Asem H. Ali

Storage of Circulating Free Fatty Acid in Adipose Tissue of Postabsorptive Humans Quantitative Measures and Implications for Body Fat Distribution

OBJECTIVE Preferential upper-body fat gain, a typical male pattern, is associated with a greater cardiometabolic risk. Regional differences in lipolysis and meal fat storage cannot explain sex differences in body fat distribution. We examined the potential role of the novel free fatty acid (FFA) storage pathway in determining body fat distribution in postabsorptive humans and whether adipocyte lipogenic proteins (CD36, acyl-CoA synthetases, and diacylglycerol acyltransferase) predict differences in FFA storage. RESEARCH DESIGN AND METHODS Rates of postabsorptive FFA (palmitate) storage into upper-body subcutaneous (UBSQ) and lower-body subcutaneous (LBSQ) fat were measured in 28 men and 53 premenopausal women. Stable and radiolabeled palmitate tracers were intravenously infused followed by subcutaneous fat biopsies. Body composition was assessed with a combination of dual-energy X-ray absorptiometry and computed tomography. RESULTS Women had greater FFA (palmitate) storage than men in both UBSQ (0.37 ± 0.15 vs. 0.27 ± 0.18 μmol · kg−1 · min−1, P = 0.0001) and LBSQ (0.42 ± 0.19 vs. 0.22 ± 0.11 μmol · kg−1 · min−1, P < 0.0001) fat. Palmitate storage rates were significantly greater in LBSQ than UBSQ fat in women, whereas the opposite was true in men. Plasma palmitate concentration positively predicted palmitate storage in both depots and sexes. Adipocyte CD36 content predicted UBSQ palmitate storage and sex-predicted storage in LBSQ fat. Palmitate storage rates per kilogram fat did not decrease as a function of fat mass, whereas lipolysis did. CONCLUSIONS The FFA storage pathway, which had remained undetected in postabsorptive humans until recently, can have considerable, long-term, and sex-specific effects on body fat distribution. It can also offer a way of protecting the body from excessive circulating FFA in obesity.

Free Fatty Acid Storage in Human Visceral and Subcutaneous Adipose Tissue: Role of Adipocyte Proteins

OBJECTIVE Because direct adipose tissue free fatty acid (FFA) storage may contribute to body fat distribution, we measured FFA (palmitate) storage rates and fatty acid (FA) storage enzymes/proteins in omental and abdominal subcutaneous fat. RESEARCH DESIGN AND METHODS Elective surgery patients received a bolus of [1-14C]palmitate followed by omental and abdominal subcutaneous fat biopsies to measure direct FFA storage. Long chain acyl-CoA synthetase (ACS) and diacylglycerol acyltransferase activities, CD36, fatty acid-binding protein, and fatty acid transport protein 1 were measured. RESULTS Palmitate tracer storage (dpm/g adipose lipid) and calculated palmitate storage rates were greater in omental than abdominal subcutaneous fat in women (1.2 ± 0.8 vs. 0.7 ± 0.4 μmol ⋅ kg adipose lipid−1 ⋅ min−1, P = 0.005) and men (0.7 ± 0.2 vs. 0.2 ± 0.1, P < 0.001), and both were greater in women than men (P < 0.0001). Abdominal subcutaneous adipose tissue palmitate storage rates correlated with ACS activity (women: r = 0.66, P = 0.001; men: r = 0.70, P = 0.007); in men, CD36 was also independently related to palmitate storage rates. The content/activity of FA storage enzymes/proteins in omental fat was dramatically lower in those with more visceral fat. In women, only omental palmitate storage rates were correlated (r = 0.54, P = 0.03) with ACS activity. CONCLUSIONS Some adipocyte FA storage factors correlate with direct FFA storage, but sex differences in this process in visceral fat do not account for sex differences in visceral fatness. The reduced storage proteins in those with greater visceral fat suggest that the storage factors we measured are not a predominant cause of visceral adipose tissue accumulation.

A novel ELISA for measuring CD36 protein in human adipose tissue

CD36 is a transmembrane protein present in many tissues that is believed to facilitate inward fatty acid transport. Western blotting is the most widely used method to measure tissue CD36 protein content, but it is time consuming, technically demanding, and semiquantitative. To more precisely measure adipose tissue CD36 content we developed an enzyme linked immunosorbent assay (ELISA) after establishing that: 1) the anti-CD36 antibodies gave a single distinct band on traditional Western blots, and 2) the vast majority of adipocyte CD36 resides in the plasma membrane. By using serial dilutions of each sample and including a calibrator sample and quality control sample on each plate, we could achieve inter- and intra-assay variability of ∼ 10%. We found that CD36 content in omental and abdominal subcutaneous adipose tissue varied over a 2–5-fold range depending upon the means of data expression (per units of tissue protein, weight, or lipid). Omental CD36 content in women decreased markedly (P = 0.01) as a function of fat cell size. For the most part, tissue CD36 content was not correlated with CD36 mRNA. This ELISA method for tissue CD36 content should enhance research into the role of this protein on tissue fatty acid uptake.

Storage Rates of Circulating Free Fatty Acid Into Adipose Tissue During Eating or Walking in Humans

We measured subcutaneous adipose tissue free fatty acid (FFA) storage rates in postprandial and walking conditions to better understand the contributions of this pathway to body fat distribution. Palmitate tracers were infused intravenously and fat biopsies collected to measure palmitate storage in upper- (UBSQ) and lower-body subcutaneous (LBSQ) fat in 41 (17 men) and 40 (16 men) volunteers under postprandial and under postabsorptive walking conditions, respectively. Postprandial palmitate storage was greater in women than men in UBSQ (0.50 ± 0.25 vs. 0.33 ± 0.37 μmol ⋅ kg fat−1 ⋅ min−1; P = 0.007) and LBSQ fat (0.37 ± 0.25 vs. 0.22 ± 0.20 μmol ⋅ kg fat−1 ⋅ min−1; P = 0.005); storage rates were significantly greater in UBSQ than LBSQ fat in both sexes. During walking, UBSQ palmitate storage did not differ between sexes, whereas LBSQ storage was greater in women than men (0.40 ± 0.22 vs. 0.25 ± 0.15 μmol ⋅ kg fat−1 ⋅ min−1; P = 0.01). In women only, walking palmitate storage was significantly greater in LBSQ than UBSQ fat. Adipocyte CD36 and diacylglycerol acyltransferase (DGAT) correlated with LBSQ palmitate storage in the postprandial and walking condition, respectively. We conclude that UBSQ fat is the preferred postprandial FFA storage depot for both sexes, whereas walking favors storage in LBSQ fat in women. Transmembrane transport (CD36) and esterification into triglycerides (DGAT) may be rate-limiting steps for LBSQ FFA storage during feeding and exercise.

Fatty acid metabolism in the elderly: effects of dehydroepiandrosterone and testosterone replacement in hormonally deficient men and women.

CONTEXT Aging, low dehydroepiandrosterone (DHEA), and testosterone are associated with increased adiposity and metabolic risk. Treatment with these hormones may improve these abnormalities. OBJECTIVE The objective of the study was to determine effects of aging, DHEA, or testosterone replacement on adiposity, meal fat partitioning, and postabsorptive lipolysis. DESIGN This was a cross-sectional, 2-yr, double-blind, randomized, placebo-controlled trial. SETTING The study was conducted in the general community. PATIENTS Elderly women and men (>or=60 yr) with low DHEA sulfate (women and men) and bioavailable testosterone (men) concentrations and young adults. INTERVENTIONS Thirty elderly women each received 50 mg DHEA or placebo daily for 2 yr. Thirty elderly men received 75 mg DHEA, 29 received 5 mg testosterone (patch), and 32 received placebo daily for 2 yr. Thirty young women and 32 young men served as controls. MAIN OUTCOME MEASURES In vivo measures of meal fat storage into sc fat, postabsorptive lipolysis, and regional adiposity at baseline and after treatment. RESULTS At baseline, the elderly had more body fat, greater systemic lipolysis (women, P = 0.0003; men, P < 0.0001) adjusted for resting energy expenditure, greater meal fat oxidation (women, P = 0.026; men, P = 0.0025), and less meal fat storage in sc fat (women, P = 0.0139; men, P= 0.0006). Although testosterone treatment increased meal fat storage into upper- vs. lower-body fat in elderly men, neither hormone affected regional adiposity, meal fat oxidation, or systemic lipolysis. CONCLUSIONS Aging, in the context of low DHEA sulfate (women and men) and bioavailable testosterone (men) concentrations, is associated with changes in meal fat partitioning and postabsorptive lipolysis that are not corrected by DHEA and only partly corrected by testosterone replacement.

Acute effects of betahistine hydrochloride on food intake and appetite in obese women: a randomized, placebo-controlled trial

BACKGROUND Central nervous system histaminergic tone is thought to play a role in appetite regulation. In animal models, histamine receptor 1 (HRH1) agonists and histamine receptor 3 (HRH3) antagonists decrease food intake. OBJECTIVE The objective of this study was to examine the acute effects of betahistine hydrochloride (an HRH1 agonist and HRH3 antagonist) on food intakes and appetites. DESIGN The study was a proof-of-concept, randomized, double-blinded, placebo-controlled, dose-ranging study performed to examine the effects of betahistine in women with class I or II obesity [body mass index (BMI; in kg/m²) of 30-39.99]. After a 24-h placebo run-in period, subjects received a placebo (n = 19) or 48 (n = 19), 96 (n = 17), or 144 (n = 21) mg betahistine/d for 24 h. Treatment was followed by a buffet test meal to assess energy intake. Hunger, satiety, and desire to eat were measured after consuming the meal by using visual analog scales. Data were analyzed by using regression models with the assumption that there would be an increasing effect of betahistine doses. Analyses were adjusted for age, log fat and lean mass, food preferences, and intake during a buffet test meal obtained during the placebo run-in period. RESULTS Of the 79 obese women (mean ± SD age: 42 ± 11 y; BMI: 35 ± 3) enrolled in the study, 76 women completed the study. The betahistine dose did not significantly change intakes from those observed during the run-in period of the buffet test meal (P = 0.78). Hunger, fullness, and desire to eat (all P > 0.62) similarly showed no differences according to the betahistine dose. CONCLUSIONS Betahistine did not produce an effect on food intakes or appetites. More potent histaminergic modulators may be required to elucidate the possible role of histaminergic pathways in human obesity. This trial was registered at clinicaltrials.gov as NCT00459992.

Sex-Associated Differences in Free Fatty Acid Flux of Obese Adolescents

CONTEXT In obesity, increases in free fatty acid (FFA) flux can predict development of insulin resistance. Adult women release more FFA relative to resting energy expenditure (REE) and have greater FFA clearance rates than men. In adolescents, it is unknown whether sex differences in FFA flux occur. OBJECTIVE Our objective was to determine the associations of sex, REE, and body composition with FFA kinetics in obese adolescents. PARTICIPANTS Participants were from a convenience sample of 112 non-Hispanic white and black adolescents (31% male; age range, 12-18 years; body mass index SD score range, 1.6-3.1) studied before initiating obesity treatment. MAIN OUTCOME MEASURES Glucose, insulin, and FFA were measured during insulin-modified frequently sampled iv glucose tolerance tests. Minimal models for glucose and FFA calculated insulin sensitivity index (SI) and FFA kinetics, including maximum (l0 + l2) and insulin-suppressed (l2) lipolysis rates, clearance rate constant (cf), and insulin concentration for 50% lipolysis suppression (ED50). Relationships of FFA measures to sex, REE, fat mass (FM), lean body mass (LBM) and visceral adipose tissue (VAT) were examined. RESULTS In models accounting for age, race, pubertal status, height, FM, and LBM, we found sex, pubertal status, age, and REE independently contributed to the prediction of l2 and l0 + l2 (P < .05). Sex and REE independently predicted ED50 (P < .05). Sex, FM/VAT, and LBM were independent predictors of cf. Girls had greater l2, l0 + l2 and ED50 (P < .05, adjusted for REE) and greater cf (P < .05, adjusted for FM or VAT) than boys. CONCLUSION Independent of the effects of REE and FM, FFA kinetics differ significantly in obese adolescent girls and boys, suggesting greater FFA flux among girls.

Systemic Free Fatty Acid Disposal Into Very Low-Density Lipoprotein Triglycerides

We measured the incorporation of systemic free fatty acids (FFA) into circulating very low-density lipoprotein triglycerides (VLDL-TGs) under postabsorptive, postprandial, and walking conditions in humans. Fifty-five men and 85 premenopausal women with BMI 18–24 (lean) and 27–36 kg/m2 (overweight/obese) received an intravenous bolus injection of [1,1,2,3,3-2H5]glycerol (to measure VLDL-TG kinetics) and either [1-14C]palmitate or [9,10-3H]palmitate to determine the proportion of systemic FFA that is converted to VLDL-TG. Experiments started at 0630 h after a 12-h overnight fast. In the postabsorptive protocol, participants rested and remained fasted until 1330 h. In the postprandial protocol, volunteers ingested frequent portions of a fat-free smoothie. In the walking protocol, participants walked on a treadmill for 5.5 h at ∼3× resting energy expenditure. Approximately 7% of circulating FFA was converted into VLDL-TG. VLDL-TG secretion rates (SRs) were not statistically different among protocols. Visceral fat mass was the only independent predictor of VLDL-TG secretion, explaining 33–57% of the variance. The small proportion of systemic FFA that is converted to VLDL-TG can confound the expected relationship between plasma FFA concentration and VLDL-TG SRs. Regulation of VLDL-TG secretion is complex in that, despite a broad spectrum of physiological FFA concentrations, VLDL-TG SRs did not vary based on different acute substrate availability.

Adipose Tissue Free Fatty Acid Storage in vivo – Effects of Insulin versus Niacin as a Control for Suppression of Lipolysis

Insulin stimulates the translocation fatty acid transport protein 1 (FATP1) to plasma membrane, and thus greater free fatty acid (FFA) uptake, in adipocyte cell models. Whether insulin stimulates greater FFA clearance into adipose tissue in vivo is unknown. We tested this hypothesis by comparing direct FFA storage in subcutaneous adipose tissue during insulin versus niacin-medicated suppression of lipolysis. We measured direct FFA storage in abdominal and femoral subcutaneous fat in 10 and 11 adults, respectively, during euglycemic hyperinsulinemia or after oral niacin to suppress FFA compared with 11 saline control experiments. Direct palmitate storage was assessed using a [U-13C]palmitate infusion to measure palmitate kinetics and an intravenous palmitate radiotracer bolus/timed biopsy. Plasma palmitate concentrations and flux were suppressed to 23 ± 3 and 26 ± 5 µmol ⋅ L−1 (P = 0.91) and 44 ± 4 and 39 ± 5 µmol ⋅ min−1 (P = 0.41) in the insulin and niacin groups, respectively, much less (P < 0.001) than the saline control group (102 ± 8 and 104 ± 12 µmol ⋅ min−1, respectively). In the insulin, niacin, and saline groups, abdominal palmitate storage rates were 0.25 ± 0.05 vs. 0.25 ± 0.07 vs. 0.32 ± 0.05 µmol ⋅ kg adipose lipid−1 ⋅ min−1, respectively (P = NS), and femoral adipose storage rates were 0.19 ± 0.06 vs. 0.20 ± 0.05 vs. 0.31 ± 0.05 µmol ⋅ kg adipose lipid−1 ⋅ min−1, respectively (P = NS). In conclusion, insulin does not increase FFA storage in adipose tissue compared with niacin, which suppresses lipolysis via a different pathway.

Measuring Plasma Fatty Acid Oxidation with Intravenous Bolus Injection of 3H- and 14C-Fatty Acid

Accurate measures of plasma FA oxidation can improve our understanding of diseases characterized by impaired FA oxidation. We describe and compare the 24 h time-courses of FA oxidation using bolus injections of [1-14C]palmitate versus [9,10-3H]palmitate under postabsorptive, postprandial, and walking conditions. Fifty-one men and 95 premenopausal women participated in one condition (postabsorptive, postprandial, or walking), one tracer (14C- or 3H-labeled), and an acetate or palmitate study. Groups were matched for sex, age, and body mass index (BMI). At 24 h, cumulative [3H]acetate recovery as 3H2O was 80 ± 6%, 78 ± 2%, and 81 ± 6% in the postabsorptive, postprandial, and walking conditions, respectively (not significant). Model-predicted maximum [1-14C]acetate recovery as expired 14CO2 was 59 ± 12%, 52 ± 8%, and 65 ± 10% in the postabsorptive, postprandial, and walking condition, respectively (one way ANOVA, P = 0.12). When corrected with the corresponding acetate recovery factors, 24 h time-courses of FFA oxidation were similar between [1-14C]palmitate and [9,10-3H]palmitate in all three conditions. In contrast to previous meal ingestion studies, an acetate-hydrogen recovery factor was needed to achieve comparable oxidation rates using an intravenous bolus of [3H]palmitate. In conclusion, intravenous boluses of [9,10-3H]palmitate versus [1-14C]palmitate gave similar estimates of 24 h cumulative FFA oxidation in age-, sex- and BMI-matched individuals.