosis, uncontrolled hypertension, severe neuropathy, arthritis, prior history of diabetes mellitus or fasting plasma glucose of greater than or equal to 126 mg/dl (6.99 mmol/liter), current substance abuse, or other contraindication to exercise were also excluded. In addition, subjects requiring parenteral nutrition, parenteral or oral glucocorticoid therapy (⬎7.5 mg daily), estrogen, progesterone derivatives, supraphysiological testosterone, or ketoconazole within 3 months of enrollment were excluded. Of the 84 subjects screened, 42 were ineligible, five declined to participate, 12 withdrew, and 25 completed the protocol. All subjects gave written informed consent, and the study was approved by the Human Research Committee at the Massachusetts General Hospital and the Committee on Use of Human Subjects at the Massachusetts Institute of Technology. Data on limited body composition parameters, strength, cardiovascular risk markers, and insulin and glucose responses to oral glucose tolerance testing (OGTT) were previously published (32). We subsequently analyzed the change from baseline in anterior thigh muscle attenuation and sc leg fat by CT in relationship to changes in insulin and other metabolic indices, including adiponectin and FFA.

Protocol At baseline, eligible subjects underwent an OGTT for insulin responses, HIV viral load, and CD4 count after a 12-h fast. Anthropometric measurements were obtained, including WHR. Single-slice crosssectional CT scans at mid-thigh and abdomen and a total-body dualenergy x-ray absorptiometry (DEXA) were performed. On completion of the 3-month intervention, subjects returned for testing identical to that at baseline. Eligible subjects were randomly assigned to one of two treatment groups (metformin alone or metformin and exercise) at baseline. Randomization was performed by the Biostatistics Center of the Massachusetts General Hospital Clinical Research Center, using a permuted block algorithm. The metformin-only treatment group received 500 mg metformin twice a day, with a dose increase to 850 mg twice a day after 2 wk, provided that resting lactic acid levels were within normal limits and no serious side effects were reported. Subjects randomized to exercise training received metformin, as described above, and also began an aerobic and resistance exercise training program. The exercise training session began with a 5-min warm-up on a stationary bicycle and a standard flexibility routine to minimize the risk of injury (33), followed by an aerobic component set for 20 min during the first 2 wk and 30 min thereafter. The intensity of the aerobic component was 60% of maximal heart rate during the first 2 wk and 75% thereafter. The aerobic training program followed the general guidelines established by the American College of Sports Medicine (34). On completion of the aerobic component, a resistance training program was performed, using constant external resistance Life Circuit equipment, alternating upper and lower body exercises in the following order: 1) hip extension, 2) lateral pull down, 3) knee extension, 4) elbow flexion, 5) knee flexion, and 6) chest press. Resistance training was based on the

Driscoll et al. • Muscle Adiposity and Insulin in HIV

progressive resistance exercise concept originally proposed by De Lorme and Watkins (35). Subjects performed three sets of 10 repetitions each for every muscle group, resting 2–3 sec between repetitions, 2 min between sets, and 4 min between muscle groups. The initial intensity of exercise was 60% of the one-repetition maximum (1 RM). After 2 wk, the relative intensity was increased to 70% of the 1 RM and after an additional 2 wk to 80% of the 1 RM. The 1 RM was measured every other week, and the absolute load was adjusted accordingly to maintain the relative intensity at 80% of the 1 RM. Each training session was supervised by a physical therapist from the Physical Therapy Services of the Massachusetts General Hospital and conducted at the Massachusetts General Hospital affiliated Charles River Park Health Club.

Experimental methods Body composition analysis. CT scans were performed with a LightSpeed CT scanner (General Electric, Milwaukee, WI). A lateral scout image of the abdomen was obtained to identify the L4 pedicle, which served as a landmark for a single-slice image at this level. Scan parameters for each image were standardized (144-cm table height, 80 kV, 70 mA, 2 sec, 1-cm slice thickness, and 48-cm field of view). The single cross-sectional CT image at L4 was used to assess distribution of sc and visceral abdominal fat. Fat attenuation values were set at ⫺50 to ⫺250 Hounsfield units as described by Borkan et al. (36) and intraabdominal visceral and sc fat areas were determined on the basis of tracings obtained using graphical analysis software (Alice; Parexel, Waltham, MA). A coronal scout image was used to prescribe an axial CT image of the left thigh. The image was obtained at midpoint between the articular surface of the femoral head and medial femoral condyle using 120 kV, 170 mA, 2 sec, 1-cm slice thickness, and 36-cm field of view. The sc tissue area and muscle attenuation were measured using graphical analysis software (Alice; Parexel). Manual tracings separated sc fat and anterior and posterior muscle compartments. Subfascial adipose tissue interspersed between the muscle was excluded from the determination of muscle area and muscle attenuation and included in the determination of the sc fat compartment. Muscle attenuation in Hounsfield units was determined in the anterior left thigh muscle compartment. The coefficient of variation for the measurement of mid-femur thigh muscle attenuation in our laboratory is 2.4%. Whole-body DEXA (DXA; QDR-4500 densitometer, Hologic, Waltham, MA) was performed to assess fat mass, with a precision error of 3% for fat (37). BMI was calculated from fasting weight and height measurements at baseline and 3 months. WHR was determined by fasting measurements of waist circumference at the umbilicus and hip circumference at the level of the iliac crest with the patient in an upright, standing position. Laboratory methods. Insulin levels were measured in serum samples using RIA (Diagnostic Products Corp., Los Angeles, CA). Intra- and interassay coefficients of variation ranged from 3.1–9.3% and 4.9 –10%, respectively. CD4 counts were measured by flow cytometry (FACS scan analyzer, Becton Dickinson and Co., San Jose, CA). 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. Nonesterified or FFA concentrations were measured using an in vitro enzymatic colorimetric assay kit (Wako Chemicals USA, Inc., Richmond, VA). The intraassay coefficient of variation for FFA ranged from 1.1– 2.7%. The published normal range for FFA is 0.1– 0.6 mmol/liter. Adiponectin levels were measured in serum samples using a RIA kit (LINCO Research, Inc., St. Charles, MO). Intraassay coefficients of variation were 3.59, 6.21, and 1.78% for samples that target a low (1.5 ␮g/ml), middle (3 ␮g/ml), and high (7.5 ␮g/ml) concentration of adiponectin, respectively. Interassay coefficients of variation were 9.25, 6.90, and 9.25% for low (1.5 ␮g/ml), middle (3 ␮g/ml), and high (7.5 ␮g/ml) concentration of adiponectin, respectively. Statistical analysis. Baseline characteristics were compared between randomization groups using the Wilcoxon’s rank-sum test and the Fisher’s exact test for noncontinuous variables. Median change from baseline between treatment groups was compared using the Wilcoxon’s rank-sum test. Simple linear regression analysis was used to compare changes from baseline in fasting and 2-h insulin levels with changes from baseline for various



measures of body composition. A best-fit multivariate regression analysis model was developed using various body composition measures, specifically WHR, DEXA trunk fat, CT visceral abdominal fat, CT sc leg fat, and CT anterior thigh muscle attenuation, to explain changes in insulin. Statistical analysis was performed using JMP Statistical Database Software (version 4, SAS Institute, Cary, NC). Statistical significance was defined as P ⱕ 0.05. All data are presented as median values with interquartile ranges, except where otherwise indicated.

Results

Thirty-seven patients were randomized, and 25 completed the protocol, 11 in the metformin and exercise group and 14 in the metformin-only group. Baseline data are shown in Table 1. Subjects were similar at baseline for age, gender, racial demographics, duration of HIV infection, CD4 count, HIV viral load, and medication use (Table 1). Measures of body composition by DEXA and CT scan were not significantly different between groups at baseline (Table 2). OGTT measures of insulin were not statistically different between treatment groups at baseline (Table 2). Eight patients in the metformin and exercise group vs. four in the metformin-only group withdrew from the study, as previously described (32). Four patients, two from each group, were withdrawn from the study because of minor elevations in resting lactic acid or aspartate aminotransferase above prespecified safety parameters. Two, one from each treatment group, were lost to follow-up. Six patients in the metformin and exercise group withdrew, three because of personal issues; one lived too far away, and two were unwilling to comply with exercise. Among subjects completing the protocol, the mean percentage compliance ⫾ sem with exercise sessions was 93 ⫾ 2%, time on the stationary bicycle was 97 ⫾ 2% of expected, and completion of weight repetitions was 97 ⫾ 1%. Metformin compliance determined by pill count was 99.1 ⫾ 1% (mean ⫾ sem) in the metformin and exercise training group and 93 ⫾ 5% in the metformin-only group. As previously reported, fasting insulin and insulin area under the curve improved significantly more and 2-h insulin tended to improve more in the metformin and exercise group in comparison with the metformin-only group (Table 2) (32). There was a statistically significant (P ⬍ 0.05) difference in the change from baseline in CT anterior thigh muscle attenuation between treatment groups (Fig. 1 and Table 2). Muscle

2173

attenuation increased to a greater extent in the combined treatment group, demonstrating reduced muscle adiposity in response to metformin and exercise. In contrast, muscle attenuation decreased in the metformin-only group, suggesting an increase in intramuscular adiposity (Fig. 1). In addition, there was an improvement in thigh muscle crosssectional area and a trend toward a greater reduction in CT sc leg fat in the metformin and exercise group in comparison with the metformin-only group (Table 2). WHR decreased significantly more in the metformin and exercise group in comparison with the metformin-only group (Table 2). Adiponectin tended to decrease more in the metformin and exercise group in comparison with the metformin-only group, and FFA concentrations did not change significantly between treatment groups (Table 2). In univariate regression analysis among all subjects, the change from baseline in fasting insulin correlated significantly with the change in DEXA trunk fat (Table 3). In addition, the change in 2-h insulin correlated positively with the change in DEXA trunk fat and negatively with the change in CT anterior thigh muscle attenuation (Table 3). Multivariate regression analysis was performed among all subjects with change in fasting and 2-h insulin as the dependent variables and change in WHR, CT anterior thigh muscle attenuation, CT sc leg fat, CT visceral abdominal fat, and DEXA trunk fat as independent variables (Table 3). Changes in sc leg fat, trunk fat, and thigh muscle attenuation were significant in the multivariate model for fasting insulin. The parameter estimates demonstrated that increased muscle attenuation (decreased adiposity) and reduced trunk fat were associated with improved fasting insulin, whereas reduced sc leg fat was associated with an increase in insulin (Table 3). Reduced trunk fat and reduced muscle adiposity were associated with reduced 2-h insulin (Table 3). The models explained a large percentage of the variability in the change in insulin in response to treatment (R2 ⫽ 0.84 for fasting insulin and R2 ⫽ 0.44 for 2-h insulin). The overall improvement in fasting insulin with exercise and metformin compared with metformin alone (Tables 2 and 3) occurred despite trends toward greater reductions in sc fat and adiponectin. Neither the addition of adiponectin nor FFA to the multivariate models changed the results (data not shown).

TABLE 1. Clinical characteristics of subjects at baseline

Age Gender (M/F) Race (Cauc./Hisp./AA/other) Duration HIV (months) CD4 count (no./mm3) HIV viral load (copies/ml) Current/past PI use (%) Months current/past PI use Current/past NRTI use (%) Months current/past NRTI use Current/past NNRTI use (%) Months current/past NNRTI use

Metformin only (n ⫽ 14)

Metformin and exercise (n ⫽ 11)

42 (34, 46) 12/2 11/2/1/0 99 (48, 161) 429 (295, 545) 38 (0, 11776) 50/36 23 (13, 42) 86/7 30 (24, 50) 36/7 15 (13, 48)

43 (36, 49) 8/3 7/1/2/1 122 (76, 151) 557 (375, 814) 25 (0, 587) 67/17 32 (10, 49) 92/0 48 (26, 81) 33/25 17 (10, 42)

Data are expressed as median with (25–75 percentile) interquartile range, number (n), or percentages, as indicated. Baseline comparisons were not significantly different (P ⬎ 0.05) by Wilcoxon or Fisher’s exact test. M/F, Male/female; Cauc., Caucasian; Hisp., Hispanic; AA, AfricanAmerican; PI, protease inhibitor; NRTI, nucleoside reverse transcriptase inhibitor; NNRTI, nonnucleoside reverse transcriptase inhibitor.

2174



TABLE 2. Median baseline and change from baseline for both treatment groups Metformin (n ⫽ 14) Median baseline*

Body composition 27.2 (24.8, 30.4) BMI (kg/m2) WHR 0.98 (1.00, 0.95) DEXA total fat (kg) 18.9 (14.8, 22.6) DEXA trunk fat (kg) 12.0 (8.6, 15.0) CT visceral abdominal 159 (92, 195) fat (cm2) CT sc leg fatc (cm2) 36.6 (20.5, 62.4) CT mid-thigh leg musclec 169 (140, 188) (cm2) CT anterior thigh muscle 55.0 (50.0, 56.5) attenuationc (HU) Metabolic Fasting insulin (␮IU/ml) 12.2 (9.2, 17.2) Insulin 2-h (␮IU/ml) 58 (36, 144) Insulin AUC (␮IU/ml ⫻ 7,660 (4,342, 11,898) 120 min) Adiponectin (␮g/ml) 2.86 (1.46, 3.36) FFA (mEq/liter) 0.66 (0.49, 0.81)

Metformin and exercise (n ⫽ 11)

Median change

Median baselinea

Median change

⫺0.8 (⫺1.1, 0) ⫺0.01 (⫺0.03, 0.02) 0 (⫺0.01, ⫺0.03) ⫺0.7 (⫺1.7, ⫺0.1) 3 (⫺26, 16)

27.7 (25.9, 32.6) 1.03 (0.98, 1.05) 18.6 (12.9, 29.6) 11.7 (8.6, 16.8) 199 (152, 237)

0.8 (⫺2.1, 9.5) ⫺7 (⫺11, 0)

42.0 (22.2, 99.8) 162 (139, 189)

⫺3.3 (⫺7.5, 4.3) 3 (⫺3, 12)

0.06 0.02

54.0 (45.5, 55.0)

2.0 (0.5, 5.0)

0.04

⫺6.23 (⫺13.2, ⫺3.2) ⫺14.4 (⫺70, 13.4) ⫺4,292 (⫺7,896, ⫺221)

0.03 0.23 0.04

⫺1.0 (⫺3.5, 0) ⫺0.2 (⫺4.64, 4.42) ⫺0.2 (⫺24.4, 31.9) ⫺1,284 (⫺2,611, 1,243)

19.9 (14.3, 24.2) 47.6 (38.2, 175) 12,192 (6,813, 14,174)

0.48 (⫺1.03, 1.23) ⫺0.05 (⫺0.13, 0.08)

2.18 (1.35, 3.53) 0.65 (0.4, 0.86)

⫺0.5 (⫺1.2, ⫺0.1) ⫺0.02 (⫺0.06, ⫺0.01) ⫺0.01 (⫺0.03, 0.01) ⫺1.3 (⫺2.1, ⫺0.5) ⫺17 (⫺35, 3)

P valueb

⫺0.51 (⫺1.67, ⫺0.03) 0.04 (⫺0.07, 0.18)

0.96 0.03 0.49 0.17 0.06

0.08 0.23

Data are expressed as median with (25–75 percentile) interquartile ranges. HU, Hounsfield units; AUC, area under the curve. SI conversion for insulin is 6.945 for ␮IU/ml to pmol/liter; 1 mEq/liter ⫽ 1 mmol/liter for FFA. a P ⬎ 0.05 for baseline comparisons, except for WHR (P ⫽ 0.03). b P value for comparison of median change from baseline between treatment groups using the Wilcoxon’s rank-sum test. c One data point missing in metformin group and two in metformin and exercise group.

FIG. 1. Comparison of change from baseline to end of intervention between treatment groups for CT anterior thigh muscle attenuation. Data are expressed as median (dark line) with (25–75 percentile) interquartile ranges (box). P value is for comparison of median change between groups by the Wilcoxon’s rank-sum test.

Discussion

HIV-lipodystrophy is a syndrome characterized by fat redistribution and metabolic abnormalities, such as insulin resistance (1– 8). Determination of the physiological mechanism of insulin resistance and the relationships of fat redistribution and intramuscular lipid to insulin resistance is an important research question with clinical implications for this population. Several studies in non-HIV-infected populations have demonstrated that muscle adiposity, measured by a variety of methods, is strongly related to insulin resistance (16 –22). A similar correlation has also been demonstrated in studies of HIV-lipodystrophy patients (23–25). However, previous studies have not determined the effects of treatment strategies on muscle adiposity nor investigated

how changes in muscle adiposity might affect insulin while simultaneously controlling for changes in other important regional fat depots. The results from our previous study demonstrate that a combined aerobic and resistance exercise training program given together with metformin improves strength, aerobic endurance, WHR, CT thigh muscle cross-sectional area, blood pressure, and insulin more than metformin alone in HIV-infected patients with fat redistribution and insulin resistance (32). In this study, we evaluated anterior thigh muscle attenuation to determine whether anterior thigh muscle attenuation improved in the exercise treatment group and whether such a change affected insulin, controlling for specific changes in other fat depots. Our data demonstrate a significant increase in muscle attenuation in the metformin and exercise treatment group vs. the metformin-only group, indicative of reduced muscle adiposity. Torriani et al. (25) found higher psoas muscle adiposity by CT attenuation measurements in a previous cross-sectional study of HIV-lipodystrophy patients compared with nonlipodystrophic HIV-infected patients and healthy control subjects. In addition, muscle adiposity was found to be highly associated with hyperinsulinemia (25). Our present study further validates and extends these findings in a different population of HIV-infected patients, showing the degree to which changes in muscle adiposity determine effects on insulin in response to different insulin-sensitizing strategies. Furthermore, we used data on muscle attenuation from the anterior thigh muscles, a region that was exercised in the treatment protocol and for which we had specific information on sc fat and muscle cross-sectional area. The association between decreased thigh muscle adiposity and decreased insulin found in the current study remained significant in regression models controlling for other important regional fat depots. The models explained a significant



2175

TABLE 3. Univariate and multivariate regression analysis Median change from baseline Fasting insulin (␮IU/ml) Median change from baseline

WHR CT anterior leg attenuation (HU) CT sc leg fat (cm2) CT visceral abdominal fat (cm2) DEXA trunk fat (kg)

2-hr insulin (␮IU/ml)

Multivariatea

Univariate R value

P value

Parameter estimate

0.357 ⫺0.172 0.332 0.225 0.868

0.08 0.44 0.13 0.28 ⬍0.01

⫺0.329 ⫺0.422 ⫺0.003 ⬍0.001 0.008

Univariate

Adjusted P value

R value

0.99 0.05 0.05 0.51 ⬍0.01

0.308 ⫺0.427 0.108 ⫺0.017 0.396

Multivariateb

P value

Parameter estimate

Adjusted P value

0.14 0.05 0.63 0.94 0.05

348.471 ⫺6.422 ⫺0.029 ⬍0.001 0.029

0.40 0.04 0.19 0.99 0.04

HU, Hounsfield units. R2 ⫽ 0.84 for model. b R2 ⫽ 0.44 for model. a

degree of the variance in fasting and 2-h insulin. Of note, combined treatment with metformin and exercise, more than metformin alone, tended to decrease leg sc fat. Previously, Mynarcik et al. (14) showed an inverse relationship between extremity fat and insulin sensitivity in HIV-infected patients. We show a similar relationship using leg CT, a direct measure of leg sc fat area, in multivariate modeling. These data in a human lipodystrophic model further support previous animal data suggesting the importance of sc fat to maintain normal insulin sensitivity (38). Taken together, our data suggest that the improvement in insulin in the metformin and exercise group occurs, in part, because of the beneficial effects of exercise on muscle adiposity and trunk fat, despite a relatively greater loss of sc fat in this group. Additional studies are necessary to verify these results in larger studies of insulin-resistant patients. Magnetic resonance spectroscopy is a commonly used and validated measure of intramyocellular lipid concentration. Intramyocellular lipid concentration, as determined by magnetic resonance spectroscopy, correlates with lipid content on muscle biopsy and insulin sensitivity in non-HIV-infected patients (16 –18, 39 – 42). In contrast to intramyocellular lipid concentration measurements using magnetic resonance spectroscopy, CT muscle attenuation measurements assess regional muscle adiposity or the relative fat content of muscle in a given area. However, CT attenuation measurements are much easier to perform than intramyocellular lipid concentration measurements and do not require spectroscopy. CT attenuation measurements of muscle tissue content have been validated through comparison with muscle biopsy (43) and magnetic resonance imaging (44) in non-HIV populations. Additional work is necessary to validate this technique against biopsy results in the HIV population, but the strong associations with insulin in our study suggest that determination of muscle attenuation is biologically meaningful and useful as a technique to further characterize regional adiposity in this population. Several studies link the accumulation of intramuscular lipid with reduced insulin sensitivity, but the mechanism is not completely understood. Intramyocellular lipid may decrease muscle glucose uptake. FFAs are known to decrease phosphatidyl inositol (PI)-3 kinase-associated insulin signaling (45, 46) and may contribute to insulin resistance among patients with increased intramuscular lipid. Several studies in type II diabetics have shown a correlation between higher

concentrations of plasma FFA, intramyocellular triglyceride content, and muscle insulin resistance (47, 48). HIV-infected patients with fat redistribution and insulin resistance demonstrate higher plasma FFA concentrations (30, 31). In this study, we did not see significant changes in FFA between treatment groups, suggesting that the changes in insulin were related more directly to other factors in this intervention study. Adiponectin is an adipokine that has also been implicated as a contributor to insulin resistance in HIV-lipodystrophy. Several studies in HIV populations show that patients with HIV-lipodystrophy have lower plasma and adipose adiponectin levels than HIV-infected patients without lipodystrophy and normal control subjects (26 –28). Adiponectin levels have been shown to negatively correlate with insulin sensitivity in this population (26 –29). We have previously shown that adiponectin is positively associated with extremity fat and negatively associated with trunk fat in patients with HIV-lipodystrophy (28). In this study, adiponectin tended to decrease more in the metformin and exercise group in comparison with the metformin-only group. Despite reductions in sc fat and adiponectin, there was an overall improvement in insulin sensitivity in response to combined treatment with metformin and exercise. The phenomenon of improved insulin sensitivity with both acute and chronic exercise training has been well documented in several non-HIV populations (49 –55); however, the specific physiological mechanism is not well understood. Limited physiological studies of exercise in normal, insulinresistant, and type II diabetic subjects demonstrate an upregulation of GLUT-4 and PI-3 kinase expression, which may explain the increased glucose transport and phosphorylation by insulin observed after exercise training (52, 56 – 62). More recently, activation of atypical protein kinase C, which operates downstream of PI-3 kinase, has been shown with exercise training (63). To date, there have been no investigations looking at the biochemical mechanisms behind improved insulin sensitivity with exercise training in HIVinfected patients. Furthermore, we are not aware of previous studies investigating the effects of exercise on muscle adiposity and regional body composition parameters in relationship to insulin sensitivity in HIV or non-HIV populations. The mechanisms by which metformin improves insulin sensitivity are also not entirely understood, but it is thought

2176


that metformin primarily acts to improve hepatic glucose production. In a recent study, Zhou et al. (64) showed that metformin may act to improve AMP kinase in the liver and inhibit hepatic glucose production. Recent studies suggest that metformin may also increase AMP kinase activity in the muscle, which may stimulate glucose uptake and also increase muscle fatty acid oxidation (65). To our knowledge, no previous study has investigated the effects of metformin on muscle adiposity and the specific effects of metformin on regional fat depots, including muscle, in humans. Our data demonstrate that metformin does not improve muscle adiposity in patients with HIV-lipodystrophy. In contrast, muscle adiposity tended to increase in the metformin-only patients. This increase in muscle adiposity may be due, in part, to the natural history of disease progression or a unique interaction in the HIV population. Additional studies are necessary to investigate the effects of metformin on muscle adiposity in this and other populations. In addition, we did not assess liver fat, which may also affect insulin sensitivity and may be an additional determinant of insulin sensitivity in this population. In previous studies, lifestyle modification treatment strategies, including weight loss and physical activity, improved glucose more than metformin (66, 67). Our study population cannot be directly compared with non-HIV-infected populations with glucose intolerance and insulin resistance; however, it is intriguing to speculate that the observed differential effects of exercise and metformin on muscle adiposity may in part explain such results. Additional studies are necessary to investigate whether the differential effects of metformin and exercise on regional adiposity and insulin sensitivity observed in this study extend to other populations with insulin resistance. The fat redistribution seen in HIV-lipodystrophy is heterogeneous, and the changes may not be mechanistically linked (68). The predominant feature may be loss of sc fat, but visceral fat accumulation may also occur, as seen in the patient population we selected. The changes in fat distribution seen in our patient population, namely increased visceral fat, increased muscle adiposity, and decreased sc fat, may simultaneously contribute to insulin resistance. Our results demonstrate that various treatment strategies will affect such depots differentially, accounting for greater or lesser effects on insulin. Our results suggest that exercise may be beneficial because it reduces muscle adiposity, but this effect is partially negated by a further reduction of sc fat in the HIV population. A more ideal strategy might be one that decreases muscle adiposity while increasing sc fat and decreasing abdominal fat. In contrast to the regimen of exercise and metformin, preliminary studies of the thiazolidinediones in insulin-resistant patients with HIV-lipodystrophy have shown significantly increased adiponectin in association with improved sc fat (69, 70), but the effect of the thiazolidinediones on intramuscular lipid content is not known. Because metformin and exercise work to decrease intramuscular adiposity and trunk fat, at the expense of sc fat, it is not known whether this strategy would be as effective in patients with primary lipoatrophy, in whom the thiazolidinediones might be a more logical therapeutic strategy. Larger longitudinal treatment studies investigating the


relationship between intramuscular lipid content and insulin sensitivity are needed to establish the mechanisms and optimal treatment strategies for insulin resistance in HIVlipodystrophy. Our data further link higher muscle adiposity and insulin resistance in HIV-lipodystrophy. Exercise in combination with metformin reduces muscle adiposity as measured by CT attenuation in concert with decreased abdominal fat and decreased sc fat. The reduction in sc fat may not be beneficial with respect to insulin resistance, but the net effect of combined aerobic and progressive resistance training and metformin on critical regional fat depots is beneficial with respect to insulin. Our data provide new insight into the mechanisms and potential efficacy of treatment strategies for insulin resistance in the HIV population and strongly suggest that therapies reducing muscle adiposity will improve insulin sensitivity. Acknowledgments We thank both the Massachusetts Institute of Technology and Massachusetts General Hospital Clinical Research Center nursing and bionutrition staff and members of the Massachusetts General Hospital Physical Therapy Services for their dedicated patient care. We also thank the staff at the Charles River Park Health Club for the use of their exercise facilities. Received October 27, 2003. Accepted January 27, 2004. Address all correspondence and requests for reprints to: Steven Grinspoon, M.D., Program in Nutritional Metabolism, 55 Fruit Street, LON207, Boston, Massachusetts 02144-2696. E-mail: [email protected]. This work was funded in part by National Institutes of Health Grants DK49302, RR300088, and RR01066.

References 1. Hadigan C, Meigs JB, Corcoran C, Rietschel P, Piecuch S, Basgoz N, Da B, Sax P, Stanley T, Wilson PW, D’Agostino RB, Grinspoon S 2001 Metabolic abnormalities and cardiovascular disease risk factors in adults with human immunodeficiency virus infection and lipodystrophy. Clin Infect Dis 32: 130 –139 2. Carr A, Samaras K, Burton S, Law M, Freund J, Chisholm DJ, Cooper DA 1998 A syndrome of peripheral lipodystrophy, hyperlipidaemia and insulin resistance in patients receiving HIV protease inhibitors. AIDS 12:F51–F58 3. Miller KD, Jones E, Yanovski JA, Shankar R, Feuerstein I, Falloon J 1998 Visceral abdominal-fat accumulation associated with use of indinavir. Lancet 351:871– 875 4. Saint-Marc T, Partisani M, Poizot-Martin I, Rouviere O, Bruno F, Avellaneda R, Lang JM, Gastaut JA, Touraine JL 2000 Fat distribution evaluated by computed tomography and metabolic abnormalities in patients undergoing antiretroviral therapy: preliminary results of the LIPOCO study. AIDS 14: 37– 49 5. Carr A, Samaras K, Thorisdottir A, Kaufmann GR, Chisholm DJ, Cooper DA 1999 Diagnosis, prediction, and natural course of HIV-1 protease-inhibitorassociated lipodystrophy, hyperlipidaemia, and diabetes mellitus: a cohort study. Lancet 353:2093–2099 6. Martinez E, Gatell J 1998 Metabolic abnormalities and use of HIV-1 protease inhibitors. Lancet 352:821– 822 7. Dube MP, Edmondson-Melancon H, Qian D, Aqeel R, Johnson D, Buchanan TA 2001 Prospective evaluation of the effect of initiating indinavir-based therapy on insulin sensitivity and B-cell function in HIV-infected patients. J Acquir Immune Defic Syndr 27:130 –134 8. Walli R, Herfort O, Michl GM, Demant T, Jager H, Dieterle C, Bogner JR, Landgraf R, Goebel FD 1998 Treatment with protease inhibitors associated with peripheral insulin resistance and impaired oral glucose tolerance in HIV-1-infected patients. AIDS 12:F167–F173 9. Hong Y, Rice T, Gagnon J, Despres JP, Nadeau A, Perusse L, Bouchard C, Leon AS, Skinner JS, Wilmore JH, Rao DC 1998 Familial clustering of insulin and abdominal visceral fat: the HERITAGE Family Study. J Clin Endocrinol Metab 83:4239 – 4245 10. Marcus MA, Murphy L, Pi-Sunyer FX, Albu JB 1999 Insulin sensitivity and serum triglyceride level in obese white and black women: relationship to visceral and truncal subcutaneous fat. Metabolism 48:194 –199 11. Seidell JC, Bjorntorp P, Sjostrom L, Kvist H, Sannerstedt R 1990 Visceral fat


12. 13.

14.

15. 16.

17.

18.

19. 20. 21. 22. 23.

24.

25.

26.

27.

28.

29.

30.

31.

32.

accumulation in men is positively associated with insulin, glucose, and cpeptide levels, but negatively with testosterone levels. Metabolism 9:897–901 Mauriege P, Klein Kranenbarg WM, Prud’homme D, Lamarche B, Tremblay A, Bouchard C, Nadeau A, Despres JP 1996 Insulin and glucagon responses to adrenaline infusion in abdominal obese men. Int J Obes 20:668 – 676 Expert Panel on Detection, Evaluation, and Treatment of High Blood Cholesterol in Adults 2001 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). JAMA 285:2486 –2497 Mynarcik DC, McNurlan MA, Steigbigel RT, Fuhrer J, Gelato MC 2000 Association of severe insulin resistance with both loss of limb fat and elevated serum tumor necrosis factor receptor levels in HIV lipodystrophy. J Acquir Immune Defic Syndr 25:312–321 Meininger G, Hadigan C, Rietschel P, Grinspoon S 2002 Body composition measurements as predictors of glucose and insulin abnormalities in HIVpositive men. Am J Clin Nutr 76:460 – 465 Kautzky-Willer A, Krssak M, Winzer C, Pacini G, Tura A, Farhan S, Wagner O, Brabant G, Horn R, Stingl H, Schneider B, Waldhausl W, Roden M 2003 Increased intramyocellular lipid concentration identifies impaired glucose metabolism in women with previous gestational diabetes. Diabetes 52:244 –251 Jacob S, Machann J, Rett K, Brechtel K, Volk A, Renn W, Maerker E, Matthaei S, Schick F, Claussen CD, Haring HU 1999 Association of increased intramyocellular lipid content with insulin resistance in lean nondiabetic offspring of type 2 diabetic subjects. Diabetes 48:1113–1119 Thamer C, Machann J, Bachmann O, Haap M, Dahl D, Wietek B, Tschritter O, Niess A, Brechtel K, Fritsche A, Clausseen C, Jacob S, Schick F, Haring HU, Stumvoll M 2003 Intramyocellular lipids: anthropometric determinants and relationships with maximal aerobic capacity and insulin sensitivity. J Clin Endocrinol Metab 88:1785–1791 Pan DA, Lillioja S, Kriketos AD, Milner MR, Baur LA, Bogardus C, Jenkins AB, Storlien LH 1997 Skeletal muscle triglyceride levels are inversely related to insulin action. Diabetes 46:983–988 Simoneau JA, Colberg SR, Thaete FL, Kelley DE 1995 Skeletal muscle glycolytic and oxidative enzyme capacities are determinants of insulin sensitivity and muscle composition in obese women. FASEB J 9:273–278 Goodpaster BH, Thaete FL, Simoneau JA, Kelley DE 1997 Subcutaneous abdominal fat and thigh muscle composition predict insulin sensitivity independently of visceral fat. Diabetes 46:1579 –1585 Goodpaster BH, Thaete FL, Kelley DE 2000 Thigh adipose tissue distribution is associated with insulin resistance in obesity and in type 2 diabetes mellitus. Am J Clin Nutr 71:885– 892 Gan SK, Samaras K, Thompson CH, Kraegen EW, Carr A, Cooper DA, Chisholm DJ 2002 Altered myocellular and abdominal fat partitioning predict disturbance in insulin action in HIV protease inhibitor-related lipodystrophy. Diabetes 51:3163–3169 Luzi L, Perseghin G, Tambussi G, Meneghini E, Scifo P, Pagliato E, Del Maschio A, Testolin G, Lazzarin A 2003 Intramyocellular lipid accumulation and reduced whole body lipid oxidation in HIV lipodystrophy. Am J Physiol Endocrinol Metab 284:E274 –E280 Torriani M, Hadigan C, Jensen ME, Grinspoon S 2003 Psoas muscle attenuation measurement with computed tomography indicates intramuscular fat accumulation in patients with the HIV-lipodystrophy syndrome. J Appl Physiol 95:1005–1010 Sutinen J, Korsheninnikova E, Funahashi T, Matsuzawa Y, Nyman T, YkiJarvinen H 2003 Circulating concentration of adiponectin and its expression in subcutaneous adipose tissue in patients with highly active antiretroviral therapy-associated lipodystrophy. J Clin Endocrinol Metab 88:1907–1910 Lihn AS, Richelsen B, Pedersen SB, Haugaard SB, Rathje GS, Madsbad S, Andersen O 2003 Increased expression of TNF-{␣}, IL-6, and IL-8 in HALS: implications for the reduced expression and plasma levels of adiponectin. Am J Physiol Endocrinol Metab 285:E1072–E1080 Tong Q, Sankale JL, Hadigan CM, Tan G, Rosenberg ES, Kanki PJ, Grinspoon SK, Hotamisligil GS 2003 Regulation of adiponectin in human immunodeficiency virus-infected patients: relationship to body composition and metabolic indices. J Clin Endocrinol Metab 88:1559 –1564 Vigouroux C, Maachi M, Nguyen TH, Coussieu C, Gharakhanian S, Funahashi T, Matsuzawa Y, Shimomura I, Rozenbaum W, Capeau J, Bastard JP 2003 Serum adipocytokines are related to lipodystrophy and metabolic disorders in HIV-infected men under antiretroviral therapy. AIDS 17:1503– 1511 Meininger G, Hadigan C, Laposata M, Brown J, Rabe J, Louca J, Aliabadi N, Grinspoon S 2002 Elevated concentrations of free fatty acids are associated with increased insulin response to standard glucose challenge in human immunodeficiency virus-infected subjects with fat redistribution. Metabolism 51:260 –266 Vigouroux C, Gharakhanian S, Salhi Y, Nguyen TH, Chevenne D, Capeau J, Rozenbaum W 1999 Diabetes, insulin resistance and dyslipidaemia in lipodystrophic HIV-infected patients on highly active antiretroviral therapy (HAART). Diabetes Metab 25:225–232 Driscoll SD, Meininger GE, Lareau MT, Dolan SE, Killilea KM, Hadigan CM, Lloyd-Jones DM, Klibanski A, Frontera WR, Grinspoon S 2004 Effects


33. 34. 35. 36. 37. 38. 39. 40. 41. 42.

43. 44. 45. 46. 47. 48.

49. 50. 51. 52.

53. 54.

55.

56.

57.

2177

of exercise training and metformin on body composition and cardiovascular indices in HIV infected patients. AIDS 18:465– 473 Hartig DE, Henderson JM 1999 Increasing hamstring flexibility decreases lower extremity overuse injuries in military basic trainees. Am J Sports Med 27:173–176 LaPierre A, Klimas N, Major P, Perry A 1997 Acquired immunodeficiency syndrome. In: Exercise management for persons with chronic diseases and disabilities. Champaign, IL: Human Kinetic Books; 132–136 De Lorme TL, Watkins AL 1948 Techniques of progressive resistance exercise. Arch Phys Med 29:263–273 Borkan GA, Gerzof SG, Robbins AH, Hults DE, Silbert CK, Silbert JE 1982 Assessment of abdominal fat content by computed tomography. Am J Clin Nutr 36:172–177 Mazess RB, Barden HS, Bisek JP, Hanson J 1990 Dual-energy x-ray absorptiometry for total-body and regional bone-mineral and soft-tissue composition. Am J Clin Nutr 51:1106 –1112 Gavrilova O, Marcus-Samuels B, Graham D, Kim JK, Shulman GI, Cas AL, Vinson C, Eckhaus M, Reitman ML 2000 Surgical implantation of adipose tissue reverses diabetes in lipoatrophic mice. J Clin Invest 105:271–278 Schick F, Eismann B, Jung WI, Bongers H, Bunse M, Lutz O 1993 Comparison of localized proton NMR signals of skeletal muscle and fat tissue in vivo: two lipid compartments in muscle tissue. Magn Reson Med 29:158 –167 Boesch C, Slotboom J, Hoppeler H, Kreis R 1997 In vivo determination of intra-myocellular lipids in human muscle by means of localized 1H-MRspectroscopy. Magn Reson Med 37:484 – 493 Szczepaniak LS, Babcock EE, Schick F, Dobbins RL, Garg A, Burns DK, McGarry JD, Stein DT 1999 Measurement of intracellular triglyceride stores by H spectroscopy: validation in vivo. Am J Physiol 276:E977–E989 Howald H, Boesch C, Kreis R, Matter S, Billeter R, Essen-Gustavsson B, Hoppeler H 2002 Content of intramyocellular lipids derived by electron microscopy, biochemical assays, and (1)H-MR spectroscopy. J Appl Physiol 92: 2264 –2272 Goodpaster BH, Kelley DE, Thaete FL, He J, Ross R 2000 Skeletal muscle attenuation determined by computed tomography is associated with skeletal muscle lipid content. J Appl Physiol 89:104 –110 Mitsiopoulos N, Baumgartner RN, Heymsfield SB, Lyons W, Gallagher D, Ross R 1998 Cadaver validation of skeletal muscle measurement by magnetic resonance imaging and computerized tomography. J Appl Physiol 85:115–122 Shulman GI 2000 Cellular mechanisms of insulin resistance. J Clin Invest 106:171–176 Hegarty BD, Furler SM, Ye J, Cooney GJ, Kraegen EW 2003 The role of intramuscular lipid in insulin resistance. Acta Physiol Scand 178:373–383 Kelley DE, Williams KV, Price JC, McKolanis TM, Goodpaster BH, Thaete FL 2001 Plasma fatty acids, adiposity, and variance of skeletal muscle insulin resistance in type 2 diabetes mellitus. J Clin Endocrinol Metab 86:5412–5419 Perseghin G, Scifo P, De Cobelli F, Pagliato E, Battezzati A, Arcelloni C, Vanzulli A, Testolin G, Pozza G, Del Maschio A, Luzi L 1999 Intramyocellular triglyceride content is a determinant of in vivo insulin resistance in humans: a 1H–13C nuclear magnetic resonance spectroscopy assessment in offspring of type 2 diabetic parents. Diabetes 48:1600 –1606 Larsen JJ, Dela F, Kjaer M, Galbo H 1997 The effect of moderate exercise on postprandial glucose homeostasis in NIDDM patients. Diabetologia 40: 447– 453 Devlin JT, Hirshman M, Horton ED, Horton ES 1987 Enhanced peripheral and splanchnic insulin sensitivity in NIDDM men after single bout of exercise. Diabetes 36:434 – 439 Minuk HL, Vranic M, Marliss EB, Hanna AK, Albisser AM, Zinman B 1981 Glucoregulatory and metabolic response to exercise in obese noninsulindependent diabetes. Am J Physiol 240:E458 –E464 Hughes VA, Fiatarone MA, Fielding RA, Kahn BB, Ferrara CM, Shepherd P, Fisher EC, Wolfe RR, Elahi D, Evans WJ 1993 Exercise increases muscle GLUT-4 levels and insulin action in subjects with impaired glucose tolerance. Am J Physiol 264:E855–E862 Dela F, Larsen JJ, Mikines KJ, Ploug T, Petersen LN, Galbo H 1995 Insulinstimulated muscle glucose clearance in patients with NIDDM: effects of onelegged physical training. Diabetes 44:1010 –1020 Eriksson J, Tuominen J, Valle T, Sundberg S, Sovijarvi A, Linsholm H, Tuomilehto J, Koivisto V 1998 Aerobic endurance exercise or circuit-type resistance training for individuals with impaired glucose tolerance? Horm Metab Res 30:37– 41 Mayer-Davis EJ, D’Agostino R, Jr., Karter AJ, Haffner SM, Rewers MJ, Saad M, Bergman RN 1998 Intensity and amount of physical activity in relation to insulin sensitivity: the Insulin Resistance Atherosclerosis Study. JAMA 279: 669 – 674 Kennedy JW, Hirshman MF, Gervino EV, Ocel JV, Forse RA, Hoenig S, Aronson D, Goodyear LJ, Horton ES 1999 Acute exercise induces GLUT4 translocation in skeletal muscle of normal human subjects and subjects with type 2 diabetes. Diabetes 48:1192–1197 Perseghin G, Price TB, Petersen KF, Roden M, Cline GW, Gerow K, Rothman DL, Shulman GI 1996 Increased glucose transport-phosphorylation and muscle glycogen synthesis after exercise training in insulin-resistant subjects. N Engl J Med 335:1357–1362

2178


58. Houmard JA, Egan PC, Neufer PD, Friedman JE, Wheeler WS, Israel RG, Dohm GL 1991 Elevated skeletal muscle glucose transporter levels in exercisetrained middle-aged men. Am J Physiol 261:E437–E443 59. Houmard JA, Shinebarger MH, Dolan PL, Leggett-Frazier N, Bruner RK, McCammon MR, Israel RG, Dohm GL 1993 Exercise training increases GLUT-4 protein concentration in previously sedentary middle-aged men. Am J Physiol 264:E896 –E901 60. Dela F, Ploug T, Handberg A, Petersen LN, Larsen JJ, Mikines KJ, Galbo H 1994 Physical training increases muscle GLUT4 protein and mRNA in patients with NIDDM. Diabetes 43:862– 865 61. Houmard JA, Shaw CD, Hickey MS, Tanner CJ 1999 Effect of short-term exercise training on insulin-stimulated PI 3-kinase activity in human skeletal muscle. Am J Physiol 277:E1055–E1060 62. Kirwan JP, del Aguila LF, Hernandez JM, Williamson DL, O’Gorman, I, Lewis R, Krishnan RK 2000 Regular exercise enhances insulin activation of IRS-1-associated PI3-kinase in human skeletal muscle. J Appl Physiol 88: 797– 803 63. Beeson M, Sajan MP, Dizon M, Grebenev D, Gomez-Daspet J, Miura A, Kanoh Y, Powe J, Bandyopadhyay G, Standaert ML, Farese RV 2003 Activation of protein kinase C-␨ by insulin and phosphatidylinositol-3,4,5-(PO4)3 is defective in muscle in type 2 diabetes and impaired glucose tolerance: amelioration by rosiglitazone and exercise. Diabetes 52:1926 –1934 64. Zhou G, Myers R, Li Y, Chen Y, Shen X, Fenyk-Melody J, Wu M, Ventre J, Doebber T, Fujii N, Musi N, Hirshman MF, Goodyear LJ, Moller DE 2001


65.

66.

67.

68. 69.

70.

Role of AMP-activated protein kinase in mechanism of metformin action. J Clin Invest 108:1167–1174 Musi N, Hirshman MF, Nygren J, Svanfeldt M, Bavenholm P, Rooyackers O, Zhou G, Williamson JM, Ljunqvist O, efendic S, Moller DE, Thorell A, Goodyear LJ 2002 Metformin increases AMP-activated protein kinase activity in skeletal muscle of subjects with type 2 diabetes. Diabetes 51:2074 –2081 Knowler WC, Barrett-Connor E, Fowler SE, Hamman RF, Lachin JM, Walker EA, Nathan DM, Diabetes Prevention Program Research Group 2002 Reduction in the incidence of type 2 diabetes with lifestyle intervention or metformin. N Engl J Med 346:393– 403 Tuomilehto J, Lindstrom J, Eriksson JG, Valle TT, Hamalainen H, IlanneParikka P, Keinanen-Kiukaanniemi S, Laakso M, Louheranta A, Rastas M, Salminen V, Uusitupa M, Finnish Diabetes Prevention Study Group 2001 Prevention of type 2 diabetes mellitus by changes in lifestyle among subjects with impaired glucose tolerance. N Engl J Med 344:1343–1350 Grunfeld C, Basic science and metabolic disturbances. XIV International AIDS Conference, Barcelona, Spain, 2002, p 81 (Oral Presentation TuOr158) Gelato MC, Mynarcik DC, Quick JL, Steigbigel RT, Fuhrer J, Brathwal CE, Brebbia JS, Wax MR, McNurlan MA 2002 Improved insulin sensitivity and body fat distribution in HIV-infected patients treated with rosiglitazone: a pilot study. J Acquir Immune Defic Syndr 31:163–170 Hadigan C, Yawetz S, Thomas A, Havers F, Sax PE, Grinspoon S, A randomized, double-blind, placebo-controlled study of rosiglitazone for patients with HIV lipodystrophy. 5th International Workshop on Adverse Drug Reactions and Lipodystrophy in HIV, Paris, France, 2003, p L12

JCEM is published monthly by The Endocrine Society (http://www.endo-society.org), the foremost professional society serving the endocrine community.