At physiologic albumin/oleate concentrations oleate uptake by isolated hepatocytes, cardiac myocytes, and adipocytes is a saturable function of the unbound oleate concentration. Uptake kinetics are consistent with the conventional theory.

Sorrentino, Dario; Robinson, Richard B.; Kiang, Chih-Li; Berk, Paul D.

doi:10.1172/jci114301

Cited by 128 publications

(85 citation statements)

References 63 publications

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“…23,[37][38][39][40] At least four proteins have been proposed as LCFA transporters: plasma membrane fatty acid-binding protein (FABPpm), 32,37-39 fatty acid translocase (FAT), 30 the fatty acid transport polypeptide family (FATP) [41][42][43] and caveolin-1. 44,45 While there is good evidence supporting a role in LCFA transport for these proteins, there are also unresolved issues that have led to persistent doubts about each.…”

Section: Discussionmentioning

confidence: 99%

Differences in adipocyte long chain fatty acid uptake in Osborne–Mendel and S5B/Pl rats in response to high-fat diets

et al. 2008

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Objective: To determine whether strain differences in adipocyte uptake of long chain fatty acids (LCFAs) contribute to differences in weight gain by Osborne-Mendel (OM) and S5B/Pl rats (S) fed a high-fat diet (HFD). Subjects: Ninety-four adult (12-14-week old) male OM and S rats. Measurements: Body weight; epididymal fat pad weight; adipocyte size, number, LCFA uptake kinetics; and plasma insulin and leptin during administration of HFD or chow diets (CDs). Results: In both strains, rate of weight gain (RWG) was greater on an HFD than a CD; RWG on an HFD was greater, overall, in OM than S. A significant RWG increase occurred on days 1 and 2 in both strains. It was normalized in S by days 6-9 but persisted at least till day 14 in OM. RWGs were significantly correlated (Po0.001) with the V max for saturable adipocyte LCFA uptake (V max ). In S, an increase in V max on day 1 returned to baseline by day 7 and was correlated with both plasma insulin and leptin levels throughout. In OM, a greater increase in V max was evident by day 2, and persisted for at least 14 days, during which both insulin and leptin levels remained elevated. Growth in epididymal fat pads on the HFD correlated with body weight, reflecting hypertrophy in OM and both hypertrophy and hyperplasia in S. Conclusions: (a) Changes in V max contribute significantly to changes in RWG on HFDs. (b) There are important strain differences in circulating insulin and leptin responses to an HFD. (c) Both insulin and leptin responses to an HFD are closely correlated with V max of adipocyte fatty acid uptake in S animals, but suggest early onset of insulin resistance in OM. Thus, differences in hormonal regulation of adipocyte LCFA uptake may underlie the different responses of OM and S to HFD.

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Section: Discussionmentioning

confidence: 99%

Differences in adipocyte long chain fatty acid uptake in Osborne–Mendel and S5B/Pl rats in response to high-fat diets

et al. 2008

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“…, to determine the dissociation and fluorometric constants of Equation 2 which relates [FFA u ] to R. (2) In Equation 2, R 0 is the value of R when no FFA is present, Q is the ratio of the intensity at 457 nm when no FFA is present to that when the probe is saturated with FFA, K d is the dissociation constant, and R m is R at saturation. Probe calibration was a two-step process involving the titration of probes with (i) un-buffered and (ii) BSA-buffered FFA stocks with known total ([FA t ] = [FFA bound ] + [FFA u ]) and unbound FFA concentrations, respectively.…”

Section: Probe Calibrationmentioning

confidence: 99%

“…with targets such as protein receptors and cell membranes (for example (2)). A profiling of unbound metabolite concentrations should therefore provide the most accurate measure of physiologic health.…”

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Fatty Acid-Specific Fluorescent Probes and Their Use in Resolving Mixtures of Unbound Free Fatty Acids in Equilibrium with Albumin

et al. 2006

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We report the first measurements to profile mixtures of unbound free fatty acids. Measurements utilized fluorescent probes with distinctly different response profiles for different free fatty acids (FFA). These probes were constructed by labeling site-specific mutants of the rat intestinal fatty acid binding protein (rI-FABP) with acrylodan. The probes were produced and screened by high throughput methods and from more than 30,000 such probes we selected 6 that together have sufficient specificity and sensitivity to resolve the profile of unbound FFA (FFA u ) in mixtures of different FFA u . We developed analytical methods to determine the FFA u profile from the fluorescence (ratio) response of the different probes and used these methods to determine FFA u profiles for mixtures of arachidonate, linoleate, oleate, palmitate and stearate in equilibrium with bovine serum albumin (BSA). Measurements were performed using mixtures with a range of total FFA u concentrations, including 0.9 nM, which is similar to normal plasma levels. We also measured single FFA binding isotherms for BSA and found that binding was well described by 6-7 sites with the same binding constants (K d ). The K d values for the FFA (4 to 38 nM) were inversely related to the aqueous solubility of the FFA. We constructed a model with these parameters to predict the FFA u profile in equilibrium with BSA and found excellent agreement between the profiles measured using the FFA probes and those calculated with this model. These results should lead to a better understanding of albumin's role in buffering FFA u and to profiling FFA u in intra and extracellular biological fluids. Keywordsunbound free fatty acids; fatty acid binding proteins; fluorescent probes; site-specific mutants; high throughput screening; fatty acid profiling; binding affinities; bovine serum albumin Metabolomics is advancing rapidly as a result of new technologies and the expanding interest in systems biology (1). This advance is being driven by the recognition that physiologic phenotype is essentially a reflection of the metabolic profile, and therefore, metabolic profiling should provide an accurate representation of the states of health and disease. The activity of a given metabolite is frequently dictated by its solubility as a "free" or unbound molecule in aqueous bodily fluids. For many metabolites the unbound concentration represents a small fraction of the total, with most of the total metabolite bound in carrier complexes. Total metabolite concentrations are typically measured, but it is the unbound metabolite that interacts

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“…After conversion to micrometers (m) (1 U ¼ 9.6 mm), the corresponding mean cell surface area (SA), in m 2 , was calculated. 10 LCFA uptake studies Cell aliquots from each preparation were incubated at 371C in DMEM containing 500 mM BSA 11,12 and one of five different concentrations of OA, such that the OA:BSA molar ratio (n) was 0.25, 0.5, 1.0, 1.5, or 2.0 : 1. The initial velocity (V 0 ) of cellular oleate uptake from each test solution was determined by a standard, rapid filtration technique [7][8][9]13 from four samples obtained in triplicate over the initial 30 s of incubation, during which uptake was a linear function of time.…”

Section: Preparation Of Isolated Adipocytesmentioning

confidence: 99%

Long-chain fatty acid uptake is upregulated in omental adipocytes from patients undergoing bariatric surgery for obesity

et al. 2004

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OBJECTIVE:To determine the impact of obesity on adipocyte cell size and long-chain fatty acid (LCFA) uptake kinetics in human subjects undergoing laparoscopic abdominal surgery. SUBJECTS: A total of 10 obese patients (BMI 49.8711.9 (s.d.) kg/m 2 ) undergoing laparoscopic bariatric surgery, and 10 nonobese subjects (BMI 24.272.3 kg/m 2 ) undergoing other clinically indicated laparoscopic abdominal surgical procedures. MEASUREMENTS: Cell size distribution and [ 3 H]oleic acid uptake kinetics were studied in adipocytes isolated from omental fat biopsies obtained during surgery. Adipocyte surface area (SA) was calculated from the measured cell diameters. Plasma leptin and insulin concentrations were measured by RIA in fasting blood samples obtained on the morning of surgery. RESULTS: The mean SA of obese adipocytes (41 50875381 m 2 /cell) was increased 2.4-fold compared to that of nonobese adipocytes (16 92876529 m 2 /cell; Po0.01). LCFA uptake in each group was the sum of saturable and nonsaturable components. Both the V max of the saturable component (21.376.3 vs 5.171.9 pmol/s/50 000 cells) and the rate constant k of the nonsaturable component (0.01570.002 vs 0.006670.0023 ml/s/50 000 cells) were increased (Po0.001) in obese adipocytes compared with nonobese controls. When expressed relative to cell size, V max /m 2 SA was greater in obese than nonobese adipocytes (Po0.05), whereas k/m 2 SA did not differ between the groups. CONCLUSION: The data support the concepts that (1) adipocyte LCFA uptake consists of distinct facilitated (saturable) and diffusive processes; (2) increased saturable LCFA uptake in obese adipocytes is not simply a consequence of increased cell size, but rather reflects upregulation of a facilitated transport process; and (3) the permeability of adipocyte plasma membranes to LCFA is not appreciably altered by obesity, and increased nonsaturable uptake in obese adipocytes principally reflects an increase in cell SA. Regulation of saturable LCFA uptake by adipocytes may be an important control point for body adiposity.

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At physiologic albumin/oleate concentrations oleate uptake by isolated hepatocytes, cardiac myocytes, and adipocytes is a saturable function of the unbound oleate concentration. Uptake kinetics are consistent with the conventional theory.

Cited by 128 publications

References 63 publications

Differences in adipocyte long chain fatty acid uptake in Osborne–Mendel and S5B/Pl rats in response to high-fat diets

Differences in adipocyte long chain fatty acid uptake in Osborne–Mendel and S5B/Pl rats in response to high-fat diets

Fatty Acid-Specific Fluorescent Probes and Their Use in Resolving Mixtures of Unbound Free Fatty Acids in Equilibrium with Albumin

Long-chain fatty acid uptake is upregulated in omental adipocytes from patients undergoing bariatric surgery for obesity

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