scholarly journals A modified perifused incubation system for isolated fat-cells. Investigation of intermediary metabolism by perifusion

1984 ◽  
Vol 224 (1) ◽  
pp. 235-239
Author(s):  
R D Harper
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Fatty Acid Synthesis ◽  
Acid Synthesis ◽  
Fat Cells ◽  
Isolated Fat Cells ◽  
Glycerol Synthesis ◽  
Perifusion System ◽  
Net Release

Perifused fat-cells showed similar values for acylglycerol glycerol synthesis from glucose with insulin and for the effects of added palmitate to those in normal incubations and those reported in the literature. Fatty acid synthesis was lower in perifused cells compared with normal incubations, and there was a net release of fatty acids only with the perifused fat-cells. Hence fluxes of metabolites were different in the two incubation systems, and the perifusion system enables the investigation of the flux of metabolites under conditions which may more closely resemble those in vivo.

scholarly journals The regulation of glyceride synthesis in isolated white-fat cells. The effects of palmitate and lipolytic agents

1972 ◽  
Vol 128 (5) ◽  
pp. 1057-1067 ◽  
Author(s):  
E. D Saggerson
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Free Fatty Acids ◽  
Fatty Acid Synthesis ◽  
Acid Synthesis ◽  
Fat Cells ◽  
Glyceride Synthesis ◽  
Glucose Incorporation ◽  
High Concentrations ◽  
Stimulation Of

1. 0.5mm-Palmitate stimulated incorporation of [U-14C]glucose into glyceride glycerol and fatty acids in normal fat cells in a manner dependent upon the glucose concentration. 2. In the presence of insulin the incorporation of 5mm-glucose into glyceride fatty acids was increased by concentrations of palmitate, adrenaline and 6-N-2′-O-dibutyryladenosine 3′:5′-cyclic monophosphate up to 0.5mm, 0.5μm and 0.5mm respectively. Higher concentrations of these agents produced progressive decreases in the rate of glucose incorporation into fatty acids. 3. The effects of palmitate and lipolytic agents upon the measured parameters of glucose utilization were similar, suggesting that the effects of lipolytic agents are mediated through increased concentrations of free fatty acids. 4. In fat cells from 24h-starved rats, maximal stimulation of glucose incorporation into fatty acids was achieved with 0.25mm-palmitate. Higher concentrations of palmitate were inhibitory. In fat cells from 72h-starved rats, palmitate only stimulated glucose incorporation into fatty acids at high concentrations of palmitate (1mm and above). 5. The ability of fat cells to incorporate glucose into glyceride glycerol in the presence of palmitate decreased with increasing periods of starvation. 6. It is suggested that low concentrations of free fatty acids stimulate fatty acid synthesis from glucose by increasing the utilization of ATP and cytoplasmic NADH for esterification of these free fatty acids. When esterification of free fatty acids does not keep pace with their provision, inhibition of fatty acid synthesis occurs. Provision of free fatty acids far in excess of the esterification capacity of the cells leads to uncoupling of oxidative phosphorylation and a secondary stimulation of fatty acid synthesis from glucose.

scholarly journals Studies on the role of insulin in the regulation of glyceride synthesis in rat epididymal adipose tissue

1975 ◽  
Vol 150 (3) ◽  
pp. 441-451 ◽  
Author(s):  
S R Sooranna ◽  
E D Saggerson
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Fatty Acid Synthesis ◽  
Acid Synthesis ◽  
Synthesis Process ◽  
Epididymal Adipose Tissue ◽  
Fat Cells ◽  
Fat Cell ◽  
Glyceride Synthesis

1. When rat isolated fat-cells were incubated with fructose and palmitate, insulin significantly stimulated glyceride synthesis as measured by either [14C]fructose incorporation into the glycerol moiety or of [3H]palmitate incorporation into the acyl moiety of tissue glycerides. Under certain conditions the effect of insulin on glyceride synthesis was greater than the effect of insulin on fructose uptake. 2. In the presence of palmitate, insulin slightly stimulated (a) [14C]pyruvate incorporation into glyceride glycerol of fat-cells and (b) 3H2O incorporation into glyceride glycerol of incubated fat-pads. 3. At low extracellular total concentrations of fatty acids (in the presence of albumin), insulin stimulated [14C]fructose, [14C]pyruvate and 3H2O incorporation into fat-cell fatty acids. Increasing the extracellular fatty acid concentration greatly inhibited fatty acid synthesis from these precursors and also greatly decreased the extent of apparent stimulation of fatty acid synthesis by insulin. 4. These results are discussed in relation to the suggestion [A.P. Halestrap & R.M Denton (1974) Biochem. J. 142, 365-377] that the tissue may contain a specific acyl-binding protein which is subject to regulation. It is suggested that an insulin-sensitive enzyme component of the glyceride-synthesis process may play such a role.

scholarly journals The regulation of glyceride synthesis in isolated white-fat cells. The effects of acetate, pyruvate, lactate, palmitate, electron-acceptors, uncoupling agents and oligomycin

1972 ◽  
Vol 128 (5) ◽  
pp. 1069-1078 ◽  
Author(s):  
E. D Saggerson
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Fatty Acid Synthesis ◽  
Acid Synthesis ◽  
Fat Cells ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Normal Cells ◽  
Glyceride Synthesis ◽  
Glucose Incorporation

1. The incorporation of 5mm-[U-14C]glucose into glyceride fatty acids by fat cells from normal rats incubated in the presence of 20munits of insulin/ml was increased by acetate, pyruvate, palmitate, NNN′N′-tetramethyl-p-phenylenediamine, phenazine methosulphate, dinitrophenol, tetrachlorotrifluoromethyl benzimidazole and oligomycin. Lactate did not stimulate glucose incorporation into fatty acids. The effects of these agents were concentration-dependent. 2. In the presence of 5mm-glucose+insulin, [U-14C]acetate, [U-14C]pyruvate and [U-14C]lactate were incorporated into fatty acids in a concentration-dependent manner, thereby further increasing the total rate of fatty acid synthesis. 3. NNN′N′-tetramethyl-p-phenylenediamine decreased the incorporation of [U-14C]pyruvate into fatty acids in normal cells and increased the incorporation of [U-14C]lactate into fatty acids. 4. In fact cells from 72h-starved rats the stimulatory effects of NNN′N′-tetramethyl-p-phenylenediamine upon glucose and lactate incorporation into fatty acids were totally and partially abolished respectively whereas the stimulatory effects of acetate upon glucose incorporation were retained. 5. Combinations of the optimum concentrations of the substances that stimulate glucose incorporation into fatty acids were tested and compared. The effects of acetate+NNN′N′-tetramethyl-p-phenylenediamine and acetate+palmitate upon normal cells were additive. The effects of NNN′N′-tetramethyl-p-phenylenediamine+palmitate were not additive. It was found that total fatty acid synthesis in the presence of glucose was most effectively increased by raising the concentration of pyruvate in the incubation system. 6. The significance of these results in supporting the proposal that fatty acid synthesis from glucose in adipose tissue is a ‘self-limiting process’ is discussed.

scholarly journals Fatty acid elongation by a particulate fraction from germinating pea

1980 ◽  
Vol 191 (3) ◽  
pp. 791-797 ◽  
Author(s):  
B R Jordan ◽  
J L Harwood
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Fatty Acid Synthesis ◽  
High Speed ◽  
Saturated Fatty Acids ◽  
Acyl Chain ◽  
Fatty Acid Synthetase ◽  
Acid Synthesis ◽  
Particulate Fraction

The synthesis of fatty acids from [14C]malonyl-CoA was studied with a high-speed particulate fraction from germinating pea (Pisum sativum). The variety used (Feltham First) produced mainly saturated fatty acids with palmitate (30–40%) and stearate (40–60%) predominating. Several palmitate-containing lipids stimulated overall synthesis and, in addition, increased the percentage of label in stearate. The production of stearate was severely inhibited by preincubation of the microsomal fraction with snake venom phospholipase A2 or by incubation with Rhizopus arrhizus lipase. Addition of a series of di-saturated phosphatidylcholines, with different acyl constituents, resulted in stimulation of overall fatty acid synthesis as well as an increase in the radiolabelling of the fatty acid two carbon atoms longer than the acyl chain added. This chain lengthening of fatty acids donated from phosphatidylcholine was due to the action of both fatty acid synthetase and palmitate elongase. The latter would utilize dipalmitoyl phosphatidylcholine and was sensitive to arsenite whereas fatty acid synthetase would use dilauroyl phosphatidylcholine and was sensitive to cerulenin. The results are discussed in relation to previous data obtained in vivo on plant fatty acid synthesis and current suggestions for the role of phosphatidylcholine in this process.

scholarly journals Regulation of lipogenesis. Stimulation of fatty acid synthesis in vivo and in vitro in the liver of the newly hatched chick

1970 ◽  
Vol 118 (2) ◽  
pp. 259-263 ◽  
Author(s):  
Alan G. Goodridge
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Fatty Acid Synthesis ◽  
Acid Synthesis ◽  
Latent Form ◽  
Triose Phosphate ◽  
Liver Slices ◽  
Stimulation Of

1. A single glucose meal stimulated the incorporation of acetate into fatty acids in liver slices. If the glucose was added in vitro, it had no effect. Fructose and glycerol in vitro markedly stimulated fatty acid synthesis from acetate. Fructose and glycerol probably by-passed a rate-controlling reaction between glucose and triose phosphate. This reaction may have been stimulated by glucose administered in vivo. 2. The stimulation of fatty acid synthesis caused by fructose did not require the synthesis of enzyme, thus indicating that fatty acid-synthesizing enzymes were present in a latent form in the livers from unfed chicks.

scholarly journals Interrelationship and control of glucose metabolism and lipogenesis in isolated fat-cells. Effect of the amount of glucose uptake on the rates of the pentose phosphate cycle and of fatty acid synthesis

1972 ◽  
Vol 128 (5) ◽  
pp. 1089-1096 ◽  
Author(s):  
H. Kather ◽  
M. Rivera ◽  
K. Brand
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Glucose Metabolism ◽  
Glucose Uptake ◽  
Fatty Acid Synthesis ◽  
Acid Synthesis ◽  
Pentose Phosphate ◽  
Fat Cells ◽  
Pentose Phosphate Cycle ◽  
Wide Range

In order to study the quantitative relationship between fatty acid synthesis and pentose phosphate-cycle activity under different hormonal and dietary conditions affecting the extent of glucose uptake, cells isolated from rat epididymal adipose tissue were incubated in bicarbonate buffer containing [U-14C]-, [1-14C]- or [6-14C]-glucose. From the amount of glucose taken up, the production of lactate and pyruvate, and the incorporation of 14C from differently labelled [14C]glucose into CO2, fatty acids and glyceride glycerol, the rates of glucose metabolism via different pathways and the extent of lipogenesis under various experimental conditions were determined. The contribution of the pentose phosphate-cycle to glucose metabolism under normal conditions was calculated to be 8%. Starvation and re-feeding, and the presence of insulin, caused an enhancement of glucose uptake, pentose phosphate-cycle activity and fatty acid synthesis. Plots of both pentose phosphate-cycle activity and fatty acid synthesis versus glucose uptake revealed that the extent of glucose uptake, over a wide range, determines the rates of fatty acid synthesis and glucose metabolism via the pentose phosphate cycle. A balance of formation and production of nicotinamide nucleotides in the cytoplasm was established. The total amount of cytoplasmic NADH and NADPH formed was only in slight excess over the hydrogen equivalents required for the synthesis of fatty acids, glyceride glycerol and lactate. Except in cells from starved animals, the pentose phosphate cycle was found to provide only about 60% of the NADPH required for fatty acid synthesis. The results are discussed with respect to an overall control of the different metabolic and biosynthetic reactions in the fat-cells by the amount of glucose transported into the cell.

scholarly journals Interrelationship and control of glucose metabolism and lipogenesis in isolated fat-cells. Control of pentose phosphate-cycle activity by cellular requirement for reduced nicotinamide adenine dinucleotide phosphate

1972 ◽  
Vol 128 (5) ◽  
pp. 1097-1102 ◽  
Author(s):  
H. Kather ◽  
M. Rivera ◽  
K. Brand
Keyword(s):  
Fatty Acid ◽  
Glucose Metabolism ◽  
Fatty Acid Synthesis ◽  
Nicotinamide Adenine Dinucleotide Phosphate ◽  
Acid Synthesis ◽  
Pentose Phosphate ◽  
Fat Cells ◽  
Isolated Fat Cells ◽  
Pentose Phosphate Cycle ◽  
Phenazine Methosulphate

By using inhibitors and stimulators of different metabolic pathways the interdependence of the pentose phosphate cycle and lipogenesis in isolated fat-cells was studied. Rotenone, which is known to inhibit electron transport in the respiratory chain, blocked glucose breakdown at the site of pyruvate dehydrogenase. Consequently, because of the lack of acetyl-CoA, fatty acid synthesis was almost abolished. A concomitant decrease in pentose phosphate-cycle activity was observed. Phenazine methosulphate stimulated pentose phosphate-cycle activity about five- to ten-fold without a considerable effect on fatty acid synthesis. The influence of rotenone on both the pentose phosphate cycle and lipogenesis could be overcome by addition of phenazine methosulphate, indicating that rotenone has no direct effect on these pathways. The decreased rate of the pentose phosphate cycle in the presence of rotenone therefore has to be considered as a consequence of decreased fatty acid synthesis. The rate of glucose catabolism via the pentose phosphate cycle in adipocytes appears to be determined by the requirement of NADPH for lipogenesis. Treatment of cells with 6-aminonicotinamide caused an accumulation of 6-phosphogluconate, indicating an inhibition of 6-phosphogluconate dehydrogenase. The rate of glucose metabolism via the pentose phosphate cycle as well as the rate of fatty acid synthesis, however, was not affected by 6-aminonicotinamide treatment and could still be stimulated by addition of insulin. Since even in cells from starved animals, in which the pentose phosphate-cycle activity is extremely low, no accumulation of 6-phosphogluconate was observed, it is concluded that the control of this pathway is achieved by the rate of regeneration of NADP at the site of glucose 6-phosphate dehydrogenase.

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