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Endocrinology 142(12):5220 –5225
Copyright © 2001 by The Endocrine Society
Angiotensinogen-Deficient Mice Exhibit Impairment of
Diet-Induced Weight Gain with Alteration in Adipose
Tissue Development and Increased Locomotor Activity
Centre National de la Recherche Scientifique 6543, Centre de Biochimie (F.M., P.S.-M., R.N., G.A., M.T.), Nice 06108,
France; University Medical Center, Department of Physiology, Faculty of Medicine (J.S.), Genève, 1211, Switzerland;
INSERM, U-352, Institut National des Sciences Appliquées (A.G.), Villeurbanne 69100, France; INSERM, U-465 (A.Q.-B.,
S.T.), Paris 75270, France; and University of Tsukuba (A.F.), Tsukuba, Ibaraki 305, Japan
White adipose tissue is known to contain the components of
the renin-angiotensin system, which gives rise to angiotensin
II from angiotensinogen (AGT). Recent evidence obtained in
vitro and ex vivo is in favor of angiotensin II acting as a trophic
factor of adipose tissue development. To determine whether
AGT plays a role in vivo in this process, comparative studies
were performed in AGT-deficient (agtⴚ/ⴚ) mice and control
wild-type mice. The results showed that agtⴚ/ⴚ mice gain less
weight than wild-type mice in response to a chow or high fat
diet. Adipose tissue mass from weaning to adulthood appeared altered rather specifically, as both the size and the
weight of other organs were almost unchanged. Food intake
was similar for both genotypes, suggesting a decreased metabolic efficiency in agtⴚ/ⴚ mice. Consistent with this hypothesis, cellularity measurement indicated hypotrophy of adipocytes in agtⴚ/ⴚ mice with a parallel decrease in the fatty acid
synthase activity. Moreover, AGT-deficient mice exhibited a
significantly increased locomotor activity, whereas metabolic
rate and mRNA levels of uncoupling proteins remained similar in both genotypes. Thus, AGT appears to be involved in the
regulation of fat mass through a combination of decreased
lipogenesis and increased locomotor activity that may be centrally mediated. (Endocrinology 142: 5220 –5225, 2001)
A
3T3-L1 preadipose cells and human adipocytes (10), consistent with the observation that rats treated with an oral AngII
receptor antagonist (losartan) exhibit a decrease in adipocyte
size (11). In contrast to liver cells, in which numerous hormones have been shown to enhance AGT mRNA levels and
AGT secretion, adipose cells respond only to fatty acids and
glucocorticoids, which are known to be implicated in the
hyperplastic and hypertrophic growth of WAT (12, 13). Collectively, these studies indicate that AngII plays a local role
in the development of adipose tissue and its cellularity, i.e.
fat cell number and size. To gain a better understanding of
the effects of AngII on adipose tissue growth, we examined
the comparative development of WAT and brown adipose
tissue (BAT) in wild-type (WT) and AGT-deficient (agt⫺/⫺)
mice in response to a standard chow or a high fat diet.
NGIOTENSINOGEN (AGT), the unique substrate of
renin, is the precursor of angiotensin I (AngI) that
gives rise to active angiotensin II (AngII) through the action
of AngI-converting enzyme. The renin-angiotensin system is
known to have a major role in the regulation of blood pressure and fluid and sodium homeostasis (1). White adipose
tissue (WAT) is an important extrahepatic production site of
AGT (2), and several reports have suggested the existence of
a functional renin-angiotensin system in this tissue. In isolated adipocytes and cultured adipose cells from rodents and
human, recent data have shown the presence of 1) renin, by
RT-PCR, and renin-like activity; 2) AngI-converting enzyme,
by RT-PCR and Western blot; and 3) AngII production (3–5).
In addition to the systemic effect of AngII in the regulation
of blood pressure, various roles of AGT via locally produced
AngII have been proposed: 1) at the time of adipose tissue
development, AngII appears to be a trophic factor that is
involved in organogenesis of rodent, primate, and human
fetuses (6); 2) AngII has been implicated in cell cycle progression of human preadipocytes, the cell type that precedes
the formation of nondividing adipocytes (7); 3) both in vitro
and in vivo, AngII stimulates the production and release of
prostacyclin from adipocytes, which, in turn, stimulates adipogenesis of adipose precursor cells (8, 9); and 4) AngII
increases lipogenesis and triglyceride accumulation in
Abbreviations: AGT, Angiotensinogen; AngII, angiotensin II; BAT,
brown adipose tissue; FAS, fatty acid synthase; UCP-1, uncoupling
protein-1; WAT, white adipose tissue; WT, wild-type.
Materials and Methods
Mice
Generation of agt⫺/⫺ mice has been previously described. Briefly,
chimeric mice were backcrossed with ICR mice for at least 10 generations
(14), then 5 agt⫺/⫺ males and 15 agt⫺/⫺ females were bred to generate
further generations. AGT-deficient mice reproduced normally. Although some perinatal lethality occurred, as previously reported (15),
their life expectancy after weaning was not different from that of WT
mice.
Only male agt⫺/⫺ mice were used in the experiments herein, and
ICR-CD1 control WT mice were purchased from Harlan (Gammat,
France). Animals were housed five per cage and had free access to food
and water in a controlled environment with a 12-h light, 12-h dark cycle
and constant temperature (22 C). At weaning, the mice were fed either
5220
Downloaded from https://academic.oup.com/endo/article-abstract/142/12/5220/2988805 by guest on 21 May 2020
FLORENCE MASSIERA, JOSIANE SEYDOUX, ALAIN GELOEN, ANNIE QUIGNARD-BOULANGE,
SOPHIE TURBAN, PERLA SAINT-MARC, AKIYOSHI FUKAMIZU, RAYMOND NEGREL,
GÉRARD AILHAUD, AND MICHÈLE TEBOUL
Massiera et al. • Fat Mass Alterations in Angiotensinogen-Deficient Mice
a standard laboratory chow diet or a high fat diet containing 1% cholesterol, 30% corn oil (representing 65% of calories as fat), 27% carbohydrates, 11.5% proteins, and 1.9% minerals (UAR, Villemoisson,
France). Body weight was assessed weekly for up to 46 wk. At the
indicated times mice were killed by cervical dislocation according to
French Centre National de la Recherche Scientifique ethical guidelines.
Epididymal WAT, BAT, and hind limb skeletal muscle were rapidly
removed and immediately used for RNA preparation.
Food consumption and feces analysis
Body weight and body composition
Body weight was measured at the same time each day. For body
composition, mice were killed by cervical dislocation, and the whole
carcasses were incised, dried to a constant weight at 70 C, then subsequently homogenized. Total body fat content was determined by the
Soxhlet extraction method as described above. The results are presented
as absolute weight (grams) and as percentage of total body weight. The
fat-free mass, which includes mineral content (which accounts for ⬃2%
of fat-free mass in mice) was obtained by subtraction of body fat content
from dry weight.
Adipose tissue cellularity
The size and number of adipocytes were determined as previously
described (16). Briefly, fat cell size was determined by a procedure
derived from a microphotometric method; micrographs of isolated cells
were taken with a light microscope, and measurement of cell diameters
was performed using a computer equipped with an image analyzer. Fat
cell number was estimated from a portion of adipose tissue by dividing
the lipid content by the average fat cell weight.
Fatty acid synthase (FAS) activities
FAS activities were assayed spectrophotometrically in crude cytosolic
extracts of epididymal fat pads by measuring the oxidation of NAPDH
in the presence of acetyl coenzyme A and malonyl coenzyme A (17). Data
are expressed as nanomoles of NAPDH oxidized per min/mg, i.e. milliunits per mg cytosolic proteins, which were assayed by the method of
Bradford (17).
5221
equation and expressed in terms of watts per kg BW to the 0.75 power.
For each mouse, the mean metabolic rate was calculated for the last 23 h.
For locomotor activity, the system used has been previously described (19). The home-cage traveled distance was measured during
either the lights on or lights off cycle. When placed in this experimental
set-up, mice did not have access to food or water. Quantitative analyses
of the distance traveled during the entire period were made off-line. The
fraction of time spent in activity was calculated by measuring the time
during which the animal showed a displacement of its center of mass of
at least 1 cm. All calculations were made using 386-Matlab (Mathworks,
Sherborn, MA).
Blood parameters
Mice were anesthetized 2 h after lights on with 60 l xylene/ketamine
(1:4, vol/vol). Blood was collected by eye puncture into tubes containing
citrate at a final concentration of 0.01 m. After 10 min in ice, plasma was
separated by centrifugation at 10,000 ⫻ g for 10 min and stored at ⫺20
C. A volume of 100 l of a 1:2 dilution was used for mouse leptin assays
using a commercial kit (R&D Systems, Inc., London, UK). Glucose,
triglycerides, total cholesterol, and free T3 were determined using standard laboratory procedures. Insulin was determined by RIA using a
commercial kit with a rat insulin standard (CIS Biointernational,
Gif-sur-Yvette, France).
Statistical analysis
All data are expressed as the mean ⫾ sem. The values were examined
by the one-way ANOVA or t test with the computer software STATISTIX, version 4.0 (Analytical Software, Tallahassee, FL).
Results
AGT-deficient mice gain less weight than WT mice in
response to diets
Male agt⫺/⫺ and WT mice were fed a chow diet or a high
fat diet from weaning up to 46 wk of age. Figure 1 shows that
agt⫺/⫺ mice exhibited, at weaning, a lower body weight than
WT mice. This difference remained throughout development
and was still observed at adulthood. At 6 wk of age and
thereafter, agt⫺/⫺ mice fed a chow diet weighed 21% less
than WT mice. At that age, the epididymal fat pad mass of
chow-fed agt⫺/⫺ mice was 2-fold lower than that of WT mice
(see Table 2), and at 20 wk of age (Table 1, Exp 1), the total
fat mass of agt⫺/⫺ mice, determined on whole carcasses, was
Isolation and analysis of RNA
RNA was extracted using the RNeasy Midi kit according to the
manufacturer’s protocol (QIAGEN, Cortaboeuf, France). Northern blot
analysis was performed as described previously (18). Autoradiographs
were quantified using a Fujix PhosphorImager (Tokyo, Japan). All
results were normalized to -actin signals.
Measurements of metabolic rate and locomotor activity
For locomotor activity and metabolic rate measurements, mice were
randomly and alternatively placed into the respective experimental
chambers; at least 1 wk separated successive testing. Metabolic rate was
measured by indirect calorimetry during 24 h. An open circuit calorimeter, as described in detail previously (19), equipped with a sensitive
mass flow meter (model 5875, Brooks Instrument, Veenendaal, The
Netherlands) was used. Food and water were available during testing.
The ambient temperature was set at 22 C. The data were recorded every
5 sec by an on-line computerized data acquisition system (SICMU, CMU,
Geneva, Switzerland). The metabolic rate was calculated using Weir’s
FIG. 1. Body weights of WT (〫 and ⽧) and AGT-deficient (䡺 and f)
mice fed a chow diet (〫 and 䡺) or a high fat diet (⽧ and f) from
weaning onward (n ⫽ 30).
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Mice were housed individually in metabolic cages (Marty Technology, Marcilly-sur-Eure, France) for 1 wk, fed ad libitum with a standard
or high fat diet, and given free access to water in a controlled environment at 22 C with a 12-h light, 12-h dark cycle. Food consumption was
measured during the last 4 d as the difference between the amount of
food given and that removed from the cage after the amount of any food
spilled was taken into account. Similarly, feces were collected during the
last 4 d of feeding, and the weight of pooled feces was determined after
drying at 70 C to a constant weight. The fat content of the feces was
determined by the Soxhlet extraction method using petroleum benzine.
Endocrinology, December 2001, 142(12):5220 –5225
5222 Endocrinology, December 2001, 142(12):5220 –5225
Massiera et al. • Fat Mass Alterations in Angiotensinogen-Deficient Mice
intestinal absorption. As shown in Table 1, this was excluded,
as food intake was higher in chow-fed agt⫺/⫺ than in WT
mice and even higher in agt⫺/⫺ mice when corrected for
body mass to the 0.75 power, consistent with similar levels
of circulating leptin (see Table 3). In high fat-fed animals,
food intake was slightly lower in agt⫺/⫺ mice, but, when
expressed per body mass to the 0.75 power, both values were
similar. Feces analysis did not show any difference in daily
quantity or fat content expressed as a percentage of dry
weight between WT and agt⫺/⫺ mice. Because the body
composition data in Table 1 indicate that the lower body
weight was partly due to a lower fat mass, a detailed analysis
of WAT cellularity was performed.
AGT deficiency leads to alterations in WAT cellularity
TABLE 1. Body weight, food intake, body composition, and feces
analysis in wild-type and AGT-deficient mice (16 –20 wk old)
Exp 1: chow diet
BW (g)
Food intake (g/day)
Food intake (g/day䡠g
BW0.75)
Fat mass (g)
Adiposity (% of BW)
Fat-free mass (g)
Leanness (% of BW)
Feces (g/day)
Feces fat content
(% of dry wt)
Exp 2: high fat diet
BW (g)
Food intake (g/day)
Food intake (g/day䡠g
BW0.75)
Feces (g/day)
Feces fat content
(% of dry wt)
Wild-type
(n ⫽ 6)
AGT-deficient
(n ⫽ 9)
P
48.5 ⫾ 1.7
5.3 ⫾ 0.3
0.3 ⫾ 0.01
34.2 ⫾ 1.2
5.4 ⫾ 0.2
0.4 ⫾ 0.02
⬍0.005
NS
⬍0.05
4.5 ⫾ 0.7
9.1 ⫾ 1.2
11.2 ⫾ 0.3
23.1 ⫾ 0.3
1.3 ⫾ 0.1
2.4 ⫾ 0.2
2.4 ⫾ 0.3
6.9 ⫾ 0.6
8.1 ⫾ 0.2
23.7 ⫾ 0.2
1.1 ⫾ 0.1
2.5 ⫾ 0.2
⬍0.01
⬍0.05
⬍0.001
NS
⬍0.01
NS
Wild-type
(n ⫽ 8)
AGT-deficient
(n ⫽ 8)
P
48.5 ⫾ 1.2
5.0 ⫾ 0.2
0.3 ⫾ 0.01
30.7 ⫾ 1.3
4.0 ⫾ 0.45
0.3 ⫾ 0.02
⬍0.001
⬍0.01
NS
1.1 ⫾ 0.1
4.1 ⫾ 0.2
1.1 ⫾ 0.1
3.9 ⫾ 0.2
NS
NS
Mice were fed either a chow diet (Exp 1) or a high fat diet (Exp 2)
from weaning onward. For body composition, mice were weighed and
killed at 20 weeks of age, and total carcass fat content and fat-free dry
mass were determined as described in Materials and Methods. All
results are the means ⫾ SE. n, Number of animals.
When fed a standard chow diet, a 2-fold difference was
observed in the weight of epididymal fat pads between WT
mice and agt⫺/⫺ mice. This was due to hypotrophy of adipocytes in agt⫺/⫺ mice, as the number of adipocytes was
similar in both genotypes. Upon high fat feeding, i.e. under
conditions of increased supply of exogenous fatty acids to
adipose tissue, the size of adipocytes increased in both genotypes, but to a greater extent in WT mice. A significantly
lower weight of epididymal fat pads and fat cell size persisted in agt⫺/⫺ mice (Table 2). Importantly, in agt⫺/⫺ mice
only, adipose tissue hypoplasia was observed in high fat-fed
mice compared with chow-fed mice.
To gain some insights on the metabolic pathways leading
to triglyceride accumulation in adipocytes, measurement of
endogenous fatty acid synthesis was performed by determining FAS activities of cytosolic extracts of epididymal fat
pads of WT and agt⫺/⫺ mice. Data from Table 2 show that
FAS activity was 2.2-fold higher in extracts from WT mice
than in those from agt⫺/⫺ mice, consistent with the 2.6-fold
increase observed in adipocyte weight. This observation is
also in accordance with a report showing that AngII regulates lipogenesis by increasing FAS activity (10). Upon high
fat feeding, it is known that the exogenous supply of fatty
acids from chylomicrons increases dramatically, leading to a
down-regulation of FAS activity. As shown in Table 2, this
down-regulation was taking place in WT mice, but not in
agt⫺/⫺ mice, suggesting that this modulation was AngII
related.
TABLE 2. Cellularity of epididymal fat in 6-wk-old mice fed a chow or a high fat diet after weaning
Chow diet
BW (g)
Epididymal fat (mg)
Fat cell no. (⫻106)e
Cell diameter (m)
Cell weight (ng)
FAS activity (mU/mg)
AGT-deficient
n
Wild-type
AGT-deficient
n
30.4 ⫾ 0.2
248.8 ⫾ 13.0
4.6 ⫾ 0.5
48 ⫾ 1
55.8 ⫾ 4.4
28.3 ⫾ 3.5
24.3 ⫾ 0.3a
124.5 ⫾ 7.0a
5.9 ⫾ 0.7
31.2 ⫾ 1a
21.2 ⫾ 1.5a
12.8 ⫾ 1.5a
50
36
10
10
10
5
33.3 ⫾ 0.6b
365.0 ⫾ 32.0b
3.6 ⫾ 1.4
53 ⫾ 2.6b
81.3 ⫾ 6.9a
11.7 ⫾ 3a
23.8 ⫾ 0.6c
251.5 ⫾ 17d,g
3.9 ⫾ 0.5d
48 ⫾ 2.6d
56.8 ⫾ 8.4f,g
10.5 ⫾ 4.3d
33
8
6
6
6
6
FAS, Fatty acid synthase.
P ⬍ 0.001 vs. chow-fed wild-type.
b
P ⬍ 0.01 vs. chow-fed wild-type.
c
P ⬍ 0.001 vs. high fat-fed wild-type.
d
P ⬍ 0.005 vs. chow-fed AGT-deficient.
e
Fat cell number is expressed per two epididymal fat pads.
f
P ⬍ 0.001 vs. chow-fed AGT-deficient.
g
P ⬍ 0.01 vs. high fat-fed wild-type.
a
High fat diet
Wild-type
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1.9-fold lower. Due to the trophic role of AngII in organogenesis, it is of interest to note that the effect of AGT deficiency was mainly confined to adipose tissue. At 6 wk of age,
no effect on body length was observed (8.8 ⫾ 0.4 cm for WT
vs. 9.0 ⫾ 0.5 for agt⫺/⫺ mice; n ⫽ 20), and only a modest effect
(⬍1.2-fold) was detected on the weights of heart, kidney, and
liver. Although fat-free mass was lower in agt⫺/⫺ mice, leanness (expressed as protein mass per g BW) was similar in
agt⫺/⫺ and WT mice in adulthood (Table 1). In WT mice, a
weight gain of 10 –15% was induced by high fat feeding; in
contrast, no weight gain but, instead, a slight decrease (6%)
was observed in agt⫺/⫺ mice compared with mice fed a chow
diet (Fig. 1). The lack of weight gain in agt⫺/⫺ mice fed a high
fat diet could be due to lower food intake and/or defective
Massiera et al. • Fat Mass Alterations in Angiotensinogen-Deficient Mice
AGT deficiency and thermogenesis-related parameters
5223
increased by 69% and 32% compared with that in WT mice,
respectively. The time spent in activity during the lights on
periods was also increased in agt⫺/⫺ mice compared with
WT mice (Table 4).
Discussion
To determine the role of AGT in body weight and composition, we performed a detailed comparison of agt⫺/⫺ vs.
control WT mice. The phenotype of the AGT-null mouse line
offers clues to the function of AGT as a secretory product
from adipocytes. It is known that AngII plays a trophic role
in fetal and/or postnatal development in rodents and humans (6). Our results provide evidence that in the absence of
AGT, WAT development from weaning to adulthood is impaired, whereas both the length of the mice and the weight
of other organs are only weakly altered, if at all. Contributing
in part to the lower body weight are the smaller fat stores in
agt⫺/⫺ mice compared with WT mice. Cellularity measurements of epididymal fat pads indicate that this was mainly
AGT-deficient mice exhibit increased locomotor activity
To investigate whether changes in energy expenditure
could account for the lower metabolic efficiency in AGTdeficient mice compared with WT mice, metabolic rate and
locomotor activity were measured in chow-fed mice. When
oxygen consumption was corrected for body weight, similar
mean metabolic rates were obtained for both genotypes. Locomotor activity was significantly enhanced in agt⫺/⫺ mice;
the distance covered during lights on and lights off was
FIG. 2. UCP RNA content of interscapular BAT, epididymal WAT,
and skeletal muscle (SM) of 6-wk-old WT and AGT-deficient (agt⫺/⫺)
mice.
TABLE 3. Blood parameters in 6-wk-old mice fed a chow or a high fat diet after weaning
Chow diet
Triglycerides (g/liter)
Cholesterol (g/liter)
Glucose (g/liter)
Insulin (U/ml)
Leptin (ng/ml)
T3 (pmol/liter)
a
AGT-deficient
n
Wild-type
AGT-deficient
n
1.04 ⫾ 0.01
0.96 ⫾ 0.03
2.3 ⫾ 0.09
34.3 ⫾ 5.2
2.09 ⫾ 0.24
4.43 ⫾ 0.18
0.67 ⫾ 0.07a
0.74 ⫾ 0.03a
2.65 ⫾ 0.14c
28.3 ⫾ 5.7
2.01 ⫾ 0.22
4.88 ⫾ 0.40
18
18
18
12
18
10
0.28 ⫾ 0.03a
1.76 ⫾ 0.08a
3.09 ⫾ 0.16a
30.4 ⫾ 7.2
1.9 ⫾ 0.29
5.18 ⫾ 0.27c
0.27 ⫾ 0.05b
1.57 ⫾ 0.01b
2.94 ⫾ 0.14
28.9 ⫾ 5.5
2.53 ⫾ 0.49
6.74 ⫾ 0.54a,d
18
18
18
12
12
6
P ⬍ 0.001 vs. chow-fed wild-type.
P ⬍ 0.001 vs. chow-fed AGT-deficient.
c
P ⬍ 0.01 vs. chow-fed wild-type.
d
P ⬍ 0.005 vs. chow-fed AGT-deficient.
b
High fat diet
Wild-type
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Comparative analysis of the main metabolic blood parameters is shown in Table 3. Compared with WT mice, agt⫺/⫺
mice exhibited a moderate decrease in circulating levels of
cholesterol and triglycerides. Although statistically significant, the increase observed in glucose levels in agt⫺/⫺ mice
was slight and could not be considered physiologically important. In addition, insulin levels were similar. Upon high
fat feeding, some interesting features emerged. First, glucose
levels of agt⫺/⫺ mice remained unchanged, thus abolishing
the slight difference observed between the two genotypes.
Second, triglyceride levels were significantly decreased,
whereas cholesterol levels were increased, in agreement with
the responsiveness to dietary fat and cholesterol reported in
various strains of inbred mice (20). Free T3 was also determined, as this hormone has been long known to be involved
in thermogenesis (21). Table 3 shows similar levels of free T3
in chow-fed WT and agt⫺/⫺ mice. However, upon high fat
feeding, the levels were significantly increased in both genotypes. Interestingly, a significant increase (1.3-fold) was
observed in the level of free T3 in agt⫺/⫺ mice compared with
that in WT mice. The levels of insulin and leptin were similar
in both genotypes when fed a standard chow or a high fat
diet. Moreover, these data are in agreement with the fact that
at 6 wk of age, no statistically significant difference between
the two genotypes was observed with respect to interscapular BAT weight (59.4 ⫾ 2.9 mg for WT mice vs. 61.9 ⫾ 2.2
mg for agt⫺/⫺ mice; n ⫽ 12) and uncoupling protein-1
(UCP-1) mRNA levels (Fig. 2). Northern blot analysis was
also performed for UCP-2 from epididymal fat pads and for
UCP-3 from skeletal muscle. Using -actin mRNA levels as
an internal standard and taking an arbitrary unit of 1 for WT
mice, the values for agt⫺/⫺ mice were, respectively, 1.09 for
UCP-1 (n ⫽ 9; P ⫽ NS), 1.13 for UCP-2 (n ⫽ 6; P ⫽ NS), and
1.24 for UCP-3 (n ⫽ 3; P ⫽ NS).
Endocrinology, December 2001, 142(12):5220 –5225
5224 Endocrinology, December 2001, 142(12):5220 –5225
Massiera et al. • Fat Mass Alterations in Angiotensinogen-Deficient Mice
TABLE 4. Metabolic rate and locomotor activity in 16-wk-old
wild-type and AGT-deficient mice fed a chow diet
0.75
Metabolic rate (watts/kg )
Distance covered (lights on; m)
Distance covered (lights off; m)
Activity (lights on; % of time)
Activity (lights off; % of time)
Wild-type
(n ⫽ 6)
AGT-deficient
(n ⫽ 9)
P
5.1 ⫾ 0.2
29.5 ⫾ 5.5
83.6 ⫾ 5.7
31.6 ⫾ 4.5
55.0 ⫾ 4.5
5.1 ⫾ 0.2
49.8 ⫾ 3.8
110.5 ⫾ 6.8
45.2 ⫾ 3.0
68.6 ⫾ 4.3
NS
⬍0.01
⬍0.02
⬍0.02
NS
Acknowledgments
The authors thank Dr. Marie-France Masseyeff-Elbaz and Mr. JeanJacques René (Institut Arnault Tzanck, St. Laurent du Var, France) for
performing assays of blood parameters. Thanks are also due to Dr. B.
Phillips for careful reading of the manuscript, and to Mrs. Geneviève
Oillaux for skillful secretarial assistance.
Received May 7, 2001. Accepted August 20, 2001.
Address all correspondence and requests for reprints to: Dr. Gérard
Ailhaud, Centre de Biochimie, UMR 6543, Centre National de la
Recherche Scientifique, Université de Nice-Sophia Antipolis, Faculté des
Sciences, Parc Valrose, 06108 Nice Cedex 2, France. E-mail: ailhaud@
unice.fr.
This work was supported by a special grant from Bristol-Myers
Squibb Foundation (to G.A.), Grant 31-57-129.99 from the Swiss National
Science Foundation (to J.S.), a grant from Comité Française de Coordination des Recherches sur l’Athérosclérose et le Cholestérol (to M.T.),
and a fellowship from the French Ministère de la Recherche (to F.M.).
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due to adipocyte hypotrophy in agt⫺/⫺ mice compared with
WT mice. The lower triglyceride content of adipocytes in
agt⫺/⫺ mice was tightly correlated with decreased endogenous lipogenesis through decreased FAS activity. This is in
accordance with a report showing that AngII enhances triglyceride accumulation by stimulating FAS and glycerol-3phosphate dehydrogenase activities (10).
In contrast to WT mice, which gained weight between 12
and 46 wk of age on high fat feeding, the body weight of
agt⫺/⫺ mice remained stable. Under the conditions of augmented exogenous fatty acid supply, the weight of epididymal fat pads remained 1.8-fold higher in WT mice than in
agt⫺/⫺ mice at 6 wk of age, and this increase became more
evident at 16 wk of age. As anticipated, during high fat
feeding, hypertrophy of adipocytes occurred in agt⫺/⫺ mice,
but a large difference persisted, as the weights of adipocytes
of WT mice remained much higher than those of agt⫺/⫺ mice.
The stable body weight of agt⫺/⫺ mice appears at odds with
the increase in adipocyte size observed in the epididymal
depot. However, it cannot be ruled out that the growth of this
depot may not be identical to that of the other adipose depots,
in a way similar to the differential growth of different adipose depots in response to nutritional or environmental stimuli (22).
In search of additional factors that could explain the lower
body weight of agt⫺/⫺ mice compared with WT mice in
response to a chow or a high fat diet, deficient mice show a
decrease in food efficiency, as estimated by the ratio of the
mean 23-h metabolic rate divided by the energy content of
the food eaten by the same animal. This ratio is significantly
(P ⬍ 0.001) lower in agt⫺/⫺ mice than in WT mice. This
finding suggests the occurrence of activation of futile cycles
in the metabolic pathways of agt⫺/⫺ mice. In looking for
possible mechanisms to explain the increased energy dissipation of these mice, our data probably exclude a difference
in intestinal absorption, as both the amount and the fat content of feces were similar. In addition, leptin, which is known
to increase sympathetic activity (23) and energy expenditure
in ob/ob mice (24), is also excluded, as circulating leptin levels
were similar in the two genotypes fed either a standard or a
high fat diet. In agreement with this assumption, the sympathetic pathway did not seem to be altered, as shown indirectly by UCP-1 expression levels in BAT. Free T3 levels
were similar in agt⫺/⫺ mice and WT mice fed a chow diet.
Upon high fat feeding, although both genotypes increased
their levels of free T3 compared with those in chow-fed animals, a significant hyperthyroidism was seen in agt⫺/⫺ mice
compared with WT mice. This additional component may
contribute to lower the metabolic efficiency of AGT-deficient
mice.
Locomotor activity, as expressed by the distance covered,
was clearly and significantly increased in agt⫺/⫺ mice compared with WT mice and may participate to some extent in
the higher energy dissipation (19). Therefore, it is assumed
that the more frequent and longer activity periods of AGTdeficient mice, in addition to decreased lipogenesis, are responsible for the decreased fat deposition. It has been reported in rats that brain AGT participates in a central
regulation of blood pressure (25), and it can be hypothesized
that AngII affects similarly the central pathway(s) leading to
increased locomotor activity. In summary, our results show
that, compared with WT mice, agt⫺/⫺ mice do not gain
weight in response to a high fat diet and exhibit alterations
in WAT development and locomotor activity, supporting the
involvement of AngII in the regulation of body fat mass.
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