In the present study, we exposed to cold b3-adrenoceptor knockout mice b3KO on a C57BL/6J genetic background and measured in white adipose tissue the density of multilocular cells and th
Trang 1b3-Adrenoceptor knockout in C57BL/6J mice depresses
the occurrence of brown adipocytes in white fat
Maria Jimenez1, Giorgio Barbatelli3, Roberta Allevi3, Saverio Cinti3, Josiane Seydoux2,
Jean-Paul Giacobino1, Patrick Muzzin1and Fre´de´ric Preitner1
De´partements de Biochimie Me´dicale1et Physiologie2, Centre Me´dical Universitaire, Gene`ve, Switzerland;
3
Istituto di Morfologia Umana Normale, Universita` di Ancona, Italy
White and brown adipocytes are usually located in distinct
depots; however, in response to cold, brown adipocytes
appear in white fat This response is mediated by
b-adrenoceptors but there is a controversy about the
sub-type(s) involved In the present study, we exposed to cold
b3-adrenoceptor knockout mice (b3KO) on a C57BL/6J
genetic background and measured in white adipose tissue the
density of multilocular cells and the expression of the brown
adipocyte marker uncoupling protein-1 (UCP1) In brown
fat of b3KO mice, UCP1 expression levels were normal at
24C as well as after a 10-day cold exposure Strikingly, under both conditions, in the white fat of b3KO mice the levels of UCP1 mRNA and protein as well as the density of multilocular cells were decreased These results indicate that
b3-adrenoceptors play a major role in the appearance of brown adipocytes in white fat and suggest that the brown adipocytes present in white fat differ from those in brown fat Keywords: adipose tissue; cold exposure; differentiation; knockout; uncoupling protein 1
Brown adipose tissue (BAT) and white adipose tissue
(WAT) both play an important role in the control of energy
balance in mammals Indeed, BAT is involved in the control
of body temperature and body weight via nonshivering and
diet-induced thermogenesis [1,2] whereas the primary
func-tion of WAT is to store and release energy BAT funcfunc-tion is
particularly important for thermoregulation in small
mam-mals in which the surface to volume ratio is unfavorable [1]
Moreover, activation of BAT prevents obesity induced by
overfeeding in rodents [2]
Cold exposure, via activation of the sympathetic nervous
system, induces heat production in BAT [3,4] and lipolysis
in WAT [5] The effects of norepinephrine released at the
nerve endings are mediated in brown and white adipocytes
mainly by the b3-adrenoceptor subtype [6]
Brown and white adipocytes are usually located in
distinct depots [4] and can be distinguished morphologically
[7]: brown adipocytes contain multiple (multilocular) lipid
droplets and numerous mitochondria whereas white
adipo-cytes display a main (unilocular) lipid droplet and few
mitochondria Nevertheless, the morphological aspect
alone is not sufficient to distinguish between these two
cell types [8] In fact, the only criterion is the specific
expression in mitochondria of brown adipocytes, of the
uncoupling protein-1 (UCP1), which uncouples the oxidative
phosphorylations from electron transport, thus dissipating energy as heat [9]
An observation made initially by Young et al [10] was the presence of multilocular mitochondria-rich brown adipocytes in depots previously defined as typical WAT The emergence of these ectopic cells was found to be induced by cold acclimation in WAT of rats [11,12] and of mice [13,14] The new cells were found to be sympathetically innervated [15] and to remain present as long as a sympathetic nervous system stimulation persisted [16] Several reports also showed that the administration of selective b3-adrenoceptor agonists like CL 316, 243 induced the emergence of brown adipocytes in WAT depots [14,17–19]; and that this phenomenon was strongly depend-ent on the genetic background [14,18,20] It was, however, recently shown that a transgenic over-expression of a human b1-adrenoceptor in WAT of mice also induced the appearance of abundant brown adipocytes in this tissue [21] These results suggested that the b3-adrenoceptor might not
be the only b-subtype controlling the emergence of brown adipocytes in WAT
Another open question is the origin and the real nature of the multilocular cells expressing UCP1 that appeared in WAT upon cold acclimation or b3-adrenoceptor stimula-tion Himms–Hagen et al [8], studying the effect of
CL 316, 243 in rats, suggested that the multilocular cells expressing UCP1 that appeared in WAT were different from the classical brown adipocytes and postulated that these cells might derive, at least in part, from preexisting unilocular adipocytes
Mice with a targeted disruption of their b3-adrenoceptor (b3KO) have been generated [22,23] Our b3KO model, first generated on a mixed background (C57BL/6J· 129Sv/ev) [23], was then backcrossed to obtain the mutation on the C57BL/6J genetic background In the present work, we studied the effect of b-adrenoceptor-deficiency on the
Correspondence to M Jimenez, De´pt de Biochimie Me´dicale Centre
Me´dical Universitaire 1, rue Michel-Servet CH-1211 Gene`ve 4,
Switzerland Fax: + 41 22 702 55 02, Tel.: + 41 22 702 54 88,
E-mail: Maria.Jimenez@medecine.unige.ch
Abbreviations: b 3 KO, b 3 -adrenoceptor knockout; UCP1, uncoupling
protein-1; BAT, brown adipose tissue; WAT, white adipose tisue.
(Received 23 August 2002, revised 26 November 2002,
accepted 10 December 2002)
Trang 2occurrence of brown adipocytes in WAT of C57BL/6J mice
at 24C and after cold exposure We therefore measured (a)
the density of multilocular cells in WAT, and (b) the levels
of UCP1 mRNA and protein in WAT and BAT, as
compared to wild-type (WT) controls
Materials and methods
Materials
All organic and inorganic chemicals of analytical or
molecular biology grade were purchased from Merck
(Darmstadt, Germany), Boehringer Mannheim
(Mann-heim, Germany), Sigma Chemical Co (St Louis, MI) and
Fluka (Bucks, Switzerland) Trizol and the RNA ladder
were purchased form Gibco BRL (New York, NY),
Electran Nylon membranes from BDH Laboratory
Sup-plies (Poole, UK), Quikhyb from Stratagene Inc (La Jolla,
CA), Hyperfilm ECL films and [a-32P]dCTP from
Amer-sham International (Bucks, UK), cytochrome oxidase
subunit IV from Molecular probes (Eugene, OR) Rabbit
antisheep IgG, avidin and biotin complex (ABC) peroxidase
conjugated were purchased from Vector Laboratories
Burlingame (CA)
Mice
b3KO mice were initially generated on a mixed 129Sv/
ev· C57BL/6J background Then, the b3KO mice were
backcrossed six times with C57BL/6J (C57) mice (obtained
from BRL Ltd, Fu¨llinsdorf, Switzerland), in order to get
mutated mice with the purified C57 genetic background
(98.4% homogeneity) Mice used in this study were
offsprings of WT and b3KO founders of the C57 purified
genetic background The genotypes were determined by
Southern blots as previously described [23] Three- to
four-month-old female C57 WT or b3KO mice with free access to
water and standard laboratory chow diet (Nordos, Cergy,
France) were used The average weights of the mice were
22.5 ± 0.3 and 22.8 ± 0.3 for the C57 WT and b3KO,
respectively They were housed individually for 10 days at
24 ± 1C on a 12-h light : 12-h dark cycle (7:00–19:00 h)
before the beginning of each experiment They were then
divided into three groups, one maintained at 24C and the
other two exposed to cold (6C) for either two or 10 days
The fat pads along the uterus and fallopian tube
(param-etrial fat) and those around the ovaries (periovarian) were
dissected and pooled They are referred to in the text as
genital WAT
Morphometry and immunochemistry
Mice were killed by CO2 inhalation and perfused
intra-cardially with 4% paraformaldehyde in 0.1M phosphate
buffer, pH 7.4 Genital WAT was dissected, further fixed
by immersion in the 4% paraformaldehyde solution
overnight at 4C and then embedded in paraffin blocks
In each animal, two sections, at a distance of 200 lm from
each other in the central part of the parametrial or
periovarian depots were studied The adipocytes were
identified as unilocular (a unique prevalent vacuole) or
multilocular (more than five small lipid droplets) and the
respective number of these cells in each section counted The average number of adipocytes counted per section and per animal were 8173 and 10774, respectively, in a total of 237036 cells In some experiments, the mean area
of 100 unilocular adipocytes randomly chosen on the total surface of a section was measured using a Nikon Eclipse E800 connected with a digital Camera DXM1200 at a magnification of 20· The surface area of the cells was measured using the Nikon LUCIA IMAGE Version 4.61 program
Three-micrometer sections of tissue were deparaffinizated and rehydrated with distilled water and immunohistochem-ical detection of UCP1 was performed as follows: (a) 0.3% hydrogen peroxide in methanol for 30 min to inactivate endogenous peroxidase; (b) normal rabbit serum 1 : 75 in NaCl/Pi for 20 min to reduce non-specific background staining; (c) polyclonal sheep antirat UCP1 diluted 1 : 5000
in NaCl/Pi, at 4C, overnight; (d) NaCl/Pi, twice for
15 min; (e) biotinylated secondary antibody, rabbit anti-sheep IgG 1 : 200, 30 min at room temperature; (f) NaCl/Pi, twice for 15 min; (g) ABC complex for 1 h; (h) NaCl/Pi twice for 15 min; (i) enzymatic development of peroxidase with 0.05% diaminobenzidine hydrochloride and 0.02% hydrogen peroxide in 0.05MTris/HCl pH 7.6, for 4 min Negative controls were performed by substituting the primary antibody by sheep IgG No cross-reaction with UCP2 and UCP3 were observed in tissues expressing the highest levels of UCP2 (liver) and UCP3 (skeletal muscle) The specificity of the UCP1 antibody used in this study has been recently confirmed [24]
Northern blots The mice were killed by cervical dislocation followed by decapitation and the interscapular BAT as well as genital WAT were dissected and quickly frozen in liquid nitrogen Total RNA was isolated using the Trizol technique and
15 lg of RNA were electrophoresed in a 1% agarose gel containing formaldehyde and transferred according to standard protocols The probes used were a full-length mouse UCP1 cDNA (GenBank accession number U63418),
a b1-adrenoceptor PCR probe (position 297–1053, GenBank accession number L10084) and a PGC-1 PCR probe (posi-tion 297–2308; GenBank accession number AF049330) They were labelled by random priming with [a-32P]dCTP to
a specific activity of approximately 1· 109d.p.m.Ælg)1 DNA Hybridizations were performed using QuikhybTM solution as previously described [25] Blots were exposed at )80 C to Hyperfilm ECL films RNA levels were quanti-fied by scanning photodensitometry of the autoradiograms using IMAGEQUANT Software version 3.3 of Molecular Dynamics (Sunnyvale, CA) Subsequent hybridization of the blots with a [a-32P]ATP labelled synthetic oligonucleo-tide specific for the 18S rRNA subunit was used to correct for the differences in the amounts of RNA loaded onto the gels
Isolation of mitochondria Other groups of mice were used for Western blot experi-ments BAT and WAT mitochondria were prepared as previously described [26] and the pellet containing heavy
Trang 3and light mitochondria resuspended in 400 lL distilled
water Mitochondrial protein concentrations were
deter-mined according to Bradford et al [27] using the Bio-Rad
Protein Assay, with bovine serum albumin as a standard
Isolated mitochondria were stored at )20 C as 15 lg
mitochondrial protein aliquots
Western blots
Fifteen micrograms of purified BAT and WAT
mitochon-dria were dried under vacuum and resuspended in 10 lL of
a loading buffer containing 50% glycerol, 5% SDS, 2.5%
bromophenol blue and 0.5MTris/HCl pH 6.8 The samples
were electrophoresed on a 12% polyacrylamide, 0.1% SDS
gel, and transferred to a poly(vinylidene difluoride)
mem-brane using the standard dry transfer method with a buffer
containing 10% methanol, 25 mM Tris/HCIpH 6.8 and
190 mMglycine The membrane was blocked with a NaCl/
Pibuffer containing 0.1% Tween and 2% nonfat dry milk
This same buffer was used for all subsequent hybridizations
UCP1 protein was detected using a sheep polyclonal
primary antibody raised against rat UCP1 protein
(gener-ously provided by D Ricquier, Meudon, France) at a
concentration of 0.5 lgÆmL)1and a 1 : 4000 diluted sheep
monoclonal anti-mouse peroxidase-labelled secondary
antibody The cytochrome oxidase protein was detected as
above using a 1 : 1000 diluted monoclonal antibody specific
for cytochrome oxidase subunit IV, and a 1 : 1000 diluted
goat anti-mouse peroxidase-labelled secondary antibody
The signals were detected by chemiluminescence using a
standard ECL kit, and developed on a Hyperfilm ECL
They were quantified by scanning photodensitometry
Cytochromec oxidase activity
BAT homogenate was prepared as previously described [26]
and 10 lg of homogenate proteins were used to measure the
cytochrome c oxidase activity by the method of Yonetani
and Ray [28]
Statistics
Data are presented as mean ± SE and were analyzed using
a two-factor analysis of variance (ANOVA< 0.05) for the
main effects of group (b3KO vs WT) and treatment
condition (24C vs 6 C 2 or 10 days) as well as for the
group· treatment condition interaction effect, using the
computer softwareSTATISTIX, version 4.0 (Analytical
Soft-ware, St Paul, MN)
The non parametric Kruskal–Wallis test was used to
analyze morphometric studies
Results
Morphometry and immunohistochemistry in the WAT
The density of multilocular cells was measured in genital
WAT sections As shown in Fig 1, in the WAT of C57 WT
mice maintained at 24C, only 1.1% of the total adipocytes
was multilocular The effects of the lack of b3-adrenoceptor
were striking In mice kept at 24C, the density of
multilocular cells was 36-fold (P < 0.01) lower in the
WAT of b3KO mice as compared to WT mice Ten days of cold-exposure increased the density of multilocular cells in both WT and b3KO mouse WAT (11-fold, P < 0.03 and ninefold, P < 0.01, respectively) However, the density of multilocular cells was 42-fold lower (P < 0.03) in the WAT
of b3KO as compared to WT mice UCP1 protein expression in multilocular cells was assessed by immuno-histochemistry No UCP1 could be detected in the WAT multilocular cells of WT or b3KO mice either maintained at
24C or exposed to 6 C for 2 days A 10-day cold exposure, however, induced the emergence of UCP1 expression in WAT multilocular cells Figure 2A and B show multilocular cells and UCP1 positive multilocular cells
in WT and b3KO mouse parametrial depots Note that the intensity of the UCP1 expression in multilocular cells varies from cell to cell In b3KO mouse, UCP1-positive multilo-cular cells were heterogenously distributed in the WAT depot and it was therefore not possible to assess the number
of these cells and to compare it to that in WT mouse WAT Genital pads of b3KO mice were 1.5-fold heavier than those of WT (388 ± 33 vs 251 ± 14, P < 0.005, n¼ 12) despite a normal number of cells, as assessed by DNA content (1142 ± 93 vs 1098 ± 69 lg per genital WAT,
n¼ 12), suggesting an increase in cell size Consistent with this hypothesis, a morphometric study showed an increase
of 2.4-fold of the unilocular adipocyte mean area in the genital WAT of b3KO as compared to WT mice (1594 ± 42 vs 656 ± 49 mm2, n¼ 4 and 3 sections, respectively)
UCP1 mRNA expression in WAT and BAT Quantitative measurements were then performed on whole BAT and WAT The expression of UCP1 mRNA was measured in genital WAT, and in interscapular BAT of C57
WT and b3KO mice (Fig 3) In genital WAT of WT mice (Fig 3A), the basal level of UCP1 mRNA at 24C, measured by Northern blot, was found to be increased
Fig 1 Morphometric analysis of genital WAT from C57BL/6J wild-type (WT) or b 3 KO female mice, maintained at 24 °C or exposed to
6 °C for 2 or 10 days (2 d and 10 d, respectively) The number of multilocular (ML) cells is expressed as a percentage of the total amount
of adipocytes in the pooled sections of the two WAT depots n ¼ 3–4.
*P < 0.05 and **P < 0.01 vs respective WT; #P < 0.05 and
## P < 0.01 vs respective control at 24 C.
Trang 42.6-fold (P < 0.05) and 4.8-fold (P < 0.001) after 2- and
10-day cold exposure, respectively The effects of a lack of
b3-adrenoceptor were striking The UCP1 mRNA
expres-sion levels in WAT of b3KO mice were strongly depressed
by 20-fold (P < 0.01), 33-fold (P < 0.005) and 2.9-fold
(P < 0.005) as compared to WT mice kept at 24C or
exposed to 6C for 2 or 10 days, respectively An effect of
cold exposure on UCP1 mRNA expression was also seen in
WAT of b3KO mice but only after 10 days (P < 0.005)
In BAT of C57 mice (Fig 3B), 2- or 10-day cold exposure
increased UCP1 mRNA expression 1.8-fold (P < 0.05
and P < 0.005, respectively) in the WT, and 1.6-fold
(P < 0.05) and 1.9-fold (P < 0.01) in the b3KO mice It is
noteworthy that the b3-adrenoceptor deficiency did not
affect UCP1 mRNA expression under all conditions
studied
In WT C57 mice, the expression level of UCP1 mRNA in
WAT relative to that in BAT, assessed on a same Northern
blot, was approximately 3% in mice kept at 24C and 11% after a 10-day cold exposure (data not shown)
UCP1 protein expression in WAT and BAT The level of UCP1 protein was measured in isolated mitochondria from genital WAT and BAT of C57 WT and b3KO mice The results represent the amount of UCP1 in a given amount of mitochondrial proteins (specific value) The mitochondrial marker cytochrome c oxidase was chosen as a standard to normalize UCP1 protein level
As illustrated in Fig 4, whereas a 2-day cold-exposure did not increase UCP1 protein level in WAT mitochondria
of WT mice, 10-day cold-exposure increased it 6.7-fold (P < 0.02) I n WAT, the effects of b3KO on UCP1 protein levels were comparable with those observed on UCP1 mRNA Indeed, the UCP1 protein levels in bKO mice kept
Fig 2 Genital WAT of C57BL/6J wild-type (WT) (A) and b 3 KO (B)
mice exposed to 6 °C for 10 days Note the presence of multilocular
cells and of UCP1 positive multilocular cells Bar, 31.74 lm The mean
area of the adipocytes in these sections was 2.4-fold higher in b 3 KO
than in wild-type mice (P < 0.05).
Fig 3 Northern blot quantification of UCP1 mRNA expression in adipose tissues of C57BL/6J wild-type (WT) or b 3 KO female mice, maintained at 24 °C or exposed to 6 °C for 2 or 10 days (2 d and 10 d, respectively) (A), Genital WAT; (B), brown adipose tissue (BAT) The results are expressed as means ± SE of arbitrary values normalized using the corresponding 18S mRNA values The ratio of the control values is considered as 1.0 n ¼ 6 **P < 0.01 and ***P < 0.005
vs respective WT;#P < 0.05 and###P < 0.005 vs respective control
at 24 C.
Trang 5at 24C or exposed to 6 C for 10-days were by 7.7-fold
(P < 0.005) and 11-fold (P < 0.01) lower than those in
WT animals
It is interesting to note the parallel effects of the b3KO on
the percentage of multilocular cells, and on the degree of
UCP1 expression at the mRNA and protein levels This
suggests that UCP1 is mainly expressed in multilocular cells
In BAT of C57 mice, 10-day cold-exposure increased
UCP1 protein level 1.9-fold (P < 0.05) in WT mice and
2.8-fold (P < 0.05) in b3KO mice In agreement with
the UCP1 mRNA results, b3KO did not affect the
UCP1 protein level under all conditions studied (data not
shown) Furthermore, cytochrome c oxidase total activity
(as expressed per BAT) showed no difference between
wild-type and b3KO C57 mice kept at 6C (170 ± 1 vs
140 ± 5 lUÆmg)1) This observation suggests that the total
amount of mitochondria per BAT was not modified by the
b3KO
The expression level of UCP1 protein in WAT relative to
that in BAT, assessed on a same Western blot, was
approximately 5% in mice kept at 24C and 11% after a
10-day cold exposure (data not shown)
b1-adrenoceptor mRNA expression in WAT of C57
The possibility of a compensation by the b1-adrenoceptor in
the b3KO mice was tested This receptor is the second
most abundant b subtype in WAT [29] The b1
-adreno-ceptor mRNA was not significantly affected by the b3KO in
either the BAT or the WAT of C57 (100 ± 15 vs 78 ± 8;
100 ± 9 vs 88 ± 7)
PGC-1 mRNA expression in WAT and BAT of C57
PGC-1 is a transcriptional coactivator induced by
cold-exposure via the b-adrenergic signaling pathway in BAT
and in skeletal muscle [30] PGC-1 stimulates the
transcrip-tional activity of PPARc, of the thyroid hormone receptors and of UCP1 in BAT The expression of PGC-1 mRNA was measured in interscapular BAT and in genital WAT, using the same Northern blots as for UCP1 expression measurements (Fig 5) In WT mice, PGC-1 was expressed
in WAT but approximately 100-fold less than in BAT (data not shown) Moreover, a 2-day cold exposure increased PGC-1 mRNA expression in WAT by 1.8-fold (P < 0.001) Strikingly, the b3KO WAT exhibited normal PGC-1 mRNA levels in both conditions
Discussion Morphometric analyzes showed that the number of multi-locular cells present in WAT was strongly depressed in the
b3KO mice maintained at 24C as compared to WT mice The absence of UCP1 expression in multilocular cells under this condition might represent a cellular phenotype inter-mediary between white and brown adipocytes or might be due to the heterogeneity of the content of multilocular cells
in different regions of a given WAT depot [11] Ten days of cold-exposure increased the number of multilocular cells and induced the expression of UCP1 in some of them in both WT and b3KO mice
Mice maintained singly housed at 24C are under a mild cold stimulus It is interesting to note in the WAT of C57 mice at 24C a significant level of expression of UCP1 mRNA reaching 3% of that observed in BAT Our study shows for the first time that the lack of b3-adrenoceptor strongly depresses UCP1 mRNA and protein expression in WAT I n mice kept at 24C the UCP1 mRNA and protein levels in WAT are strongly depressed by the b3KO Ten days of cold-exposure increased UCP1 mRNA and protein
in WAT of both WT and b3KO mice but the differences between the two genotypes was conserved
I n BAT of C57 mice, the lack of b3-adrenoceptor did not affect the basal or cold-induced expression of UCP1 mRNA and protein The b3-adrenoceptor independency
of UCP1 expression in BAT suggests that in brown
Fig 4 Western blot quantification of UCP1 protein expression in
gen-ital WAT mitochondria of C57BL/6J wild-type (WT) or b 3 KO female
mice, maintained at 24 °C or exposed to 6 °C for 2 or 10 days (2d and
10d, respectively) The results are expressed as means ± SE of
arbi-trary values normalized using the corresponding cytochrome c oxidase
values The ratio of the control value is considered to be 1.0 n ¼ 6.
**P < 0.01 and ***P < 0.005 vs respective WT; ## P < 0.01 vs.
respective control at 24 C.
Fig 5 Northern blot quantification of PGC-1 mRNA expression in genital WAT of C57BL/6J wild-type (WT) or b 3 KO female mice, maintained at 24 °C or exposed to 6 °C for 2 (2d) (A), Genital WAT The results are expressed as means ± SE of arbitrary values nor-malized using the corresponding 18S mRNA values The ratio of the control values is considered as 1.0 n ¼ 6 # P < 0.05 and
###
P < 0.005 vs respective control at 24 C.
Trang 6adipocytes, one or both of the other b-adrenoceptor
subtypes are able to substitute and confirms previous
findings in which we demonstrated that the lack of b3
-adrenoceptor did not affect BAT metabolism [23,31] Our
study shows that there is no compensation at the level of
the b1-adrenoceptor mRNA in BAT or WAT Our in vivo
studies are in contrast with in vitro experiments performed
in cultivated adipocytes of male mice suggesting that
UCP1 mRNA expression is under strict control of the b3
-adrenoceptor [32,33] A compensation by a adrenoceptors
cannot be excluded Nevertheless, we and others, recently
showed that, at least in BAT, a total lack of b-adrenergic
signaling is not compensated by other adrenergic
sub-types Indeed, mice lacking the whole b-adrenoceptor
family are dramatically cold intolerant with an abnormal
BAT morphology [34,35]
The PPARc-1 coactivator, PGC-1, is induced by cold via
b-adrenergic stimulation It has been shown to stimulate
brown adipocyte differentiation including UCP1 expression
and was undetectable in WAT [30] In our study, PGC-1
was expressed in genital WAT of WT mice, although at a
much lower level than in BAT, and its expression was
increased by cold exposure In WT WAT, PGC-1
expres-sion varied in parallel with UCP1 expresexpres-sion, consistent
with the reported control of UCP1 by PGC-1 in brown
adipocytes Strikingly, in b3KO WAT where UCP1
expres-sion was depressed, PGC-1 levels were normal This
suggests that b3-adrenoceptors are not essential for a
normal expression of PGC-1 b3-adrenergic signaling might
stimulate UCP1 expression in WAT by modulating a target
molecule downstream PGC-1 This hypothesis deserves
further studies
Together, our results suggest that the brown
adipocyte-like cells present in WAT and the brown adipocytes
constituting BAT are subjected to different control systems
The hypothesis of a different nature of BAT and WAT
multilocular cells has already been proposed by
Himms-Hagen et al [8], who suggested that most of the multilocular
cells appearing in WAT upon b3-adrenoceptor agonist
stimulation derived from preexisting convertible unilocular
white adipocytes
The results of the present study show that the number of
multilocular cells and the expression of UCP1 in WAT are
under the selective control of the b3-adrenoceptor This
notion should be useful to develop new drugs, which might
stimulate the recruitment of UCP1 expressing cells in
human WAT and transform the latter into an energy
dissipating tissue
Acknowledgements
We are greatly indebted to Daniel Ricquier for the generous gift of the
anti-UCP1 Ig, and to Philippe Vallet for his technical help This work
was supported by the Swiss National Science Foundation grant n31–
54306 98, by Ricerca Scientifica d’Ateneo 2001 and by EU grant: ERB
CHRX CT94-0490 to S C and Jules Thorn Charitable Overseas Trust.
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