168

BBALIP

53516

Lipogenesis in genetically diabetic ( db/db) mice: developmental changes in brown adipose tissue, white adipose tissue and the liver Paul Trayhurn*

and Monica C. Wusteman

(Received

Key words:

Lipogenesis;

Brown adipose

10 April 1990)

tissue; White adipose

tissue; Insulin

resistance:

(Mouse liver)

Developmental changes in lipogenesis have been examined in interscapular brown adipose tissue (BAT), epididymal white adipose tissue and the liver of genetically diabetic (&/db) mice and their normal siblings. Lipogenesis was measured in vivo with 3H,0, from weaning (21 days of age) until 20 weeks of age. Hyperinsulinaemia was evident in db/db mice at all ages. Low rates of lipogenesis were observed at weaning in tissues of both groups of mice, but the rate rose rapidly in the first few days post-weaning. In normal mice, peak lipogenesis was obtained in each tissue at 4-5 weeks of age, and there were no major changes (on a whole-tissue basis) thereafter. A different develo~en~ pattern was apparent in db/db mice. The rate of lipogenesis in BAT rose sharply after weaning, reaching a peak at 26 days of age (several times higher than normal mice), and then falling rapidly such that by 45 days of age it was lower than in normal mice; at age 20 weeks lipogenesis in BAT of the diabetic animals was negligible. In white adipose tissue of the db/db mutants lipogenesis (per tissue) reached a maximum at 5 weeks of age, and fell substantially between 10 and 20 weeks of age. Hepatic lipogenesis in the db/& mice rose pro~essiveIy from weaning until 8 weeks of age, and then decreased. Except at weaning, hepatic lipogenesis (per tissue) was much greater in db/db mice than in normal mice, and the liver was a more important site of lipogenesis in diabetic mice than in normals, accounting for up to 60% of the whole-body total. In contrast, BAT accounted for a considerably smaller proportion of whole-body lipogenesis in db/db mice than in normal mice. It is concluded that there are major developmental differences in lipogenesis between tissues of db/& mice, and between diabetic and normal animals. The data suggest that there is an early and preferential development of insulin resistance in BAT of the db/db mutant.

Introduction Brown adipose tissue (BAT) is the principal site of non-shivering thermogenesis in many mammals (see Ref. 1,2), and in some species it also plays an important role in the regulation of energy balance (see Refs. 2,3). Recent studies have established that fatty acids are the main substrate for thermogenesis in BAT, glucose being only of minor significance [4-61. The fatty acids utilized by the tissue may be obtained either from circulating triacylglycerols, or by de nova synthesis. Several reports indicate that BAT has a considerable capacity for lipogenesis, particularly in animals adapted to the cold where it is a major site of the conversion of carbohydrate to lipid [7-lo].

P. Trayhum, DiGsion of Correspondence and * Present address: Biochemical Sciences, Roweii Research Institute, Bucksbum, Aberdeen AB2 9SB, Scotland. U.K.

0005-2760/90/$03.50

0 1990 Elsevier Science Publishers

B.V. (Biomedical

In some physiological conditions the rate of lipogenesis in BAT parallels the thermogenic activity of the tissue 19-111. However, studies on suckling animals and on cold-acclimated animals fed high-fat diets have shown that the rate of lipogenesis in BAT is not necessarily linked to thermogenesis [10,12,13]. A ‘dissociation’ between the rates of thermogenesis and lipogenesis has also been observed in the genetically obese (ub/ob) mouse and the fatty (fu/fa) rat; relative to lean siblings these obese animals exhibit high rates of lipogenesis, but low levels of thermogenesis [2,3,14,15]. Developmental studies have indicated that the hyperhpogenesis in BAT of ob/ob mice is transitory, occurring only at weaning and the immediate post-weaning period 1151. In contrast, hyperlipogenesis persists in both white adipose tissue and the liver of the obese mutant [15]. The initial hyperlipogenesis in BAT results from the hyperinsulinaemia of the ob/ob mouse, lipogenesis in BAT being stimulated by insulin [7,S]. The sharp fall in lipogenic rate shortly after weaning has been attributed to the development of insulin resistance Division)

169 in BAT [15,16], and this appears to lead to a defective thermogenic response to cold in the tissue [16,17]. The diabetic (db/db) mouse is characterized by severe diabetes and moderate obesity, disorders which result from a single gene defect [3,18]. Previous work has demonstrated that the thermogenic activity and capacity of BAT is impaired in the db/db mutant [19]. In the present study the rates of lipogenesis in interscapular BAT of db/db mice have been investigated at different ages, from weaning onwards. The primary aim was to determine the effect of hyperinsulinaemia, which is evident from before weaning [18], and subsequent diabetes on lipogenesis in BAT of the db/db mutant. We were also concerned to compare the developmental changes in BAT with those occurring in white adipose tissue (WAT) and the liver, and to assess the relative importance of each of these organs to whole-body lipogenesis during the establishment of obesity and diabetes. Materials and Methods

Animals The animals were from a colony of diabetic (db/db) mice with the ‘db’ gene on the C57Bl/Ks background, maintained at the Dunn Nutrition Laboratory [19]. Diabetic (db/db) animals were bred from heterozygous (db/ + ) breeding pairs. Wea~ng was at 21 days of age, and diabetic individuals were identified by their mildly obese appearance. On weaning, the mice were paired (1 db/db and 1 normal animal) in plastic cages with sawdust bedding. Only males were used, with non-diabetic siblings (db/ + or + / + ) as controls. Lipogenesis was measured at the following ages: 21, 25/26, 30, 36/37 and 45 days, and at 8, 10 and 20 weeks. The mice were housed at 21 k 2OC, with a 12 h light/l2 h dark cycle (lights from 07.00 h). Water and a low fat/high carbohydrate rodent diet (LAD-l, K&K Greef Chemicals, -Croydon, U.K.) were available ad libitum. The diet contained (w/w) 21.3% protein, 3.4% lipid, 41.8% starch and 2.8% sucrose. Lipogenesis Plasma insulin levels were measured by radioimmunoassay, using a rat insulin standard [15]. Rates of lipogenesis were determined in vivo by measuring the incorporation into fatty acids of tritium from ‘H,O, essentially as previously [9-111. Each study was conducted at between 09.00 and 11.00 h, at the same temperature as the animal house, with the mice having been previously fed ad libitum. Each mouse was injected intraperitoneally with 400 PCi of 3H,0 (Amersham International, Amersham, U.K.) and returned to its home cage. After 60 min, blood was collected from the jugular veins into tubes containing anticoagulant, and the mice killed by cervical dislocation Interscapular BAT, the liver, and the epididymal

WAT were rapidly removed and frozen in liquid nitrogen, along with the remaining carcass. The tissues were stored at -2O”C, until analysis. Plasma was collected from whole blood by centrifugation, and deproteinized with 10% (w/v) trichloracetic acid [lo]. The amount of tritium in the deproteinized plasma was then measured, to determine the specific activity of the precursor pool. Tissue samples were saponified with ethanolic KOH, acidified, and fatty acids extracted with light petroleum, b.p. 40-60 o C [20]. The light petroleum extracts were evaporated to dryness in pre-weighed scintillation vials, and reweighed to obtain the fatty acid content of the tissues. In one set of studies the effect of insulin on rates of lipogenesis was examined, Mice were injected subcutaneously with ‘Neuphane’ isophane insulin B.P. (Wellcome Foundation, Beckenbam, U.K.), at a dose of 10 units/kg body wt. [16]. After 30 min, 3H,0 was administered intraperitoneally, and the experiment continued as above. Measurement of radioactivity and data analysis A toluene/Triton X-100 (2 : 1, v/v)-based scintillation solution was used for the measurement of radioactivity, in a Packard Tri-Carb 2425 liquid scintillation spectrometer. The counting efficiency was approximately 25%, and corrections were made for quench (external standard) and for background radioactivity. Rates of lipogenesis were calculated as ‘pg-atoms of ‘H’ incorporated per h’, without correction for the isotope discrimination effect [lo]. The statistical. significance of differences between normal and db/db mice at each age was assessed by Student’s t-test. Results

Tissue weights, fatty acid contents, and plasma insulin At weaning at 21 days of age the body weights of the db/db and normal mice were similar, and there was no significant difference in weight between the two groups until 45 days of age (Table I). At this stage, and thereafter, the diabetic animals were heavier than the controls. The amount of interscapular BAT was significantly greater in the db/db mice than in the controls at all ages (Table I). Epididymal WAT weigbt was also greater at each age in the db/db mutants than in the controls (Table I). Hypertrophy of the liver was evident in the db/db animals by 5 weeks of age (Table I). During the measurements of lipogenesis the fatty acid content of the main lipogenic organs was determined (Table II). In normal mice, 25% of the weight of the interscapular BAT pads at weaning was due to fatty acids, while in db/db animals a figure of 50% was obtained. Following weaning there was a rapid increase in the fatty acid content of the tissue in both groups of mice. The normal animals reached a plateau of 50-60%

170 of tissue weight by 30 days of age, with the db/db animals rising to a level of 7580% at 36 days of age. At most ages the fatty acid content of epididymal WAT was similar in normal and db/db mice, rising to 80-90% of tissue weight by 6-7 weeks of age. As expected, the fatty acid content of the liver of the normal mice was very low, ranging from 2 to 4% of organ weight at all ages. In contrast, although the fatty acid content of the liver of the db/db mice was similar to that of the normal animals at weaning, at all other ages it was much higher, averaging approx. 10% of organ weight. The db/db mice were markedly hyperinsulinaemic at weaning, and hyperinsulinaemia was maintained to a varying extent at all other ages (Fig. 1).

0-O 10

’ 110

B 130

’ 150

synthesis rates ‘per tissue’, whole-body synthesis and the contribution of each tissue can be assessed. At weaning, the rate of lipogenesis in interscapular BAT of the db/db mice was greater than in the normal animals, ‘per g tissue’ and ‘per tissue’ (Figs. 2A and 3A). In the first few days after weaning, both groups

I

Developmental

changes in bo& weight and tissue weights of normal and diabetic (db/db)

mice

are given as mean values& S.E., for six to nine mice in each group. Interscapular brown tissue wt. (mg)

Body wt. (g)

Age

21 days 25/26 days 30 days 36/37 days 45 days 8 weeks 10 weeks 20 weeks

normal

db/db

nortnal

1.7 + 0.4 11.5*0.7 16.1* 0.7 20.3 f 0.5 22.0 f 0.9 24.6kO.l 25.8 f 0.6 28.2 * 1.4

8.6f0.4 11.7*0.5 16.2kl.O 20.6fl.l 30.5 + 0.7 33.9* 1.1 34.7 f 1.2 46.7 f 2.0

21* 48+ 63rt 75+ 73* 103f 116+11 114*12

c C = c

’ P < 0.05, b P < 0.01, ’ P < 0.001 compared TABLE



90

Age (days)

Rates of lipogenesis were calculated ‘per g of tissue’ (wet wt.) and ‘per whole tissue’. The former provides a basis for comparing between different tissues. It does not, however, take into account the considerable variation in the lipid content of adipose tissues. From the

Results



70

.

50

Fig. 1. Plasma insulin levels in normal (0) and db/db ( 0) mice. Blood was collected from mice at different ages and the plasma insulin concentration measured by radioimmunoassay. Results are given as mean values f SE. (bars), for five to eight mice at each age

Lipogenesis

TABLE

*

30

4 4 4 3 7 4

with normal

adipose

Epididymal white adipose tissue wt. (mg)

Liver wt.

(g)

db/db

normal

db/db

normal

db/db

66+ 4c 129+ 8” 173*12’ 231+15’ 206kll’ 323+14’ 231k14’ 250+11=

17* 5 44+ 8 90* 14 166+ 14 222* 29 321+ 20 402* 33 543*113

70+ 15b 116+ 17b 226k 45 a 375* 42’ 888f 63’ 1168k 66’ 16OOf106 c 2011* 9’

0.32 + 0.04 0.65 k 0.06 0.98 zb0.03 1.27+0.03 1.32 + 0.05 1.32+0.06 1.43 f 0.08 1.51+0.03

0.31+ 0.02 0.68 + 0.04 1.09 + 0.08 1.66+0.15 2.53 + 0.14 3.03 + 0.22 2.34+0.10’ 3.39+0.1s

B ’ c ‘

mice of the same age.

II

Developmentalchangesin the fatty acid content of interscapular diabetic (db/db)

The fatty acid content mice in each group. Age

of the tissues was determined as described ’ P -C 0.05, b P < 0.01, ’ P < 0.001, compared Fatty acid content Interscapular

21 days 25/26 days 30 days 36/37 days 45 days 8 weeks 10 weeks 20 weeks

brown adipose tissue, epididymal

white adipose tissue and the liver of normal and

mice

brown

in the Materials and Methods. Results with normal mice of the same age.

are given as mean values + SE., for 6-9

of tissue (X of wet wt.) adipose

normal

db/db

25.2 f 4.2 38.5f5.2 50.2 f 1.7 52.1 f 2.3 50.8 rt 4.2 59.4* 1.1 53.9f2.2 53.7 f 3.4

49.6 f 50.4 f 67.6 + 74.7 f 19.3 f 78.8 f 75.9 f 79.4*

tissue

Epididiymal normal

6.2 b 4.4 1.5 = 2.1= 0.1= 1.2 ’ 2.1= 3.0 =

60.2 + 55.2 k 71.2 k 75.6+ 78.8 f 85.3 f 84.4 + 85.6 +

1.3 3.3 2.7 1.1 1.3 1.6 1.0 0.5

white adipose

tissue

Liver

db/db

normal

db/db

69.9k4.1 66.7 f 2.0 a 74.4* 3.9 80.2 f 1.6 a 84.5 * 2.4 88.4 f 1.8 83.0 f 4.0 a5.9*0.5

2.8 f 0.3 3.6 + 0.3 3.8 * 0.2 3.5 *0.1 3.7 + 0.1 3.7 f 0.1 3.6kO.2 4.2 f 0.3

3.1 f 0.7 5.1+0.4 7.6 + 0.7 9.6+1.1’ 10.2 f 1.5 12.4* 1.9 6.4kO.7 9.9 f 1.2

a ’ C ’ ’ b

171

showed a marked rise in lipogenic rate, irrespective of the mode of expressing the results. In the normal mice the rate peaked at 5 weeks of age, with little change over the subsequent 15 weeks. A very different pattern was, however, apparent in the db/db mutants, the peak in lipogenesis occurring at 26 days of age with a sharp fall over the following 2-3 weeks. This pattern is evident ‘per g’ and ‘per tissue’, but the differences between db/db and normal mice are particularly clear when expressed on a whole-tissue basis. At 26 days of age the lipogenic rate in BAT ‘per tissue’ of the db/db animals was 4.4-times that of the normal mice, but by 45 days of age it had declined such that it was lower than the normals (66% of normals). From 45 days to 10 weeks of age lipogenesis in BAT of the db/db mice was essentially constant, and below that of the normal animals. Although there was no change in rate in the normal mice between 10 and 20 weeks of age, there was a

c d

150

Age(days) Fig. 3. Developmental changes in the rate of lipogenesis ‘per tissue’ in normal (9) and db/db (0) mice. (A) Interscapular brown adipose tissue, (B) epididymal white adipose tissue, (C) liver. Lipogenesis was measured in viva with 3H,0. Results, which were calculated from the synthesis rates ‘per g of tissue’ (Fig. 2) and the tissue weight (Table I), are given as mean values* S.E. (bars), for six to nine animals at each age. ’ P < 0.05, b P -z 0.01, ‘P -z 0.001 compared of the same age.

90 q f EF 8

110

300;;-; c 200.

O

100.

ii

130

16.6

.c k E s 0 s

\

0 10

\ 0.

30

l

!_____,

-@./-----o

50

70

90

110

mice

150 CC)

c

with normal

130

150

Age (days) Fig. 2. Developmental changes in the rate of lipogenesis ‘per g of tissue’ in normal (0) and db/db (0) mice. (A) Interscapular brown adipose tissue, (B) epididymal white adipose tissue, (C) liver. Lipogenesis was measured in vivo with 3H,0; for full details see Materials and Methods. Results are given as mean values f SE. (bars), for six to nine animals at each age. ’ P < 0.05, b P < 0.01, ‘P < 0.001 compared with normal mice of the same age.

substantial reduction in the diabetic animals over the same period. Lower rates of lipogenesis occurred for both groups of mice in WAT than in BAT. As with BAT, the lipogenic rate in epididymal WAT at weaning was greater in the db/db mice than in the normal animals (Figs. 2B and 3B). In both genotypes the rate increased rapidly after weaning, reaching a peak (per g and per tissue) in the normals at around 30 days of age. In the db/db group the peak lipogenic rate occurred at 36 days of age. At this point the rate of lipogenesis per tissue in the db/db mice was 6.4times that of the normal animals. From 5 to 10 weeks of age there were no major changes in synthesis ‘per tissue’ in either group, but between 10 and 20 weeks there was a substantial fall in lipogenesis in the epididymal pads of the diabetic mice, similar to BAT.

172 80 1

(A)

bT

TABLE

111

Effect of insulin administration diabetic (dh/db) mice

on the rute of lipogenesrs rn normal und

Mice. aged 40 days, were injected subcutaneously with insulin units/kg body weight) and ‘H,O administered intraperitoneally mm later. For full details, see Materials and Methods. Results given as mean values? SE.. for five mice in each group.

10

30

50

70

90

110

130

Lipogenesis (pg atoms ‘H’ incorporated/h per tissue)

150

control Brown adipose normal db/db Epididymal normal db/db

10

30

50

70

90

110

130

150

Age (days)

Fig. 4. Developmental changes in lipogenesis in the carcass (A), and in the whole-body (B), of normal (0) and db/db (0) mice. Lipogenesis was measured in vivo with 3H,0. Whole-body synthesis was obtained by su mming the total synthesis in individual tissues (Fig. 3) and the carcass fraction. Results are given as mean values+ S.E. (bars), for six to nine animals at each age. a P < 0.05, b P -C0.01. ‘P i 0.001 compared with normal mice of the same age.

In contrast to the adipose tissues, there was no difference between normal and db/db mice in the rate of lipogenesis in the liver at weaning, either ‘per g’ or on a whole organ basis (Figs. 2C and 3C). Following weaning, a rapid increase in lipogenesis was apparent in both groups. The peak in lipogenesis (per g and per tissue) in the normal mice occurred at 30 days of age, but in the db/db animals there was an almost linear increase from weaning until a peak was reached (per tissue) at 8 weeks of age (Fig. 3C). At this time, total lipogenesis in the liver was 13.6-times greater in the diabetic mice than in the normal animals. There was a sharp fall in hepatic lipogenesis from 8 to 10 weeks of age, but no further reduction was apparent between 10 and 20 weeks, the rate per tissue in the diabetic animals remaining several times greater than in normal mice. There was little difference at weaning in the rate of lipogenesis in the carcass fraction of normal and db/db animals (Fig. 4A). Thereafter, the pattern for both genotypes was similar to that in BAT. Whole-body lipogenesis was calculated from the rate in the individual tissues, and the carcass fraction. It peaked in both types of mice at 30-36 days of age, and changed little thereafter in the normal animals (Fig. 4B). However, in the diabetic mice there was a marked fall in whole-body lipogenesis between 10 and 20 weeks of age. At all ages, total lipogenesis was higher in the db/db mice than in normal animals.

(10 30 are

Liver: normal db/db Whole-body: normal db/db

+ insulin

tissue: 7.7+ 14.8+

2.5 1.8

white adipose tissue: 3.2+ 0.8 48.Ok 4.3

42.8t 6.1 679 +69

216 1281

+32 &72

a P < 0.05. b P -c 0.01. C P i 0.001. compared

23.6 * 21.oi

5.4 d 3.3

12.3* 45.8+

1.7 h 5.5

89.2i 3.3 ’ 546 *74

632 1231

+90 h +83

with normal

mice.

Effects of insulin on lipogenesis In a separate set of experiments the effect of exogenous insulin on the rates of lipogenesis was examined in mice aged 40 days. High doses of insulin were used to give a maximum stimulation of lipogenesis [15,16]. Insulin administration to normal mice induced a 3-4-fold increase in total tissue lipogenesis in BAT and epididyma1 WAT, and whole-body lipogenesis was stimulated similarly (Table III). Hepatic lipogenesis was also significantly increased by insulin, but to a lesser extent than in the other tissues. In contrast to the normal animals, there was no significant effect of insulin on total tissue lipogenesis in BAT, WAT, or the liver of the db/db mice (Table III). Whole-body lipogenesis was also unaltered following the administration of insulin. Discussion Thermogenesis in BAT is fuelled primarily by fatty acids [4-61, which may be obtained either from circulating triacylglycerols or by de novo synthesis within the tissue. The present study has examined the developmental changes in fatty acid synthesis in BAT of normal and diabetic (db/db) mice, and compared these changes with those in the other major lipogenic organs. The db/db mouse has severe diabetes, a moderate obesity [18], and impaired thermogenesis [19], and as such provides a valuable model for investigating the relationship between diabetes and BAT function.

173 After weaning at 21 days of age there was a rapid increase in the rate of lipogenesis in BAT, and in each of the other tissues examined. In normal mice, peak rates of lipogenesis were observed at 30-36 days of age, with no major changes in the subsequent period up to 20 weeks of age. Rates of lipogenesis increased particularly rapidly after weaning in the db/db mice, the increases being more marked than in normal animals. There were, however, major differences in the developmental profile of different tissues in the db/db mutants, and between the diabetic and normal animals. In each of the tissues of db/db mice the post-weaning peak in lipogenesis was followed by a substantial reduction. This pattern was particularly marked in BAT, which exhibited a sharp peak in lipogenic rate 5 days after weaning, followed by a rapid fall. The reduction in lipogenesis in BAT was so substantial that by 20 weeks of age the rate ‘per tissue’ was reduced to only 8% of that of normal animals, and to a mere 2% of the peak at 26 days of age. Indeed, with respect to lipogenesis, BAT had effectively ‘atrophied’ in the diabetic mice by 20 weeks of age. ‘Hyperlipogenesis’ was sustained, however, in the liver of the diabetic animals, while in the epididymal WAT it was only at 20 weeks of age that the lipogenic rate was not greater in the diabetic mice than in the normal animals. The rapid development of hyperlipogenesis in tissues of db/db mice on weaning presumably reflects their marked hyperinsulinaemia. Hyperinsulinaemia is established in the db/db mutant by weaning, and lipogenesis in BAT is stimulated by insulin [7,8]. The substantial fall in lipogenesis in BAT of the diabetic animals, to below that of the normal siblings by 45 days of age, occurred despite continuing hyperinsulinaemia, suggesting that the tissue develops insulin resistance. This appears to be preferential to BAT since hyperlipogenesis was maintained for much longer in both WAT and the liver. Exogenous insulin did not increase lipogenesis in tissues of db/db mice at 40 days of age, in contrast to normal animals. This indicates that tissues of the diabetic mutant are either insulin resistant, or are being maximally stimulated by the endogenous levels of the hormone. At 40 days of age there is, even after the injection of insulin, a substantial hyperlipogenesis in both liver and WAT of the diabetic animals, relative to normal mice. This is not, however, the case with BAT, which is consistent with the proposition that there is a selective development of insulin resistance in BAT of the db/db mutant. The preferential development of insulin resistance in BAT has been observed previously in the obese (ob/ob) mouse [15,16], and in mice made obese with gold thioglucose [21]. Insulin resistance in BAT has also been reported in the Zucker (fa/fa) rat [22]. Insulin resistance in the tissue appears, therefore, to be a common

early feature of animal models of obesity and diabetes [15,21,23]. The defect is one of several in BAT of obese/diabetic animals (see Refs. 2,3), but the implications for thermogenesis have not been widely explored. Insulin resistance does not appear to affect the basal activity of the thermogenic mitochondrial proton conductance pathway, but studies on the ob/ob mouse suggest that it does impair the acute cold-induced activation of the pathway [16,17]. Insulin resistance in BAT may also inhibit mitochondrial proliferation, and the induction of uncoupling protein, on cold exposure [24]. Regulation of the level of uncoupling protein is an important effect of insulin on BAT [25]. Insulin resistance in obese/diabetic animals may result primarily from hyperinsulinaemia, but this does not satisfactorily explain the early and preferential appearance of the abnormality in BAT. Obese animals, including db/db mice, generally exhibit low levels of thermogenesis in BAT under normal conditions (see Refs. 2,3), resulting in a reduced substrate requirement. The decrease in thermogenesis stems from a reduction in sympathetic activity (see Refs. 2,3), and it is possible that the low level of sympathetic stimulation may lead to a loss of insulin sensitivity in BAT, with subsequent overt insulin resistance [26]. Insulin resistance, together with the reduced substrate requirement for thermogenesis, reduces the importance of BAT as a site of lipogenesis and of glucose uptake. This in turn may contribute to the induction of glucose intolerance and the initiation of marked diabetes in diabetic/obese animals [27-291. BAT is a highly insulin sensitive tissue [23,28], and has been reported to account for up to 12% of glucose uptake from blood [22,30]. Previous studies using tritiated water have indicated that BAT is a major site of lipogenesis in rats and mice [7,9,10]. In the present study, interscapular BAT accounted for approximately 4% of whole-body lipogenesis at most ages in normal animals (Fig. 5A). Since the interscapular site is a quarter of all the BAT in rats and mice [lo], it is estimated that the total tissue can account for some 16% of whole-body lipogenesis. This figure is close to that obtained previously for other mice [lo]. A more complex picture is evident in the db/db mutant (Fig. 5A). At weaning the interscapular depot accounted for 5% of whole-body lipogenesis, which on the same basis as the normal mice suggests that 20% of total lipogenesis is attributable to BAT. However, the contribution declines rapidly with age, and by 20 weeks interscapular BAT accounts for only 0.2%, and total BAT an estimated 0.8% of whole-body lipogenesis. The liver is only a minor site of lipogenesis at weaning, both in normal and in db/db mice (Fig. 5B). By 25 days of age, however, it had risen to 25% of whole-body lipogenesis in normal mice, with no significant increase thereafter. In contrast, in the diabetic mice the contri-

174 References

10

30

50

30

50

70

90

110

130

150

90

110

130

150

60 . 50 40 30. 20. 10.

0’

10



3

70 Age

(days)

Fig. 5. Developmental changes in the relative importance of interscapular brown adipose tissue (A), and the liver (B) to whole-body Iipogenesis in normal (0) and db/db (0) mice. Calculated from the data in Figs. 3 and 4. Results are given as mean values* SE. (bars), for six to nine animals at each age. a P < 0.05, b P i 0.01, ‘P -C 0.001 compared with normal mice of the same age.

bution of the liver to total lipogenesis rose progressively, to a figure of almost 60% by 8 weeks of age. It is concluded that there are major developmental changes in the db/db mutant in the relative importance of different tissues to whole-body lipogenesis. It is apparent from the present results that BAT is unlikely to be a major source of fatty acids for deposition in other tissues during the development of obesity in the db/db mutant, in contrast to the suggestion made for Zucker (fu/fa) rats [14].

Acknowledgements We are grateful advice.

to Dr Stewart

Mercer

for his helpful

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db) mice: developmental changes in brown adipose tissue, white adipose tissue and the liver.

Developmental changes in lipogenesis have been examined in interscapular brown adipose tissue (BAT), epididymal white adipose tissue and the liver of ...
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