Adipose-tissue plasticity in health and disease.

Adipose tissue, colloquially known as "fat," is an extraordinarily flexible and heterogeneous organ. While historically viewed as a passive site for energy storage, we now appreciate that adipose tissue regulates many aspects of whole-body physiology, including food intake, maintenance of energy levels, insulin sensitivity, body temperature, and immune responses. A crucial property of adipose tissue is its high degree of plasticity. Physiologic stimuli induce dramatic alterations in adipose-tissue metabolism, structure, and phenotype to meet the needs of the organism. Limitations to this plasticity cause diminished or aberrant responses to physiologic cues and drive the progression of cardiometabolic disease along with other pathological consequences of obesity.

White adipocyte dysfunction and obesity-associated pathologies in humans.

The prevalence of obesity and associated chronic diseases continues to increase worldwide, negatively impacting on societies and economies. Whereas the association between excess body weight and increased risk for developing a multitude of diseases is well established, the initiating mechanisms by which weight gain impairs our metabolic health remain surprisingly contested. In order to better address the myriad of disease states associated with obesity, it is essential to understand adipose tissue dysfunction and develop strategies for reinforcing adipocyte health. In this Review we outline the diverse physiological functions and pathological roles of human white adipocytes, examining our current knowledge of why white adipocytes are vital for systemic metabolic control, yet poorly adapted to our current obesogenic environment.

ZFP423 controls EBF2 coactivator recruitment and PPARγ occupancy to determine the thermogenic plasticity of adipocytes.

Energy-storing white adipocytes maintain their identity by suppressing the energy-burning thermogenic gene program of brown and beige adipocytes. Here, we reveal that the protein-protein interaction between the transcriptional coregulator ZFP423 and brown fat determination factor EBF2 is essential for restraining the thermogenic phenotype of white adipose tissue (WAT). Disruption of the ZFP423-EBF2 protein interaction through CRISPR-Cas9 gene editing triggers widespread "browning" of WAT in adult mice. Mechanistically, ZFP423 recruits the NuRD corepressor complex to EBF2-bound thermogenic gene enhancers. Loss of adipocyte Zfp423 induces an EBF2 NuRD-to-BAF coregulator switch and a shift in PPARγ occupancy to thermogenic genes. This shift in PPARγ occupancy increases the antidiabetic efficacy of the PPARγ agonist rosiglitazone in obesity while diminishing the unwanted weight-gaining effect of the drug. These data indicate that ZFP423 controls EBF2 coactivator recruitment and PPARγ occupancy to determine the thermogenic plasticity of adipocytes and highlight the potential of therapeutically targeting transcriptional brakes to induce beige adipocyte biogenesis in obesity.

Beige adipocytes are a distinct type of thermogenic fat cell in mouse and human.

Brown fat generates heat via the mitochondrial uncoupling protein UCP1, defending against hypothermia and obesity. Recent data suggest that there are two distinct types of brown fat: classical brown fat derived from a myf-5 cellular lineage and UCP1-positive cells that emerge in white fat from a non-myf-5 lineage. Here, we report the isolation of "beige" cells from murine white fat depots. Beige cells resemble white fat cells in having extremely low basal expression of UCP1, but, like classical brown fat, they respond to cyclic AMP stimulation with high UCP1 expression and respiration rates. Beige cells have a gene expression pattern distinct from either white or brown fat and are preferentially sensitive to the polypeptide hormone irisin. Finally, we provide evidence that previously identified brown fat deposits in adult humans are composed of beige adipocytes. These data provide a foundation for studying this mammalian cell type with therapeutic potential. PAPERCLIP:

Supraclavicular brown adipocytes originate from Tbx1+ myoprogenitors.

Brown adipose tissue (BAT) dissipates energy as heat, contributing to temperature control, energy expenditure, and systemic homeostasis. In adult humans, BAT mainly exists in supraclavicular areas and its prevalence is associated with cardiometabolic health. However, the developmental origin of supraclavicular BAT remains unknown. Here, using genetic cell marking in mice, we demonstrate that supraclavicular brown adipocytes do not develop from the Pax3+/Myf5+ epaxial dermomyotome that gives rise to interscapular BAT (iBAT). Instead, the Tbx1+ lineage that specifies the pharyngeal mesoderm marks the majority of supraclavicular brown adipocytes. Tbx1Cre-mediated ablation of peroxisome proliferator-activated receptor gamma (PPARγ) or PR/SET Domain 16 (PRDM16), components of the transcriptional complex for brown fat determination, leads to supraclavicular BAT paucity or dysfunction, thus rendering mice more sensitive to cold exposure. Moreover, human deep neck BAT expresses higher levels of the TBX1 gene than subcutaneous neck white adipocytes. Taken together, our observations reveal location-specific developmental origins of BAT depots and call attention to Tbx1+ lineage cells when investigating human relevant supraclavicular BAT.

Drosophila genome-wide obesity screen reveals hedgehog as a determinant of brown versus white adipose cell fate.

Over 1 billion people are estimated to be overweight, placing them at risk for diabetes, cardiovascular disease, and cancer. We performed a systems-level genetic dissection of adiposity regulation using genome-wide RNAi screening in adult Drosophila. As a follow-up, the resulting approximately 500 candidate obesity genes were functionally classified using muscle-, oenocyte-, fat-body-, and neuronal-specific knockdown in vivo and revealed hedgehog signaling as the top-scoring fat-body-specific pathway. To extrapolate these findings into mammals, we generated fat-specific hedgehog-activation mutant mice. Intriguingly, these mice displayed near total loss of white, but not brown, fat compartments. Mechanistically, activation of hedgehog signaling irreversibly blocked differentiation of white adipocytes through direct, coordinate modulation of early adipogenic factors. These findings identify a role for hedgehog signaling in white/brown adipocyte determination and link in vivo RNAi-based scanning of the Drosophila genome to regulation of adipocyte cell fate in mammals.

IL-4 activates the futile triacylglyceride cycle for glucose utilization in white adipocytes.

The development of cardiometabolic complications during obesity is strongly associated with chronic latent inflammation in hypertrophied adipose tissue (AT). IL-4 is an anti-inflammatory cytokine, playing a protective role against insulin resistance, glucose intolerance and weight gain. The positive effects of IL-4 are associated not only with the activation of anti-inflammatory immune cells in AT, but also with the modulation of adipocyte metabolism. IL-4 is known to activate lipolysis and glucose uptake in adipocytes, but the precise regulatory mechanisms and physiological significance of these processes remain unclear. In this study, we detail IL-4 effects on glucose and triacylglycerides (TAGs) metabolism and propose mechanisms of IL-4 metabolic action in adipocytes. We have shown that IL-4 activates glucose oxidation, lipid droplet (LD) fragmentation, lipolysis and thermogenesis in mature 3T3-L1 adipocytes. We found that lipolysis was not accompanied by fatty acids (FAs) release from adipocytes, suggesting FA re-esterification. Moreover, glucose oxidation and thermogenesis stimulation depended on adipocyte triglyceride lipase (ATGL) activity, but not the uncoupling protein (UCP1) expression. Based on these data, IL-4 may activate the futile TAG-FA cycle in adipocytes, which enhances the oxidative activity of cells and heat production. Thus, the positive effect of IL-4 on systemic metabolism can be the result of the activation of non-canonical thermogenic mechanism in AT, increasing TAG turnover and utilization of excessive glucose.

Sorting out adipocyte precursors and their role in physiology and disease.

The ability to maintain and expand the pool of adipocytes in adults is integral to the regulation of energy balance, tissue/stem cell homeostasis, and disease pathogenesis. For decades, our knowledge of adipocyte precursors has relied on cellular models. The identity of native adipocyte precursors has remained unclear. Recent studies have identified distinct adipocyte precursor populations that are physiologically regulated and contribute to the development, maintenance, and expansion of adipocyte pools in mice. With new tools available, the properties of adipocyte precursors can now be defined, and the regulation and function of adipose plasticity in development and physiology can be explored.

The molecular and metabolic program by which white adipocytes adapt to cool physiologic temperatures.

Although visceral adipocytes located within the body's central core are maintained at approximately 37°C, adipocytes within bone marrow, subcutaneous, and dermal depots are found primarily within the peripheral shell and generally exist at cooler temperatures. Responses of brown and beige/brite adipocytes to cold stress are well studied; however, comparatively little is known about mechanisms by which white adipocytes adapt to temperatures below 37°C. Here, we report that adaptation of cultured adipocytes to 31°C, the temperature at which distal marrow adipose tissues and subcutaneous adipose tissues often reside, increases anabolic and catabolic lipid metabolism, and elevates oxygen consumption. Cool adipocytes rely less on glucose and more on pyruvate, glutamine, and, especially, fatty acids as energy sources. Exposure of cultured adipocytes and gluteal white adipose tissue (WAT) to cool temperatures activates a shared program of gene expression. Cool temperatures induce stearoyl-CoA desaturase-1 (SCD1) expression and monounsaturated lipid levels in cultured adipocytes and distal bone marrow adipose tissues (BMATs), and SCD1 activity is required for acquisition of maximal oxygen consumption at 31°C.

Promote lipolysis in white adipocytes by magnetic hyperthermia therapy with Fe3O4microsphere-doped hydrogel.

Obesity has become an ongoing global crisis, since it increases the risks of cardiovascular disease, type 2 diabetes, fatty liver, cognitive decline, and some cancers. Adipose tissue is closely associated with the disorder of lipid metabolism. Several efforts have been made toward the modulation of lipid accumulation, but have been hindered by poor efficiency of cellular uptake, low safety, and uncertain effective dosage. Herein, we design an Fe3O4microsphere-doped composite hydrogel (Fe3O4microspheres @chitosan/ß-glycerophosphate/collagen), termed as Fe3O4@Gel, as the magnetocaloric agent for magnetic hyperthermia therapy (MHT), aiming to promote lipolysis in white adipocytes. The experimental results show that the obtained Fe3O4@Gel displays a series of advantages, such as fast sol-gel transition, high biocompatibility, and excellent magneto-thermal performance. MHT, which is realized by Fe3O4@Gel subjected to an alternating magnetic field, leads to reduced lipid accumulation, lower triglyceride content, and increased mitochondrial activity in white adipocytes. This work shows that Fe3O4@Gel-mediated MHT can effectively promote lipolysis in white adipocytesin vitro, which provides a potential approach to treat obesity and associated metabolic disorders.

Human visceral adipose tissue microvascular endothelial cell isolation and establishment of co-culture with white adipocytes to analyze cell-cell communication.

Communication between adipocytes and endothelial cells (EC) is suggested to play an important role in the metabolic function of white adipose tissue. In order to generate tools to investigate in detail the physiology and communication of EC and adipocytes, a method for isolation of adipose microvascular EC from visceral adipose tissue (VAT) biopsies of subjects with obesity was developed. Moreover, mature white adipocytes were isolated from the VAT biopsies by a method adapted from a previously published Membrane aggregate adipocytes culture (MAAC) protocol. The identity and functionality of the cultivated and isolated adipose microvascular EC (AMvEC) was validated by imaging their morphology, analyses of mRNA expression, fluorescence activated cell sorting (FACS), immunostaining, low-density lipoprotein (LDL) uptake, and in vitro angiogenesis assays. Finally, we established a new trans filter co-culture system (membrane aggregate adipocyte and endothelial co-culture, MAAECC) for the analysis of communication between the two cell types. EC-adipocyte communication in this system was validated by omics analyses, revealing several altered proteins belonging to pathways such as metabolism, intracellular transport and signal transduction in adipocytes co-cultured with AMvEC. In reverse experiments, induction of several pathways including endothelial development and functions was found in AMvEC co-cultured with adipocytes. In conclusion, we developed a robust method to isolate EC from small quantities of human VAT. Furthermore, the MAAECC system established during the study enables one to study the communication between primary white adipocytes and EC or vice-versa and could also be employed for drug screening.

Regulatory mechanisms underlying interleukin-6 expression in murine brown adipocytes.

Three types of adipocytes, white, brown, and beige, regulate the systemic energy balance through the storage and expenditure of chemical energy. In addition, adipocytes produce various bioactive molecules known as adipokines. In contrast to white adipocyte-derived molecules, less information is available on the adipokines produced by brown adipocytes (batokine). This study explored the regulatory expression of interleukin (IL)-6 in cell culture studies. Norepinephrine or a nonselective ß-adrenergic receptor agonist increased the expression of IL-6 in primary brown adipocytes and HB2 brown adipocytes. Treatment with forskolin (Fsk), an activator of the cAMP-dependent protein kinase (PKA) pathway (downstream signaling of the ß-adrenergic receptor), efficiently stimulated IL-6 expression in brown adipocytes and myotubes. Phosphorylated CREB and phosphorylated p38 MAP kinase levels were increased in Fsk-treated brown adipocytes within 5 min. In contrast, a long-term (∼60 min and ∼4 h) treatment with Fsk was required for increase in STAT3 phosphorylation and C/EBPß expression, respectively. The PKA, p38 MAP kinase, STAT3, and C/EBPß pathways are required for the maximal IL-6 expression induced by Fsk, which were verified by use of various inhibitors of these signal pathways. Vitamin C enhanced Fsk-induced IL-6 expression through the extracellular signal-regulated kinase activity. The present study provides basic information on the regulatory expression of IL-6 in activated brown adipocytes.

Perfluorooctanoate (PFOA) cell-autonomously promotes thermogenic and adipogenic differentiation of brown and white adipocytes.

Perfluorooctanoic acid (PFOA) is a synthetic organofluoride surfactant associated with several toxic effects in humans and animals. Particularly, it has been observed that PFOA treatment of mice results in weight loss associated with recruited brown adipose tissue (BAT), including an increased amount of uncoupling protein 1 (UCP1). The molecular mechanism behind this BAT recruitment is presently unknown. To investigate the existence of possible cell-autonomous effects of PFOA, we treated primary cultures of brown and white (inguinal) adipocytes with PFOA, or with the non-fluorinated equivalent octanoate, or with vehicle, for 48 h (from day 5 to day 7 of differentiation). PFOA in itself increased the gene expression (mRNA levels) of UCP1 and carnitine palmitoyltransferase 1A (CPT1α) (thermogenesis-related genes) in both brown and white adipocytes. In addition, PFOA increased the expression of fatty acid binding protein 4 (FABP4) and peroxisome proliferator-activated receptor α (PPARα) (adipogenesis-related genes). Also the protein levels of UCP1 were increased in brown adipocytes exposed to PFOA. This increase was more due to an increase in the fraction of cells that expressed UCP1 than to an increase in UCP1 levels per cell. The PFOA-induced changes were even more pronounced under simultaneous adrenergic stimulation. Octanoate induced less pronounced effects on adipocytes than did PFOA. Thus, PFOA in itself increased the levels of thermogenic markers in brown and white adipocytes. This could enhance the energy metabolism of animals (and humans) exposed to the compound, resulting in a negative energy balance, leading to diminished fitness.

Engineered allele substitution at PPARGC1A rs8192678 alters human white adipocyte differentiation, lipogenesis, and PGC-1α content and turnover.

AIMS/HYPOTHESIS: PPARGC1A encodes peroxisome proliferator-activated receptor γ coactivator 1-α (PGC-1α), a central regulator of energy metabolism and mitochondrial function. A common polymorphism in PPARGC1A (rs8192678, C/T, Gly482Ser) has been associated with obesity and related metabolic disorders, but no published functional studies have investigated direct allele-specific effects in adipocyte biology. We examined whether rs8192678 is a causal variant and reveal its biological function in human white adipose cells. METHODS: We used CRISPR-Cas9 genome editing to perform an allelic switch (C-to-T or T-to-C) at rs8192678 in an isogenic human pre-adipocyte white adipose tissue (hWAs) cell line. Allele-edited single-cell clones were expanded and screened to obtain homozygous T/T (Ser482Ser), C/C (Gly482Gly) and heterozygous C/T (Gly482Ser) isogenic cell populations, followed by functional studies of the allele-dependent effects on white adipocyte differentiation and mitochondrial function. RESULTS: After differentiation, the C/C adipocytes were visibly less BODIPY-positive than T/T and C/T adipocytes, and had significantly lower triacylglycerol content. The C allele presented a dose-dependent lowering effect on lipogenesis, as well as lower expression of genes critical for adipogenesis, lipid catabolism, lipogenesis and lipolysis. Moreover, C/C adipocytes had decreased oxygen consumption rate (OCR) at basal and maximal respiration, and lower ATP-linked OCR. We determined that these effects were a consequence of a C-allele-driven dysregulation of PGC-1α protein content, turnover rate and transcriptional coactivator activity. CONCLUSIONS/INTERPRETATION: Our data show allele-specific causal effects of the rs8192678 variant on adipogenic differentiation. The C allele confers lower levels of PPARGC1A mRNA and PGC-1α protein, as well as disrupted dynamics of PGC-1α turnover and activity, with downstream effects on cellular differentiation and mitochondrial function. Our study provides the first experimentally deduced insights on the effects of rs8192678 on adipocyte phenotype.

Proto-oncogene FAM83A contributes to casein kinase 1-mediated mitochondrial maintenance and white adipocyte differentiation.

Family with sequence similarity 83 A (FAM83A) is a newly discovered proto-oncogene that has been shown to play key roles in various cancers. However, the function of FAM83A in other physiological processes is not well known. Here, we report a novel function of FAM83A in adipocyte differentiation. We used an adipocyte-targeting fusion oligopeptide (FITC-ATS-9R) to deliver a FAM83A-sgRNA/Cas9 plasmid to knockdown Fam83a (ATS/sg-FAM83A) in white adipose tissue in mice, which resulted in reduced white adipose tissue mass, smaller adipocytes, and mitochondrial damage that was aggravated by a high-fat diet. In cultured 3T3-L1 adipocytes, we found loss or knockdown of Fam83a significantly repressed lipid droplet formation and downregulated the expression of lipogenic genes and proteins. Furthermore, inhibition of Fam83a decreased mitochondrial ATP production through blockage of the electron transport chain, associated with enhanced apoptosis. Mechanistically, we demonstrate FAM83A interacts with casein kinase 1 (CK1) and promotes the permeability of the mitochondrial outer membrane. Furthermore, loss of Fam83a in adipocytes hampered the formation of the TOM40 complex and impeded CK1-driven lipogenesis. Taken together, these results establish FAM83A as a critical regulator of mitochondria maintenance during adipogenesis.

Exercise-regulated white adipocyte differentitation: An insight into its role and mechanism.

White adipocytes play a key role in the regulation of fat mass amount and energy balance. An appropriate level of white adipocyte differentiation is important for maintaining metabolic homeostasis. Exercise, an important way to improve metabolic health, can regulate white adipocyte differentiation. In this review, the effect of exercise on the differentiation of white adipocytes is summarized. Exercise could regulate adipocyte differentiation in multiple ways, such as exerkines, metabolites, microRNAs, and so on. The potential mechanism underlying the role of exercise in adipocyte differentiation is also reviewed and discussed. In-depth investigation of the role and mechanism of exercise in white adipocyte differentiation would provide new insights into exercise-mediated improvement of metabolism and facilitate the application of exercise-based strategy against obesity.

Developmental and functional heterogeneity of white adipocytes within a single fat depot.

Recent studies suggest that, even within a single adipose depot, there may be distinct subpopulations of adipocytes. To investigate this cellular heterogeneity, we have developed multiple conditionally immortalized clonal preadipocyte lines from white adipose tissue of mice. Analysis of these clones reveals at least three white adipocyte subpopulations. These subpopulations have differences in metabolism and differentially respond to inflammatory cytokines, insulin, and growth hormones. These also have distinct gene expression profiles and can be tracked by differential expression of three marker genes: Wilms' tumor 1, transgelin, and myxovirus 1. Lineage tracing analysis with dual-fluorescent reporter mice indicates that these adipocyte subpopulations have differences in gene expression and metabolism that mirror those observed in the clonal cell lines. Furthermore, preadipocytes and adipocytes from these subpopulations differ in their abundance in different fat depots. Thus, white adipose tissue, even in a single depot, is comprised of distinct subpopulations of white adipocytes with different physiological phenotypes. These differences in adipocyte composition may contribute to the differences in metabolic behavior and physiology of different fat depots.

Differences in the metabolic profiles of brown and white adipocytes based on secretomics.

Brown adipocytes and white adipocytes play important roles in systemic metabolism and energy homeostasis. Recent studies have demonstrated that white adipocytes and brown adipocytes secrete numerous adipokines and thus act as endocrine cells. However, differences in the metabolites secreted from white adipocytes and brown adipocytes have never been reported. In this study, we assessed the metabolites secreted from white adipocytes and brown adipocytes. In total, the levels of 47 metabolites in brown adipocytes were significantly different from those in white adipocytes, with 31 high and 16 low in brown adipocytes as compared with those in white adipocytes. We classified these secreted metabolites as amino acids and peptides, fatty acids, and conjugates, glycerophosphocholines, furanones, and trichloroacetic acids. In addition, we identified the glycerophospholipid metabolism activated in white adipocytes, and these differentially expressed metabolites were associated with the mitogen-activated protein kinase pathway and Janus kinase-signal transducer and activator of transcription signaling pathway according to the Ingenuity Pathway Analysis (IPA) software analysis. This study revealed novel metabolites secreted from brown adipocytes and white adipocytes, and these metabolites from adipocytes may perform specific biological functions based on the type of adipocyte that secretes them, and this forms the material basis of the interaction between adipocytes and other cells.

Malonate induces the browning of white adipose tissue in high-fat diet induced obesity model.

Obesity increases the risk of various diseases, and many studies have examined prevention and treatment strategies. Browning of white adipocytes promotes triglyceride (TG) metabolism and is the new focus for treating obesity. This study investigated the role of malonate-a modulator of mitochondrial function-in adipocyte browning, and its potential as a therapeutic agent in obesity. Our findings revealed that malonate increased oxygen consumption without inhibiting ATP synthesis. Malonate induced expression of PRDM16-an important transcription factor for browning-and uncoupling protein 1 (beige adipocyte marker), suggesting that malonate induces browning in white adipocytes. In an obesity mouse model induced by a high-fat diet, malonate significantly reduced body weight and white adipose tissue weight, as well as improved insulin resistance. Importantly, malonate stimulated browning in white adipose tissue and maintained the mass of brown adipose tissue in the high-fat diet-induced obesity mouse model. We propose that manipulation of mitochondrial function by malonate is a promising therapeutic approach for obesity.

Transcriptomics profiling reveal the heterogeneity of white and brown adipocyte.

The marker genes associated with white adipocytes and brown adipocytes have been previously identified; however, these markers have not been updated in several years, and the differentiation process of preadipocytes remains relatively fixed. Consequently, there has been a lack of exploration into alternative differentiation schemes. In this particular study, we present a transcriptional signature specific to brown adipocytes and white adipocytes. Notably, our findings reveal that ZNF497, ZIC1, ZFY, UTY, USP9Y, TXLNGY, TTTY14, TNNT3, TNNT2, TNNT1, TNNI1, TNNC1, TDRD15, SOX11, SLN, SFRP2, PRKY, PAX3KLHL40, PAX3, INKA2-AS1, SOX11, and TDRD15 exhibit high expression levels in brown adipocytes. XIST, HOXA10, PCAT19, HOXA7, PLSCR3, and AVPR1A exhibited high expression levels in white adipocytes, suggesting their potential as novel marker genes for the transition from white to brown adipocytes. Furthermore, our analysis revealed the coordinated activation of several pathways, including the PPAR signaling pathway, focal adhesion, retrograde endocannabinoid signaling, oxidative phosphorylation, PI3K-Akt signaling pathway, and thermogenesis pathways, in brown adipocytes. Moreover, in contrast to prevailing culture techniques, we conducted a comparative analysis of the differentiation protocols for white preadipocytes and brown preadipocytes, revealing that the differentiation outcome remained unaffected by the diverse culture schemes employed. However, the expression levels of certain marker genes in both adipocyte types were found to be altered. This investigation not only identified potential novel marker genes for adipocytes but also examined the impact of different differentiation methods on preadipocyte maturation. Consequently, these findings offer significant insights for further research on the differentiation processes of diverse adipocyte subtypes.