Metabolism of genetically obese rats on normal or high-fat diet

Diabetologia, Nov 1974

Summary Adult genetically obesefafa rats showed a high level of lipogenesis from glucose in liver but not in adipose tissue; pancreatic content and serum levels of insulin were elevated. Glucose uptake and insulin sensitivity were decreased in muscle,fafa rats and their lean littermates fed a high-fat diet showed increased fat deposits. Serum insulin levels were not significantly affected by diet in either group. The larger the fat cells were, the more actively they utilised glucose; insulin sensitivity was influenced both by diet and cell size. Control rats made obese by a high caloric diet did not show insulin resistance in muscle. — The data indicate that in adult obesity in these rats, even in the presence of marked hyperinsulinism, increased lipogenesis in adipose tissue is not a prerequisite. Rather, fat storage is a consequence of increased uptake of circulating triglycérides. On a diet rich in carbohydrate, adipose tissue fatty acids were mainly of hepatic origin; on a high-fat diet they were of dietary origin.

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Metabolism of genetically obese rats on normal or high-fat diet

Metabolism of Genetically Obese Rats on N o r m a l or H i g h - F a t Diet D. Lemonnier 0 R. Aubert 0 J.-P. Suquet 0 G. Rosselin 0 0 Laboratoire de Nutrition Humaine de l'Institut Scientifique et Technique de l'Alimentation, et Unit6 de Recherche de Diab6tologie et d'Etudes Radioimmunologiques des I-Iormones Prot6iques, U.55 I.N.S.E.R.M. , Paris , France Summary. Adult genetically obese fafa rats showed a high level of lipogenesis from glucose in liver but not in adipose tissue; pancreatic content and serum levels of insulin were elevated. Glucose uptake anO insulin sensitivity were decreased in muscle, fafa rats and their lean littermates fed a high-fat diet showed increased fat deposits. Serum insulin levels were not significantly affected by diet in either group. The larger the fat cells were, the more actively they utilised glucose; insulin sensitivity was influenced both by diet and cell size. Control rats made obese by a high caloric diet did not show insulin resistance in muscle. -- The data indicate that in adult obesity in these rats, even in the presence of marked hyperinsulinism, increased lipogenesis in adipose tissue is not a prerequisite. Rather, fat storage is a consequence of increased uptake of circulating triglycerides. On a diet rich in carbohydrate, adipose tissue fatty acids were mainly of hepatic origin; on a high-fat diet they were of dietary origin. High-fat diet; rat; nutritional obesity; genetic obesity; adipose tissue; liver; muscle metabolism; insulin sensitivity; pancreatic glucagon; pancreatic and plasmatic insulin - Obesity is associated with hyperinsulinism i.e. increased insulin secretion and high plasma insulin levels. This hyperinsulinism is partly related to a high carbohydrate intake [l]. Long term feeding of a high fat diet to genetically obese obob mice (a) markedly increases their obesity, (b) has no effect on their hyperglycemia, but (c) decreases serum insulin levels [ 2 ]. Moreover, obesity can be induced in lean animals b y feeding a high-fat-low-carbohydrate diet given ad lib. [ 3 ]. This kind of experimental obesity is characterized b y normal levels of circulating insulin in vivo, and i n vitro b y a decreased insulin response to glucose [ 4 ]. These data indicate t h a t obesity per #e is not closely related to hyperinsulinism. I n the present investigation, genetically obese f a f a rats were studied. They are known to present with a high degree of hyperinsulinism [ 5, 6 ], high levels of serum lipids [7] and an elevated lipogenesis of adipose tissue [ 8 ]. The aim of the study was to investigate the Materials and Methods Five m o n t h old male genetically obese f a f a rats [ 7 ] and their lean littermates (FaFa or Fafa) were fed ad lib for 7 months either a control diet T (9% of calories as lipid, 22% as protein, and 69% as carbohydrate), or a high-fat (72% of calories as lipid, 22% as protein, and 16% as carbohydrate) diet S [ 3 ]. Four groups of rats were obtained. Animals were killed b y deeapita- 9 tion in the fed state in the morning. Blood was collected and serum glucose was assayed b y the glucose oxidase method (Boehringer Mannheim Test). Serum insulin was measured by the method of Rosselin et al. [ 9 ]. The pancreatic content of insulin was determined as previously described [ 4 ]. Pancreatic glueagon was assayed according to Jarrousse et al. [ 10 ]. The incorporation of glueose-U14C into C02, lipids or glycogen in liver slices, hemidiaphragms and periGroups Fa T Fa S f a T fa S Body weight (g) 5 3 0 = ~ 1 2 . 2 5664-16.1 672i36.4 885-L31.0 b Serum glucose (mg/lOO ml) 132~=5.6 119 134-4-4.7 121-4-3.1 Serum insulin (vU/ml) 138 1 1 . 5 167 42.3 536~= 101 422~= 6 9 . 9 Pancreatic weight (nag) 14044-77.7 1 3 4 1 = = 6 5 . 2 1 3 1 4 = ~ 9 7 . 5 1 4 1 4 ~ = 6 4 . 0 Stored insulin Stored glucagon (U per pancreas) (~zgper pancreas) 1.85+0.149 8.37 1.48~0.185 7.06 3.22~0.549 5.22i0.400 3.15 6.58=[=0.38a6 locus of fat synthesis in the obese animals and their lean littermates, and the possible influence of different diets. Special attention was given to the role of insulin and of fat cell size in this model of hereditary and/or dietary obesity. genital adipose tissue pieces, was measured as described elsewhere [i1]. Insulin (1 mU/ml) was added to the incubation medium in the case of adipose tissue and muscle. Measurements of adipose tissue cellularity have been described [ 3 ]. D. Lemonnier et al. : Metabolism of Genetically Obese l~ats on Normal or High-Fat Diet Body weight was significantly increased b y the combination of high-fat diet and the obesity genes of fafa rats (Table 1). Serum glucose tended to decrease slightly in rats on high fat diet. Pooling obese (fafa) and nonobese (FaFa) t y p e rats, the high fat diet reduced blood glucose levels significantly ( p ~ 0.01). Serum insulin was not significantly changed b y high fat diet. The most striking observation was a four-fold increase in circulating insulin levels in the obese group on control diet. As shown in Table 1, the pancreas of obese rats contained twice as much insulin as those of lean animals ( p ~ 0 . 0 1 ) ; this was not affected b y diet. Pancreatic glucagon content was similar in all groups. Adipose Tissue Metabolism F a t cell volume was 2.2 fold increased in the epidid y m a l adipose tissue of genetically obese animals regardless of diet composition. I n the lean rats however, high fat diet induced a 50% increase in fat cell volume above control. Changes in cell number were not observed in this site [ 12 ]. Fig. 1 shows the basal incorporation of glucose -U~4C into CO2, t o t a l glycerol and t o t a l f a t t y acids (FA) of adipose tissue fragments. All results are expressed per fat cell. Total d p m recovered as 14C02 and labeled lipids were twice as high ( p ~ 0 . 0 1 ) in the obese rats indicating an increased glucose u p t a k e in these animals. High fat diet increased this value as well (p ~ 0.01) but to a lesser extent. Whereas FA incorporated only 5--10~ of t o t a l 14C uptake, the bulk of radioactivity was found in the glycerol moiety. Fig. 1 also shows t h a t in the basal state, labeled glycerol was augmented b y b o t h genetic obesity and high fat diet. B y contrast, 14C incorporation into FA was increased only b y genetic obesity, but not b y diet. The effect of insulin added in vitro (1 mU/m]) was most m a r k e d for FA synthesis (Fig. l c). Adipose tissue of animals on control diet was significantly (p ~ 0 . 0 5 ) more responsive. The s a m e was true for 14C0~ production. B y contrast, insulin-induced glycerol synthesis was insignificant whatever the group. When the results were related to fat cell surface, m o s t of these differences were maintained. Liver Metabolism , CO2 c 8O nN Fatty acids FaT FaS faT faS The addition of I mU insulin per ml of medium increased glucose u p t a k e and glycogen synthesis in all groups. The effect of insulin on muscle was reduced b y obesity but not b y diet. I Triglyceride-g[ycero[ Triglyceride-fatty-acids 290+51.6 FaT FaS faT faS high lot content increased not only the fat stores of the genetically obese, but also of the lean rats [ 12 ]. Thus, b o d y weight appears to be influenced b y at least two factors, the genetic background and the composition of the diet. Similar results have been described in obob mice [ 2 ]. Studies done with Swiss mice and Wistar rats showed a more m a r k e d effect of diet [ 3, 13 ], Diabetologia, Suppl. to Vol. 10 D i s c u s s i o n The cause of obesity is still poorly understood. Various animal models have been studied to shed light onto possible pathogenic mechanisms. The present s t u d y deals with the obesity of the genetically obese fafa rats and the influence of diet composition. As Table 1 shows, the genetically obese animals were heavier t h a n their lean siblings. A diet with a 104d )m/liver into: C02 p a r t l y because in those animals, the diet was begun earlier in life t h a n in the present s t u d y [ 13, 14 ]. I n order to analyse the possible mechanisms involved in the pathogenesis of obesity in these fafa rats, the three m a i n organ systems involved in diabetic metabolism have been studied in vitro, namely adipose tissue, muscle and liver. Adipose tissue of the adult genetically obese rats showed a much higher glucose uptake, the bulk of C02 Production(dpm) T Glycogensynthesis {dpm) T I --:.. xx x\ ,N "E r ~F o o or o o s Z IT 5 1000 0 40000'L 30000 which was incorporated into glycerol (Fig. 1). FA synthesis was not particularly marked. However, the effect of insulin added in vitro was m o s t striking in increasing FA synthesis and b y far the m o s t impressive in the animals (lean or obese genetically) on control diet. I n muscle, as one might expect, a b o u t 90% of glucose t a k e n up was incorporated into glycogen. High fat diet decreased glucose u p t a k e and, thereby, glycogen synthesis and CO2 production. Liver slices of genetically obese rats showed a six fold incorporation rate into TG-FA compared to t h a t of lean animals. Therefore, the main site for lipogenesis in adultfafa rats appears to be the liver and not adipose tissue. On the high fat diet, lipogenesis was reduced b y more t h a n 90% in lean and obese animals, indicating t h a t diet is an i m p o r t a n t regulator of hepatic lipogenesis. I t was of particular interest to s t u d y the conditions in vivo which m a y have influenced the behaviour of the organs in vitro. Hyperinsulinemia appears to be a characteristic of genetically obese fafa rats (Table 1). However, high fat diet reduced serum insulin levels slightly whereas fat stores increased m a r k e d l y [ 12 ]. This m a y be due p a r t l y to the lower carbohydrate intake which could not be avoided with the high fat diet, as long as protein supply had to be constant. I t is quite possible t h a t the insulin levels in vivo were directly responsible for the behaviour of the isolated organs in vitro. Indeed, adipose tissue of fafa rats exposed to very high insulin levels in vivo, showed higher rates of glucose metabolism (Fig. 1). This was all the more m a r k e d in liver slices where up to sixfold differences could be observed (Fig. 2). The connection between hyperinsutinemia and hepatic lipogenesis has been suggested b y Steiner [ 15 ] and L e t a r t e [ 16 ]. I n muscle, interestingly, fafa rats showed less glucose metabolism. By contrast, although no dramatic changes in insulin levels were obtained b y feeding the high fat diet, the glucose u p t a k e decreased m a r k e d l y in liver and muscle tissue. I n adipose tissue such an effect was found only for the insulin response in respect to FA synthesis. Although the main FA synthesis occurs in the liver, fafa rats have an increased adipose mass where glucose is mainly incorporated into ~-glyeerophosphate to form triglyceride. The FA m o i e t y comes from circulating triglycerides as suggested b y the doubled lipoprotein lipase activity found in the fat cells of these fafa rats [ 17 ]. A similar argument applies to the higher glucose incorporation into triglyceride glycerol of adipose tissue of obese and lean animals fed the high fat diet, i.e. the increased supply of FA (in this case from exogenous sources) needed more ~-glycerophosphate for triglyceride synthesis. Thus, feeding a high fat diet seems to speed the effects of ageing [ 18 ]. F a t t y acid composition of adipose tissue reveals t h a t dietary lat~ty acids are stored as such in adipose tissue [1 i]. H u m a n studies with adult obese and lean subjects have lead to similar conclusions: lipogenesis is unsignificant in adipose tissue [ 19, 20 ]. The role of fat cell size deserves special consideration. Since fafa rats have adipocytes of twice the volume of lean rats, one might implicate this fact in the higher glucose u p t a k e of fafa adipose tissue. Indeed, when adipocytes of lean r a t s were increased in volume b y high fat diet, glucose u p t a k e of adipose tissue in vitro was equally increased. Similar results have been obtained b y Smith in h u m a n obesity [ 20 ] and b y Stern in fafa rats [ 21 ]. An additional increase of glucose utilisation was observed in the fafa rat fed a high-fat diet although cell size did not change any further. Insulin resistance has been suggested to be related to the increased fat cell size b y a decreased number of binding sites [ 22 ]. The present data confirm this a r g u m e n t in p a r t : in vitro added insulin had a smaller effect on tissue of fafa rats which have larger adipocytes. However, although high fat diet increased fat cell size and decreased insulin sensitivity in the lean rats, in the obese rats the insulin effect was diminished b y high fat diet with no further change in fat cell size. I t appears, therefore, t h a t basal glucose u p t a k e and insulin sensit i v i t y are not dependent on fat cell size only. I n conclusion, the data presented here show the importance of diet composition in relation to obesity of genetically obesefafa rats. I n adults the bulk of adipose tissue FA appears to derive from either dietary f a t t y acids or from f a t t y acids synthetized in the liver. Hyperinsulinemia is p r o b a b l y an i m p o r t a n t factor contributing to obesity in this animal as well as to insulin resistance in muscle. Also, fat cell size induced by either genetic background or b y high fat diet appears to be related to increased glucose metabolism and p a r t l y to insulin resistance. Acknowledgements. This work was supported in part, by Research grants (Contrat Libre 71.5.219.12) from the Institut National de la Sent6 et de la Recherche M6dicale (France). The authors are indebted to Claudine Boissard, Andr6 Alexiu and Robert Loeb for their expert technical assistance. They are most grateful to Dr. A.E. Renold and Dr. W.A. Muller for their precious advice and help in the preparation of the manuscript. 1. Grey , N. , Kipnis , D.M. : Effect of diet composition on the hyperinsulinemia of obesity . New Engl. J. Med . 285 , 827 ( 1971 ) 2. Lemonnier , D. , Winand , J. , Furnelle , J. , Christophe , J. : Effects of a high-fat diet on obese-hyperglycemic and non-obese Bar-Harbor mice . Diabetologia 7 , 328 ( 1971 ) 3. Lemormier , D. : Effect of age, sex and site on the cellularity of the adipose tissue in mice and rats rendered obese b y a high-fat diet . J. din. Invest . 51 , 2907 ( 1972 ) 4. 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Stern , J. , Johnson, P.R. , Greenwood , M. 1 %.C., Zucker , L.M. , Hirsch , J.: Insulin resistance and pancreatic insulin release in the genetically obese Zueker rat . Proc. Soc. exp. Biol . (N.Y.) 139 , 66 ( 1972 ) 22. Freychet , P. , Laudat , M.-I{. , Laudat , P. , Rosselin , G. , Kahn , C. 1 %., Gorden , P. , Roth , J.: I m p a i r m e n t of insulin binding to the fat cell plasma membrane in the obese hyperglycemic mouse . FEBS Letters 25 , 339 ( 1972 ) 292, rue Saint Martin F-75141 Paris Cedex 03 France Responsible for the text: Prof . Dr. W. C~ EV~ZFET .DT,3s Universit/itsklinik, Humboldtallee 1, D - 34 GSttlngen/F.R.G. Prof. Dr. K. SCtrS~ FLING , Zentrum der Innerenl~edizin,Theodor-Stern- Kai7 , D- 6 Frankfurt 70/F.1%.G. Responsiblefor advertisements: L. SIeGeL, G. MARTIN,D- 1000Berlin 15, Kurfiirstendamm 237 . Springer-Verlag,Berlin, Heidelberg,New York. Printed in Gerrnanyby aprinta, Wemding/Sehwaben. Copyright 9 by Springer-VerlagBerlin. Heidelberg 1974


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D. Lemonnier, R. Aubert, J. P. Suquet, G. Rosselin. Metabolism of genetically obese rats on normal or high-fat diet, Diabetologia, 1974, 697-701, DOI: 10.1007/BF01222007