Annals of Medicine, 2014; Early Online: 1–9
© 2014 Informa UK, Ltd.
ISSN 0785-3890 print/ISSN 1365-2060 online
DOI: 10.3109/07853890.2014.952328
SPECIAL SELECTION: BROWN FAT
Thermogenic brown and beige/brite adipogenesis in humans
Rubén Cereijo, Marta Giralt & Francesc Villarroya
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Departament de Bioquímica i Biologia Molecular, Institute of Biomedicine (IBUB), University of Barcelona, and CIBER Fisiopatología de la
Obesidad y Nutrición, Barcelona, Catalonia, Spain
Evidence from rodents established an important role of brown
adipose tissue (BAT) in energy expenditure. Moreover, to sustain
thermogenesis, BAT has been shown to be a powerful sink for
draining and oxidation of glucose and triglycerides from blood.
The potential of BAT activity in protection against obesity and
metabolic syndrome is recognized. Recently, an unexpected
presence and activity of BAT has been found in adult humans.
Here we review the most recent research in this field and, specifically, how new findings apply to humans. Moreover, we seek to
clarify the underlying biological processes occurring beyond the
burst of new nomenclature in the field. The cell type responsible
for thermogenesis, the brown adipocyte, arises from complex developmental processes. In addition to ‘classical’ brown adipocytes,
present in developmentally programmed BAT depots, there are
brown adipocytes, named ‘brite’ (from ‘brown-in-white’) or ‘beige’,
which appear in response to thermogenic stimuli in white fat
due to the so-called ‘browning’ process. Beige/brite cells appear
to be important components of BAT depots in adult humans. In
addition to the known control of BAT activity by the sympathetic
nervous system, metabolic and hormonal signals originating in
muscle or liver (e.g. irisin, FGF21) are recognized as activators of
BAT and beige/brite adipocytes.
Key words: Antiobesity agents, brown adipose tissue,
metabolic syndrome X, obesity, white adipose tissue
Introduction: the shifting history of human brown
adipose tissue
There is a long history of evidence for the existence of brown
adipose tissue (BAT) in humans. Classical anatomists and
histologists reported the presence of masses of brown adipose
tissue containing the typical multilocular adipocyte morphology in several anatomical sites in humans. The prevalence of
BAT depots in human neonates and the progressive decrease in
the amount of BAT in adults were commonly recognized, and it
was assumed that BAT progressively involutes with age. Given
the recognized role of BAT as a thermogenic tissue, it was also
assumed that BAT function in humans was probably restricted
to the neonatal period, during which it would provide the heat
needed to cope with the thermal stress associated with birth (1).
Key messages
• Thermogenic, energy-dissipating, beige/brite adipocytes
are a subtype of brown adipocytes with a cell lineage
origin distinct from ‘classical’ brown adipocytes.
• Human adult adipose tissue possesses a remarkable
plasticity and may contain thermogenic, energydissipating, beige/brite adipocytes.
• The energy-dissipating and metabolite oxidation
properties of beige/brite adipocytes (like ‘classical’
brown adipocytes) make them attractive candidates to
be stimulated in order to prevent obesity and ameliorate
hyperglycemia and hyperlipidemia.
However, studies in rodents reported by Rothwell and Stock in
the late 1970s proposed that BAT was not only a site of coldinduced thermogenesis but was also a main site of diet-induced
thermogenesis (2), reigniting interest in the role of human BAT
owing to its obvious potential to promote energy expenditure
and protect against obesity. The subsequent discovery of UCP1
(uncoupling protein-1) in rodents as the key protein that conferred specific thermogenic properties to BAT mitochondria
was soon followed by the identification and characterization of
human UCP1, confirming that the molecular mechanisms necessary for BAT-associated thermogenesis were present not only
in rodents but also in humans (3). However, skepticism about
the physiological relevance of BAT in humans prevailed (4). Nevertheless, some lines of evidence continued to provide support
for the physiological role of BAT, including genetic association
studies that related polymorphisms of the UCP1 gene with altered energy balance and metabolism in adult human patients. A
single-base polymorphism (A-3826G) in the 5’ regulatory region
of the UCP1 gene is relatively frequent in the human population.
Multiple studies in the late 1990s reported significant associations of this polymorphic variant with metabolic abnormalities
in distinct adult human populations. After reviewing 12 independent reports, Gonzalez-Barroso et al. (5) concluded that the
A-3826G polymorphism is not a major contributor to obesity
development, but it is related to an increased propensity toward
Correspondence: Francesc Villarroya, Departament de Bioquimica i Biologia Molecular, Facultat de Biologia, Universitat de Barcelona, Av. Diagonal 643,
08028-Barcelona, Catalonia, Spain. E-mail: fvillarroya@ub.edu
(Received 6 June 2014; accepted 4 August 2014)
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R. Cereijo et al.
weight gain over time, especially in persons with a higher risk
of obesity. Since that time, more than 20 additional studies of
distinct adult human populations have confirmed the association
of this UCP1 gene polymorphism with metabolic and energy balance alterations in human adults. Of course, these genetic data
could be interpreted as evidence for a biological role of UCP1
activity in neonatal BAT that has consequences in adulthood,
but the possibility remained that UCP1-related BAT activity was
physiologically relevant in adults.
The concepts governing thinking on the subject of BAT in
human adults have recently undergone a radical shift, propelled
initially by the serendipitous finding of metabolically active
‘adipose tissue’ in patients suspected of cancer—a finding made
possible by the introduction of [18F]-fluorodeoxyglucose-based
positron emission tomography (PET) assays for diagnostic
purposes. Active metabolic sites revealed by PET, unrelated to
tumors, were found mainly in neck and shoulder areas of adults
(6). Three independent studies in 2009 demonstrated that these
sites corresponded to metabolically active BAT, leading to the
current concept that the majority of human adults possess
metabolically active BAT (7–9). Notably, recent studies that have
assessed the association of UCP1 -3826A/G and PET-determined
BAT activity established an association between this UCP1 gene
polymorphism and the extent of age-related decreases in BAT
activity (10). In recent years, multiple studies have addressed
studies on BAT activity in distinct human populations, confirming several conclusions: BAT activity is higher in women than
men; BAT activity progressively decreases with age; BAT activity
is induced by cold exposure; and BAT activity is reduced in obese
patients. Recent reviews provide updated information on the PET
scan-based assessment of BAT activity in humans (11,12).
Beige/brite: nomenclature, tissues,
and cellular identities
The story of the recent rediscovery of BAT in adult humans has
coincided with important advances in our understanding of the
cell biology of the brown adipocyte and a growing awareness of its
unexpectedly high degree of complexity.
In rodents, classical anatomical brown fat depots (i.e. interscapular) are present throughout life. BAT develops during fetal
life, whereas white adipose tissue (WAT) grows mainly after birth.
However, this apparent transformation of adipose depots that occurs in mice after birth, from predominantly brown to predominantly white, does not rely on the conversion of mature brown
adipocytes to white adipocytes, but is instead based primarily on
the differentiation of white adipocytes from precursor cells (13).
During embryonic development, adipocytes develop from
mesenchyme of mesodermal origin, with the exception of the
mesenchyme in the cephalic region, which has an ectodermal origin (14). Numerous studies have sought to establish the identity
of adipocyte precursor cells (15). It is now well established that
brown and white adipocytes have distinct developmental origins.
At least two distinct adipocyte lineages have been defined based
on expression of the myogenic lineage marker Myf5 (myogenic
factor 5). Brown adipocytes present in BAT depots originate from
Myf5-expressing precursors, which are also the source of skeletal
myogenic cells and a subpopulation of white adipocytes (16–18).
In contrast, most white adipocytes are derived from Myf5-negative
precursors; this also appears to be the case for brown adipocytes
that develop in WAT as a consequence of the ‘browning’ process
(16,19,20).
The so-called browning of WAT is defined as the appearance of
functional brown adipocytes—also named ‘brite’ (‘brown-in-white’)
or ‘beige’ (see below)—in WAT depots after thermogenic stimuli.
In rodents, it is now well established that this process, together
with the ‘recruitment’ of existing BAT depots, is required for
optimal adaptive energy expenditure (21). As discussed below,
cold exposure and adrenergic signaling are the main inducers of
these processes, although other factors, shared to varying degrees,
might also contribute. Increased browning of WAT is also often
found in genetically engineered rodent models resulting in suppressed ‘classical’ BAT activity (22), possibly as a compensatory
process.
Adipocyte cells were formerly classified into two types: unilocular, fat-storing, white adipocytes; and multilocular, thermogenic brown adipocytes. The current classification scheme
now includes a third category of adipocytes, the so-called beige or
brite adipocytes, also sometimes referred to as ‘inducible’ brown
adipocyte-like cells (23–25). These cells have phenotypic and
functional characteristics of brown adipocytes, but are found in
anatomical white adipose depots. In contrast, brown adipocytes
located in defined anatomical BAT depots are often referred to as
‘classical’, ‘constitutive’, or ‘developmentally programmed’ brown
adipocytes. This terminology is clearly applicable in rodents, but,
as described below, its application to adipocyte cell biology in
humans is not consolidated yet.
There is genetic evidence that the capacity to induce the appearance of beige/brite adipocytes in WAT depots is highly
relevant for protection against obesity in rodents. Accordingly,
differences in beige/brite adipocyte abundance in WAT reported
between mice strains correlate positively with their resistance to
induce obesity by diet (26). There are also significant differences
in the number of these beige/brite cells between WAT depots;
they are most abundant in subcutaneous inguinal WAT and least
abundant in visceral perigonadal WAT (27).
Recent lineage-tracing studies using transgenic mice indicate that beige/brite adipocytes arise from a cell lineage (Myf5negative) different from that leading to brown adipocytes
(Myf5-positive) in anatomically defined BAT depots (16).
However, research to clarify the precise origin of distinct cells
found in WAT—genuine white adipocytes and beige/brite adipocytes—remains ongoing. In fact, whether beige/brite adipocytes
derive from pre-existing white adipocytes through a process of
transdifferentiation or through de novo adipogenesis from a
specific subgroup of precursor cells remains a matter of debate
(28,29). These processes are not mutually exclusive, and it is possible that their relative importance in different WAT depots may
vary. Indeed, it has been proposed that β3-adrenergic activation
induces browning in epididymal WAT through proliferation and
further differentiation of precursors, whereas in inguinal WAT it
acts through white-to-brown transdifferentiation (19).
Support for the white-to-brown adipocyte transdifferentiation process was first proposed based on the observed lack of
induced cellular proliferation and the presence of morphological intermediate forms of adipocytes (‘paucilocular’) in the
transition from large unilocular white adipocytes to UCP1positive multilocular beige/brite adipocytes in response to cold
exposure or β3-adrenergic receptor stimulation (30–32). Recent
lineage-tracing experiments in adult mice directly demonstrated that mature white adipocytes in inguinal WAT have the
potential to convert to beige/brite adipocytes and, furthermore,
that this is a reversible process that depends on environmental
temperature (33). However, another recent study in mice using
a pulse-chase fate-mapping technique to mark mature white
adipocytes reached the conclusion that most beige/brite adipocytes arise from precursor cells during browning in inguinal
WAT (34).
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Brown and beige/brite adipogenesis in humans 3
Another question is whether there are specific precursor cell
types for beige/brite adipocytes distinct from those for white
adipocytes in WAT. As discussed below, gene-profiling analyses
of primary WAT preadipocytes differentiated in vitro have suggested two types of preadipocytes (25). In contrast, in vivo mouse
studies have been less conclusive, due in part to the difficulty of
defining a true white adipocyte specific marker, or even unique
gene markers for tracing WAT adipocyte developmental origins.
So, the actual identity of beige/brite precursor cells (if they exist)
is poorly known, in addition to the fact that they are expected
to be Myf5-negative. Interestingly, Spiegelman and colleagues
have recently identified a smooth muscle-like origin for at least a
subset of beige/brite cells (35).
In any case, it should be kept in mind that a difference in the
developmental origin of adipocytes does not necessarily imply a
functional difference. How important is the activity of the new
beige/brite cells that appear as a consequence of browning of
WAT relative to the activity of the classical BAT depots in rodents? First data indicated that cultured cells representative of
the ‘classical’ brown and beige/brite lineages have similar rates of
basal and uncoupled respiration (25). Moreover, a recent study
that directly determined the uncoupling activity and metaboliteoxidizing capacity in mitochondria from beige/brite adipocytes
compared with ‘classical’ brown adipocytes concluded that the
two types of adipocytes showed very similar behavior (36). Only
lower capacity of utilization of glycerol-3-phosphate in beige/
brite mitochondria relative to ‘classical’ brown mitochondria was
reported. Theoretical calculations have suggested that the total
amount of beige/brite cells cannot account for a large portion
of energy expenditure relative to the capacity elicited by classical BAT depots (36). However, studies using different strains of
mice characterized by distinct WAT browning capacities but a
similar capacity to activate classical BAT have shown that browning capacity is highly correlated with protection against highfat-diet-induced obesity (37). In fact, a tendency toward lower
thermogenic capacity was reported in mitochondria obtained
from an obesity-prone mouse strain (C57Bl/6) compared with
those from an obesity-resistant mouse strain (129Sv) (36). Further
studies will be needed to reconcile the minor bioenergetic impact
on the organism expected from browned WAT, determined on
the basis of biochemical considerations, with the importance of
the process evidenced by the genetic studies described above.
Moreover, a potential endocrine role of BAT beyond its intrinsic
thermogenic activity has been recently proposed (38); to date,
there are no data indicating important differences in the bioactive
factors (e.g. FGF21) released by beige/brite cells relative to ‘classical’ brown adipocytes (see below).
Molecular controllers of differentiation of distinct
adipocyte cell types
Despite originating from different lineages, adipocytes undergo
adipogenic differentiation processes that share common transcriptional cascades (39). Indeed, the nuclear receptor PPARγ
(peroxisome proliferator-activated receptor-γ) is indispensable
for the development of all types of adipose cells (40,41). The
other master gene that determines adipogenic differentiation is
C/EBPα (CCAAT/enhancer-binding protein-α). C/EBPα acts
to maintain PPARγ expression, and both factors promote and
maintain the differentiated state of adipocytes by co-operatively
regulating the transcription of genes involved in processes such
as lipid and glucose metabolism and insulin sensitivity (42). The
absence of C/EBPα in mice prevents the development of all white,
but not brown, adipose depots, indicating that a lack of C/EBPα
can be compensated for in brown fat development, probably by
C/EBPβ (43,44). In fact, C/EBPβ has been shown to play a key
role in brown adipogenesis through direct interaction with the
co-regulator PRDM16 (PR-domain containing protein-16) (45).
PRDM16, together with PGC-1α (PPARγ-coactivator-1α),
have been identified as the main regulators of the phenotype
of both ‘classical’ brown adipocytes and beige/brite adipocytes.
In particular, PGC-1α, which coactivates PPARγ and PPARα
(46,47), is involved in the regulation of mitochondrial biogenesis,
oxidative metabolism, and thermogenesis. In contrast, PRDM16,
but not PGC-1α, has been shown specifically to confer brown
fat cell identity (48,49). In fact, PRDM16 acts primarily through
co-regulating C/EBPβ, PPARγ, PPARα, and PGC-1α to induce
expression of brown fat-specific genes (16,45,49,50). MiR-133a/b
and miR-155 are microRNA species that target PRDM16 and C/
EBPβ, respectively, thus having an inhibitory role on brown and
beige/brite adipogenesis and function. In contrast, miR-196a
appears to induce specifically beige/brite adipogenesis via inhibition of the homeobox protein HOXC8 (51). On the other hand,
the co-regulator TLE3 competes with PRDM16 for binding to
PPARγ, thus impairing brown adipocyte thermogenic gene expression and promoting a more white adipocyte gene expression
signature (52). Recent studies of adipocyte-specific PRDM16knockout mice showed that ‘classical’ BAT remains functionally
intact whereas there is a loss of the browning capacity (i.e. beige/
brite adipocyte function) in response to cold and β3-adrenergic
stimulation. When exposed to a high-fat diet, those mice show an
increase of subcutaneous WAT mass, but this WAT depot acquires
physiological characteristics more related to visceral WAT (53).
In contrast, selective ablation of PRDM16 in the brown (Myf5positive)—but not beige/brite (Myf5-negative)—adipose lineage
results in altered ‘classical’ brown adipocyte identity and function
in adult mice (54).
Genetic markers of the beige/brite phenotype
Several studies using differential gene expression profiling in cell
cultures have identified specific genes whose expression may be
used to distinguish beige/brite from brown adipocytes despite
common expression of the thermogenic genes, such as UCP1
(29,55). For instance, differential high expression of the zinc
finger transcription factor Zic-1 and LIM homeobox 8 protein
(Lhx8) is commonly found in ‘classical’ brown adipocytes relative
to beige/brite adipocytes in most studies (23,56). Expression of
TNF receptor superfamily member 9 (CD137), T-box associated
transcription factor (TBX1), transmembrane 26 (TMEM26), and
short stature homeobox-2 (SHOX2) are considered indicative of
beige/brite cell identity (25,56). The assessment of the expression of these genes in adipose tissue depots from mice confirmed
partially that the differential gene expression signature occurs
in vivo, (e.g. a high and distinctive expression of Zic-1 in ‘classical’ BAT depots but absence in beige/brite-prone WAT depots, or
preferential expression of SHOX2 in inguinal WAT, a site highly
sensitive to ‘browning’). Thus, it appears that marker gene expression profiles allows to distinguish a predominant ‘classical’ brown
adipocyte phenotype in the BAT depots versus a predominant
beige/brite phenotype in WAT depots in which browning had
been induced (27). However, considering that several members
of the gene panels that distinguish ‘classical’ BAT versus beige/
brite adipose tissue are homeobox genes or other genes involved
in positional determination during mammalian development, it
is unclear to what extent the expression of these marker genes
in tissues is reflecting cell identity or just positional location of
adipose cells across whole-body anatomy. In this sense, a recent
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approach aimed to identify a comprehensive molecular description of brown versus beige/brite gene expression using ribosomal
profiling concluded that several candidate genes may in fact
reflect much more the anatomical location in which adipocytes
reside rather than intrinsic adipocyte type-specific identity (35).
On the other hand, recent reports also claim that gene expression
signature in adipose depots is highly dependent on the developmental stage even in the same anatomical depot (57).
Finally, some authors have questioned the definition of
beige/brite as an independent cell type with respect to classical
brown adipocytes, and instead consider these cells should be
named just brown adipocytes (58). In fact, white adipocytes from
subcutaneous versus visceral depots are considered the same cell
type, notwithstanding reported differences in developmental
lineage, gene expression signature, and/or hormonal responsiveness (59–61). In the same sense, myocytes from different skeletal
muscle fiber-types also constitute unique cell types (62). At this
point, it may happen that, in fact, issues related to cell biology
and those related to the preferred use of a given terminology to
classify cells overlap.
Signals that control beige/brite cell appearance
and function
Because prolonged thermogenic stimulation appears to be the
main context in which the browning of WAT occurs in rodent
models, the classically recognized regulatory axis of sympathetic
activation and subsequent noradrenergic signaling is considered
the main mechanism that elicits the appearance of beige/brite
cells in WAT depots (63). However, a number of novel factors
capable of inducing the browning process have been identified
in recent years. Most of these factors also induce the activation of
existing brown adipocytes and promote the recruitment of classical BAT anatomical depots. These new factors, which are capable
of controlling the browning of WAT through mechanisms totally or partially independent of sympathetic activation, originate
from distinct tissues. The discovery of one of these factors, irisin,
is especially remarkable. Exercise has been reported to induce
browning of WAT (64). Bostrom et al. (65) observed that overexpression in muscle of PGC-1α, a transcriptional co-regulator
involved in muscular bioenergetics and metabolism, also caused
browning of WAT. A search for the mechanisms responsible for
this effect led to the identification of irisin, a bioactive protein released by skeletal muscle after cleavage of the membrane protein
FNDC5 (fibronectin type III domain containing 5), and showed
that this bioactive protein is capable of inducing the browning
of WAT in response to muscle contraction. Another factor that
promotes browning and activation of BAT is FGF21 (fibroblast
growth factor-21) (66,67), a hormone released mainly by the liver
in response to lipid availability (68). FGF21 is also released by
white and brown adipocytes in rodents and may have both an
endocrine and a paracrine action in promoting the browning and
thermogenic activity of BAT (67,69). A number of other factors
have been reported to promote browning and BAT activation
in rodents, as recently reviewed (70). They include cardiac peptides (71), vitamin A derivatives (72) or even metabolites such as
β-aminoisobutyric acid (73) and lactate (74).
Human brown adipose tissue: ‘classical’ versus
beige/brite
How do the above concepts and findings, which have been
developed mainly using cell culture models and rodents, apply
to humans? Cell culture analysis to explore adipocyte lineage, as
mentioned above for rodents, provided limited data for human
brown-versus-beige/brite identity distinction on the basis of
marker gene expression analysis. Table I summarizes the current
findings relative to marker genes for the different subsets of adipocytes specifically in humans. Remarkable similarities in marker
genes useful to distinguish adipocyte identity in adipose depots
have been found relative to rodent studies and, also as in rodents,
there is a wide range of biological roles for the corresponding
gene products, most of them unrelated to the current knowledge
of specific brown and beige/brite biological function.
The discovery of active BAT in adult humans at the anatomical
sites mentioned above has spurred interest in directly characterizing the features of this tissue through analysis of biopsies. In
fact, in one of the original studies published in the New England
Journal of Medicine (8), the authors reported high expression of
UCP1 and other marker genes of BAT ⫹ beige/brite versus WAT
molecular identity. The typical brown adipocyte morphology and
expression of UCP1 were confirmed by an independent study in
neck adipose tissue from adult humans (75). Recent studies have
also attempted to determine whether adult BAT at upper trunk
anatomical locations is ‘classical’ BAT or beige/brite BAT by direct
analysis of fat at these anatomical sites.
The first reports of these analyses yielded the surprising finding
that BAT in the supraclavicular region of adult humans expressed
the same marker genes representative of the beige/brite type previously identified in rodents and human cell culture studies (25).
An additional study, performed using a set of samples of supraclavicular as well as visceral fat depots in neonates and children,
also found indications of gene expression typical of beige/brite
adipocytes, but no evidence for expression of markers of classical,
developmentally programmed BAT (56). These findings raised
the surprising possibility that all BAT in humans might be the
beige/brite type.
However, subsequent reports have challenged this concept.
Lidell et al. (76) found that samples of BAT from the adult supraclavicular and periadrenal area expressed beige/brite markers
genes, but an analysis of neonatal interscapular BAT, an anatomical site equivalent to the interscapular area in rodents, revealed
differential expression of the classical BAT marker, ZIC1. In fact,
on the basis of the expression of marker genes for classical BAT
(i.e. ZIC1) and beige/brite BAT (i.e. CD137), we found that human fetuses at term simultaneously contain both types of brown
adipocytes depending on the anatomical region: ‘classical’ BAT
in the interscapular area and beige/brite in the omentum (77). At
this point, the question becomes: does ‘classical’ BAT exist only in
human fetuses and neonates, with all BAT in human adults being
beige/brite? Even this formulation is challenged by recent findings. Cypess et al. (78) found that the deep fat layers in the necks
of adults contain brown adipocytes (based on morphology and
UCP1 expression) that show a pattern of gene expression (high
ZIC1 expression) characteristic of classical BAT. In a separate
study of neck adipose tissue from adults, the authors reached a
similar conclusion, suggesting that human supraclavicular BAT
might consist of both ‘classical’ brown and beige/brite adipocytes,
also on the basis of marker gene expression (79).
In summary, clarifying the identity of human brown adipocytes through direct studies of human biopsies and samples has
proven to be a difficult task, first because of the limited availability and difficulty of obtaining such samples, and second because
of inherent doubts about the suitability of directly applying profiles of ‘classical’ and beige/brite marker genes, derived mainly
from rodent studies (and not totally agreed upon in the literature) to human BAT. Expression of the FGF21 gene highlights
these difficulties. In rodents, FGF21 is substantially expressed in
Brown and beige/brite adipogenesis in humans 5
Table I. Confirmed marker genes for different subsets of adipocytes in humans.
Phenotype
‘Classical’ brown
Gene symbol
Biological function
Zic family member 1
LHX8
LIM homeobox 8
miR206
MicroRNA 206
miR133b
HOXC4
MicroRNA 133b
Homeobox C4
HOXA1
UCP1
Homeobox A1
Uncoupling protein 1
DIO2
PRDM16
Deiodinase, iodothyronine, type
II
Peroxisome proliferator-activated
receptor gamma, coactivator 1
alpha
PR domain containing 16
ADRB3
FGF21
β3-adrenergic receptor
Fibroblast growth factor 21
CIDEA
Cell death-inducing DFFA-like
effector A
CITED1
TMEM26
SHOX2
Cbp/p300-interacting
transactivator, with Glu/
Asp-rich carboxy-terminal
domain, 1
T-box 1
Tumor necrosis factor receptor
superfamily, member 9
Transmembrane protein 26
Short stature homeobox 2
Beige/brite ⫹ white
HOXC8
Homeobox C8
White
HOXC9
LEP
Homeobox C9
Leptin
‘Classical’ brown ⫹
beige/brite
PPARGC1A (PGC-1a)
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Gene name
ZIC1
Beige/brite
TBX1
TNFRSF9 (CD137)
both WAT and BAT (although FGF21 has been proposed to be
preferentially expressed in beige/brite adipocytes in rodent cell
culture studies) (25,56). However, in humans, FGF21 expression
is negligible in WAT (both subcutaneous and visceral) (80), but
is substantial in both classical and beige/brite fat depots from human neonates (77).
BAT in pheochromocytoma patients: a human model
of the browning of WAT
Pheochromocytoma is a neuroendocrine tumor that secretes
large amounts of catecholamines. More than 50 years ago, Feyrter identified the presence of adipocytes with the multilocular
lipid droplet morphology typical of brown adipocytes in the
fat depots close to pheochromocytoma tumors (81). Further
research confirmed that these cells possessed the morphological (cytoplasm filled with mitochondria with numerous tightly
packed cristae) and biochemical (mitochondrial guanosine
diphosphate-sensitive, loose respiratory coupling) features
of bona fide brown adipocytes (Figure 1) (82). Expression of
the UCP1 gene and high amounts of UCP1 in mitochondria,
an unequivocal sign of brown adipocyte identity, was also
confirmed (83,84). In addition to UCP1, other BAT ⫹ beige/
brite marker genes (e.g. PRDM16, β3-adrenoreceptor) are also
highly expressed in omental adipose tissue from pheochromocytoma patients (85). Moreover, recent PET scan studies have
consistently reported hypermetabolism in the omental and
Multifunctional C2H2-type zinc finger
transcription factor
Cysteine-rich double-zinc finger transcription
factor
Inhibition of muscular transcription factors
myogenin and MyoD in brown adipocytes
See miR206; regulation of PRDM16 expression
Homeodomain-containing DNA-binding
transcription factor
See HOXC4
Uncoupling of oxidative metabolism from ATP
production, allowing heat production
Conversion of thyroxine (T4) to 3,3’,5triiodothyronine (T3)
Transcriptional co-activator controlling the
expression of key metabolism-related genes
Transcriptional co-regulator, control of the
brown phenotype transcriptional network
Norepinephrine receptor
Increase of glucose uptake and oxidation,
induction of UCP1
Activator of apoptosis upon DNA
fragmentation; lipolysis and thermogenesis
regulator
Transcriptional co-activator for estrogen
receptors
T-box-associated transcription factor
Clonal expansion, survival, and development of
T cells, induction of a Th1 program
Unknown
Homeodomain-containing DNA-binding
transcription factor
Homeodomain-containing DNA-binding
protein
See HOXC8
Regulation of body weight by inhibiting food
intake and energy expenditure in adipocytes
(Ref.)
(55, 76, 78, 79)
(55, 76, 78, 79)
(27, 79)
(79)
(9)
(9)
(25, 55, 76, 78, 79)
(76)
(21, 76, 79, 108)
(76, 79, 111)
(76)
(66, 77, 108)
(79)
(56, 79)
(25, 55, 76, 79)
(25)
(25, 78, 79, 108)
(76, 78, 112)
(27, 79, 113)
(27, 79, 108, 113)
(78)
mesenteric adipose tissue from pheochromocytoma patients,
indicating the functionality of brown adipocytes that develop
at those regions (86–89).
According to current concepts, the brown adipocytes that
appear in pheochromocytoma patients are, by definition, the
consequence of a browning process because they appear at sites
in which only WAT is present in control conditions and their
appearance is dependent on a particular stimulus—in this case,
continuous adrenergic stimulation caused by the tumor. Notably,
under basal conditions, visceral adipose tissue, the anatomical
site where BAT appears as a consequence of pheochromocytoma,
is considered much less prone to browning than subcutaneous fat
(a concept developed largely based on rodent studies), and remnant UCP1 expression levels in WAT from healthy adult humans
is more prevalent in visceral than in subcutaneous fat (90). In any
case, under this scenario, brown adipocytes that appeared in the
visceral fat of pheochromocytoma patients would be expected to
be beige/brite, but, to date, no extensive analysis of the molecular expression signature—‘classical’ versus beige/brite—has been
reported in BAT from pheochromocytoma patients.
How these adipocytes develop is the subject of a current
controversy, fueled by results obtained in rodent models. Some
authors suggest that precursor cells differentiate into a brown
(beige/brite) phenotype, whereas others claim that the existence of morphologically intermediate forms between white and
adipocytes—so-called paucilocular adipocytes, which express
UCP1—supports the idea that a transdifferentiation process from
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6
R. Cereijo et al.
Figure 1. Presence of beige/brite adipocytes in visceral adipose tissue from a patient with pheochromocytoma. Left: light microscopy, indicating
the presence of multilocular beige/brite adipocytes among other unilocular white adipocytes. Right: electron microscopy of the area indicated in the left,
high mitochondrial content characteristic of beige/brite adipocyte phenotype is shown, as well as poor mitochondria presence in the cytoplasmic rim of
white adipocytes.
white to beige/brite adipocytes takes place in adipose tissue from
these patients (85).
In any case, the appearance of brown adipocytes in patients
with pheochromocytoma confirms the existence of browning
potential in adipose tissues of adult humans. The availability of
samples as a byproduct of surgical treatment procedures provides
unique research opportunities that could be exploited to further
elucidate the biological processes underlying adipose browning
in humans.
How is browning controlled in humans?
Recent advances in our understanding of the neuro-hormonal
control of the browning process in rodents (70,91,92) are beginning to be extended to humans, with the obvious limitations associated with human studies. As in rodents, a cold environment
appears as a major inducer of human BAT activity (9,93). The
pheochromocytoma-associated appearance of brown adipocytes
in BAT depots is possibly the clearest evidence that in humans,
as in rodents, chronic adrenergic stimulation is a major inducer
of browning. Notably, the adrenergic receptor antagonist propanolol suppresses BAT activity in the neck and cervical areas
of humans, as assessed by PET scans (94). This indicates that
if most neck brown adipocytes are of the beige/brite type, as
noted above, their activity is controlled by sympathetic-driven,
adrenergic-mediated action. Some researchers have reported
that the sympathomimetic ephedrine activates BAT in humans
(95), whereas others were unable to replicate these findings (96),
perhaps owing to differences in the dose administered.
Do other, non-sympathetic-related, factors control BAT activity and the browning process in humans? Among the factors
identified in previous rodent studies, irisin and FGF21 possibly
deserve the most attention in humans. Investigations into the
capacity of exercise to increase irisin levels in humans have led
to conflicting results, with an initial report of higher irisin levels
after endurance training (65) not being systematically confirmed
by other independent studies (97–100). Whether exercise was
even effective in promoting browning in human adipose tissue
has been questioned (101). Some studies have indicated that irisin
is unable to induce the expression of BAT-related marker genes
in human white adipocytes (98), whereas others have reported
that FNDC5 activates the thermogenic activity of neck (mostly
beige/brite) adipocytes (102). It should be noted, however, that
the reliability of current methods for measuring human irisin in
blood has been a recent subject of debate, making it difficult to
compare data from different laboratories (103,104). In summary,
it appears that clear-cut evidence that irisin is able to induce the
browning of human WAT is still lacking, and further research will
be required (105,106). However, the recent finding that irisin is
expressed and released by human WAT itself (107) and thus has
a potential autocrine role in promoting browning may open new
research directions that help to establish the actual function of
irisin in human adipose tissue and its potential role in browning.
With respect to FGF21, it should first be noted that, unlike
the case in rodents, FGF21 in humans is very poorly expressed
in WAT, but is highly expressed in BAT—both classical (interscapular BAT from neonates) and beige/brite (neck BAT from
healthy humans, omental fat from pheochromocytoma patients)
(77,108). There is evidence that human adipocytes are sensitive
to FGF21 action, and treatment of human preadipocytes with
FGF21 activates beige/brite genes involved in the browning program (102,108), suggesting that FGF21 may be an endocrine as
well as an autocrine factor in controlling beige/brite appearance
and activity. A recent report found a strong association between a
rise in circulating FGF21 levels in response to cold and activation
of BAT (mostly beige/brite) in the neck in humans, consistent
with a role for FGF21 in promoting the activity of beige/brite
cells (102).
Conclusions and perspective
Clarifying the cellular identity of distinct types of brown adipocytes may at a first glance appear to be somewhat of an academic
exercise in the larger context of cell biology research. However,
the current awareness of the potential value of promoting BAT
activity in humans so as to enhance energy expenditure (thus
protecting against obesity) and drain glucose and fat from the
circulation (thus protecting against diabetes and hyperlipidemia)
stresses the relevance of this line of research. Experiments in
rodents suggest that the neurohormonal processes that control
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Brown and beige/brite adipogenesis in humans 7
the activation and perhaps recruitment of existing BAT may
not be identical to those that promote the enrichment of brown
adipocytes in former WAT depots. Whereas some classical activators (e.g. β-adrenergic activation) and more recent activators
(FGF21) appear to induce both the activity of existing BAT and
the promotion of browning (66,67), others (e.g. irisin) (65) appear more selective for the browning process. However, the role
of these factors in the control of BAT activity and browning of
WAT specifically in humans has not yet been definitively established. Identification of cell targets in humans and determining
their relative sensitivity to neurohormonal modulators is of
utmost relevance to the identification of drug targets and development of pharmacological tools for future treatment of obesity
and metabolic diseases. Moreover, although currently available
data support common thermogenic roles for ‘classical’ brown
and beige/brite adipocytes, their different gene expression
profiles suggest that, beyond their current role as experimental
tools, these cell types may potentially serve distinct, and as yet
unidentified, physiological functions. Finally, new roles of BAT
and/or beige/brite activation that could be used for metabolic
therapy purposes are increasingly being proposed, ranging from
the control of hyperlipidemia and hyperglycemia (109,110) to
the alleviation of redox pressure (74). The fascinating story of
adipose plasticity in humans and the prospects for pharmacological intervention are only just beginning.
11.
12.
13.
14.
15.
16.
17.
18.
19.
20.
Acknowledgements
21.
This work was supported by MINECO (grant SAF2011-23636),
Instituto de Salud Carlos III (grant PI11/00376), EU (FP7 project
BETABAT, grant HEALTH-F2-2011-277713), and Generalitat
de Catalunya (2009SGR-284). Thanks are given to A. Goday and
J. M. Gallego-Escuredo for help with microscopy images from
pheochromocytoma patients.
22.
Declaration of interest: The authors report no conflicts of
interest.
24.
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