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Unsaturated fatty acids potentiate insulin action on adipocytes
Prog, Lipid Res. Vol. 25, pp. 511-513, 1986 Printed in Great Britain. All rights reserved
0163-7827/86/$0.00 + 0.50 © 1986 Pergamon Journals Ltd
UNSATURATED FATTY ACIDS POTENTIATE INSULIN ACTION ON ADIPOCYTES F. LOMEO,M. A. K H o ~
and P. DANDON^
Metabolic Unit, Department of Chemical Pathology and Human Metabolism, Royal Free Hospital and School of Medicine, London NW3 2QG, U.K.
INTRODUCTION
We have recently shown that plasma nonesterified fatty acid (NEFA) concentrations are markedly lowered in healthy and diabetic subjects to whom dihomo-T-linolenic acid (DHLA) has been administered. This suggested that DHLAs may exert a pro-insulin effect, contrary to the concept that fatty acids are antagonistic to insulin action. 7 We have thus investigated the effects of DHLA and other fatty acids on insulin action on isolated rat adipocytes. METHODS
Solubilization of Fatty Acids NEFAs were dissolved in small volumes of ethanol and added to Krebs Ringer Bicarbonate Buffer (KRB) containing 3% NEFA free bovine serum albumin. The final volume of ethanol present in the incubates was 30 ml/mol NEFA/I. Unsaturated NEFAs used were: oleic (9-18:1), linoleic (9,12-18:2), linolenic (9,12,15-18:3), DHLA (8, 11, 14-20:3) and arachidonic (5, 8, 11, 14-20:4). Saturated NEFAs used were: palmitic (16:0) and stearic (18:0).
Assay for Triglyceride Synthesizing Adipocytes Fat cell suspensions were prepared from epididymal fat pads and incubated in KRB (pH 7.4) in the presence of test substances and [U-14C]glucose.I Lipids were then extracted and portions counted to give total triglyceride synthesis, while further portions were hydrolyzed and the glycerol and NEFA fractions counted separately. 3 Experiments were performed at glucose concentrations of 0.55 mmol/l (0.7 Ci/mol) and 3 mmol/l (2 Ci/mol). In this paper, data obtained using linolenic acid as a representative unsaturated NEFA are presented. Similar results were obtained using the other unsaturated NEFAs. RESULTS
Addition of the various unsaturated NEFA solutions to incubates did not alter the incorporation of glucose into triglyceride at 0.55 mmol/l of glucose. However, glucose incorporation into the fatty acid moiety of triglyceride was inhibited, whereas incorporation into the glycerol moiety was stimulated (Table 1). Glucose incorporation into both the fatty acid and glycerol moieties of triglyceride was increased by the unsaturated NEFAs at 3 mmol/l of glucose (Table 2). Concomitant incorporation of insulin and unsaturated NEFA resulted in a potentiation of insulin-stimulated glucose incorporation into both the fatty acid and glycerol moieties of triglyceride (Tables 1 and 2). However, at the higher glucose concentration, the increase in incorporation into the fatty acid moiety was more marked (Table 2). No stimulatory effect of saturated NEFA was observed, nor did these NEFAs produce a potentiation of insulin action. Ethanol produced a mild inhibition of glucose incorporation into glycerol and fatty acids at concentrations used to "carry" fatty acids into aqueous solution. 511
F. Lomeo
512
et
al.
TABLE 1. Percent Stimulation Over Basal of Triglyceride, Triglyceride Fatty Acid and Triglyceride Glycerol by Insulin and Linolenic Acid, Alone and in Combination, in the Presence of 0.55 mmol
Glucose % Stimulation over basal (mean 4-SEM. n = 6)
Additions I0 mU/1 insulin
Triglyceride
Triglyceride fatty acid
Triglyceride glycerol
136 5:9
204 + 10
97 + 6
0.5 mmol/l
linolenic acid
17 4- 4
- 4 5 5:3
78 + 8
10 mU/l insulin + 0.5 mmol/l
linoenic acid
363 _ 12 885:16
251 5:11
449 _ 16
Basal metabolic rate (nmol glUCO~/I0 7 cells/½hr) 365:8 36+8
TABLE 2. Percent Stimulation Over Basal of Triglyceride, Triglyceride Fatty Acid and Triglyceride Glycerol Synthesis by Insulin and Linolenic Acid, Alone and in Combination; in
the Presence of 3 mmol Glucose % Stimulation over basal (mean + SEM, n = 6)
Additions 10 mU/1 insulin
Triglyceride
Triglyceride fatty acid
Triglyceride glycerol
148 5:10
143 + 17
77 + 10
153 + I0
79 + 6
195 + 17
743 5:23
706 + 34
504 + 12
0.5 mmol/l
linolenic acid 10 mU/l insulin + 0.5 mmol/l
linolenic acid
Basal metabolic rate (nmol glucose/107 cells/~ hr) 260 + 60
192 + 44
84 5:20
DISCUSSION
Our data show that the concomitant incubation of unsaturated NEFA with adipocytes potentiates the stimulatory action of insulin on lipogenesis. This confirms the suggestion that NEFAs, in the presence of insulin, stimulate the incorporation of glucose into triglyceride giycerolJ However, our data also show that unsaturated NEFAs potentiate insulin-stimulated de n o v o fatty acid synthesis and that this effect becomes more marked on raising the glucose concentration to physiological levels. The discrepancy between these data and the principle that NEFAs are antagonistic to insulin action is explicable as the concept suggested by Randle e t al. 7 was based on the effects of NEFAs on muscle metabolism. Adipocyte metabolism, especially adipocyte lipogenesis, was not included in the investigation. Unsaturated NEFAs (arachidonic 2 and oleic and va~enic acids ~) enhance membrane fluidity in artificial systems and accelerate the passive diffusion and carrier-mediated uptake of glucose. 6 It is possible that the unsaturated NEFAs also get incorporated into cell membranes, and thus alter the permeability and uptake of glucose under the influence of insulin. It is of interest that oleic acid has been reported to exert glucagon-like effects upon hepatic NEFA metabolism, 4 which contrasts with the insulin-like effects reported here. Thus, in the light of present knowledge, unsaturated NEFAs would enhance synthesis and stores of lipids in adipose tissue (the present data); diminish lipolysis; 8 diminish NEFA concentrations in plasma ~ and accelerate fl-oxidation and keotogenesis in the liver, thus reducing the release of NEFAs from adipose tissue and accelerating their metabolism in the liver. Unsaturated NEFAs may therefore provide a feedback mechanism for the control of their own concentration.
Unsaturated fatty acids potentiate insulin action
513
Clinically, the administration of unsaturated NEFAs may provide a means of increasing insulin sensitivity in insulin-dependent diabetics and in diminishing the tendency to ketogenesis in ketosis-prone diabetics. REFERENCES 1. 2. 3. 4. 5. 6. 7. 8.
Cout$roN, L. G. and DANDONA,P. Diabetes 29, 665-667 (1982). DIPLOCK,A. T., LucY, J. A., VnmND~R, M. and 7AF.LgANtn~Srd,A. FEBS Left. 82, 341-344 (1977). DoI~, V. and Mwa~tTS, H..I. Biol. Chem. 235, 2595-2599 (1960). McGAgRY, J. D. and FOSTER,D. W. Diabetes 29, 236-240 (1980). MIKHAILIDIS,D. P., KIRTLAND,S., BARRADAS,M. A. and DAh'DONA,P. Diabetologia 27, 311A (1984). PILCH, P. F., THOMPSON,P. A. and CZECH, M. Proc. Natl. Acad. Sci. U.S.A. 77, 915-918 (1980). RANDL~,P. J., GARLAND,P. B., HALESC. N. and NeWSHOLME,E. A. Lancet i, 785-789 (1963). RODar~L, M. Ann. N.Y. Acad. Sci. 131, 302-314 (1965).
0163-7827/86/$0.00 + 0.50 © 1986 Pergamon Journals Ltd
UNSATURATED FATTY ACIDS POTENTIATE INSULIN ACTION ON ADIPOCYTES F. LOMEO,M. A. K H o ~
and P. DANDON^
Metabolic Unit, Department of Chemical Pathology and Human Metabolism, Royal Free Hospital and School of Medicine, London NW3 2QG, U.K.
INTRODUCTION
We have recently shown that plasma nonesterified fatty acid (NEFA) concentrations are markedly lowered in healthy and diabetic subjects to whom dihomo-T-linolenic acid (DHLA) has been administered. This suggested that DHLAs may exert a pro-insulin effect, contrary to the concept that fatty acids are antagonistic to insulin action. 7 We have thus investigated the effects of DHLA and other fatty acids on insulin action on isolated rat adipocytes. METHODS
Solubilization of Fatty Acids NEFAs were dissolved in small volumes of ethanol and added to Krebs Ringer Bicarbonate Buffer (KRB) containing 3% NEFA free bovine serum albumin. The final volume of ethanol present in the incubates was 30 ml/mol NEFA/I. Unsaturated NEFAs used were: oleic (9-18:1), linoleic (9,12-18:2), linolenic (9,12,15-18:3), DHLA (8, 11, 14-20:3) and arachidonic (5, 8, 11, 14-20:4). Saturated NEFAs used were: palmitic (16:0) and stearic (18:0).
Assay for Triglyceride Synthesizing Adipocytes Fat cell suspensions were prepared from epididymal fat pads and incubated in KRB (pH 7.4) in the presence of test substances and [U-14C]glucose.I Lipids were then extracted and portions counted to give total triglyceride synthesis, while further portions were hydrolyzed and the glycerol and NEFA fractions counted separately. 3 Experiments were performed at glucose concentrations of 0.55 mmol/l (0.7 Ci/mol) and 3 mmol/l (2 Ci/mol). In this paper, data obtained using linolenic acid as a representative unsaturated NEFA are presented. Similar results were obtained using the other unsaturated NEFAs. RESULTS
Addition of the various unsaturated NEFA solutions to incubates did not alter the incorporation of glucose into triglyceride at 0.55 mmol/l of glucose. However, glucose incorporation into the fatty acid moiety of triglyceride was inhibited, whereas incorporation into the glycerol moiety was stimulated (Table 1). Glucose incorporation into both the fatty acid and glycerol moieties of triglyceride was increased by the unsaturated NEFAs at 3 mmol/l of glucose (Table 2). Concomitant incorporation of insulin and unsaturated NEFA resulted in a potentiation of insulin-stimulated glucose incorporation into both the fatty acid and glycerol moieties of triglyceride (Tables 1 and 2). However, at the higher glucose concentration, the increase in incorporation into the fatty acid moiety was more marked (Table 2). No stimulatory effect of saturated NEFA was observed, nor did these NEFAs produce a potentiation of insulin action. Ethanol produced a mild inhibition of glucose incorporation into glycerol and fatty acids at concentrations used to "carry" fatty acids into aqueous solution. 511
F. Lomeo
512
et
al.
TABLE 1. Percent Stimulation Over Basal of Triglyceride, Triglyceride Fatty Acid and Triglyceride Glycerol by Insulin and Linolenic Acid, Alone and in Combination, in the Presence of 0.55 mmol
Glucose % Stimulation over basal (mean 4-SEM. n = 6)
Additions I0 mU/1 insulin
Triglyceride
Triglyceride fatty acid
Triglyceride glycerol
136 5:9
204 + 10
97 + 6
0.5 mmol/l
linolenic acid
17 4- 4
- 4 5 5:3
78 + 8
10 mU/l insulin + 0.5 mmol/l
linoenic acid
363 _ 12 885:16
251 5:11
449 _ 16
Basal metabolic rate (nmol glUCO~/I0 7 cells/½hr) 365:8 36+8
TABLE 2. Percent Stimulation Over Basal of Triglyceride, Triglyceride Fatty Acid and Triglyceride Glycerol Synthesis by Insulin and Linolenic Acid, Alone and in Combination; in
the Presence of 3 mmol Glucose % Stimulation over basal (mean + SEM, n = 6)
Additions 10 mU/1 insulin
Triglyceride
Triglyceride fatty acid
Triglyceride glycerol
148 5:10
143 + 17
77 + 10
153 + I0
79 + 6
195 + 17
743 5:23
706 + 34
504 + 12
0.5 mmol/l
linolenic acid 10 mU/l insulin + 0.5 mmol/l
linolenic acid
Basal metabolic rate (nmol glucose/107 cells/~ hr) 260 + 60
192 + 44
84 5:20
DISCUSSION
Our data show that the concomitant incubation of unsaturated NEFA with adipocytes potentiates the stimulatory action of insulin on lipogenesis. This confirms the suggestion that NEFAs, in the presence of insulin, stimulate the incorporation of glucose into triglyceride giycerolJ However, our data also show that unsaturated NEFAs potentiate insulin-stimulated de n o v o fatty acid synthesis and that this effect becomes more marked on raising the glucose concentration to physiological levels. The discrepancy between these data and the principle that NEFAs are antagonistic to insulin action is explicable as the concept suggested by Randle e t al. 7 was based on the effects of NEFAs on muscle metabolism. Adipocyte metabolism, especially adipocyte lipogenesis, was not included in the investigation. Unsaturated NEFAs (arachidonic 2 and oleic and va~enic acids ~) enhance membrane fluidity in artificial systems and accelerate the passive diffusion and carrier-mediated uptake of glucose. 6 It is possible that the unsaturated NEFAs also get incorporated into cell membranes, and thus alter the permeability and uptake of glucose under the influence of insulin. It is of interest that oleic acid has been reported to exert glucagon-like effects upon hepatic NEFA metabolism, 4 which contrasts with the insulin-like effects reported here. Thus, in the light of present knowledge, unsaturated NEFAs would enhance synthesis and stores of lipids in adipose tissue (the present data); diminish lipolysis; 8 diminish NEFA concentrations in plasma ~ and accelerate fl-oxidation and keotogenesis in the liver, thus reducing the release of NEFAs from adipose tissue and accelerating their metabolism in the liver. Unsaturated NEFAs may therefore provide a feedback mechanism for the control of their own concentration.
Unsaturated fatty acids potentiate insulin action
513
Clinically, the administration of unsaturated NEFAs may provide a means of increasing insulin sensitivity in insulin-dependent diabetics and in diminishing the tendency to ketogenesis in ketosis-prone diabetics. REFERENCES 1. 2. 3. 4. 5. 6. 7. 8.
Cout$roN, L. G. and DANDONA,P. Diabetes 29, 665-667 (1982). DIPLOCK,A. T., LucY, J. A., VnmND~R, M. and 7AF.LgANtn~Srd,A. FEBS Left. 82, 341-344 (1977). DoI~, V. and Mwa~tTS, H..I. Biol. Chem. 235, 2595-2599 (1960). McGAgRY, J. D. and FOSTER,D. W. Diabetes 29, 236-240 (1980). MIKHAILIDIS,D. P., KIRTLAND,S., BARRADAS,M. A. and DAh'DONA,P. Diabetologia 27, 311A (1984). PILCH, P. F., THOMPSON,P. A. and CZECH, M. Proc. Natl. Acad. Sci. U.S.A. 77, 915-918 (1980). RANDL~,P. J., GARLAND,P. B., HALESC. N. and NeWSHOLME,E. A. Lancet i, 785-789 (1963). RODar~L, M. Ann. N.Y. Acad. Sci. 131, 302-314 (1965).