L. Herberg and H.K. Kley
Bierman, E.L., D. Porte Jr.: Carbohydrate intolerance and lipaemia. Ann.lntern.Med. 68: 926-933 (1968) Björntorp, P., L. Sjöström: Number and size of adipose tissue fat cells in relation to metabolism in human obesity. Metabolism 20: 703-713 (1971) Björntorp, p" A. Gustafson, B. Persson: Adipose tissue fat cell size and number in relation to metabolism in endogenous hypertriglyceridaemia. Acta Med.Scand. 190: 363-367 (1971) Bray, G.A., M.B. Davidson, E.J. Drenick: Obesity, a serious symptom. Ann.lnt.Med. 77: 779-795 (1972) Brown, W. V., H. Greten: Type 1 hyperlipoproteinaemia. Clinics in Endocr. and Metab. 2: 73-80 (1973) Car/son, L.A., L.E. Böttiger: Ischaemic heart disease in relation to fasting values of plasma triglycerides and cholesterol. Lancet 1: 865-868 (1972) Cramp, D., G. Robertson: The fluaromctric assay of triglyceride by a semiautomated method. Anal.Biochem. 25: 246-251 (1968) Di Girolamo, M., S. Mendlinger, J. W. Fertig: A simple method to determine fat cell size and number in four mammalian species. Amer.J.Physiol. 221: 850-857 (1971) Feldman, E.B., P. Benkel, R. V. Nayar: Physiological factors influencing circulating triglyceride concentration in wo-
men: age, weight gain and ovarian function. J.Lab.Clin. Med. 62: 437-477 (\963) Galton, D.J., J,N. Fain: Effects of prolonged incubation of isolated fat cells on their response to hormones stimulating lipolysis and glucose metabolism. Biochem.J. 98: 557-561 (1966) Glueck, c.J., R./. Levy, D.S. Fredrickson: Immunoreactive insulin, glucose tolerance and carbohydrate inducibility in type 11, IIJ, IV and V hyperlipoproteinaemia. Diabetes 18: 739-749 (1969) Goldstein, J.L., W.R. Hazzard, H.G. Schrott, E.L. Bierman, A. G. Motulsky: Hyperlipidaemia in coronary heart disease; I. Lipid levels in 500 survivors of myocardial infarction. J.Clin.lnvest. 52: 1533-1543 (1973) Hume, R.: Prediction of lean body mass from height and weight. J.Clin.Path. 19: 389-391 (1966) Reaven, G.M., R.L. Lerner, M.P. Stern, J, W. Farquhar: Role of insulin in endogenous hypertriglyceridaemia. J.Clin. Invest. 46: 1756-1767 (1967) Rodbell, M.: Metabolism of isolated fat cells. 1. Effects of hormones on glucose metabolism and Iipolysis. J .Biol. Chem. 239: 375-380 (1964) Smith, u., L. Sjöström, P. Björntorp: Comparison of two methods for determining human adipose cell size. J.Lipid Res. 13: 822-824 (1972)
Requests for reprints should be addressed to: Dr. J.P.D. Reckless, Diabetes and Lipid Res. Lab., London E.C. 1 (England)
st. Bartholomew's Hospital,
Harm. Metab. Res. 7 (1975) 410-415
© Georg Thieme Verlag Stuttgart
Adrenal Function and the Effect of a High-Fat Diet on C57BL/6J and C57BL/6J-ob/ob Mice L. Herberg and H. K. Kley Diabetes Forschungsinstitut und 2. Medizinische und Poliklinik Universität Düsseldorf. Germany
Summary In C57BL/6J mice and the ob/+ and ob/ob mutants total plasma corticosterone levels were found to be statistically different. In C57BL/6J mice the level was 1.9 ± 0.2 pg/lOO ml plasma, in ob/+ mice 8.6 ± 1.6 J..Ig/I00 ml and in ob/ob mice 13.7 ± 1.5 J..Ig/lOO ml. The percentage of protein-bound wrticosterone as weil as the free endogenous corticosterone levels were also different. Feeding a high-fat diet to young C57BL/6J and C57BL/6Jol/ob mice for aperiod of 4 weeks had no effect upon blood glucose, plasma insulin and plasma corticosterone levels. The significantly higher increase in body weight of the high-fat diet groups of both Iines of mice was mainly due to fat cell hypertrophy. Key-Words: C57BL/6J, ob/+, ob/ob Mice - Plasma Corti· costerone - High·Fat Diet - Adipose Tissue Cellularity
Introduction Experiments with small rodents have revealed that inherited adipose tissue cellularity can be modified Received: 10 Mar. 1975
Accepted: 18 June 1975
either by the ad lib. feeding of diets of different composition of nutrients or by the over- or underfeeding of identical diets. Such dietary induced modifications have been reported in metabolically intact as weil as in genetically obese rats and mice (Lemonnier 1970a, 1970b, 1972, Lemonnier, Winand, Furnel/e and ehristophe 1971, Lemonnier and Alexiu 1974, Johnson, Stern, Greenwood, Zucker and Hirsch 1973, Herberg, Döppen, Major and Gries 1974). However, feeding a high-fat diet had no effect upon blood glucose nur upon insulin concentrations in metabolically intact rats or NMRI miee, nor in genetically obese NZO mice which represent the hypertrophie type of obesity (Malaisse, Lemonnier, Malaisse-Lagae and Mandelbaum 1969, Herberg et al. 1974).
Downloaded by: National University of Singapore. Copyrighted material.
410
We now were interested in the effect of a high-fat diet upon CS7BL/6J-ob/ob mice which in adulthood exhibit the hypertrophie-hyperplastic type of obcsity and which in different breeding Iines are thc most commonly used obese-hyperglyccm ie- hyperinsu \incl11ie
Adrenal Funetion and the Effeet of a High-Fat Diet on C57BL/6J and C57BL/6J-ob/ob Mice
Materials and Methods Animals and diets: Male C57BL/6J and male C57BL/6J-ob/ob mice were obtained from the Jackson Laboratories, Bar Harbor, Maine, USA. The animals were obtained at 5-6 weeks of life and were housed, 6 to a cage, in a temperature-controlled room (25 0 C) with a 12 hr light-dark eyele (6 a.m. unti16 p.m.) and had free access to food and tap water. When two months old, the animals were weighed and each line was divided into two groups from which one group remained on the standard pelleted mouse chow whereas the other was changed to a high-fat diet. The diets were not isocaloric. The feeding period lasted for 4 weeks. Per cent composition (in calories) of the standard diet was: carbohydrate 63, fat 13, protein 24; percent composition (in calories) of the high-fat diet was: carbohydrate 19, fat 63, protein 18. (For further details see: Herberg et al. 1974).
of protein-bound corticosterone was determined in diluted plasma (1:5 with phosphate buffer) by equilibrium dialysis at 370 C and the fraction of free endogenous corticosterone was calculated as described by Westphal (1969). Statisties: Statistical analyses were made using Student's ttest.
Results I. Plasma corticosterone (Table I).
Total plasma corticosterone levels in C57BL/61-ob/ob and C57BL/61-ob/+ mice exceeded those in C57BL/61 mice by 7 and 4 times, respectively, the differences between the 3 lines being statistically significant. The percentage of protein-bound corticosterone as weIl as the free endogenous corticosterone levels were smaller in both lines of lean animals than in the obese mice. Il. Feeding ex.periment (Tables 2 and 3)
In C57BL/61 mice on the standard or the high-fat diet body weight increased about 13% or 28%, respectively. The final body weight of the mice on the highfat diet was 18% higher than that of the control group. Blood glucose and plasma insulin levels were the same for both groups as was the plasma corticosterone concentration. In C57BL/61-ob/ob mice the increase in body weight was 35% for the control and 55% for the high-fat diet group. At the end of the feeding period the body weight of the mice on the high-fat diet was about 17% higher than that of the controls. Blood glucose and plasma IRI concentrations were lower in the highfat diet group, the difference for insulin concentrations being highly significant. Plasma corticosterone levels were nearly the same for both groups.
Downloaded by: National University of Singapore. Copyrighted material.
rodents. Especially in animals of the Swedish colony an enlargement of the cortical part of the adrenals as weil as an increase in the in vitro biosynthesis of corticosterone as compared with metabolically intact controls has been described (Hellerström, Hellman and Larsson 1962, Carstensen, HeIlman and Larsson 1961, Hellman 1965). Whereas size and microscopical appearance of the adrenal cortex was observed to be normalized by a restricted diet (Hellerström, Hel/man and Larsson 1962), adrenalectomy does not prevent the excessive body weight gain (Naeser 1973). We therefore, initially determined plasma corticosterone levels in homozygous lean C57BL/61, heterozygous lean C57BL/61-ob/+, and homozygous obese C57BL/ 61-ob/ob mice. In a second series of experiments we examined the influence of a high-fat diet on body weight, adipose tissue cellularity, basal blood glucose, plasma insulin and plasma corticosterone concentration in C57BL/61 and C57BL/61-ob/ob mice.
411
In C57BL/61 mice on the high-fat diet the wet weight of the epididymal adipose tissue was 117% and the mean cell volume 115% higher than that of the controls. The wet weight of the subcutaneous adipose tissue of the high-fat diet group was 88% higher and the Handling o[ animals and analytical tests: The animals were weighed the day before sacrifice. Between mean fat cell volume 128% larger than that of the 9 and 9: 15 they were decapitated. Removal from the cage controls. The statistical differences between the two and blood collection were done within 10 to 15 seconds. groups were highly significant for both wet weight Blood was collected in small heparinized plastic vessels. Gluand me an fat cell volume for both tissue localizations. cose determinations were performed immediatelyon 25 J.Il In epididymal as weil as in subcutaneous adipose tisof whole blood by a reduction method with neocuproine sue mean fat cell number was the same for both groups. utilizing an Autoanalyzer II (Bittner and Manning 1966). For insulin and corticosterone determinations blood was cen- In C57BL/61-ob/ob mice the wet weight and the mean trifuged and the plasma stored at -18 0 C until assayed. Insufat cell volume of the epididymal adipose tissue were lin was measured in 100 J.Il of plasma by solid phase immusignificantly higher in the high-fat diet group, the difnoassay (Deutsche Pharmaeia, Frankfurt/Main, Germany). ference accounted for 27% or 18%, respectively. The Corticosterone was determined by a eompetitive pro tein binding teehnique similar to that previously described for fat cell number was slightly but not significantly highcortisol (Kley and Krüskemper 1971) and adapted to small er in the high-fat diet group. The wet weight of the volumes 00-50 J.Il plasma). The eoefficient of variation from day to day for a plasma sampie with low corticosterone con- subcutaneous adipose tissue was about 32% higher in the high-fat diet group whereas the me an fat cell voleentration (2.53 ± 0.24 pg/lOO ml) was 9.5% and that of urne was only 7% higher. In this group of mice, fat another plasma 00.5 ± 0.9 1Jg/100 ml) 8.8%. No corticoids other than eorticosterone eould be detected (concentration cell number, however, exceeded that observed in the smaller than 0.1 ng/ml) when eolumn ehromatography on controls by about 26%, the difference between the Sephadex LH-20 (Murphy 1971) was used. The percentage two groups being statistically significant.
412
L. Herberg and H.K. Kley
Table 1. Total plasma corticosterone levels, the percentage of binding in plasma corticosterone after dialysis and the free endogenous corticosterone in C57BL/6J mice, ob/+, and ob/ob mutants mice
percentage of protein-bound corticosterone (after dialysis at 37 0 C)x
total plasma corticosterone (.Ug/100 ml plasma)
C57BL/6J (10)
1.9 ± 0.2
C57BL/6J-ob/+ (0)
8.6 ± 1.6
C57BL/6J-ob/ob (0)
13.7 ± 1.5
.
free endogenous corticosterone (.Ug/100 ml plasma)
71.9
0.12
74.5
0.52
80.5
0.68
6
X The
percentage of protein-bound and the free endogenous corticosterone were measured from the pooled plasma of the 10 animals of each group.
Table 2. Body weight and adipose tissue cellularity in C57BL/6J mice and the ob/ob mutants fed different diets ad !ib. for 4 weeks mice
C57BL/6J
C57BL/6J-ob/ob
body weight epididymal adipose tissue gm wet weight fat cell number volume mg ~3x103 10 4
subcutaneous adipose tissue wet weight fat cell volume number rng ~3x103 104
diet
26.1 ± 0.6 (14)
.
337.0 ± 16.1
.
150 ± 12
.
201 ± 9
693.4 ± 53.2
.
54 ± 4
836 ± 49 standard
30.7 ± 0.9 (10)
731.0 ± 59.3
323 ± 27
213 ± 9
1302.0 ± 105.4
123 ± 16
839 ± 50 high-fat
43.4 ± 0.9 2589.3 ± 91.6 (14)
.
1192 ± 33
..
220 ± 7
8497.2 ± 272.5
.
611 ± 21
1304 ± 72 11 standard
50.6 ± 0.9 3292.2 ± 90.5
1407 ± 43
240 ± 11
11179.4 ± 403.6
653 ± 33
1651 ± 62 high-fat
.
.
Number of anirnals in parentheses. Mean values ± S.E.M. Statistical differences within the two groups of C57BL/6J or C57BL/6J-ob/ob mice, respectively ... = p < 0.001,6 = p < 0.005.
Table 3. Body weight, blood glucose, plasma IRI and corticosterone in C57BL/6J mice and the ob/ob mutants fed different diets ad !ib. for 4 weeks. mice
C57BL/6J
body weight gm initial final
plasma IRI ~U/ml
plasma corticosterone J.Ig/100 ml
23.1 ±0.9 (14) 6
26.1 ± 0.6 (14)
8.690 ± 0.165 (14)
37 ± 5 (14)
3.0 ±0.3 (20)
24.0 ±0.5 (10)
30.7 ± 0.9 (10)
8.305 ±0.330 (10)
43 ± 3 (10)
3.2 ±0.2 (19)
32.2 ± 1.3 (14)
43.4 ±0.9 (14)
..
C57BL/6J-ob/ob
blood glucose mM
..
32.7 ±4.7 (12)
.
.
diet
standard
high-fat
.
50.6 ±0.9 (12)
12.210 ± 1.265 (14) 10.605 ± 0.605 (12)
320 ± 34 (14)
..
185 ± 16 (12)
15.3 ± 0.9 (18) standard 16.7 ± 1.3 (19) high-fat
Number of anirnals in parentheses. Mean values and S.E.M. Statistical differences between the initial and final body weight of each of the groups and within the two groups of C57BL/6J or C57BL/6J-ob/ob mice, as weil as in plasma IRI levels of C57BL/ 6J-ob/ob mice fed the standard or the high-fat diet ... = p < 0.001,6 = p < 0.005.
Downloaded by: National University of Singapore. Copyrighted material.
Number of anirnals in parentheses. Mean values ± S.E.M . .. = P < 0.001, 6= p
Bierman, E.L., D. Porte Jr.: Carbohydrate intolerance and lipaemia. Ann.lntern.Med. 68: 926-933 (1968) Björntorp, P., L. Sjöström: Number and size of adipose tissue fat cells in relation to metabolism in human obesity. Metabolism 20: 703-713 (1971) Björntorp, p" A. Gustafson, B. Persson: Adipose tissue fat cell size and number in relation to metabolism in endogenous hypertriglyceridaemia. Acta Med.Scand. 190: 363-367 (1971) Bray, G.A., M.B. Davidson, E.J. Drenick: Obesity, a serious symptom. Ann.lnt.Med. 77: 779-795 (1972) Brown, W. V., H. Greten: Type 1 hyperlipoproteinaemia. Clinics in Endocr. and Metab. 2: 73-80 (1973) Car/son, L.A., L.E. Böttiger: Ischaemic heart disease in relation to fasting values of plasma triglycerides and cholesterol. Lancet 1: 865-868 (1972) Cramp, D., G. Robertson: The fluaromctric assay of triglyceride by a semiautomated method. Anal.Biochem. 25: 246-251 (1968) Di Girolamo, M., S. Mendlinger, J. W. Fertig: A simple method to determine fat cell size and number in four mammalian species. Amer.J.Physiol. 221: 850-857 (1971) Feldman, E.B., P. Benkel, R. V. Nayar: Physiological factors influencing circulating triglyceride concentration in wo-
men: age, weight gain and ovarian function. J.Lab.Clin. Med. 62: 437-477 (\963) Galton, D.J., J,N. Fain: Effects of prolonged incubation of isolated fat cells on their response to hormones stimulating lipolysis and glucose metabolism. Biochem.J. 98: 557-561 (1966) Glueck, c.J., R./. Levy, D.S. Fredrickson: Immunoreactive insulin, glucose tolerance and carbohydrate inducibility in type 11, IIJ, IV and V hyperlipoproteinaemia. Diabetes 18: 739-749 (1969) Goldstein, J.L., W.R. Hazzard, H.G. Schrott, E.L. Bierman, A. G. Motulsky: Hyperlipidaemia in coronary heart disease; I. Lipid levels in 500 survivors of myocardial infarction. J.Clin.lnvest. 52: 1533-1543 (1973) Hume, R.: Prediction of lean body mass from height and weight. J.Clin.Path. 19: 389-391 (1966) Reaven, G.M., R.L. Lerner, M.P. Stern, J, W. Farquhar: Role of insulin in endogenous hypertriglyceridaemia. J.Clin. Invest. 46: 1756-1767 (1967) Rodbell, M.: Metabolism of isolated fat cells. 1. Effects of hormones on glucose metabolism and Iipolysis. J .Biol. Chem. 239: 375-380 (1964) Smith, u., L. Sjöström, P. Björntorp: Comparison of two methods for determining human adipose cell size. J.Lipid Res. 13: 822-824 (1972)
Requests for reprints should be addressed to: Dr. J.P.D. Reckless, Diabetes and Lipid Res. Lab., London E.C. 1 (England)
st. Bartholomew's Hospital,
Harm. Metab. Res. 7 (1975) 410-415
© Georg Thieme Verlag Stuttgart
Adrenal Function and the Effect of a High-Fat Diet on C57BL/6J and C57BL/6J-ob/ob Mice L. Herberg and H. K. Kley Diabetes Forschungsinstitut und 2. Medizinische und Poliklinik Universität Düsseldorf. Germany
Summary In C57BL/6J mice and the ob/+ and ob/ob mutants total plasma corticosterone levels were found to be statistically different. In C57BL/6J mice the level was 1.9 ± 0.2 pg/lOO ml plasma, in ob/+ mice 8.6 ± 1.6 J..Ig/I00 ml and in ob/ob mice 13.7 ± 1.5 J..Ig/lOO ml. The percentage of protein-bound wrticosterone as weil as the free endogenous corticosterone levels were also different. Feeding a high-fat diet to young C57BL/6J and C57BL/6Jol/ob mice for aperiod of 4 weeks had no effect upon blood glucose, plasma insulin and plasma corticosterone levels. The significantly higher increase in body weight of the high-fat diet groups of both Iines of mice was mainly due to fat cell hypertrophy. Key-Words: C57BL/6J, ob/+, ob/ob Mice - Plasma Corti· costerone - High·Fat Diet - Adipose Tissue Cellularity
Introduction Experiments with small rodents have revealed that inherited adipose tissue cellularity can be modified Received: 10 Mar. 1975
Accepted: 18 June 1975
either by the ad lib. feeding of diets of different composition of nutrients or by the over- or underfeeding of identical diets. Such dietary induced modifications have been reported in metabolically intact as weil as in genetically obese rats and mice (Lemonnier 1970a, 1970b, 1972, Lemonnier, Winand, Furnel/e and ehristophe 1971, Lemonnier and Alexiu 1974, Johnson, Stern, Greenwood, Zucker and Hirsch 1973, Herberg, Döppen, Major and Gries 1974). However, feeding a high-fat diet had no effect upon blood glucose nur upon insulin concentrations in metabolically intact rats or NMRI miee, nor in genetically obese NZO mice which represent the hypertrophie type of obesity (Malaisse, Lemonnier, Malaisse-Lagae and Mandelbaum 1969, Herberg et al. 1974).
Downloaded by: National University of Singapore. Copyrighted material.
410
We now were interested in the effect of a high-fat diet upon CS7BL/6J-ob/ob mice which in adulthood exhibit the hypertrophie-hyperplastic type of obcsity and which in different breeding Iines are thc most commonly used obese-hyperglyccm ie- hyperinsu \incl11ie
Adrenal Funetion and the Effeet of a High-Fat Diet on C57BL/6J and C57BL/6J-ob/ob Mice
Materials and Methods Animals and diets: Male C57BL/6J and male C57BL/6J-ob/ob mice were obtained from the Jackson Laboratories, Bar Harbor, Maine, USA. The animals were obtained at 5-6 weeks of life and were housed, 6 to a cage, in a temperature-controlled room (25 0 C) with a 12 hr light-dark eyele (6 a.m. unti16 p.m.) and had free access to food and tap water. When two months old, the animals were weighed and each line was divided into two groups from which one group remained on the standard pelleted mouse chow whereas the other was changed to a high-fat diet. The diets were not isocaloric. The feeding period lasted for 4 weeks. Per cent composition (in calories) of the standard diet was: carbohydrate 63, fat 13, protein 24; percent composition (in calories) of the high-fat diet was: carbohydrate 19, fat 63, protein 18. (For further details see: Herberg et al. 1974).
of protein-bound corticosterone was determined in diluted plasma (1:5 with phosphate buffer) by equilibrium dialysis at 370 C and the fraction of free endogenous corticosterone was calculated as described by Westphal (1969). Statisties: Statistical analyses were made using Student's ttest.
Results I. Plasma corticosterone (Table I).
Total plasma corticosterone levels in C57BL/61-ob/ob and C57BL/61-ob/+ mice exceeded those in C57BL/61 mice by 7 and 4 times, respectively, the differences between the 3 lines being statistically significant. The percentage of protein-bound corticosterone as weIl as the free endogenous corticosterone levels were smaller in both lines of lean animals than in the obese mice. Il. Feeding ex.periment (Tables 2 and 3)
In C57BL/61 mice on the standard or the high-fat diet body weight increased about 13% or 28%, respectively. The final body weight of the mice on the highfat diet was 18% higher than that of the control group. Blood glucose and plasma insulin levels were the same for both groups as was the plasma corticosterone concentration. In C57BL/61-ob/ob mice the increase in body weight was 35% for the control and 55% for the high-fat diet group. At the end of the feeding period the body weight of the mice on the high-fat diet was about 17% higher than that of the controls. Blood glucose and plasma IRI concentrations were lower in the highfat diet group, the difference for insulin concentrations being highly significant. Plasma corticosterone levels were nearly the same for both groups.
Downloaded by: National University of Singapore. Copyrighted material.
rodents. Especially in animals of the Swedish colony an enlargement of the cortical part of the adrenals as weil as an increase in the in vitro biosynthesis of corticosterone as compared with metabolically intact controls has been described (Hellerström, Hellman and Larsson 1962, Carstensen, HeIlman and Larsson 1961, Hellman 1965). Whereas size and microscopical appearance of the adrenal cortex was observed to be normalized by a restricted diet (Hellerström, Hel/man and Larsson 1962), adrenalectomy does not prevent the excessive body weight gain (Naeser 1973). We therefore, initially determined plasma corticosterone levels in homozygous lean C57BL/61, heterozygous lean C57BL/61-ob/+, and homozygous obese C57BL/ 61-ob/ob mice. In a second series of experiments we examined the influence of a high-fat diet on body weight, adipose tissue cellularity, basal blood glucose, plasma insulin and plasma corticosterone concentration in C57BL/61 and C57BL/61-ob/ob mice.
411
In C57BL/61 mice on the high-fat diet the wet weight of the epididymal adipose tissue was 117% and the mean cell volume 115% higher than that of the controls. The wet weight of the subcutaneous adipose tissue of the high-fat diet group was 88% higher and the Handling o[ animals and analytical tests: The animals were weighed the day before sacrifice. Between mean fat cell volume 128% larger than that of the 9 and 9: 15 they were decapitated. Removal from the cage controls. The statistical differences between the two and blood collection were done within 10 to 15 seconds. groups were highly significant for both wet weight Blood was collected in small heparinized plastic vessels. Gluand me an fat cell volume for both tissue localizations. cose determinations were performed immediatelyon 25 J.Il In epididymal as weil as in subcutaneous adipose tisof whole blood by a reduction method with neocuproine sue mean fat cell number was the same for both groups. utilizing an Autoanalyzer II (Bittner and Manning 1966). For insulin and corticosterone determinations blood was cen- In C57BL/61-ob/ob mice the wet weight and the mean trifuged and the plasma stored at -18 0 C until assayed. Insufat cell volume of the epididymal adipose tissue were lin was measured in 100 J.Il of plasma by solid phase immusignificantly higher in the high-fat diet group, the difnoassay (Deutsche Pharmaeia, Frankfurt/Main, Germany). ference accounted for 27% or 18%, respectively. The Corticosterone was determined by a eompetitive pro tein binding teehnique similar to that previously described for fat cell number was slightly but not significantly highcortisol (Kley and Krüskemper 1971) and adapted to small er in the high-fat diet group. The wet weight of the volumes 00-50 J.Il plasma). The eoefficient of variation from day to day for a plasma sampie with low corticosterone con- subcutaneous adipose tissue was about 32% higher in the high-fat diet group whereas the me an fat cell voleentration (2.53 ± 0.24 pg/lOO ml) was 9.5% and that of urne was only 7% higher. In this group of mice, fat another plasma 00.5 ± 0.9 1Jg/100 ml) 8.8%. No corticoids other than eorticosterone eould be detected (concentration cell number, however, exceeded that observed in the smaller than 0.1 ng/ml) when eolumn ehromatography on controls by about 26%, the difference between the Sephadex LH-20 (Murphy 1971) was used. The percentage two groups being statistically significant.
412
L. Herberg and H.K. Kley
Table 1. Total plasma corticosterone levels, the percentage of binding in plasma corticosterone after dialysis and the free endogenous corticosterone in C57BL/6J mice, ob/+, and ob/ob mutants mice
percentage of protein-bound corticosterone (after dialysis at 37 0 C)x
total plasma corticosterone (.Ug/100 ml plasma)
C57BL/6J (10)
1.9 ± 0.2
C57BL/6J-ob/+ (0)
8.6 ± 1.6
C57BL/6J-ob/ob (0)
13.7 ± 1.5
.
free endogenous corticosterone (.Ug/100 ml plasma)
71.9
0.12
74.5
0.52
80.5
0.68
6
X The
percentage of protein-bound and the free endogenous corticosterone were measured from the pooled plasma of the 10 animals of each group.
Table 2. Body weight and adipose tissue cellularity in C57BL/6J mice and the ob/ob mutants fed different diets ad !ib. for 4 weeks mice
C57BL/6J
C57BL/6J-ob/ob
body weight epididymal adipose tissue gm wet weight fat cell number volume mg ~3x103 10 4
subcutaneous adipose tissue wet weight fat cell volume number rng ~3x103 104
diet
26.1 ± 0.6 (14)
.
337.0 ± 16.1
.
150 ± 12
.
201 ± 9
693.4 ± 53.2
.
54 ± 4
836 ± 49 standard
30.7 ± 0.9 (10)
731.0 ± 59.3
323 ± 27
213 ± 9
1302.0 ± 105.4
123 ± 16
839 ± 50 high-fat
43.4 ± 0.9 2589.3 ± 91.6 (14)
.
1192 ± 33
..
220 ± 7
8497.2 ± 272.5
.
611 ± 21
1304 ± 72 11 standard
50.6 ± 0.9 3292.2 ± 90.5
1407 ± 43
240 ± 11
11179.4 ± 403.6
653 ± 33
1651 ± 62 high-fat
.
.
Number of anirnals in parentheses. Mean values ± S.E.M. Statistical differences within the two groups of C57BL/6J or C57BL/6J-ob/ob mice, respectively ... = p < 0.001,6 = p < 0.005.
Table 3. Body weight, blood glucose, plasma IRI and corticosterone in C57BL/6J mice and the ob/ob mutants fed different diets ad !ib. for 4 weeks. mice
C57BL/6J
body weight gm initial final
plasma IRI ~U/ml
plasma corticosterone J.Ig/100 ml
23.1 ±0.9 (14) 6
26.1 ± 0.6 (14)
8.690 ± 0.165 (14)
37 ± 5 (14)
3.0 ±0.3 (20)
24.0 ±0.5 (10)
30.7 ± 0.9 (10)
8.305 ±0.330 (10)
43 ± 3 (10)
3.2 ±0.2 (19)
32.2 ± 1.3 (14)
43.4 ±0.9 (14)
..
C57BL/6J-ob/ob
blood glucose mM
..
32.7 ±4.7 (12)
.
.
diet
standard
high-fat
.
50.6 ±0.9 (12)
12.210 ± 1.265 (14) 10.605 ± 0.605 (12)
320 ± 34 (14)
..
185 ± 16 (12)
15.3 ± 0.9 (18) standard 16.7 ± 1.3 (19) high-fat
Number of anirnals in parentheses. Mean values and S.E.M. Statistical differences between the initial and final body weight of each of the groups and within the two groups of C57BL/6J or C57BL/6J-ob/ob mice, as weil as in plasma IRI levels of C57BL/ 6J-ob/ob mice fed the standard or the high-fat diet ... = p < 0.001,6 = p < 0.005.
Downloaded by: National University of Singapore. Copyrighted material.
Number of anirnals in parentheses. Mean values ± S.E.M . .. = P < 0.001, 6= p
