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From the Clinical Research Centers |
Division of Geriatrics and Gerontology, Department of Internal Medicine, Washington University School of Medicine, St. Louis, Missouri 63110
Address all correspondence and requests for reprints to: Dr. Wendy M. Kohrt, Ph.D., Division of Geriatric Medicine, University of Colorado Health Sciences Center, 4200 East Ninth Avenue, Box B-179, Denver, Colorado 80262. E-mail: Wendy.Kohrt{at}uchsc.edu
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Abstract |
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 Top Abstract IntroductionSubjects and MethodsResultsDiscussionReferences |
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50% (LO) and 45% (MOD) of basal] than in sedentary
obese [to 72% (LO) and 63% (MOD) of basal]. The rate of appearance
of glycerol was suppressed to 36 ± 7%, 44 ± 10%, and
62 ± 7% of basal during LO in endurance-trained nonobese,
sedentary nonobese, and sedentary obese, respectively
(P < 0.05, endurance-trained nonobese <
sedentary obese), and to 34 ± 3%, 36 ± 5%, and 53 ±
8% of basal during MOD, respectively (P < 0.05,
endurance-trained nonobese < sedentary obese). There were no
between-group differences in the suppression of lipolysis in abdominal
sc AT, as evidenced by similar reductions in dialysate glycerol levels
[all to 65% (LO) and 55% (MOD) of basal]. Femoral dialysate
glycerol was reduced (P < 0.05) more in sedentary
nonobese and endurance-trained nonobese (to 75% of basal) than in
sedentary obese (to 90% of basal) during LO, but to a similar extent
(to 60% of basal) in all groups during MOD. The results indicate
that the sedentary obese women had whole body resistance to the
suppression of lipolysis by insulin. Intraabdominal AT may be the site
of resistance, as resistance was not evident in abdominal or femoral sc
AT. |
Introduction |
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TopAbstractIntroductionSubjects and MethodsResultsDiscussionReferences |
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Because of its anatomical location, the direct assessment of the metabolic activity of intraabdominal fat is not feasible in humans. However, simultaneous assessment of regional lipolysis in sc adipose tissue by microdialysis and whole body lipolysis by stable labeled isotope kinetics may provide insight into the relative roles of sc adipose tissue and other adipose tissue (presumably intraabdominal) depots in the antilipolytic response to insulin. In this context, the specific aims of this study were to evaluate the effects of obesity and physical training status on the suppression of lipolysis by insulin. To address these aims, whole body lipolysis was monitored using 2H5-glycerol infusion while regional lipolysis of femoral and abdominal sc adipose tissue was monitored using microdialysis during a hyperinsulinemic-euglycemic clamp.
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Subjects and Methods |
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TopAbstractIntroductionSubjects and MethodsResultsDiscussionReferences |
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Seventeen premenopausal women participated in the study. The subjects were six nonobese endurance-trained women [<28% body fat; maximal oxygen uptake (VO2max), >45 mL/kg·min], six nonobese sedentary women (<28% body fat; VO2max, <35 mL/kg·min), and five obese sedentary women (>35% body fat; VO2max, <35 mL/kg·min). The groups were matched for age and lean body mass. The volunteers were weight stable over the 6 months before the investigation and were in good health, with the exception of impaired glucose tolerance in three of the obese subjects. None of the subjects had a prior history of diabetes or cardiovascular disease. None of the subjects was amenorrheic, in that all had had monthly menses for the previous 12 months. Women were not studied at a standard time of the menstrual cycle, although there were the same number (n = 3) of subjects studied in the luteal phase of the cycle in each group. Subjects participated in the study after giving their informed consent according to the Institutional Review Board at Washington University School of Medicine.
Protocol
Whole body lipolysis was evaluated by 2H5-glycerol kinetics in the basal state and during low and moderate dose insulin infusions. Regional lipolysis was evaluated simultaneously by placing microdialysis probes in sc adipose tissue in the abdominal (two probes) and femoral (two probes) regions. Magnetic resonance imaging (MRI) of the abdomen was used to determine whether the degree of insulin resistance was related specifically to intraabdominal fat volume.
Body composition and fat distribution
Fat and fat-free masses were measured by dual energy x-ray absorptiometry (QDR-1000/w, Hologic, Inc., Waltham, MA), as described previously using version 5.64 of the enhanced whole body analysis program (17). Regional adiposity was also evaluated anthropometrically and by MRI. Circumference measurements were made at the waist, hip, and midthigh. The anatomical landmarks used to locate the circumference sites in a reproducible manner were those described in the Anthropometric Standardization Reference Manual (18). Nineteen cross-sectional images of the abdomen were obtained using MRI. The median slice was obtained at the level of the L3–L4 intervertebral space. The serial images were 8 mm thick with a 2-mm gap. Nineteen cross-sectional images (7 mm thick, 3-mm gap) of the thigh were also obtained using MRI. The first slice was located at the articular surface of the lateral femoral condyle. The middle nine slices were analyzed, and a continuous cylinder of the abdomen (9 cm) or thigh (9 cm) was obtained. The gap between slices was assumed to have the same fat content as the preceding analyzed slice. Images were analyzed using NIH Image software by a technician who was blinded to subject identification. The coefficient of variation for duplicate determinations of intraabdominal fat volume was 3.4 ± 0.7%, which was greater than that for determinations of sc abdominal (0.6 ± 0.3%; P < 0.01) and femoral (0.6 ± 0.2%; P < 0.01) fat volume.
VO2max
VO2max was determined on a motorized treadmill. Sedentary subjects initially walked at 3 mph for 3 min. The speed was then increased by 1.0 mph every 3 min to 5 mph. The subjects jogged at this speed (5 mph) for the remainder of the test while grade was increased by 2% every minute until exhaustion. Trained subjects jogged at 5 mph for 3 min, after which the speed was increased by 1 mph every 3 min to 7 mph. The speed was then held constant, and grade was increased by 2% every minute until exhaustion. The fractional oxygen and carbon dioxide content of expired air was measured continuously, and oxygen uptake was calculated for each 30-s interval using an automated open circuit system (14). At least two of the following criteria were met to establish that VO2max had been attained: a plateau in VO2 despite an increase in treadmill speed or grade, attainment of age-predicted maximal heart rate, and a respiratory exchange ratio greater than 1.10.
Hyperinsulinemic-euglycemic clamp and glycerol kinetics
The trained subjects were studied 16–24 h after their last
exercise bout. All subjects were instructed to consume at least
250 g carbohydrate/day for the 3 days before the clamp procedure.
The clamp studies were performed as described previously (19) at the
Washington University General Clinical Research Center after an
overnight fast. A primed (1.5 µmol/kg), constant (0.1
µmol/kg·min) infusion of
2H5-glycerol (99%; Tracer
Technologies, Newton, MA) was started and continued for 270 min (90-min
basal period, 90 min at each insulin infusion rate). The actual isotope
delivery rate was determined by measuring the enrichment of the isotope
in the infusate. Three baseline blood and dialysate samples were
obtained at 10-min intervals before insulin infusion to determine
plasma substrate and hormone concentrations and background isotope
enrichments. This 30-min basal period was followed by two sequential
90-min primed, constant insulin infusions using a Harvard infusion pump
(Harvard Apparatus, South Natick, MA). In obese subjects, insulin was
delivered at rates of 6 and 12 mU/m·min. Because a given insulin
infusion rate results in higher plasma insulin concentrations in obese
than lean individuals (4, 19, 20), the insulin infusion rates were
adjusted up (to 10 and 20 mU/m·min) in lean subjects. It was
anticipated that these infusion rates would result in plasma insulin
concentrations of approximately 15 and 35 µU/mL. These levels were
chosen because the insulin concentrations for a half-maximal
suppression of lipolysis have been shown to be about 14 and 38 µU/mL
in lean and obese subjects, respectively (4).
Blood glucose was checked every 5 min during the glucose clamp procedure. Blood glucose was maintained at approximately 5 mmol/L using a variable rate glucose infusion of 20% dextrose. The rate of glucose disposal was estimated from the steady state glucose infusion rate over the final 30 min of each insulin infusion. It is recognized that this may have resulted in an underestimate of the actual glucose disposal rate, as hepatic glucose production may not have been completely suppressed at the low insulin concentrations.
Blood samples were obtained at 10-min intervals during the final 30 min of each of the 90-min stages for the determination of plasma substrate and hormone concentrations as well as isotope enrichments. Plasma samples were stored at -80 C and subsequently analyzed for free fatty acids (NEFA kit, Wako Chemicals, Dallas, TX), glycerol (21), insulin (22), and adrenaline and noradrenaline concentrations (23).
2H5-glycerol analysis
The analysis of 2H5-glycerol was performed using a modification of the negative ion chemical ionization (NCI) gas chromatography-mass spectrometry method described by Gilker et al. (24). A sample of plasma (50 µL) was pipetted into 500 µL perchloric acid, chilled at 4 C for 20 min, and centrifuged at 2000 x g for 10 min. The supernatant was then decanted into a small glass vial and evaporated under nitrogen. Samples were reconstituted with 150 µL of a 3:1 mixture of heptafluorobutyric acid and ethyl acetate to form the tris-heptafluorobutyrl ester derivatives, placed in a heating block at 70 C for 10 min, and evaporated under nitrogen. Samples were then reconstituted with 100 µL ethyl acetate before analysis. The NCI spectrum of the heptafluorobutyric acid derivative was obtained using a model 5988A Hewlett-Packard Co. gas chromatography-mass spectrometry instrument. For gas chromatography, a DB-17 column was used (30 m, 0.25 mm id, 0.25 µm film thickness; J and W Scientific, Folsom, CA) with a helium flow rate of 0.5 mL/min, a column temperature of 100 C for 1 min, increasing 45 C/min to 280 C, and a split ratio of 20:1. The NCI mass spectrometer conditions were: source temperature, 120 C; injector port and detector temperature, 250 C; emission, 300 µA; and ion source pressure, 0.5–0.6 torr. Methane was used as the Cl- reactant gas. The [M]- ions corresponding to nominal mass 680 for unlabeled glycerol and mass 685 for1,1,2,3,3-2H5-glycerol were monitored. The rate of appearance (Ra) of glycerol over the last 30 min of each stage was calculated using the nonsteady state equations of Steele (25), assuming a volume of distribution for glycerol of 270 mL/kg (26). The coefficient of variation for isotopic enrichment was 7.1 ± 2.2% as calculated from day to day reproducibility of enrichment determinations (n = 6).
Assessment of regional lipolysis by microdialysis
The microdialysis probes (DL-3, Bioanalytical Systems, Inc., West Lafayette, IN) consisted of inlet tubing (30 cm) and outlet tubing (15 cm) separated by a polyacrylonitrile dialysis membrane (3 cm in length, 0.25 mm id, 0.35 mm od). Probes were sterilized using ethylene oxide gas and were soaked for a minimum of 16 h overnight in 5% ethanol while perfused with Ringer’s solution to remove the glycerol coating that was placed on the dialysis membrane during the manufacturing process. The absence of measurable glycerol on the microdialysis membrane was verified before the experiments.
Microdialysis probes were inserted under sterile technique and local
anesthesia (0.1 mL lidocaine without epinephrine) into the abdominal
(two probes, bilaterally 3 cm lateral to the umbilicus) and femoral
(two probes, 3 cm apart, midthigh) sc adipose tissue in each
subject. This was accomplished by inserting a 14-gauge catheter through
the skin, advancing it through the sc adipose tissue parallel to the
skin, and then exiting through the skin. The needle was withdrawn, the
microdialysis probe was threaded through the catheter, and the catheter
was then removed, leaving the membrane portion of the probe embedded in
the adipose tissue.
Microdialysis probes were perfused (Harvard infusion pump, model 22, Harvard Apparatus) at 2.0 µL/min with Ringer’s solution containing 2.5 mmol/L glucose and 5 mmol/L ethanol for 60 min. No samples were collected during this time to allow for equilibration of the microdialysis system and to allow the initial trauma of probe insertion to subside (27, 28). Glucose (2.5 mmol/L) was added to the perfusate to minimize the drainage of glucose from the interstitial fluid. No significant net loss or gain of perfusate fluid occurred over the dialysis membrane during the microdialysis process under the conditions of this experiment, as verified by weighing dialysate samples. The inclusion of ethanol in the perfusate permitted the detection of changes in nutritive blood flow in the region surrounding the probe (29, 30). This method has been validated as a measure of adipose tissue nutritive blood flow in humans by comparison to the 133xenon clearance technique (31). Ethanol at the concentration used (5 mmol/L) has been shown to have no effect on lipolysis in sc adipose tissue (30). The ethanol data can be presented as the outflow/inflow ratio ([ethanol]dialysate/[ethanol]perfusate), which is inversely related to nutritive blood flow in the region surrounding the microdialysis probe (28). The coefficient of variation in ethanol outflow/inflow ratio between probes in a given individual was 14.4 ± 3.5% (duplicate determinations in 17 subjects). The outgoing dialysate was collected in 10-min (20 µL) fractions for baseline determinations and 15-min fractions (30 µL) during insulin infusion. The dialysate samples were stored at 4 C and analyzed within 48 h for glycerol (21) and ethanol (28). Although in vivo calibration of the microdialysis probes was not performed in this study, we have previously determined the relative recovery of glycerol over this type of microdialysis membrane to be approximately 60% in the abdominal sc adipose tissue of premenopausal women (32). It should be noted that this value of 60% for relative recovery was obtained in a different group of nonobese premenopausal women and is provided here only as an approximation of recovery. Relative recovery may be reduced in individuals with increased fat mass (33, 34), indicating that recovery may have been lower in the obese than in the nonobese group in this study. Interstitial glycerol concentrations were therefore not calculated in this study.
Statistics
Differences among groups were analyzed by one-way ANOVA. If data were not normally distributed, the Kruskal-Wallis one-way ANOVA on ranks was conducted. If significance was obtained, the Newman-Keuls post-hoc analysis was used to identify group differences. The effects of insulin infusion on blood or dialysate parameters within a group were evaluated using two-tailed paired Student’s t tests. Linear regression analysis was used to determine the correlation between fat mass and indicators of lipolysis. All data are expressed as the mean ± SE. The level of significance was set at P < 0.05.
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Results |
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TopAbstractIntroductionSubjects and MethodsResultsDiscussionReferences |
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Subject characteristics with respect to age,
VO2max, body composition, and fat distribution
are presented in Table 1
. Body weight and
indexes of fatness were higher and VO2max
(milliliters per kg/min) was lower in the obese group than in the other
two groups. VO2max was also lower in the
sedentary nonobese group than in trained subjects.
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Anthropometric data with respect to skinfold thickness of the
abdomen and thigh, circumferences of the waist, hip, and thigh, and
waist to hip ratio are presented in Table 2. There were no differences in these
parameters between trained and sedentary nonobese subjects. Obese
subjects had larger skinfold thicknesses and circumferences than both
nonobese groups (P < 0.05).
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Obese subjects had more intraabdominal fat than sedentary and trained nonobese individuals (931 ± 244, 270 ± 36, and 240 ± 38 cm3, respectively; P < 0.01). Obese subjects also had more abdominal sc fat than sedentary and trained nonobese subjects (3987 ± 753, 896 ± 190, and 942 ± 200 cm3, respectively; P < 0.01), and more femoral sc fat than sedentary and trained nonobese individuals (1540 ± 261, 599 ± 86, and 460 ± 42 cm3, respectively; P < 0.01).
Glucose parameters and circulating hormone concentrations
The fasting plasma insulin concentration was 3-fold higher in
obese than nonobese individuals; however, there were no differences in
the plasma insulin concentration during insulin infusion (Table 3). There were no significant differences
among the groups in basal serum free fatty acid or adrenaline levels,
but noradrenaline levels were higher in obese subjects than in the
other two groups (Table 3). Both noradrenaline and free fatty acids
remained elevated in obese subjects during both insulin infusions.
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). Plasma glucose in all subjects was maintained at
5.0 ± 0.4 mmol/L, with a coefficient of variation of 3.0 ±
0.6% over the final 30 min of the clamp. During the first insulin
infusion, the glucose infusion rate was higher (P <
0.05) in trained (3.95 ± 0.34 mg/kg FFM·min) than in sedentary
nonobese (3.38 ± 0.37 mg/kg FFM·min) and obese (2.90 ±
0.60 mg/kg FFM·min) subjects. During the second insulin infusion, the
glucose infusion rate was elevated in trained subjects (10.15 ±
0.76 mg/kg FFM·min; P < 0.05 vs. both
sedentary groups) and higher in nonobese (6.60 ± 1.01 mg/kg
FFM·min) than obese (4.64 ± 0.86 mg/kg FFM·min) sedentary
subjects. All reported differences in glucose infusion rates were also
evident when data were expressed per kg BW. Glycerol concentrations and kinetics
Basal. In the basal state, plasma glycerol concentrations were
highest in the sedentary obese group (P < 0.05
vs. sedentary nonobese and trained) and were higher in
trained than in sedentary nonobese subjects (Figs. 1 and 2).
Basal abdominal dialysate glycerol concentrations were higher in the
sedentary obese subjects (158.7 ± 29.7 µmol/L) than in the
sedentary nonobese (95.1 ± 7.2 µmol/L; P <
0.05) and trained (108.5 ± 9.4 µmol/L; P <
0.05) subjects. Basal femoral dialysate glycerol concentrations were
also higher in the sedentary obese subjects (171.9 ± 29.7
µmol/L) than in the sedentary nonobese (82.0 ± 6.4 µmol/L;
P < 0.05) and trained (96.6 ± 7.9 µmol/L;
P < 0.05) subjects.
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Hyperinsulinemic-euglycemic clamp. Maximal changes in
dialysate glycerol concentration were achieved between 60 and 90 min
after the initiation of insulin infusion (Fig. 1). The dialysate
glycerol data presented are therefore the mean glycerol concentration
from 60–90 min after the initiation of insulin infusion. With the
exception of the femoral dialysate concentration in the obese group,
plasma and dialysate glycerol concentrations were significantly reduced
in all groups in response to the first insulin infusion (Fig. 2). The
only further significant reduction in glycerol concentration during the
second insulin infusion occurred in the femoral dialysate in obese
subjects.
In relative terms, the reductions in glycerol concentrations and
Ra by insulin were similar in trained and
sedentary nonobese subjects during both insulin infusion stages (Figs. 3 and 4).
During the second insulin infusion, the relative reductions in
abdominal and femoral dialysate glycerol levels were similar in all
three groups, but plasma glycerol concentration and
Ra remained elevated in obese, compared to
trained and sedentary nonobese, subjects.
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The basal ethanol outflow/inflow ratio from probes in abdominal sc
adipose tissue were higher in obese than in trained subjects
(0.643 ± 0.088 vs. 0.418 ± 0.039;
P < 0.05) and tended to be higher than in sedentary
nonobese subjects (0.493 ± 0.037; P = 0.10),
indicating that nutritive blood flow per volume tissue was lower in the
obese group (Fig. 5). Basal ethanol
outflow/inflow ratios from probes in femoral sc adipose tissue were
0.540 ± 0.0526, 0.558 ± 0.040, and 0.606 ± 0.062 in
the trained, sedentary nonobese, and obese groups, respectively
(P = NS).
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). Nutritive blood flow also tended to
increase in trained subjects, but changes in ethanol outflow/inflow
ratios from basal were not statistically significant. There were no
significant between-group differences in the nutritive blood flow
response to insulin infusion. Correlations
Correlations between measures of total and regional adiposity in
relation to measures of whole body insulin action are presented in
Table 4. Intraabdominal, sc abdominal,
total abdominal, femoral, and total fat volumes were all significantly
correlated to plasma glycerol and glycerol Ra
response to insulin. Intraabdominal, sc abdominal, total abdominal, and
total fat volumes were significantly correlated to glucose disposal
during insulin infusion. Data from trained subjects were not included
in these analyses because of the independent effect of exercise
training on the glucoregulatory, and possibly the antilipolytic, action
of insulin.
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Discussion |
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TopAbstractIntroductionSubjects and MethodsResultsDiscussionReferences |
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The results of this study indicate that the resistance of whole body lipolysis to insulin in obesity is not due to defective regulation of adipose tissue in abdominal and femoral sc depots. A likely candidate for the site of resistance is the intraabdominal adipose tissue. In support of this contention, it has been shown that resistance to the antilipolytic action of insulin in vitro is more pronounced in adipocytes from the omental regions than sc depots (10). Because of the inaccessibility of the intraabdominal fat depot in humans, the contribution of intraabdominal fat to the whole body lipolytic response was indirectly assessed in the current study by simultaneously measuring total lipolysis using stable isotope tracers and regional sc adipose tissue lipolysis using microdialysis. During insulin infusion, there was a greater relative suppression of whole body lipolysis in nonobese than in obese individuals despite no difference in the relative degree of suppression of regional sc adipose tissue lipolysis between these groups. These data suggest that sc adipose tissue is not the site of resistance to the antilipolytic action of insulin in obesity. This is in agreement with the in vitro findings of Mauriege et al. (35), who reported a similar response to the antilipolytic action of insulin in vitro in sc adipocytes from lean and obese individuals.
During insulin infusions, plasma glycerol levels remained 4- to 5-fold higher in obese than in nonobese subjects. These differences could be due to a higher rate of appearance and/or a reduced rate of clearance of glycerol in the obese women compared to the nonobese subjects. Glycerol Ra during the insulin infusions was suppressed to a greater extent in nonobese compared to obese subjects, providing evidence that there is a reduced suppression of lipolysis with obesity. Furthermore, because at steady state Ra = Rd, glycerol clearance also remained elevated in obese subjects compared to nonobese during insulin infusion. As intraabdominal fat volume was much larger in obese than nonobese subjects, it seems possible that resistance to the antilipolytic action of insulin in the intraabdominal adipose tissue of obese subjects accounted for the resistance that was apparent at the whole body level.
Basal lipolysis, as determined by glycerol Ra, was slightly elevated in obese compared to nonobese sedentary individuals. This was apparently due to the increased fat mass in obese individuals, as the basal lipolytic rate was actually higher in nonobese than obese individuals when glycerol Ra was expressed per kg fat mass. Microdialysis glycerol data reflect lipolysis only in the area adjacent to the dialysis membrane, i.e. per given volume of fat. The higher dialysate glycerol concentrations in abdominal and femoral regions in obese individuals therefore make it appear that basal lipolysis per kg fat mass in sc adipose tissue is elevated in obese individuals. This contradiction between whole body and regional microdialysis data can be explained by a lower basal adipose tissue nutritive blood flow in obese than nonobese individuals, resulting in a reduced clearance of glycerol from the interstitial space of the adipose tissue depot in obese individuals, i.e. a higher dialysate glycerol concentration. It can be concluded that an increased rate of basal lipolysis in obese compared to nonobese sedentary individuals is due only to the increased fat mass, rather than increased lipolysis per volume of fat, in obesity. It should be noted that the hormonal milieu of the obese individuals was considerably different from that of the nonobese, with the 3-fold higher plasma insulin concentrations in obese than nonobese individuals under basal conditions, indicating a resistance to the antilipolytic action of insulin. The 2-fold higher noradrenaline concentrations in the obese individuals may, however, have counteracted the antilipolytic action of insulin to a greater extent in the obese than in the nonobese sedentary group under basal conditions.
With respect to regional differences in response to the antilipolytic action of insulin, the data demonstrated that it was the femoral sc adipose tissue that was most resistant to the effects of insulin in both obese and nonobese sedentary individuals. This was particularly evident during the low dose insulin infusion. In both groups, lipolysis was suppressed to a greater extent in abdominal than in femoral sc adipose tissue. This may indicate regional differences in the susceptibility for fat loss, although triglyceride synthesis and response to lipolytic stimuli also play a role in net lipid accumulation.
Microdialysis was used to monitor both lipolysis and nutritive blood flow in sc adipose tissue. Evidence from numerous studies in vivo in humans (30, 36, 37, 38, 39, 40) suggests that adipose tissue blood flow and lipolysis may impact upon each other. Increased adipose tissue blood flow may result in increased supply of plasma triglycerides and hormones associated with lipid metabolism. Conversely, compounds associated with increased lipolytic rate may alter adipose tissue blood flow. It is therefore important from a physiological standpoint to simultaneously monitor blood flow and lipolysis. It is also important to monitor blood flow during microdialysis studies from a methodological standpoint. In the present investigations, the dialysate glycerol concentration was not a direct measure of the interstitial glycerol concentration due to incomplete equilibration (recovery) of glycerol over the dialysis membrane. Changes in nutritive blood flow could therefore potentially affect the recovery of glycerol over the dialysate membrane independent of changes in interstitial glycerol concentration or metabolism, as has been demonstrated for glucose in skeletal muscle (29). However, changes in nutritive blood flow do not change the conclusions about lipolysis drawn from the current dialysate glycerol data for the following reasons: 1) it has recently been shown that changes in adipose tissue blood flow during insulin stimulation do not result in changes in microdialysis probe recovery for glycerol (33); 2) the changes in nutritive blood flow in response to insulin stimulation in the present study were not significantly different between sedentary obese and sedentary nonobese groups; and 3) there was a tendency for nutritive blood flow to increase more in the femoral than abdominal adipose tissue in response to insulin infusion, thereby potentially resulting in overestimation (due to increased removal of glycerol from the interstitium by the increased nutritive blood flow) of the antilipolytic response to insulin in femoral, compared to abdominal, sc adipose tissue. The conclusions drawn of a reduced regional antilipolytic response to insulin in obese compared to nonobese individuals as well as a reduced antilipolytic response to insulin in femoral compared to abdominal sc adipose tissue remain when changes in adipose tissue nutritive blood flow are considered.
The correlation coefficients for the relationship between fat volume
and indicators of whole body insulin action (plasma glycerol, glycerol
Ra, and glucose disposal) ranged from r =
0.51 to r = 0.91. The correlation coefficients were high for all
comparisons and indicate that intraabdominal fat, sc abdominal fat,
total abdominal fat, and total body fat are all related to resistance
to both antilipolytic and glucoregulatory actions of insulin. The
correlations drawn using intraabdominal fat are probably underestimated
compared to measures of other fat depots considering the 3-fold higher
coefficient of variation in the measurement of intraabdominal fat than
other fat depots. These findings are in agreement with the reports of
many investigators who have previously proposed a link between obesity
and insulin resistance (4, 5, 6, 11, 12). Although there is not a
consensus in the literature as to the relative roles of intraabdominal
and sc adipose tissue in insulin resistance (41), it is our contention
that the intraabdominal fat was the site of resistance to the
antilipolytic action of insulin in obese women, as the antilipolytic
response to insulin in sc adipose tissue was similar in obese and
nonobese subjects under nearly every condition. It may be noted that
there was a poor suppression of lipolysis in the femoral adipose tissue
depot of obese women during the low dose insulin infusion; therefore, a
resistance to the antilipolytic action of insulin in the femoral sc
adipose tissue of obese women cannot be completely ruled out. However,
the resistance to the antilipolytic action of insulin in the femoral
adipose depot in obese women was apparent only at a low insulin
infusion rate, which resulted in plasma insulin concentrations of about
17 µU/mL, a concentration similar to the EC50
for suppression of lipolysis by insulin in lean individuals (4).
Suppression of femoral dialysate glycerol was not different between
obese and lean individuals at plasma insulin concentrations (30
µU/mL) similar to the EC50 for suppression of
lipolysis by insulin in obese individuals (4). As resistance to the
suppression of lipolysis by insulin was evident at the whole body level
during both insulin infusions, intraabdominal fat is probably the
principle site of resistance to the antilipolytic action of insulin in
obese individuals. These findings are evident at plasma insulin
concentrations between approximately 15 and 30 µU/mL. A complete
dose-response curve, which could have provided more detailed
information regarding insulin sensitivity with respect to the
antilipolysis, was not obtained. The current findings may therefore not
apply when plasma insulin concentrations are less than those used in
the present experiment. The plasma insulin concentrations attained in
the current study are, however, commonly attained postprandial in both
obese and nonobese individuals.
Some of the obese women in this study had primarily upper body obesity, whereas others had lower body obesity. It would therefore be valuable to perform similar studies in two groups of obese women, one with upper body obesity and the other with lower body obesity. This would allow a more detailed description of the effects of intraabdominal fat in obese individuals.
Effect of training
This is the first study to our knowledge to report whole body (glycerol kinetics) and regional (microdialysis) lipolysis data in trained and sedentary women. The basal glycerol Ra, expressed per unit fat mass, was more than 50% higher in the trained group than in the sedentary nonobese group. This training effect on the basal lipolytic rate may be ascribed to lower plasma insulin levels in the trained group and/or the previously reported higher sensitivity to catecholamines in the trained state (35, 42). The higher basal plasma glycerol concentrations and/or basal glycerol Ra in the trained subjects may also have been due to an acute effect of the exercise that the trained subjects performed 16–24 h before the euglycemic clamp procedure.
The relative suppression of lipolysis by insulin was essentially the same in trained and sedentary nonobese individuals at both the regional and whole body levels. Our data failed to support the hypothesis that endurance-trained women are more sensitive than sedentary nonobese women to the suppression of lipolysis by insulin. Although there is limited information regarding the effects of exercise training on the antilipolytic response to insulin, Suda et al. reported an increased suppression of lipolysis by insulin in exercise-trained rats (43). The similar response to insulin in trained and untrained groups in the present study could have been related to the fact that intraabdominal fat volume was similar in the two groups. This explanation would support the hypothesis that the intraabdominal fat is the primary site of insulin resistance with respect to lipolysis. The negative results should be interpreted with caution, however, as it is possible that the cross-sectional design coupled with the small sample size limited our ability to evaluate the true effects of exercise training on the suppression of lipolysis by insulin. Further investigation using a prospective study design would be needed to fully address the question of the effect of training on the lipolytic response to insulin.
The relative suppression of lipolysis was similar in the trained and sedentary nonobese groups, with no differences in abdominal or femoral dialysate glycerol concentrations. However, plasma glycerol remained more than 2-fold higher in the trained than sedentary nonobese individuals during insulin infusion despite similar glycerol output from sc depots in these two groups. This difference in plasma glycerol was reflected in the 2-fold higher glycerol Ra per kg fat mass in the trained than sedentary nonobese groups. A remaining question is the source of this increased glycerol output in the trained state. Intraabdominal fat stores were similar in the trained and sedentary states, indicating that lipolytic rate must be higher per kg fat mass in the trained state for intraabdominal fat to be the major site of the additional glycerol release. Another source of glycerol could have been im fat depots. This hypothesis is supported by findings that im fat depots are higher in trained than untrained individuals, and that these depots are increased with endurance training (44, 45, 46). As there was no apparent insulin resistance in sedentary compared to trained individuals in sc adipose tissue, inhibition of lipolysis may occur through a different mechanism in muscle than fat. It has recently been suggested that there are different phosphodiesterase subtypes mediating antilipolysis in skeletal muscle (47).
In summary, basal lipolysis per kg fat mass was increased in endurance-trained individuals, but was similar in obese and nonobese individuals. In all groups, the order of the antilipolytic response (determined by glycerol concentrations) to insulin was femoral sc adipose tissue < abdominal sc adipose tissue < plasma. During insulin infusion, there was a greater relative suppression of whole body lipolysis in nonobese than in obese individuals; however, there was no difference in the relative degree of suppression of regional sc adipose tissue lipolysis between these groups. These data indicate that the site of insulin resistance with respect to the antilipolytic action of insulin in obesity was not the sc adipose tissue. The primary candidate for the reduced suppression of lipolysis in obesity was the intraabdominal fat, although a reduction in the antilipolytic action of insulin in muscle could not be ruled out. There was no effect of training status on the relative degree of suppression of lipolysis by insulin.
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Footnotes |
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2 Current address: East Carolina University, 371 Ward Sports Medicine
Building, Greenville, North Carolina 27858.
Received May 11, 1999.
Revised July 2, 1999.
Accepted July 12, 1999.
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References |
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TopAbstractIntroductionSubjects and MethodsResultsDiscussionReferences |
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-2 adrenoceptors in regulating lipid
mobilization from human adipose tissue. J Clin Invest. 91:1997–2003.
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, Greater than the corresponding
value for nonobese untrained (P < 0.05). 
, Less
than basal (P < 0.05).
), during a low dose insulin infusion (
; 6–10
mU/m2·min), and during a moderate dose insulin infusion
(
; 12–20 mU/m2·min) in nonobese trained, nonobese
sedentary, and obese sedentary individuals. Data are expressed as the
mean ± SE. *, Greater than the corresponding value
for nonobese trained (P < 0.05). 
), nonobese
sedentary (
), and obese sedentary (
) individuals. Data are the
mean ± SE and are expressed as a percentage of the
basal value. *, Greater than nonobese trained and sedentary groups
(P < 0.05).