Natural Bioactive Compounds as Potential Browning Agents in White Adipose Tissue

Abstract

The epidemic of overweight and obesity underlies many common metabolic diseases. Approaches aimed to reduce energy intake and/or stimulate energy expenditure represent potential strategies to control weight gain. Adipose tissue is a major energy balancing organ. It can be classified as white adipose tissue (WAT) and brown adipose tissue (BAT). While WAT stores excess metabolic energy, BAT dissipates it as heat via adaptive thermogenesis. WAT also participates in thermogenesis by providing thermogenic fuels and by directly generating heat after browning. Browned WAT resembles BAT morphologically and metabolically and is classified as beige fat. Like BAT, beige fat can produce heat. Human adults have BAT-like or beige fat. Recruitment and activation of this fat type have the potential to increase energy expenditure, thereby countering against obesity and its metabolic complications. Given this, agents capable of inducing WAT browning have recently attracted broad attention from biomedical, nutritional and pharmaceutical societies. In this review, we summarize natural bioactive compounds that have been shown to promote beige adipocyte recruitment and activation in animals and cultured cells. We also discuss potential molecular mechanisms for each compound to induce adipose browning and metabolic benefits.

INTRODUCTION

Obesity is a major public health burden in developed societies. Currently it affects more than two billion people (1). Metabolic complications of obesity include, but not limited to, dyslipidemia, insulin resistance, type 2 diabetes, fatty liver, cardiovascular disease, and cancer (2). Overall, obesity is a result of increased energy intake and/or reduced energy expenditure. While agents reducing energy intake and nutrient absorption or food intake may reduce weight gain (3), those enhancing energy dissipation may also be beneficial. A major pathway in dissipating metabolic energy is the nonshivering adaptive thermogenesis (4). In rodents, anatomically identifiable brown adipose tissue (BAT) depots are responsible for heat production through the adaptive thermogenesis. Human infants also possess visible BAT in the interscapular region. However, this BAT is lost during growth. Over a long time, scientists believed that adult humans did not have BAT. This belief was overturned in 2009 when three independent groups reported the existence of brown fat-like adipose tissues in human adults, particularly in those after cold exposure (5–7). These milestone human studies revitalized research in brown fat biology and the role of BAT thermogenesis in whole-body energy balance. BAT is composed of brown adipocytes and many other cell types. Brown adipocytes are responsible for heat production in the BAT, though other cell types play important regulatory roles. A classical brown adipocyte is characterized by multilocular lipid droplets, abundant mitochondria, and expression of uncoupling protein-1 (UCP-1). UCP-1 protein localizes at the inner mitochondrial membrane and short-circuits the mitochondrial proton gradient to produce heat via oxidative phosphorylation of metabolic fuels such as fatty acids (8). It has long been known that multilocular lipid droplet-containing adipocytes exist in the classical white fat pads in humans and rodents (9). These cells are now known as brite or beige adipocytes because they express UCP-1 and are thermogenic (10). Importantly, they can be induced or recruited to the white adipose tissue (WAT) in response to environmental and pharmacological stimuli such as cold exposure and adrenergic activators (9–11). The process of inducing beige adipocyte appearance in the WAT is called WAT browning (12). Animal and human studies have shown that increased WAT browning is associated with resistance to diet-induced obesity and metabolic disorders, and with leanness in humans (13). Therefore, targeting thermogenic adipose tissues may counter obesity and its associated metabolic sequelae.

Adipose browning is regulated by a complex interplay of endogenous and environmental factors (14, 15). A hallmark of white fat browning is the expression and activation of UCP-1, though UCP-1-independent adipose browning mechanisms also exist (16–19). UCP-1 expression is regulated predominantly at the transcriptional level (4). Peroxisome proliferator-activated receptor γ (PPARγ) coactivator-1α (PGC-1α), a master regulator of mitochondrial biogenesis and oxidative metabolism, can induce the expression of UCP-1 and other thermogenic genes in adipocytes (13, 20). It does so by complexing with PPARγ and a transcriptional coregulator PR domain zinc finger protein 16 (PRDM16) (21, 22). PRDM16 is essential in maintaining brown fat characteristics and its deficiency induces expression of muscle genes in brown preadipocytes and brown fat pads (22). PRDM16 can interact with multiple DNA-binding transcriptional factors, including PPARα, PPARγ, and several CCAAT/enhancer-binding protein family members, and this interaction is capable of inducing PGC-1α expression (23, 24). In addition to PRDM16, PGC-1α, and PPARs, other signaling pathways also induce UCP-1 expression and adipose browning. For example, BAT activation is often associated with increased phosphorylation of 5’-AMP-activated protein kinase (AMPK) under in vitro and in vivo conditions (25, 26), and prolonged treatment with the AMPK activator 5-aminoimidazole-4-carboxamide ribonucleotide enhances WAT browning in mice (27). The crosstalk between the intracellular Ca²⁺-activated calcium/calmodulin-dependent protein kinase II-AMPK signaling and Sirtuin-1 (SIRT1), a sensor of cellular metabolism and energy utilization, mediates adipose browning downstream of capsaicin-induced activation of transient receptor potential cation channel subfamily V member 1 (TRPV1) (28). Fibroblast growth factor 21 (FGF21) is capable of inducing white fat browning (29), though not essential for nonshivering thermogenesis (30). Other growth factors, such as several bone morphogenetic proteins (BMPs), also promote brown phenotypes in adipose tissues (31–35). Pharmacological approaches targeting the aforementioned pathways induce adipose browning (28, 36–40). Nonetheless, PRDM16 expression and UCP-1 induction are most consistent with the presence of beige/brite adipocytes within white fat pads (41, 42), highlighting important roles of PRDM16 and other UCP-1-inducing factors in driving thermogenic or browning phenotypes in adipose tissues.

Natural plant-derived compounds have been the backbone of many synthetic drugs that are used today. Approximately 50% of approved drugs in the last 3 decades are directly or indirectly from natural compounds (43). Emerging evidence suggests that some food compounds, phytochemicals in particular, may be promising candidates for obesity management, a proportion of which may counter obesity by promoting adipose browning (44). Several excellent review articles have summarized some bioactive nutritional compounds that are capable of increasing energy expenditure through brown and white fat thermogenesis, such as those by Concha et al. (45), Kang et al. (46), and Zhang et al. (47). Each of these reviews was focused on certain categories of nutritional agents or bioactive compounds that activate BAT and/or WAT. In this review, we provide an update to include more natural compounds, such as polyphenols, terpenoids, and alkaloids. We focus on those capable of inducing white adipose browning in cultured adipocytes and whole animals, discuss whether the compounds’ browning effect is linked to their metabolic benefits, and highlight the potential mechanisms by which the compounds induce adipose browning. The review also identifies current challenges and opportunities for translational research to evaluate the role of phytochemicals in adipose browning and metabolic health.

NATURAL BIOACTIVE COMPOUNDS INVOLVED IN ADIPOSE BROWNING

Polyphenols

Resveratrol

Resveratrol (3, 5, 4′ trihydroxystilbene) (RSV) is a polyphenolic compound enriched in Polygonum cuspidatum. A small amount of RSV is also found in red wine, peanuts, grapes, berries, red cabbage, and spinach (48, 49). In the past 15 years or so, RSV is one of the most widely studied polyphenols due to its metabolic and health benefits, including antioxidative, anti-inflammatory, and anti-cancer effects (50–55). Studies have shown that RSV induces metabolic benefits by targeting multiple pathways, including adipogenesis, fatty acid oxidation, lipolysis, and apoptosis in adipocytes (50, 51, 56–59).

It has been shown that RSV protects rodents against high fat-induced obesity and metabolic abnormalities by promoting mitochondrial oxidative capacity of BAT, skeletal muscle and liver via the AMPK-SIRT1-PGC-1α pathway (50, 51, 60). In 2012, Park and associates reported that RSV attenuates aging-related metabolic disorders by inhibiting cAMP-degrading phosphodiesterases (PDEs) and activating the cAMP effector Epac1 (61). This study identified RSV as a nonselective PDE inhibitor and provided a unifying upstream mechanism underlying the activation of the AMPK-SIRT1-PGC-1α pathway by RSV. A few recent studies suggest that RSV supplementation may reduce weight gain and improve metabolic health by inducing energy expenditure through activation of thermogenesis and WAT browning (57, 62, 63). In a study when 5-month-old CD1 female mice were challenged for 4 weeks with a high fat diet (HFD) supplemented with or without 0.1% RSV, the mice treated with RSV versus those without RSV treatment display reduced weight gain and increased browning of the inguinal subcutaneous WAT (iWAT) as evidenced by increased multilocular lipid droplets-containing adipocytes, UCP-1 expression, and fatty acid oxidation (57). This study further showed that RSV at 10 μM increases mRNA levels of genes related to adipocyte browning, including PRDM16, UCP-1, Cidea, ELOVL Fatty Acid Elongase 3 (Elovl3), PGC-1α, CD137, T-Box Transcription Factor 1 (Tbx1), and transmembrane protein 26 (TMEM26) in differentiated stromal vascular cells (SVCs) isolated from the iWAT of mice (57) (Table I). This effect was shown to be entirely AMPKα-dependent, demonstrating a critical role of AMPK in mediating RSV-induced adipose browning (57), which is consistent with an early study showing that RSV does not exert metabolic benefits in AMPK-α1 or α2-deficient mice (52) and that RSV can activate AMPK (50). In another study, Hui et al. reported that type 2 diabetic male db/db mice fed a chow diet supplemented with 0.4% RSV for 10 weeks exhibit improved glucose homeostasis, which is associated with enhanced BAT activation and WAT browning (62) (Table I). In this study, the mice treated with RSV were found to have altered plasma and fecal bile acid compositions that were enriched of lithocholic acid (LCA). LCA is a potent endogenous ligand for Takeda G protein coupled receptor 5, a membrane-bound bile acid receptor known to mediate bile acid-induced activation of BAT thermogenesis (64, 65). Bile acids are produced in the liver and subject to extensive modifications by the gut microflora (66–68). Likewise, RSV is metabolized in the liver and gut (69). In the gut, intestinal bacteria are involved in the metabolism of RSV (70). Therefore, RSV has the potential to alter the gut microbiome. Indeed, studies have shown that RSV treatments alter the gut microbiota and promote adipose browning that depends, at least in part, on the gut microbiome in db/db or high fat-fed mice (62, 63) (Table I). RSV not only improves metabolic health in adult rodents. A recent study showed that the supplement of RSV together with nicotinamide riboside in early postnatal life attenuates fat-induced metabolic abnormalities and promotes WAT browning in adult male, but not female mice (71).

Table I.

Dietary compounds shown to stimulate WAT browning in animals

Bioactive Compounds	Animals & treatments (dosage, duration)	Effects on WAT gene expression	Metabolic phenotypes	Ref
Polyphenols
RSV	HFD-fed CD1 female mice (0.1% in diet, 4 wks)	↑ UCP-1, PRDM16, Cidea, Elovl3, PGC-1α, CD137, Tbx1, and TMEM26 in SVCs of iWAT	Decreased BW and iWAT index	Wang et al. (57)
	db/db mice (0.4% in chow diet, 10 wks)	↑ UCP-1, Cidea, PGC-1α, PPARα, PPARγ, and PRDM16 in iWAT	No difference in BW gain and food intake Increased glucose tolerance Decreased fasting blood glucose and insulin levels	Hui et al. (62)
	HFD-fed C57BL/6 mice (0.4% in diet or received feces from HFD-RVS-fed mice, 4 wks)	↑ UCP-1, PPARγ, PGC-1α, and SIRT1 in eWAT and iWAT	Decreased BW gain and WAT index No change in food intake Improved glucose homeostasis	Liao et al. (63)
	HFD-fed FVB/N mice (400mg/kg/day in diet, 8 wks)	↑ UCP-1, PRDM16, PGC-1α, and SIRT1 in subcutaneous WAT	Decreased WAT index Increased BAT index Decreased plasma TG and total cholesterol	Andrade et al. (75)
Quercetin	HFD-fed C57BL/6 J mice (0.1 % in diet, 12 wks)	↑ UCP-1 and Elovl3 in iWAT	Decreased plasma TG No difference in body composition, food intake, and energy expenditure	Kuipers et al. (85)
	HFD-fed C57BL/6 mice (0.5% in diet, 8 wks)	↑ UCP-1, PRDM16, Cidea, and PGC-1α in subcutaneous WAT and retroperitoneal WAT	No difference in BW and weights of eWAT and retroperitoneal WAT	Lee et al. (86)
GTC	HFD-fed SD rats (100 mg/kg/day, oral gavage, 30 or 45 days)	↑ UCP-1 (visceral WAT and subcutaneous WAT)	Decreased BW; No difference in food intake Decreased liver-to-BW ratios Decreased liver TG content	Yan et al. (91)
Epicatechin	HFD-fed Wistar rats (1 mg/kg daily, 2 wks)	↑ UCP-1, PGC-1α, SIRT1, and Dio2 in abdominal WAT	Decreased BW gain, blood glucose and TG, and energy intake	Gutiérrez-Salmeán et al. (92)
Curcumin	C57BL/6 mice (50 mg/kg/day, oral gavage, 50 days)	↑ UCP-1, PGC-1α, PRDM16, Dio2, PPARα, Cidea, Elovl3, Nrf1, mtTfα, and ATPsyn in iWAT	Decreased body and fat weights No difference in food intake Improved cold tolerance	Wang et al. (98)
ArtC	C57BL/6 J mice (10 mg/kg in diet, 4 wks)	↑ UCP-1 and PRDM16 in iWAT	No difference in BW gain, food intake, and weights of iWAT eWAT and iBAT	Nishikawa et al. (106)
Rutin	HFD-fed C57BL/6 J mice or db/db mice (1 mg/ml in drinking water, 10 wks)	↑ UCP-1, PGC-1α, PGC-1β, CPT1α, PPARα, CD137, and TBX in iWAT	Decreased BW and adiposity; No difference in food intake Increased energy expenditure Improved cold tolerance Improved glucose homeostasis	Yuan et al. (112)
Luteolin	HFD-fed C57BL/6 mice (0.01% in diet, 12 wks)	↑ UCP-1, PGC-1α, Elovl3, SIRT1, and Cited1 in iWAT	Decreased BW gain and fat mass Improved insulin sensitivity No difference in energy intake and body temperature Increased energy expenditure	Zhang et al. (118)
Myricetin	db/db mice (400 mg/kg/day, oral gavage, 14 wks)	↑ SIRT1 and PGC-1α in iWAT	Decreased BW, adiposity, insulin resistance and hepatic steatosis Improved plasma lipid profiles Increased energy expenditure Improved cold tolerance	Hu et al. (127)
Sudachitin	HFD-fed C57BL/6 J mice (5 mg/kg/day, oral gavage, 12 wks)	↑ UCP-1 and UCP-3 in iWAT	Decreased BW gain and fat mass; No difference in food intake Increased energy expenditure and insulin sensitivity	Tsutsumi et al. (129)
C3G	db/db mice (1 mg/ml in drinking water, 16 wks)	↑ UCP-1, Cidea, PGC-1α, PGC-1β, CPT1α, PPARα, TMTM26, CD 137, and Tbx1 in subcutaneous WAT	Decreased BW gain, adiposity and hepatic steatosis; No difference in food intake Increased energy expenditure, glucose tolerance and cold tolerance	You et al. (135)
Terpenoids
Phytol	C57BL/6 J mice (500 mg/kg by gavage every other day, 7 wks)	↑ UCP-1, PRDM16, PGC-1α, PDH, and Cyto C in iWAT	Decreased BW gain and iWAT index No difference in food intake	Zhang et al. (141)
Fucoxanthin	Wistar rats (2% in diet, 4 wks)	↑ UCP-1 in abdominal WAT	Decreased BW gain and WAT; No difference in food intake	Maeda et al. (147)
Gypenoside	HFD-fed C57BL/6 J mice (100 or 300 mg/kg in diet, 12 wks)	↑ UCP-1, PGC-1α, PRDM16, AMPKα, Cidea, Cox7α, and Elovl3 in iWAT	Decreased BW and fat No difference in food intake Improved insulin resistance	Liu et al. (155)
Menthol	HFD-fed C57BL/6 mice (1 % in diet, 12 wks)	↑ UCP-1, PGC-1α, PRDM16, β3-AR, and TRPM8 in iWAT and eWAT	Decreased BW and fat mass Improved glucose metabolism	Jiang et al. (166)
Celastrol	HFD-fed C57BL/6 mice (3 mg/kg/day in diet, 3 wks)	↑ PRDM16, Cidea, Elovl3, TMEM26, CD137, SP100, Tbx1, Scl27a1, and Hsp70 in iWAT	Decreased BW and fat mass Improved hepatic steatosis and insulin sensitivity	Ma et al. (172)
	HFD-fed C57BL/6 mice (100 mg/kg, i.p., 6 days)	↑ UCP-1 in iWAT	Decreased BW and fat mass Decreased food intake	Pfuhlmann et al. (173)
Alkaloids
Capsaicin	HFD-fed TRPV1 KO and control mice (0.01% in diet, 12 wks)	↑ UCP-1, PPARγ, BMP8β, PGC-1α, SIRT1/PPARγ/PRDM16 in iWAT and eWAT of WT mice	Decreased BW gain No difference in food intake Increased energy expenditure	Baskaran et al. (28)
	LACA mice (2 mg/kg, oral gavage, alternate days, 12 wks)	↑ UCP-1, PGC-1α, BDNF and Cideain iWAT	Increased locomotor activity	Baboota et al. (182)
Berberine	C57BL/6 mice (5 mg/kg/day, i.p., 4 wks)	↑ UCP-1, Dio2, and PRDM16 in WAT	Decreased BW and hepatic steatosis Increased energy expenditure Improved lipid profiles	Sun et al. (188)
	db/db mice (5 mg/kg/day, i.p., 4 wks)	↑ UCP-1, PGC-1α, Isdp5, Cidea, and COX8b in iWAT	Decreased BW and fat mass Increased insulin sensitivity, energy expenditure, and cold tolerance	Zhang et al. (193)

This study suggests that early use of RSV may exert long-term metabolic benefits by programming the body to a different metabolic state epigenetically. Indeed, the same group reported that RSV or nicotinamide treatment modifies methylation marks in PRDM16 and Slc27α1, two genes related to WAT browning, in the iWAT of mice (72). They also showed that RSV and nicotinamide directly induce the DNA methylation machinery to display more browning features in 3 T3-L1 adipocytes. Recently, Liu et al. (73) reported that RSV enhances browning of white adipocytes in vitro possibly by mediating the activation of mammalian target of rapamycin (mTOR). They showed that brown adipocyte-specific markers are decreased after treatment with rapamycin, an inhibitor of mTOR, in 3 T3-L1 adipocytes. On the other hand, treatment with MHY1485, an activator of mTOR, recapitulates the effect of RSV on browning markers in these cells. Another study suggest that RSV may alter lipid and oxidative metabolism in a SIRT1-dependent mechanism in 3 T3-L1 adipocytes (74). In this study, RSV treatment reduces triglyceride (TG) accumulation in 3 T3-L1 cells, and this reduction is abolished by inhibiting SIRT1 and PGC-1α. The study also showed that RSV treatment induces expression of PPARγ, PGC-1α, and carnitine palmitoyltransferase-1α (CPT-1α) in 3 T3-L1 adipocytes, all of which are abolished by SIRT1 inhibition (74). Interestingly, in a study evaluating the effect of RSV on expression of genes involved in thermogenesis, Andrade et al. showed that the HFD-fed male FVB/N mice treated with RSV (400 mg/kg/day) for 8 weeks, or obese human volunteers treated with trans-RSV (500 mg/day) for 4 weeks, displayed improved glycemic and lipid profiles along with increased mRNA levels of UCP1, PRDM16, PGC1α, SIRT1 and FNDC5 in the subcutaneous adipose tissue (75).

Quercetin

Quercetin (3,3′,4′,5,7-pentahydroxyflavone) is found in onions, apples, broccoli, and berries, and is one of the major flavonoids present in human diets (76). It has been reported that quercetin improves dyslipidemia and insulin sensitivity, reduces hypertension and BW gain, and at a high dose, lowers adipose and aortic inflammation in Zucker rats (77). In the HFD-fed mice, quercetin also improves insulin sensitivity and reduces BW gain and adipose inflammation (78, 79). Dong et al. showed quercetin attenuates mast cell and macrophage infiltration into epididymal WAT (eWAT) through the AMPKα-SIRT1 pathway in the HFD-fed mice (79). Kobori et al. (78) found that quercetin suppresses adipose infiltration of macrophages and lymphocytes in mice fed a Western diet high in fat. Obesity is often associated with a state of low-grade chronic adipose inflammation, which facilitates the development of insulin resistance and type 2 diabetes (80–83). Quercetin-induced improvement in insulin sensitivity may be linked to its suppression of inflammation in addition to the reduced weight gain.

Quercetin does not seem to reduce HFD-induced weight gain in mice by reducing food intake (79, 84, 85). Increased energy expenditure may play a part because quercetin-treated mice show increased UCP-1 expression in the BAT (79) as well as elevated mitochondrial oxidative phosphorylation in the WAT (78). Consistently, quercetin and quercetin-rich red onion extract have been demonstrated to increase energy expenditure in mice fed a HFD (84). In this study, authors further showed that both quercetin and the quercetin-rich red onion extract improve skeletal muscle mitochondrial number and function, though only quercetin but not the red onion extract increases physical activity. A couple of recent studies have shown that quercetin induces expression of genes related to adipose browning in the WAT of HFD-fed mice (Table I), though not affecting weight gain and energy expenditure (85, 86). Quercetin-induced adipose browning increases adipose uptake of TG-derived fatty acids, which may explain why quercetin lowers blood TG levels (85). In differentiated 3 T3-L1 adipocytes, quercetin at 100 μM increases expression of brown adipocyte-specific genes and CPT1α (Table II), which is associated with decreased lipid accumulation and increased multilocular lipid droplets in these adipocytes (86). The quercetin-induced browning of 3 T3-L1 adipocytes may be mediated in part by the activation of AMPK since treating these cells with 5 μM dorsomorphin, an AMPK inhibitor, attenuates quercetin-induced increases in mRNA levels of PRMD16, PGC-1α, CPT1α (a downstream target of PGC-1α), and SIRT1, though not UCP-1 (86) (Table II).

Table II.

Dietary compounds shown to induce browning in cultured adipocytes

Bioactive Compounds	Cells (treatment dosage)	Effects on browning-related genes	Ref.
Polyphenols
RSV	3T3-L1 adipocyte (20 μM)	↑ UCP1, PPARγ, and PGC-1α	Liu et al. (73)
	3T3-L1 adipocyte (50 μM)	↑ SIRT1, PPARγ, CPT1α, and PGC-1α	Imamura et al. (74)
Quercetin	3T3-L1 adipocyte (100 μM)	↑ CPT1α, SIRT1, and PGC-1α	Lee et al. (86)
Curcumin	Primary white adipocytes (2 μM)	↑ UCP-1, PPARα, and PGC-1α	Song et al. (100)
	3T3-L1 and primary white adipocytes (20 μM)	↑ UCP-1, PRDM16, and PGC-1α	Lone et al. (101)
ArtC	C3H10T1/2 cells (10 μM)	↑ UCP-1, Cidea, Cox8b, Elovl3, PPARγ, and PRDM16	Nishikawa et al. (106)
Rutin	C3H10T1/2 cells (10 μM)	↑ UCP-1, PRDM16, and PGC-1α	Yuan et al. (112)
Myricetin	C3H10T1/2 cells (10 μM)	↑ UCP-1 and PRDM16	Hu et al. (127)
Genistein	3T3-L1 adipocyte (100 μM)	↑ UCP-1 and CD137	Aziz et al. (214)
Butein	C3H10T1/2 cells (20 μM)	↑ UCP-1 and PRDM4	Song et al. (215)
Chrysin	3T3-L1 adipocyte (50 μM)	↑ UCP-1, Cidea, FGF21, PGC-1α, PRDM16, Tbx1, TMEM26, etc.	Choi et al. (216)
C3G	3T3-L1 adipocyte (100 μM)	↑ UCP-1, Tbx1, and CITED1	Matsukawa et al. (217)
Terpenoids
Phytol	3T3-L1 adipocyte (100 μM)	↑ UCP-1, PRDM16, PGC-1α, Cidea, CD137, TMEM26, etc.	Zhang et al. (141)
Ginsenoside Rb1	3T3-L1 adipocyte (10 μM)	↑ UCP-1, PGC-1α, and PRDM16	Mu et al. (156)
Thymol	3T3-L1 adipocyte (20 μM)	↑ UCP-1, PPARγ, PPARδ, pAMPK, pACC, HSL, CPT1α, PGC-1α, etc.	Choi et al. (218)
Alkaloids
Capsaicin	3T3-L1 adipocyte (1 μM)	↑ UCP-1, PGC-1α, SIRT1, PRDM16, Dio2, Cidea, and PPARα	Baboota et al. (182)

It is currently unknown why quercetin reduces BW gain in obese rodents in some but not all studies, despite similar doses of quercetin are used. This discrepancy may be addressed by using a standardized treatment regimen under identical experimental conditions including animal housing environment and diet sources.

Green Tea Catechins

Green tea catechins (GTC) are polyphenolic compounds present in the unfermented dried leaves of the plant Camellia sinensis. Epigallocatechin-3-gallate is the most abundant catechin of green tea, representing 50–80% of the total catechin content, and also considered to be the most bioactive component of green tea (87). Other minor catechins include epicatechin-3-gallate, epigallocatechin, epicatechin, and catechin (87). Green tea has anti-inflammatory, anti-atherosclerotic, and anti-cancer properties (88). During recent years, a growing number of studies have confirmed the beneficial effects of green tea on obesity, glucose homeostasis, and fat mass gain in HFD-induced obese rodent models (89, 90).

A potential mechanism by which GTCs induce anti-obesity effects may relate to GTCs’ effects on energy metabolism and adipose browning. For example, oral gavage of GTC at 100 mg/kg/day for 30 or 45 days significantly reduces BW gain, liver weight, and blood TG levels in male Sprague-Dawley rats fed a HFD (91). The authors suggested that GTCs exert their anti-obesity effects, in part, by modulating PPAR signaling pathways. They found that GTCs increase PPARγ protein levels in the subcutaneous WAT, decrease PPARγ protein levels in the visceral WAT, and increase PPARδ protein levels in both BAT and WAT along with increased mRNA expression of genes involved in fatty acid oxidation such as PPARδ target genes including alternative oxidase, CPT1α, and UCP-1 in the BAT. The expression of UCP-1 mRNA is also increased in the subcutaneous WAT and visceral WAT in the chow or HFD-fed rats treated with GTCs, suggesting enhanced browning of these adipose tissues upon GTC treatment (Table I). In another study, (−)-Epicatechin (EPI), a component of green tea catechins (92), seems to increase adipose and skeletal muscle expression of UCP-1 and iodothyronine deiodinase 2 (Dio2), which are two adipose browning markers (Table I). In this study, male Wistar rats were fed a HFD for 5 weeks, followed by a 2-week EPI administration by gavage (1 mg/kg/day) under the same dietary condition. It was found that EPI treatment reduces BW gain, hyperglycemia, and blood TG levels without affecting food intake or nutrient absorption. Because protein expression levels of UCP-1, PGC-1α, SIRT1, and Dio2 are increased in the abdominal adipose tissue, the authors suggested that increased energy expenditure may have contributed to the decreased weight gain in the EPI-treated animals (Table I). A concern for this study is the basal level of UCP-1, which is not normally expressed at a significant level in the WAT and skeletal muscle. Authors detected abundant UCP-1 protein expression in these tissues under the basal condition. It is unclear if this is specific to male Wistar rats and/or the experimental environment.

In accordance with animal studies, Nirengi et al. (93) found that ingestion of a catechin-rich beverage at 540 mg catechin/day for 12 weeks significantly increased BAT density (indicative of BAT activation) and decreased extramyocellular lipid content in healthy young women. It would be interesting to determine whether this drink promotes white fat browning.

Curcumin

Curcumin, a bioactive polyphenol component derived from the turmeric rhizomes, has been investigated in prevention of obesity, insulin resistance, diabetes and dyslipidemia in mice, rabbits and rats (94–96). Curcumin may exert its metabolic benefits by regulating lipid metabolism, increasing antioxidant enzyme expression, reducing inflammatory cytokine production, and activating fatty acid oxidation (97).

Wang et al. (98) showed that curcumin administration at 50 or 100 mg/kg/day for 50 days reduces BW gain and fat mass without affecting food intake in male C57BL/6 mice fed a chow diet. These fat and BW gain changes are associated with higher body temperatures during 6 h of cold exposure (98). Consistent with body temperature changes, the animals display increased expression of many brown fat-specific genes in the iWAT, including mRNA and protein for UCP-1 and mRNAs for PGC-1α, PRDM16, Dio2, PPARα, Cidea, Elovl3, nuclear respiratory factor 1, mitochondrial transcription factor A (Tfam), and ATP synthase (Table I). The study further showed that curcumin-treated mice have elevated blood norepinephrine (NE) concentrations and increased mRNA for β−3 adrenergic receptor (β3-AR) in the iWAT. This led the authors to suggest that curcumin induces adipose browning via the NE-β3-AR pathway. Typically, it is the locally released NE from the sympathetic nerves innervating adipose tissues, not the circulating NE, that serves as the major mediator of adipose thermogenesis during cold exposure (4, 99). Increased circulating NE levels after curcumin treatment imply a role of curcumin in stimulating NE release from adrenal glands. It would be interesting to examine whether some of the cardio-metabolic effects of curcumin are mediated by adrenal NE and epinephrine. Another study by Song et al. (100) reported that male C57BL/6 mice fed a HFD containing 1% curcumin for 18 weeks have increased energy expenditure and body temperature in response to cold exposure, which is associated with increased expression of UCP-1, PRDM16 and PPARα in the BAT but not WAT. In rat primary adipocytes, these researchers observed that the increased mRNA expression of UCP-1 cannot be completely blocked by a PPARα or PPARγ antagonist, suggesting that curcumin may increase energy expenditure partially via PPARα/PPARγ-independent mechanisms (Table II). In 3 T3-L1 and primary white adipocytes, Lone et al. (101) observed that curcumin induces brown fat phenotypes via the AMPK pathway. Curcumin treatments (1, 10, or 20 μM) increase expression of UCP-1 and other brown adipocyte-specific markers (PGC-1α and PPARγ) in a dose-dependent manner in these cells. Inhibition of AMPK by dorsomorphin abolishes expression of UCP-1, PRDM16 and PGC-1α while the AMPK activator AICAR does the opposite (Table II).

Artepillin C

Artepillin C (3,5-diprenyl-4-hydroxycinnamic acid) (ArtC) is a major component of Brazilian green propolis that is collected from the plant Baccharis dracunculiforia (102). Several studies have documented potential roles of ArtC in oxidative stress prevention, immunoregulation, chemoprevention, and cancer treatment (103–105). Nishikawa et al. (106) showed that ArtC treatment for 4 weeks at 10 mg/kg/day induces appearance of brown-like adipocytes and expression of UCP-1 and PRDM16 proteins in the iWAT of male C57BL/6 J mice (Table I). Unlike curcumin, ArtC-induced browning of WAT is not associated with significant changes in plasma NE concentrations and adipose β3-AR mRNA levels (106). ArtC seems to promote adipose browning in a cell autonomous manner because ArtC at 10 μM induces expression of mRNAs for UCP-1, Cidea, cytochrome c oxidase subunit 8B (Cox8b), and Elovl3 in murine C3H10T1/2 cells and primary iWAT-derived adipocytes (Table II), and this induction largely depends on PPARγ activation and PRDM16 protein stability.

Rutin

Rutin (3,3,4,5,7-pentahydroxyflavone-3-rhamnoglucoside) is a polyphenolic bioflavonoid from various plants and fruits, such as buckwheat, apricots, oranges, lemons, grapes, limes, berries, and peaches (107, 108). Pharmacological studies have shown that rutin has antioxidative, anti-inflammatory, and antidiabetic properties, suggesting that rutin may have therapeutic potentials in many diseases (109–111). Yuan et al. (112) reported that in both genetically obese db/db male mice and the HFD-fed C57BL/6 J male mice, rutin treatment in the drinking water (1 mg/ml) for 10 weeks attenuates weight gain and adiposity, increases energy expenditure, and improves glucose homeostasis. This occurs without changes in food intake, water intake, and physical activity. Interestingly, rutin-treated animals versus controls display a significantly higher mean core body temperature during cold (4°C) exposure for 4 h. The expression of mRNAs for several BAT-specific genes, including UCP-1, PGC-1α, PGC-1β, CPT1α, and PPARα, as well as beige cell markers, such as CD137 and TBX, are significantly increased in the subcutaneous WAT after rutin treatment (Table I). In this study, rutin (10 μM) significantly increases expression of UCP-1 and other thermogenic genes, such as PRDM16 and PGC-1α in C3H10T1/2 cells (Table II). Mechanistic studies suggest that rutin directly binds and stabilizes SIRT1, leading to hypoacetylation of PGC-1α protein and Tfam transactivation. Activation of this SIRT1-PGC-1α-Tfam pathway in heat-generating adipocytes may contribute to rutin-induced adipose browning and energy expenditure.

Luteolin

Luteolin (3′,4′,5,7-tetrahydroxyflavone) is a flavone present in fruits, vegetables, and several medicinal herbs such as peppermint, thyme, and parsley (113). Luteolin has antioxidant, anti-inflammatory, antimicrobial, and anticancer effects (113). Previous animal studies have shown that luteolin protects against high glucose-induced endothelial dysfunction as well as diet-induced hepatic steatosis and insulin resistance (114–117). Zhang et al. (118) found that male C57BL/6 mice fed a HFD supplemented with 0.01% luteolin for 12 weeks are protected against HFD-induced weight gain, body fat accumulation, and insulin resistance. These animals show no changes in energy intake and body temperature, but increases in energy expenditure, respiratory exchange ratio, and expression of thermogenic genes in BAT and subcutaneous WAT tissues (118) (Table I). Compared to the mice fed the HFD only, the luteolin-treated mice have more brown-like adipocytes and UCP-1 expression in the subcutaneous WAT. In addition, the authors observed that luteolin treatment significantly increases expression of many thermogenic genes in differentiated primary brown and subcutaneous adipocytes, which is associated with increased phosphorylation of both AMPKα and acetyl-CoA carboxylase. The AMPK inhibitor Compound C (20 μM) can reverse the effects of luteolin on thermogenic gene expression, suggesting that AMPK may mediate some of luteolin’s metabolic benefits.

Myricetin

Myricetin (3,5,7,3′,4′,5′-hexahydroxyflavone cannabiscetin) is a natural flavonoid found in wine, onions, bayberries, grapes, and tea, among others (119, 120). It has been reported that myricetin possesses various pharmacological actions, including antioxidative, anti-inflammatory, and anticancer activities (121–123). In rats, myricetin can improve glucose utilization, increase insulin sensitivity, and reduce hyperglycemia associated with diet feeding and diabetes (124–126). Similar effects were also observed in mice. Hu et al. (127) reported that diabetic db/db mice orally gavaged with myricetin at 400 mg/kg/day in distilled water for 14 weeks starting at 4 weeks of age slows down weight and body fat gain and improves plasma lipid profiles and insulin sensitivity. This is associated with increased thermogenic protein expression and mitochondrial biogenesis in both BAT and iWAT, resulting in augmented energy expenditure and cold resistance (Table I). During differentiation of murine C3H10T1/2 mesenchymal stem cells to brown adipocytes, myricetin treatment at 10 μM increases mRNA expression and secretion of adiponectin (127) (Table II). Myricetin treatment also increases adiponectin mRNA levels in BAT, eWAT, and iWAT as well as plasma concentrations of adiponectin in the db/db mice. This finding led the authors to suggest that myricetin may exert its metabolic benefits by directly regulating adiponectin expression in adipocytes.

Sudachitin

Sudachitin (5,7,4′-trihydroxy-6,8,3-trimethoxyflavone) is a polymethoxyflavone found in the peel of Citrus sudachin. In addition to its anti-inflammatory effect (128), sudachitin has been shown to possess anti-obesity action (129). Using male C57BL/6 J mice fed a HFD and diabetic db/db mice fed a normal diet, Tsutsumi et al. showed that sudachitin treatment at 5 mg/kg/day for 12 weeks prevents HFD-induced weight gain, body fat deposition and adipocyte hypertrophy without altering food intake. (129). In addition, sudachitin treatment improves insulin sensitivity in the HFD-fed C57BL/6 J mice and in the chow-fed db/db mice. Interestingly, sudachitin supplementation to the HFD significantly increases UCP-1 mRNA levels in the subcutaneous WAT in addition to increasing mitochondrial biogenesis and fatty acid oxidation in the skeletal muscle, implying that sudachitin may protect overnutrition-associated metabolic disorders in part by promoting WAT browning (Table I).

Cyanidin-3-O-β-Glucoside

Cyanidin-3-O-β-glucoside (C₂₁H₂₁O₁₁⁺) (C3G) is a component in blueberries and one of the most widely distributed anthocyanins in fruits and vegetables (130, 131). C3G seems to have potent anti-obesity and anti-diabetes properties (132–134). You et al. (135) reported that obese male db/db mice treated with C3G in drinking water (1 mg/mL) for 16 weeks display reduced BW gain, hepatic steatosis, serum lipids, and insulin resistance. The animals also have increased energy expenditure, cold tolerance, and BAT activity. These effects of C3G are not associated with significant differences in food or water intake, energy absorption, or physical activity, but with enhanced white fat browning as evidenced by increased expression of genes related to thermogenesis, fatty acid oxidation and beigeing, including UCP-1, Cidea, PGC-1α, PGC-1β, CPT1α, PPARα, TMEM26, CD137, and Tbx1, in the subcutaneous WAT (Table I). C3G treatment also increases mitochondrial number and function in both BAT and subcutaneous WAT, suggest that C3G may induce adipose browning by promoting mitochondrial biogenesis.

Terpenoids

Phytol

Phytol (3,7,11,15-tetramethylhexaded-2-en-1-ol) is a plastidial isoprenoid found in plant foods such as fruits, vegetables, and grains. It is abundant in nature as a part of the chlorophyll molecule (136, 137). Animal studies suggest that phytol may have anti-inflammatory and anti-oxidative effects (138). A study reported that phytol alters fatty acid metabolism and decreases plasma fatty acid concentrations by activating retinoid-X-receptor and PPARs in mice (139, 140). Zhang et al. (141) reported that 7 weeks of phytol treatment at 500 mg/kg every other day by oral gavage reduces weight gain without affecting the food intake in male C57BL/6 J mice fed a HFD. This treatment induces iWAT browning as evidenced by increased multilocular adipocytes and thermogenic gene expression in this fat depot (Table I). The results from in vitro cell culture studies also support the role of phytol in stimulating adipocyte browning. Phytol at 100 μM stimulates browning or the formation of brown-like adipocytes in the differentiated 3 T3-L1 cells, which is associated with activation of UCP-1 gene promoter activity and upregulated expression of brown and beige adipocyte marker genes, including PRDM16, PGC-1α, Cidea, Elovl3, CD137 and TMEM26 (141) (Table II). In this study, phytol was also shown to activate AMPKα in adipose tissues and cultured 3 T3-L1 adipocytes. Using the AMPK inhibitor Compound C (100 μM), the authors demonstrated that the AMPKα signaling pathway is required for phytol to stimulate brown adipogenic differentiation.

Fucoxanthin

Fucoxanthin (C₄₂H₅₈O₆) is a marine carotenoid. It can be found in edible brown seaweeds, such as Undaria pinnatifida (Wakame), Saccharina japonica (Makonbu), and Sargassum fulvellum (Hondawara) (142). Fucoxanthin has antioxidant, anti-inflammatory, and anticancer activities (143, 144). It has attracted wide attention because of its anti-obesity and anti-diabetic effects (145, 146). Maeda et al. (147) observed that male Wistar rats and obese female KK-Ay mice fed 2% Undaria lipids have reduced weight gain and fat weight, but no changes in daily food intake compared to the control diets. KK-Ay mice fed the diet containing 2% Undaria lipids show increased BAT weight than those on the control diets. Interestingly, fucoxanthin-containing diets significantly increase UCP-1 protein and mRNA expression in the WAT composed of perirenal, gonadal, retroperitoneal and mesenteric adipose tissues, though not in the BAT (147) (Table I). It would be interesting to determine fat depot-specific effect of fucoxanthin given the distinct function of each fat depot in energy metabolism. Nonetheless, this finding suggests that WAT browning may play a role in the anti-obesity activity of seaweed carotenoid fucoxanthin in obese mice.

In contrast with the animal studies, Rebello et al. (148) reported that human adipocytes treated with fucoxanthin or fucoxanthinol at different concentrations (1 μM, 0.1 μM, 0.01 μM, 1 nM, 0.1 nM) showed no changes in the oxygen consumption rate (OCR) or expression levels of mRNAs for UCP1, CPT-1β, and GLUT4. In addition, no significant changes were observed for the mRNA levels of PGC-1α, PPARα, PPARγ, PDK4, FAS, and the lipolytic enzymes. The authors concluded that neither fucoxanthin nor its metabolite fucoxanthinol stimulates human white adipocyte browning at the dose that is expected to exist in the human plasma without being toxic to the cells. Further studies are needed to address the discrepancy in the role of fucoxanthin in WAT browning between animals and humans.

Gypenosides

Gypenosides (C₄₇H₇₆O₁₈) are triterpenoid saponins isolated from the Gynostemma pentaphyllum plant. It may have potential benefits in inflammation, cardiovascular diseases, cancer, diabetes, and obesity-associated metabolic disorders (149–154). Liu et al. (155) showed that treatment with gypenosides at 300 mg/kg BW/day for 12 weeks reduces weight gain, fat weight, and insulin resistance in the HFD-fed male C57BL/6 J mice. These effects are associated with increased protein and mRNA expression of UCP-1, PGC-1α, and PRDM16 as well as increased mRNAs for Cidea, Cox7α, and Elovl3 in the BAT and iWAT (Table I), suggesting an adipose browning-inducing role of gypenosides in mice. In addition, this study showed that gypenosides substantially alter the gut microbiome composition with a significant increase in Akkermansia muciniphila (A. muciniphila). It was proposed that gypenoside may exert metabolic benefits in part by stimulating adipose browning and regulating the gut microbiota. Mu et al. (156) reported that ginsenoside Rb1 (10 μM) induces brown-specific genes partially through induction of PPARγ in 3 T3-L1 mature adipocytes and this effect can be inhibited by the PPARγ antagonist GW9662, suggesting that ginsenoside Rb1 may induce adipose browning at least in part by activating PPARγ (Table II).

Menthol

Menthol (C₁₀H₂₀O) is rich in cornmint or peppermint oil. It is added to foods, mouthwashes, toothpastes, drugs, among others as a cooling and flavoring agent. The cooling effect of menthol is attributable to its action as an agonist of transient receptor potential melastatin 8 (TRPM8), an ion channel that detects cold stimuli in the thermosensory system (157–159). Mice deficient in the menthol receptor TRPM8 display severely attenuated sensation of cold temperature (158, 160–162). Recent studies suggest that menthol may be used as an anti-obesity compound due to its thermogenic property (163, 164). Ma et al. (163) observed that menthol increases UCP-1 levels in murine primary brown adipocytes in a TRPM8-dependent manner and critically dependent on the Ca²⁺-dependent PKA phosphorylation. They also observed that dietary supplementation of menthol (0.5%, w/w) for 28 weeks increases the locomotor activities and core temperatures in the HFD-fed wild-type, but not TRPM8-deficient mice (163). Rossato et al. (165) showed that activation of TRPM8 by menthol dose-dependently increases UCP-1 expression in human white adipocytes, leading to increased heat production, though no changes in expression of the genes regulating mitochondrial biogenesis (165). In addition, Jiang et al. (166) showed that menthol (100 μM) increases expression of thermogenic genes such as UCP-1 and PGC-1α in cultured mouse white adipocytes, which can be inhibited by BAPTA-AM (10 μM), a membrane-permeable calcium chelator, or KT5720 (10 μM), a specific protein kinase A (PKA) inhibitor. This study suggests that menthol may induce UCP-1 and PGC-1α expression by increasing intracellular Ca²⁺ concentration and PKA phosphorylation in white adipocytes. Consistently, dietary menthol treatment at 1% (w/w) for 12 weeks attenuates HFD-induced weight gain in male C57BL/6 mice (166). The reduction in weight gain is accompanied by enhanced browning and reduced adipocyte size in the subcutaneous WAT, and by improved glucose tolerance and insulin sensitivity (166). Expression levels of thermogenic genes (UCP-1, PGC-1α, PRDM16, β3-AR) and TRPM8 are significantly increased in the subcutaneous WAT and eWAT of these menthol-treated mice (Table I). The authors concluded that TRPM8 may be involved in menthol-induced WAT browning by increasing expression of genes related to thermogenesis and energy metabolism (166).

Celastrol

Celastrol (C₂₉H₃₈O₄) is a major active component in Chinese herbal medicine Tripterygium wilfordii (‘Thunder of God Vine’) (167, 168). It has diverse biological properties, such as anti-cancer, anti-obesity, anti-diabetic, and neuroprotective (169). Celastrol’s potent anti-obesity effect has been reproduced in several studies, though the underlying mechanism has yet to be settled (170–173). One study showed that celastrol suppresses food intake and increases energy expenditure by improving leptin sensitivity, leading up to 45% weight loss in hyperleptinemic diet-induced obese mice (170). Another study showed that celastrol treatment at 3 mg/kg/day for 3 weeks reduces weight gain, fat mass, hepatic steatosis and insulin resistance without affecting food intake, locomotor activity, or respiratory exchange ratio (RER) in HFD-fed mice (172). In this study, celastrol treatment increases brown (PRDM16, UCP-1, Cidea, and Elovl3) and beige (TMEM26, CD137, SP100, Tbx1, Scl27a1, and Hsp70) selective genes in the iWAT (Table I). The induction of these thermogenic genes requires heat shock factor 1 (HSF1) and PGC-1α because it is blunted in cells derived from the iWAT of HSF1-or PGC-1α-null mice (172). However, Pfuhlmann et al. (173) showed that celastrol decreases BW independently from UCP-1 and has no effect on energy expenditure. They observed that celastrol treatment at 100 mg/kg/day for 6 days increases expression of UCP-1 in the BAT and iWAT of HFD-fed mice (Table I), but transcription of PGC-1α and key thermogenic genes (e.g., PRDM16 and Cidea) is unaffected in these tissues, and there are comparable BW loss and hypophagia between UCP-1 KO and WT mice treated with celastrol. This observation implies that UCP-1-mediated thermogenesis is not a major driver for the BW-lowering effect of celastrol. Although celastrol-induced weight loss may not depend on UCP-1, one cannot exclude its dependence on thermogenic activation because UCP1-independent thermogenic programs exist in adipose and other tissues (18, 19, 174, 175). Celastrol was shown to activate HFS1 and PGC-1α-dependent thermogenic program (172). Nur77 was identified as a critical intracellular target for celastrol, and the binding of celastrol to Nur77 promotes Nur7 translocation from the nucleus to mitochondria, helping clearance of inflamed mitochondria to suppress inflammation (171). These studies indicate that celastrol may have a profound impact on mitochondrial function and dynamics, thereby critically influencing energy metabolism.

Alkaloids

Capsaicin

Capsaicin (C₁₈H₂₇NO₃) is an alkaloid compound found in pepper, which belongs to the genus Capsicum (176). It has antioxidant and anti-cancer properties (177, 178). In addition, capsaicin has been shown to have anti-obesity effects by mechanisms that include promotion of adipose lipolysis and energy expenditure (179, 180). Oral administration of capsaicin at 2 mg/kg/day was shown to alter gut microbial compositions, increase hypothalamic satiety, and induce adipose browning in male Swiss albino mice fed a HFD (181). The browning effect is supported by increased expression of genes related to thermogenesis and mitochondrial biogenesis including UCP-1, PGC-1α, PPARα, Cidea, and brain-derived neurotrophic factor (BDNF) in the BAT and subcutaneous WAT (Table I). Because the pair feeding was used in the study, the authors suggested that the induction of adipose browning may contribute in part to the reduced weight gain in the capsaicin-treated animals. In differentiated 3 T3-L1 adipocytes, Baboota et al. (182) reported that low dose (1 μM) capsaicin induces browning-specific gene expression, an effect similar to that induced by resiniferatoxin (RTX), an agonist of TRPV1. The genes induced include PPARγ and its downstream targets as well as brown fat cell markers UCP-1, PGC-1α, PRDM16, Dio2, PPARα, and FOXC2 (Table II). The study also showed that expression of UCP-1, PGC-1α, BDNF, and Cidea are significantly upregulated in the subcutaneous WAT of male Wistar rats upon oral administration of capsaicin at 2 mg/kg BW on alternate days for 3 months (Table I). Baskaran et al. (28) analyzed the effects of dietary capsaicin (0.01%) on the HFD-induced obesity in male WT and TRPV1 KO mice. They found that dietary capsaicin suppresses HFD-induced weight gain in WT, but not TRPV1 KO mice by stimulating heat production and physical activity (28). Expression of PGC-1α, SIRT1, and PRDM16 are increased in the eWAT and iWAT of WT but not TRPV1 KO mice (Table I). In addition, there is an induction of UCP-1 in the eWAT and an increase of UCP-1 and BMP8β in the iWAT in the capsaicin-treated animals. The authors suggested that capsaicin may stimulate SIRT1-dependent deacetylation of PPARγ and PRDM16, thereby facilitating PPARγ and PRDM16 interaction to induce adipose browning via a TRPV1-dependent mechanism.

Berberine

Berberine (C₂₀H₁₈NO₄⁺), a naturally occurring alkaloid, is found in certain species of flowering plants like Berberidaccae, Coptis rhizomes and Hydrastis Canadensis that are used in traditional Chinese medicine (183). Many studies have shown that berberine has a variety of benefits in health, such as anti-inflammatory, anti-cancer, hypolipidemic, hypoglycemic, and insulin-sensitizing effects (184, 185). Multiple mechanisms may have been implicated in berberine’s health and metabolic benefits, such as activation of AMPK and SIRT1 (186–188), up-regulation of LDL receptor (189), and modulation of gut hormones and microbiota (190–192).

Studies have suggested a role of berberine in adipose browning. Sun et al. (188) reported that the beneficial effects of berberine (5 mg/kg/day, i.p., 4 weeks) on weight gain and hepatic steatosis in high-fat high-fructose diet-fed C57BL/6 mice require hepatic SIRT1 because liver-specific SIRT1 deletion abolishes these benefits. Berberine treatment increases FGF21 production from liver and primary hepatocytes and induces thermogenic gene expression in white and brown adipose tissues in mice (188). These effects also require SIRT1 and are associated with increased whole-body energy expenditure. Consistently, Zhang et al. (193) reported that male db/db mice treated with berberine (5 mg/kg/day, i.p.) have increased energy expenditure, reduced weight gain, and improved insulin sensitivity without changes in physical activity. The expression of UCP-1 and other classical BAT marker genes, such as Cidea and cytochrome c oxidase subunit 8b, are induced in the BAT and iWAT in these mice. Cell culture studies suggest that berberine upregulates UCP-1 transcription by activating the AMPK-PGC-1α pathway (193). In addition to activating AMPK/PGC-1α, long-term treatment with berberine increases expression of tyrosine hydroxylase in both BAT and iWAT in WT and db/db mice, suggesting that berberine may augment the sympathetic outflow to adipose tissues, which may contribute in part to berberine’s adipose browning effect. Interestingly, berberine has been shown to promote recruitment and activation of brown fat-like tissues in mildly overweight subjects with non-alcoholic fatty liver disease (194). This study also showed that berberine promotes brown adipogenic program in chow-fed mice, HFD-fed mice, and cultured mouse and human preadipocytes. Moreover, the study demonstrated that the berberine-induced adipose browning and thermogenic program depend on adipose AMPK because berberine has no such effects in adipose AMPKα1 and AMPKα2 KO mice. These findings collectively highlight an important role of the AMPK-dependent adipose browning in mediating berberine’s metabolic benefits.

In a human study, Wu et al. (194) examined the effect of berberine intervention to combat overweight. They found that mildly overweight individuals with NAFLD treated with berberine (500 mg, 3 times per day) for 1 month have increased BAT volume and activity. It is currently unknown whether berberine promotes WAT browning in humans.

CONCLUDING REMARKS

In this review, we focus on natural bioactive compounds as potential activators of WAT browning or beiging in preclinical animal models and cell culture systems. Current findings from animal and cell culture studies suggest that many compounds may possess such an activity. Several mechanisms have been suggested to mediate bioactive compounds-associated adipose browning, such as activation of AMPK, SIRT1, and PGC-1α pathways (Fig. 1). These pathways are well known to regulate energy metabolism and promote metabolic benefits. Humans have brown fat-like or beige adipose tissues. Since these thermogenic tissues dissipate metabolic energy as heat, their recruitment and activation by bioactive compounds may help combat obesity and related metabolic diseases in humans. Generally, naturally occurring compounds hold a special interest due to their better acceptability than pharmacological drugs and invasive procedures of obesity and metabolic disorders. Unfortunately, most bioactive compounds and their metabolites have low bioavailability (195–205), which limits their bioefficacy in animals and humans. For example, resveratrol, the most widely studied nonflavonoid polyphenol, has shown many metabolic benefits in in vitro and in vivo preclinical studies, but the low bioavailability of resveratrol and its metabolites limits their applications in humans (70, 206–209). Similarly, many flavonoids have poor bioavailability despite their potential health benefits (210, 211). Substantial efforts have been made to improve the bioavailability of resveratrol and some flavonoids and the outcomes are encouraging (212, 213). Similar efforts should be made to improve the bioavailability of each natural bioactive compound because it can proceed to large scale clinical studies, which are scarce, particularly those focusing on adipose browning and associated metabolic outcomes.

Fig. 1.

A schematic of potential mechanisms for natural agent-induced adipose browning. Many natural bioactive compounds appear to induce UCP-1-mediated adipose browning via several signaling pathways known to promote UCP-1 transcription and mitochondrial oxidation in WAT. The signaling pathways involved in the up-regulation of UCP-1 and its transcriptional regulators are shown. Some bioactive compounds appear to induce adipose browning by activating the AMPK/SIRT1/PGC-1α pathway directly or indirectly. Activation of AMPK can promote mitochondrial bioenergetics by turning on PGC-1α transcription, directly phosphorylating PGC-1α, or activating PGC-1α in a SIRT1-dependent manner. SIRT1 activates PGC-1α through deacetylation. The transcription coregulator PRDM16 can interact with PGC-1α and PPARα/γ to promote UCP-1 expression. Some bioactive compounds likely induce adipose browning by stimulating TRPM8, β3-AR receptor, or TGR5, which can lead to the activation of multiple PKA-dependent signaling pathways, including those through p38 MARP, CREB, and T3. Ac, acetylation; AC, adenylyl cyclase; AMPK, 5’ AMP-activated protein kinase; ArtC, artepillin C; ATF2, activating transcription factor 2; β3-AR, beta-3 adrenergic receptor; C3G, cyanidin-3-O-β-glucoside; CREB, cAMP-response element binding protein; Dio2, iodothyronine deiodinase 2; EPI, (−)-epicatechin; LCA, lithocholic acid; P, phosphorylation; PGC-1α, peroxisome proliferator-activated receptor γ (PPARγ) co-activator-1α; PKA, protein kinase A; PKC, protein kinase C; PRDM16, PR domain zinc finger protein 16; RSV, resveratrol; SIRT1, sirtuin-1; TGR5, Takeda G protein-coupled receptor 5; TR, thyroid hormone receptor; TRPM8, transient receptor potential cation channel subfamily M member 8; TRPV1, transient receptor potential cation channel subfamily V member 1; UCP-1, uncoupling protein-1.

In addition to an urgent need to solve the pool bioavailability problem of most bioactive food compounds, other issues are also noted in the literature. 1) the majority of the studies use male mice or rats, making the results largely limited to one gender; 2) the physiological relevance of the doses applied in the studies needs to be carefully tested to avoid unwanted side or toxic effects of a compound; 3) lack of direct assessment of the effect of adipose browning on metabolic benefits such as weight loss and insulin sensitivity; 4) lack of studies with bioactive browning agents during aging; 5) scarcity of solid evidence supporting the direct involvement of a pathway in the mediation of browning effects; and 6) targeted tissue delivery and bioavailability, and importantly toxicity or safety of a compound need to be established before performing comprehensive, large-scale and randomized clinical trials in humans. Use of natural products is a promising route for obesity management. With more research, specific natural compounds may emerge as effective and safe agents for treatment and prevention of obesity and associated metabolic sequelae.

ABBREVIATIONS

AMPK

5’ AMP-activated protein kinase

ArtC

Artepillin C

BAT

Brown adipose tissue

BDNF

Brain-derived neurotrophic factor

BMPs

Bone morphogenetic proteins

C3G

Cyanidin-3-O-β-glucoside

CPT1α

Carnitine Palmitoyltransferase 1α

Dio2

Iodothyronine Deiodinase 2

Elovl3

ELOVL Fatty Acid Elongase 3

EPI

(−)-Epicatechin

eWAT

Epididymal white adipose tissue

FGF21

Fibroblast growth factor 21

GTC

Green tea catechins

HFD

High-fat diet

iWAT

Inguinal subcutaneous WAT

LCA

Lithocholic acid

mTOR

Mammalian target of rapamycin

NE

Norepinephrine

PDEs

Phosphodiesterases

PGC-1α

PPARγ co-activator-1α

PKA

Protein kinase A

PPARγ

Peroxisome proliferator-activated receptor γ

PRDM16

PR domain zinc finger protein 16

RSV

Resveratrol

SIRT1

Sirtuin-1

SVCs

Stromal vascular cells

Tbx1

T-Box Transcription Factor 1

Tfam

Mitochondrial transcription factor A

TG

Triglyceride

TMEM26

Transmembrane protein 26

TRPM8

Transient Receptor Potential Cation Channel Subfamily M Member 8

TRPV1

Transient Receptor Potential Cation Channel Subfamily V Member 1

UCP-1

Uncoupling protein-1

WAT

White adipose tissue

β3-AR

Beta-3 adrenergic receptor