1. Synopsis
The recent reports of a third type of adipose tissue, identified as beige, bright, or pink adipose tissue and a proposed role in energy balance in response to alterations in diet, environment, hormonal regulation, and potential pharmacologic strategies are reviewed and discussed, with particular reference to the role of insulin in various strains of obese and obese non-insulin-dependent diabetic (NIDDM) rats.
2. Introduction
The brown adipose tissue (BAT) of man and animals has been widely reported to play a key function in the thermal adaptation to alterations to diet and environment and is a likely contributing factor in the development of fatness vs. leanness of an animal due to its capacity to expend energy as heat rather than storage per se when in an activated state [1–3]. The presence of BAT has been observed in cadaveric dissections for many years, but its physiologic role remained unclear until studies reported in recent decades uncovered some of the molecular mechanisms inherent in thermogenic BAT actions in man and animals [2, 4–6]. The BAT is now known to play a key role in rodents and other mammalian species during recovery from states of torpor and hibernation [2]. In humans, BAT has been proposed to play a significant thermogenic role in the newborn, where it helps to generate heat to maintain homeothermy during the first stages of life outside the womb [1, 2]. BAT has also recently been proposed as a potential site for obesity therapy due to its capacity to convert stored energy into heat energy, which may be readily dissipated from bodily surfaces or to maintain homeothermy [7, 8]. Recently, the discussion of a third intermediate type of adipose tissue termed beige, bright, or pink adipose tissue has emerged, and has been proposed with the suggestion that it may be able to transdifferentiate to become thermogenically active BAT under certain environmental or hormonally activated conditions or to return to an energy storage role when the thermogenic needs of the organism become less active [8, 9]. Increases in physiologic or environmental conditions that enhance sympathetic activity have been proposed as one of the possible mechanisms to promote development of thermogenic BAT tissues, while factors that impaired thermogenic activity of BAT, including β-blockade and excess insulinogenic activity, have been associated with decreased thermogenesis activity in BAT [9–12]. In contrast, both caffeine and ephedrine, which pharmacologically bypass the normal neuroendocrine activation of BAT, have been reported to increase parameters of non-shivering thermogenesis in both lean and obese rats [13].
3. Methods
Papers were selected following PubMed and Google Scholar searches, which yielded over 100 applicable published studies to cite from since approximately 1970–2022. Search terms used included BAT, beige adipose tissue, adipocyte hyperplasia and hypertrophy, lipomas, dermatofibrosarcomas, and liposarcomas.
In the obese phenotype of several obese rodent strains summarized below, the BAT depots are well developed and appear to have undergone significant increases in mass and cellularity, consistent with early hyperplasia, hypertrophy, and lipid accretion in the BAT depots, particularly the interscapular depot where most studies have focused [8, 14–19]. The histologic features of white and brown adipocytes are markedly different: White adipocytes are readily identified in histologic specimens by their single large lipid droplet, which typically occupies up to 90% or more of the histologic structure. The single large lipid droplet is typically surrounded with a narrow rim of cytoplasmic material and an often ovoid-shaped nucleus compressed into the cytoplasmic rim [12]. White adipocytes may continue to form from preadipocytes and via cellular hypertrophy of existing adipocytes throughout much of the adulthood of mammalian species in response to imbalances in energy intake and expenditure. In contrast, brown adipocytes expand via hyperplasia and hypertrophy predominantly during early, preadolescent life, are typically smaller in circumferential diameter, contain a dense appearance, more centrally located spherical nucleus that is surrounded by multiple small lipid locules, and a metabolically active cytoplasm containing an abundance of unique, specialized mitochondria capable of generating heat from lipid and carbohydrate energy reserves. Once formed, the differentiated mature adipocytes of both white adipose tissue (WAT) and BAT appear to remain present thereafter where they may contribute to both physiologic and pathophysiologic sequelae well into adolescence and throughout much of the adult lifespan [14].
The specialized mitochondria of BAT exhibit the capacity to convert stored energy into heat energy via a biochemical uncoupling of oxidative phosphorylation [1, 2]. The conversion of stored energy into heat energy is initiated via a BAT-unique uncoupling protein (UCP1)-activated hydrolysis of the high energy phosphate bonds of Adenosine triphosphate (ATP), forming adenosine diphosphate (ADP), inorganic phosphate (iP) and approximately 7.2 kcals of releasable heat energy per mole of ATP consumed in the reaction [1, 2]. BAT is well innervated via sympathetic neurons, releasing norepinephrine, capable of initiating the thermogenic cascade, which can impinge on stereospecific specialized β3-adrenergic receptors common to BAT and to initiate the cascade of thermogenic events [10, 11]. In addition, the presence of the UCP1 is highly unique to BAT tissues [1]. In contrast, WAT lacks direct neural innervation and can undergo biosynthetic processes of lipid accumulation or mobilization in response to endocrinologic signals mediated by circulating hormones, especially insulin and catecholamines, respectively, and lacks the capacity for expression of the UCP1 protein [2, 7, 8]. Thus, the morphometric, biochemical, and metabolic characteristics of differentiated BAT and WAT tissues differ significantly, which contributes to their established and distinct primary physiologic roles in energy metabolism and energy expenditure vs. energy storage. These unique characteristics have contributed to the identification of BAT as a potential mediator of energy balance, and as a potentially useful pharmacologic target for altering the balance between energy storage in WAT and energy expenditure in BAT as a possible mechanism projected toward reducing the magnitude of adiposity and with a goal toward establishment of a leaner physique [6–8]. The definitive origins and potential roles of the beige adipose tissue are unclear, but the discovery of the presence of the UCP1 in beige adipose tissue suggests that the tissue is more directly linked to BAT than WAT since the UCP1 protein is unique to BAT [8, 9]. The mechanisms for the transdifferentiation between BAT and beige AT are unclear; however, review of the morphometric characteristics and hormonal profiles which occur in animals expressing beige AT vs. BAT suggest that disordered elements of energy metabolism processes likely contribute a decisive role in the development of the transitional AT state. WAT can also express a limited energy-wasting metabolic fatty acid cycle, albeit via a different biochemical mechanism and independent of the actions of the UCP1 protein of BAT [20]. Pharmacologic agents that bypass the sympathetic neuroendocrine activation of BAT-mediated thermogenesis including caffeine and ephedrine have also been demonstrated to enhance parameters of non-shivering thermogenesis in both lean and obese rats, suggesting that the fundamental elements of mitochondrial heat generation remain potentially operable despite impairments in β3-adrenergenic mediated pathways in addition to thermogenic mechanisms in other tissues [3, 6, 13].
Adiposity has long been associated with the progressive development of insulin resistance in man and animals at least in part via hyperinsulinemia and a consequent decreased sensitivity to the actions of circulating insulin availability and plasma concentrations. This is associated with increases in insulin secretion in a feed-forward, counterregulatory loop and implicates the cooperation of intracellular glucose transporter 4 (GLUT4) transporters from the endoplasmic reticulum to the plasma membrane, where they can effect cellular glucose uptake [21, 22]. The decreased cellular capacity to effect efficient glucose uptake and subsequent cellular oxidation contributes to a decreased plasma membrane receptor sensitivity to insulin in many tissues, and in the development of insulin resistance in skeletal muscle and WATs [21]. Thus, the functional capacity for metabolic energy expenditure in response to alterations in diet and environment diminishes as the onset of obesity progresses. Insulin resistance in association with dysregulation of glucocorticoid actions is also associated with decreased expression and intracellular translocation of the plasma membrane-associated glucose transporter protein, GLUT4, from the endoplasmic reticulum to the plasma membrane, where it normally effects efficient cellular glucose uptake from plasma origins [23]. Most notable are the damaging effects in skeletal muscle and adipose tissue, where the GLUT4 particles are an absolute essential intermediate in transitioning the cellular uptake of glucose in insulin-dependent tissues including both skeletal muscle and adipose tissue [22, 23]. In humans, the continued derangements in insulin sensitivity and insulin action in peripheral tissues in the obese state also contribute to the development of syndrome X, also termed metabolic syndrome (MetS) and to one of its most common sequelae, Type 2 diabetes mellitus (T2DM) or NIDDM [24]. The combination of overweight and obese conditions in association with T2DM is now highly prevalent in westernized populations and present a serious and costly global challenge to deliver health care resources to the affected populations [25].
Early overnutrition results in hyperplasia and hypertrophy of BAT in lean and obese rats, but the intensity of the brown coloration in the obese phenotype typically becomes less intense over time in apparent parallel to the progression of hyperinsulinemia and insulin resistance commonly found in obese rodents and the obese of other mammalian species [26]. The epigenetic expression of an obese phenotype may also develop in the absence of NIDDM in some obesity-prone strains [14, 15, 19]. The increase in BAT mass in obese rats is disproportionate to the magnitude of thermogenesis typically observed, in that resting and norepinephrine-stimulated oxygen consumption (VO2) remain depressed among the obese phenotype despite the increases in BAT mass and cellularity [14–19]. Insulin resistance occurs in the obese phenotype of the obese, non-diabetic Zucker fatty (-fa) rat and the LA/Ntul//-cp (corpulent) rat and the NIDDM strains including the T2DM Wistar fatty rat, and the obese T2DM SHR/Ntul//-cp (corpulent) rat, and the WKY/N-cp rats with the greatest magnitude of insulin resistance (IR) among the obese+diabetes prone strains [14–19]. Hyperplasia and hypertrophy of BAT occur in both non-T2DN and in obese-T2DM/NIDDM strains, however, and became visibly evident during early postweaning growth and by early adulthood hypertrophy of the BAT tissues became readily apparent in dissections, where it was associated with a less intense brown coloration as the adipocytes accumulate additional lipid reserves when insulin resistance occurs. The hypertrophied brown adipocytes retain fundamental morphometric characteristics typical of brown adipocytes including the multilocularity of small lipid droplets, and a centrally located nucleus. Also, administration of the β3 blocker propranolol was found to result in an increase in locular diameters in histologic examinations taken from lean Sprague Dawley rats, but in that study the expansion of the BAT mass and cellularity did not result in adipocyte hypertrophy or significant “unbrowning” of the brown adipocytes, perhaps because of the shorter duration of the β-blockade treatment in combination with the genetically lean phenotype of the animals and protection from cold exposure or experimental overnutrition during housing conditions [12]. A summary of the BAT characteristics is presented in Table 1 and indicates that the BAT mass and cellularity are disproportionate to the thermogenic responses in the obese phenotype of multiple genetically obese strains.
Table 1
Qualitative summary of characteristics of bat and metabolic parameters in lean and obese rats
| Strain/phenotype | IR | NIDDM | RMR | [T3] | IBAT mass | IBAT cellularity | IBAT hypertrophy | References |
|---|---|---|---|---|---|---|---|---|
| Sprague Dawley-chow diet | No | No | Normal | Normal | Normal | Normal | No | [26] |
| Sprague-Dawley-café’ diet | No | No | Incr | Incr | Incr | Incr 2‒3× | No | [26] |
| LA/Ntul//-cp | ||||||||
| Lean | No | No | Normal | Normal | Normal | Normal | No | [14, 15] |
| Obese | Yes | No | Decr | Decr | Incr ~5×* | Incr ~5× | Yes | [14, 15] |
| SHR/Ntul//-cp | ||||||||
| Lean | No | No | Normal | Normal | Normal | Normal | No | [16] |
| Obese | Yes | Yes | Decr | Decr | Incr >5×* | Incr ~5× | Yes | [16] |
| Zucker fatty (fa) rat | ||||||||
| Lean | No | No | Normal | Normal | Normal | Normal | No | [19] |
| Obese | Yes | Yes/No** | Decr | Decr | Incr ~5×* | Incr ~5× | Yes | [19] |
| Wistar fatty(fa) rat | ||||||||
| Lean | No | No | Normal | Normal | Normal | Normal | No | [18] |
| Obese | Yes | Yes | Decr | Decr | Incr ~5×* | Incr ~5× | Yes | [18] |
| WKY/N-cp rat | ||||||||
| Lean | No | No | Normal | NR | Normal | Normal | No | [17] |
| Obese | Yes | Yes | Decr | NR | Incr >2×* | Incr | Yes | [17] |
* Indicates visible unbrowing by inspection. IR as determined by an elevated insulin to glucose ratio; IBAT mass reported in grams/interscapular depot; IBAT cellularity determine as cells number per depot; IBAT hypertrophy determined by cell lipid content per cell greater than occurred in normally fed lean animals of the same strain. Correlations between IR and RMR and of IR and increased IBAT mass yielded an r2 = 1.0 in these comparisons. IR, insulin resistance; NR, not reported; RMR, resting metabolic rate; T3, plasma triiodothyronine; IBAT, interscapular brown adipose depot; Incr, increased; Decr, decreased.
The decrease in resting metabolic rate (RMR) in association with insulin resistance among the obese phenotype of the obese rat strains despite having significant increases in interscapular brown adipose depot (IBAT) mass and cellularity is suggestive of impaired activation of non-shivering thermogenesis in those animals, while in genetically lean café-fed Sprague Dawley rats, IR was not observed and RMR was increased following a café regimen. Comparing the two variables in obese strains yielded a correlation coefficient r2 of 1.0, suggestive of a link between the two parameters. Bukowiecki et al. [10, 11] observed decreased norepinephrine-stimulated thermogenesis in isolated brown adipocytes obtained from insulin resistant obese-NIDDM SHR/Ntul//-cp rats and concluded that insulin sensitivity was an essential element in the expression of BAT thermogenesis. Glucose uptake contributes to the thermic responses in IBAT and other insulin-dependent tissues and is likely an essential significant contributor to the decreases in RMR characteristic of and commonly observed among obese rodent strains [10, 11]. Circulating plasma concentrations of triiodothyronine (T3) also contribute to an optimal thermogenic response in rats, and along with RMR are also commonly decreased among the obese phenotype of epigenetically obesity-prone strains [14–19]. Café’ feeding regimens in both lean and obese rats result in increases in BAT mass, plasma T3 concentrations, vanilmandelic acid (VMA) excretion, RMR, and norepinephrine-stimulated VO2, while pharmacologic sympathomimetic blockade via α-methylparatyrosine results in decreases in VMA and the magnitude of the sympathetic component of the thermogenic responses [26–28].
4. Discussion
The observation of a decreased intensity of robust brown coloration in IBAT depots in combination with insulin resistance as it occurs in multiple obese rodent strains is consistent with a link between hyperinsulinemias-associated insulin resistance and the “unbrowning” or beigeing of BAT with a likely onset early in the lifespan of obese animals. In lean animals when offered a highly palatable café diet regimen from weaning or thereafter, RMR, plasma T3 concentrations, and IBAT mass and cellularity also became significantly increased, but the insulin resistance and the “unbrowning” effect noted among obese littermate animals was not observed, suggestive of thermogenically active IBAT in the lean animals in response to the café’ overnutrition regimen offered during their postweaning development. The genetically obese animals of the strains cited reported typically demonstrate hyperphagia in the early postweaning growth phase, which may be the likely trigger for the BAT hyperplasia observed. Insulin is a well-documented lipogenic hormone in adipose tissues, where it can facilitate both de novo biosynthesis and cellular uptake of preformed triglycerides, while suppressing both lipolysis and the glucose-dependent elements of the thermogenic response, the combined effects thereby contributing to increased lipid accretion of both white and brown adipocytes. While an undetermined proportion of the decreased brown coloration may be due to infiltration of white adipocyte types in the IBAT mass, the greater lipid content and locule diameters of the brown adipocytes indicate that those cells also play a role in the unbrowning effect. In one study, the lipid content of the IBAT mass and cellularity of the obese phenotype was found to be 1.5-fold greater than those occurred in similarly fed lean littermates, while the capacity for non-shivering thermogenesis was significantly decreased, signaling insulin actions as one of several likely possibilities to account for the increased lipid content and decreased brown coloration of the brown adipocytes in concert with the decreased capacity for BAT thermogenesis. The peptide hormone amylin is co-secreted with insulin, and in lean rats acts to aid in satiety responses by slowing the rate of gastric emptying by impinging on amylin receptors in the antrum of the stomach [29]. Obese LA/N-cp and SHR/B-cp rats also develop hyperamylinemia and amylin resistance in concert and accelerated gastric emptying, which likely contributes to the chronic hyperphagia of the obese phenotype.
In lean animals, overfeeding via the café regimen has been shown to increase both sympathetic and thyroidal activity, both of which contribute proportionately to the thermogenic responses [27–29]. At the age of weaning, there is little difference in the body weights and only minimal differences in the adiposity of lean and obese phenotypes, but the hyperphagia common to the obese phenotypes noted above has been observed during early postweaning growth, at an age during postweaning, prepubertal development where the partial thermogenic responses to exogenous norepinephrine were still apparent [14, 30]. At later ages, the thermogenic responses to diet, norepinephrine, and acute cold exposure become progressively decreased in the obese phenotypes [31, 32]. Frontini et al. have reported that pink adipocytes conveying the thermoregulatory protein UCP1 were observed in omental tissue from human adults with pheochromocytoma, suggesting that the differentiation of brown adipocytes may have been induced by the chronically elevated catecholamine hormone secretions, similar to the hormonal profiles observed during the overfeeding regimens [9, 27, 28]. Embryologically, both WAT and BAT are thought to be derived from the neural crest during early prenatal development, giving rise to mesodermal and ectodermal preadipocytes. The preadipocyte origin of the omental pink to brown adipocytes remains unclear however, as does the confirmed embryologic origins of the beige adipocytes [33]. The newfound pink to brown adipocytes is small in number, however, compared to the mass, cellularity, and distribution of commonly observed primary BAT depots [9].
Adipose tissues expand via the processes of hyperplasia and hypertrophy and combine to bring about the storage of adequate triglyceride energy reserves in times of metabolic need. During an optimal healthful state, adipocyte mass and cellularity contribute to maintain energy balance and homeostasis, while during times of caloric surplus, the adipose mass can readily expand to accommodate storage of the excess metabolic fuels to the extent of contributing the secondary pathophysiologic sequela common to obesity. Once thought to be completed by early adulthood based on rodent epididymal fat pad measurements of WAT, it is now recognized that continued hyperplasia in subcutaneous and visceral WAT depots may continue well into adulthood in man and animals, and that the maximal adipocyte lipid content of white adipocytes becomes limited at around to approximately 1.2–1.5 µg of lipid per cellular droplet, thus necessitating additional maturation of preadipocytes to accommodate additional and often excessive energy storage throughout the lifespan [2, 14]. In BAT, however, the preadipocytes appear to increase via hyperplasia in response to early nutritional and environmental signals prior to adulthood and the morphologic characteristics and tissue mass remain present thereafter, and overall the subsequent hypertrophy of brown adipocytes is modest by comparison. It should be noted however that all hyperplasia and hypertrophy of adipose tissue may not always be of a benign nature, and adipose tissue-related neoplasms should always be a consideration. As with many tissues undergoing hyperplasia, epigenetic molecular events may occur via random or via nutritional or environmental factors in adipose and nearby tissues to result in formation of both nonmalignant lipomas, liposarcomas and, less frequently, neoplastic tumor growths, which take on the morphometric and pathologic characteristics of dermatofibrosarcomas [34, 35]. Among the liposarcomas, both well-differentiated liposarcoma (WDL) or atypical lipomatous tumor and dedifferentiated liposarcomas (DDL) may occur; the DDL is associated with more aggressive clinical behavior, a greater propensity for local recurrence, and infrequently an innate capacity for metastasis resembling that of other pleomorphic sarcomas [36]. The embryologic origins of the various forms of lipid-associated tumors remain unclear and await confirmation via identification of specific adipokine cell markers and appear at least in part to serve as an energy and nutrient source for the emerging tumor [32, 37, 38]. Confirmed neoplastic growths or overt hypertrophy of adipocytes in brown, beige, or pink adipose tissues appear to be uncommon however and were not discovered or reported to date in the citations reviewed.
Thus, the hormonal and metabolic stimulus that is linked to the transdifferentiation of brown adipocytes to become beige adipocytes in obese and obese-NIDDM phenotypes, or the proliferation and differentiation of a third class of adipocytes now known as beige, bright, or pink adipocytes remains unresolved. Beige and related descriptions of adipocytes express the thermogenic UCP1 protein and are presumed capable of exhibiting thermogenesis in a manner similar to the brown adipocytes, although the magnitude of the thermogenic responses of beige adipocytes and their insulin and adrenergic sensitivity is unknown but may be less dramatic than that observed in normally active mature brown adipocytes due at least in part to their attenuation of insulin sensitivity, a likely decreased efficiency of glucose uptake, and greater lipid accretion and locule sizes [31]. Lipid locule size in adipocytes is presumed to be a critical factor, as it changes the surface area to lipid content ratios, which may decrease or diminish the sensitivity of lipid mobilization due to the relative decrease in locular surface area to lipid ratio. The metabolic roles of accessory modulators of energy metabolism of the adipokines AMPK (adenosine monophosphate activated protein kinase) and sirtuins in beige adipocytes also remain speculative at best, while their participation in enhancing the metabolic processes of energy metabolism in lean tissues is acknowledged. The GLUT4 intracellular glucose transporter activity is essential in initiating the thermogenic responses in BAT, but glucose uptake in BAT is decreased in the obese and obese+NIDDM phenotypes, suggestive of hyperinsulinemia-mediated dysregulation of the GLUT4 transporter mechanism in those animals, resulting in an impaired thermogenic response following perturbations in diet and environment [31, 39, 40].
