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Despite encouraging outcomes of the eight FDA approved drugs in lessening obesity burden, they have been associated with side effects caused by the lack of direct action on the adipose tissue. Therefore, a nanomedicine is reported that promotes the transformation of local fat‐storage white adipocytes, associated with obesity, into thermogenic adipocytes, indicative of enhanced metabolism, to both human cells and mice. This nanomedicine consisting of a lipid‐silica nanohybrid that allows accommodation of the natural supplement forskolin has resulted in successful browning of mature human adipocytes in vitro through upregulating thermogenesis biomarkers on mature human adipocytes and in vivo in mice tissue (achieving 400‐ and 80‐fold increases in UCP1 and Cox7A1, respectively), while significantly enhancing glucose uptake and lipolysis in an acute fashion and significantly preventing weight gain in high fat diet (60% HFD) mice compared to other treatments including liposomal medicine. [ABSTRACT FROM AUTHOR]

Introduction: The activation of the P2X7 receptor subtype (P2X7R) has a main role in orchestrating the cellular inflammatory response in many different tissues. Obesity is characterized by dysfunctional fat deposition leading to a tissue-specific and systemic low-grade inflammation. Androgens and estrogens contribute to the whole adipose tissue inflammatory state, but the involvement of sex steroids in the purinergic signaling modulation in adipocytes is still unknown. Methods: We performed an in vitro study to evaluate the possible role of sex hormones on the P2X7R gene expression in human adipocytes, at baseline and after stimulation with bacterial lipopolysaccharide (LPS). We evaluated P2X7R gene expression during in vitro differentiation of human adipocytes, in the absence and presence of testosterone (T) and 17β-estradiol (E2) in the presence and absence of LPS. Furthermore, we analyzed the effects of incubation with dihydrotestosterone (DHT), a non-aromatizable androgen, using the co-incubation of isolated human adipocytes with T alone or in combination with anastrozole, an inhibitor of aromatase, the enzyme responsible of T conversion to E2. Results: At baseline, incubation of adipocytes with T or E2 did not significantly affect P2X7R gene expression. On the contrary, the incubation with DHT was associated with a significant reduction of P2X7R gene expression. LPS incubation significantly increased gene expression of P2X7R with respect to baseline. Interestingly, after LPS stimulation, DHT exposure showed an additional effect, markedly increasing the P2X7R gene expression. This amplificatory effect was confirmed by the incubation of adipocytes to both anastrozole and testosterone. In these experimental conditions, while no effect was observed at baseline, an amplification of the expression of the P2X7R mRNA was observed after stimulation with LPS. Discussion: The purinergic system is involved in the inflammatory response of adipocytes, and androgens may modulate its activity. In particular DHT, a nonaromatizable androgen, amplifies the LPS-induced P2X7R gene expression in human adipocytes thus showing a gender regulated response of the expression of this purinergic receptor strongly involved in the inflammatory response in adipose tissue. [ABSTRACT FROM AUTHOR]

Abstract Background Dysfunctional adipose tissue (AT) is known to contribute to the pathophysiology of metabolic disease, including type 2 diabetes mellitus (T2DM). This dysfunction may occur, in part, as a consequence of gut-derived endotoxaemia inducing changes in adipocyte mitochondrial function and reducing the proportion of BRITE (brown-in-white) adipocytes. Therefore, the present study investigated whether endotoxin (lipopolysaccharide; LPS) directly contributes to impaired human adipocyte mitochondrial function and browning in human adipocytes, and the relevant impact of obesity status pre and post bariatric surgery. Methods Human differentiated abdominal subcutaneous (AbdSc) adipocytes from participants with obesity and normal-weight participants were treated with endotoxin to assess in vitro changes in mitochondrial function and BRITE phenotype. Ex vivo human AbdSc AT from different groups of participants (normal-weight, obesity, pre- and 6 months post-bariatric surgery) were assessed for similar analyses including circulating endotoxin levels. Results Ex vivo AT analysis (lean & obese, weight loss post-bariatric surgery) identified that systemic endotoxin negatively correlated with BAT gene expression (p

Introduction: The activation of the P2X7 receptor subtype (P2X7R) has a main role in orchestrating the cellular inflammatory response in many different tissues. Obesity is characterized by dysfunctional fat deposition leading to a tissue-specific and systemic low-grade inflammation. Androgens and estrogens contribute to the whole adipose tissue inflammatory state, but the involvement of sex steroids in the purinergic signaling modulation in adipocytes is still unknown.Methods: We performed an in vitro study to evaluate the possible role of sex hormones on the P2X7R gene expression in human adipocytes, at baseline and after stimulation with bacterial lipopolysaccharide (LPS). We evaluated P2X7R gene expression during in vitro differentiation of human adipocytes, in the absence and presence of testosterone (T) and 17β-estradiol (E2) in the presence and absence of LPS. Furthermore, we analyzed the effects of incubation with dihydrotestosterone (DHT), a non-aromatizable androgen, using the co-incubation of isolated human adipocytes with T alone or in combination with anastrozole, an inhibitor of aromatase, the enzyme responsible of T conversion to E2.Results: At baseline, incubation of adipocytes with T or E2 did not significantly affect P2X7R gene expression. On the contrary, the incubation with DHT was associated with a significant reduction of P2X7R gene expression. LPS incubation significantly increased gene expression of P2X7R with respect to baseline. Interestingly, after LPS stimulation, DHT exposure showed an additional effect, markedly increasing the P2X7R gene expression. This amplificatory effect was confirmed by the incubation of adipocytes to both anastrozole and testosterone. In these experimental conditions, while no effect was observed at baseline, an amplification of the expression of the P2X7R mRNA was observed after stimulation with LPS.Discussion: The purinergic system is involved in the inflammatory response of adipocytes, and androgens may modulate its activity. In particular DHT, a non-aromatizable androgen, amplifies the LPS-induced P2X7R gene expression in human adipocytes thus showing a gender regulated response of the expression of this purinergic receptor strongly involved in the inflammatory response in adipose tissue.

Type 2 diabetes mellitus is a chronic metabolic disease with no cure. Adipose tissue is a major site of systemic insulin resistance. Sortilin is a central component of the glucose transporter -Glut4 storage vesicles (GSV) which translocate to the plasma membrane to uptake glucose from circulation. Here, using human adipocytes we demonstrate the presence of the alternatively spliced, truncated sortilin variant (Sort_T) whose expression is significantly increased in diabetic adipose tissue. Artificial-intelligence-based modeling, molecular dynamics, intrinsically disordered region analysis, and co-immunoprecipitation demonstrated association of Sort_T with Glut4 and decreased glucose uptake in adipocytes. The results show that glucagon-like peptide-1 (GLP1) hormone decreases Sort_T. We deciphered the molecular mechanism underlying GLP1 regulation of alternative splicing of human sortilin. Using splicing minigenes and RNA-immunoprecipitation assays, the results show that GLP1 regulates Sort_T alternative splicing via the splice factor, TRA2B. We demonstrate that targeted antisense oligonucleotide morpholinos reduces Sort_T levels and improves glucose uptake in diabetic adipocytes. Thus, we demonstrate that GLP1 regulates alternative splicing of sortilin in human diabetic adipocytes. [ABSTRACT FROM AUTHOR]

Background: Dysfunctional adipose tissue (AT) is known to contribute to the pathophysiology of metabolic disease, including type 2 diabetes mellitus (T2DM). This dysfunction may occur, in part, as a consequence of gut-derived endotoxaemia inducing changes in adipocyte mitochondrial function and reducing the proportion of BRITE (brown-in-white) adipocytes. Therefore, the present study investigated whether endotoxin (lipopolysaccharide; LPS) directly contributes to impaired human adipocyte mitochondrial function and browning in human adipocytes, and the relevant impact of obesity status pre and post bariatric surgery. Methods: Human differentiated abdominal subcutaneous (AbdSc) adipocytes from participants with obesity and normal-weight participants were treated with endotoxin to assess in vitro changes in mitochondrial function and BRITE phenotype. Ex vivo human AbdSc AT from different groups of participants (normal-weight, obesity, pre- and 6 months post-bariatric surgery) were assessed for similar analyses including circulating endotoxin levels. Results: Ex vivo AT analysis (lean & obese, weight loss post-bariatric surgery) identified that systemic endotoxin negatively correlated with BAT gene expression (p < 0.05). In vitro endotoxin treatment of AbdSc adipocytes (lean & obese) reduced mitochondrial dynamics (74.6% reduction; p < 0.0001), biogenesis (81.2% reduction; p < 0.0001) and the BRITE phenotype (93.8% reduction; p < 0.0001). Lean AbdSc adipocytes were more responsive to adrenergic signalling than obese AbdSc adipocytes; although endotoxin mitigated this response (92.6% reduction; p < 0.0001). Conclusions: Taken together, these data suggest that systemic gut-derived endotoxaemia contributes to both individual adipocyte dysfunction and reduced browning capacity of the adipocyte cell population, exacerbating metabolic consequences. As bariatric surgery reduces endotoxin levels and is associated with improving adipocyte functionality, this may provide further evidence regarding the metabolic benefits of such surgical interventions. [ABSTRACT FROM AUTHOR]

Trifuhalol A, a fucol-type phlorotannin, was extracted and identified from the brown algae Agarum cribrosum. The total yield and purity of trifuhalol A from A. cribrosum were 0.98% and 86%, respectively. Trifuhalol A at 22 and 44 μM inhibited lipid accumulation in human primary adipocytes. Consistently trifuhalol A suppressed the expression of adipogenesis-related genes, such as proliferator-activated receptor-gamma (PPAR-γ), CCAAT/enhancer-binding protein-alpha (C/EBP-α), fatty acid synthase (FAS), and sterol regulatory element-binding protein-1 (SREBP-1), in a dose-dependent manner. Trifuhalol A increased the level of proteins such as wingless/integrated (Wnt)10b, nuclear-β-catenin, total-β-catenin, phospho-AMP-activated protein kinase (pAMPK), and phospho-liver kinase B1 (pLKB1) as well as the expression of genes such as Wnt10b, Frizzled 1, and low-density lipoprotein receptor-related protein 6 (LRP6). Additionally, trifuhalol A decreased the expression of the glycogen synthase kinase-3beta (GSK3β) gene. These results suggest that trifuhalol A reduces fat accumulation in human adipocytes via the Wnt/β-catenin- and AMPK-dependent pathways.

Benzylamine is a natural molecule present in food and edible plants, capable of activating hexose uptake and inhibiting lipolysis in human fat cells. These effects are dependent on its oxidation by amine oxidases present in adipocytes, and on the subsequent hydrogen peroxide production, known to exhibit insulin-like actions. Virtually, other substrates interacting with such hydrogen peroxide-releasing enzymes potentially can modulate lipid accumulation in adipose tissue. Inhibition of such enzymes has also been reported to influence lipid deposition. We have therefore studied in human adipocytes the lipolytic and lipogenic activities of pharmacological entities designed to interact with amine oxidases highly expressed in this cell type: the semicarbazide-sensitive amine oxidase (SSAO also known as PrAO or VAP-1) and the monoamine oxidases (MAO). The results showed that SZV-2016 and SZV-2017 behaved as better substrates than benzylamine, releasing hydrogen peroxide once oxidized, and reproduced or even exceeded its insulin-like metabolic effects in fat cells. Additionally, several novel SSAO inhibitors, such as SZV-2007 and SZV-1398, have been evidenced and shown to inhibit benzylamine metabolic actions. Taken as a whole, our findings reinforce the list of molecules that influence the regulation of triacylglycerol assembly/breakdown, at least in vitro in human adipocytes. The novel compounds deserve deeper investigation of their mechanisms of interaction with SSAO or MAO, and constitute potential candidates for therapeutic use in obesity and diabetes. [ABSTRACT FROM AUTHOR]

Abdominal obesity associates with cardiometabolic disease and an accumulation of lipids in the visceral adipose depot, whereas lipid accumulation in the subcutaneous depot is more benign. We aimed to further investigate whether the adipogenic properties where cell-intrinsic, or dependent on a depot-specific or obesity-produced microenvironment. We obtained visceral and subcutaneous biopsies from non-obese women (n = 14) or women living with morbid obesity (n = 14) and isolated adipose stem and progenitor cells (ASPCs) from the stromal vascular fraction of non-obese (n = 13) and obese (n = 13). Following in vitro differentiation into mature adipocytes, we observed a contrasting pattern with a lower gene expression of adipogenic markers and a higher gene expression of immunogenic markers in the visceral compared to the subcutaneous adipocytes. We identified the immunogenic factor BST2 as a marker for visceral ASPCs. The effect of obesity and insulin resistance on adipogenic and immunogenic markers in the in vitro differentiated cells was minor. In contrast, differentiation with exogenous Tumor necrosis factor resulted in increased immunogenic signatures, including increased expression of BST2, and decreased adipogenic signatures in cells from both depots. Our data, from 26 women, underscore the intrinsic differences between human visceral and subcutaneous adipose stem and progenitor cells, suggest that dysregulation of adipocytes in obesity mainly occurs at a post-progenitor stage, and highlight an inflammatory microenvironment as a major constraint of human adipogenesis.

It is increasingly recognized that hypoxia may develop in adipose tissue as its mass expands. Adipose tissue is also the main reservoir of lipophilic pollutants, including polychlorinated biphenyls (PCBs). Both hypoxia and PCBs have been shown to alter adipose tissue functions. The signaling pathways induced by hypoxia and pollutants may crosstalk, as they share a common transcription factor: aryl hydrocarbon receptor nuclear translocator (ARNT). Whether hypoxia and PCBs crosstalk and affect adipokine secretion in human adipocytes remains to be explored. Using primary human adipocytes acutely co-exposed to different levels of hypoxia (24 h) and PCB126 (48 h), we observed that hypoxia significantly inhibits the PCB126 induction of cytochrome P450 (CYP1A1) transcription in a dose-response manner, and that Acriflavine (ACF)—an HIF1α inhibitor—partially restores the PCB126 induction of CYP1A1 under hypoxia. On the other hand, exposure to PCB126 did not affect the transcription of the vascular endothelial growth factor-A (VEGFA) under hypoxia. Exposure to hypoxia increased leptin and interleukin-6 (IL-6), and decreased adiponectin levels dose-dependently, while PCB126 increased IL-6 and IL-8 secretion in a dose-dependent manner. Co-exposure to PCB126 and hypoxia did not alter the adipokine secretion pattern observed under hypoxia and PCB126 exposure alone. In conclusion, our results indicate that (1) hypoxia inhibits PCB126-induced CYP1A1 expression at least partly through ARNT-dependent means, suggesting that hypoxia could affect PCB metabolism and toxicity in adipose tissue, and (2) hypoxia and PCB126 affect leptin, adiponectin, IL-6 and IL-8 secretion differently, with no apparent crosstalk between the two factors. [ABSTRACT FROM AUTHOR]

The striking rise of obesity-related metabolic disorders has focused attention on adipocytes as critical mediators of disease phenotypes. To better understand the role played by excess adipose in metabolic dysfunction it is crucial to decipher the transcriptional underpinnings of the low-grade adipose inflammation characteristic of diseases such as type 2 diabetes. Through employing a comparative transcriptomics approach, we identified IRF1 as differentially regulated between primary and in vitro-derived genetically matched adipocytes. This suggests a role as a mediator of adipocyte inflammatory phenotypes, similar to its function in other tissues. Utilizing adipose-derived mesenchymal progenitors we subsequently demonstrated that expression of IRF1 in adipocytes indeed contributes to upregulation of inflammatory processes, both in vitro and in vivo. This highlights IRF1's relevance to obesity-related inflammation and the resultant metabolic dysregulation.

Recent studies in mice and humans demonstrated the relevance of H2S synthesising enzymes, such as CTH, CBS, and MPST, in the physiology of adipose tissue and the differentiation of preadipocyte into adipocytes. Here, our objective was to investigate the combined role of CTH, CBS, and MPST in the preservation of adipocyte protein persulfidation and adipogenesis. Combined partial CTH, CBS, and MPST gene knockdown was achieved treating fully human adipocytes with siRNAs against these transcripts (siRNA_MIX). Adipocyte protein persulfidation was analyzed using label-free quantitative mass spectrometry coupled with a dimedone-switch method for protein labeling and purification. Proteomic analysis quantified 216 proteins with statistically different levels of persulfidation in KD cells compared to control adipocytes. In fully differentiated adipocytes, CBS and MPST mRNA and protein levels were abundant, while CTH expression was very low. It is noteworthy that siRNA_MIX administration resulted in a significant decrease in CBS and MPST expression, without impacting on CTH. The combined partial knockdown of the CBS and MPST genes resulted in reduced cellular sulfide levels in parallel to decreased expression of relevant genes for adipocyte biology, including adipogenesis, mitochondrial biogenesis, and lipogenesis, but increased proinflammatory- and senescence-related genes. It should be noted that the combined partial knockdown of CBS and MPST genes also led to a significant disruption in the persulfidation pattern of the adipocyte proteins. Although among the less persulfidated proteins, we identified several relevant proteins for adipocyte adipogenesis and function, among the most persulfidated, key mediators of adipocyte inflammation and dysfunction as well as some proteins that might play a positive role in adipogenesis were found. In conclusion, the current study indicates that the combined partial elimination of CBS and MPST (but not CTH) in adipocytes affects the expression of genes related to the maintenance of adipocyte function and promotes inflammation, possibly by altering the pattern of protein persulfidation in these cells, suggesting that these enzymes were required for the functional maintenance of adipocytes. [ABSTRACT FROM AUTHOR]

It is increasingly recognized that hypoxia may develop in adipose tissue as its mass expands. Adipose tissue is also the main reservoir of lipophilic pollutants, including polychlorinated biphenyls (PCBs). Both hypoxia and PCBs have been shown to alter adipose tissue functions. The signaling pathways induced by hypoxia and pollutants may crosstalk, as they share a common transcription factor: aryl hydrocarbon receptor nuclear translocator (ARNT). Whether hypoxia and PCBs crosstalk and affect adipokine secretion in human adipocytes remains to be explored. Using primary human adipocytes acutely co-exposed to different levels of hypoxia (24 h) and PCB126 (48 h), we observed that hypoxia significantly inhibits the PCB126 induction of cytochrome P450 (CYP1A1) transcription in a dose-response manner, and that Acriflavine (ACF)—an HIF1α inhibitor—partially restores the PCB126 induction of CYP1A1 under hypoxia. On the other hand, exposure to PCB126 did not affect the transcription of the vascular endothelial growth factor-A (VEGFA) under hypoxia. Exposure to hypoxia increased leptin and interleukin-6 (IL-6), and decreased adiponectin levels dose-dependently, while PCB126 increased IL-6 and IL-8 secretion in a dose-dependent manner. Co-exposure to PCB126 and hypoxia did not alter the adipokine secretion pattern observed under hypoxia and PCB126 exposure alone. In conclusion, our results indicate that (1) hypoxia inhibits PCB126-induced CYP1A1 expression at least partly through ARNT-dependent means, suggesting that hypoxia could affect PCB metabolism and toxicity in adipose tissue, and (2) hypoxia and PCB126 affect leptin, adiponectin, IL-6 and IL-8 secretion differently, with no apparent crosstalk between the two factors.

Recent studies in mice and humans demonstrated the relevance of H2S synthesising enzymes, such as CTH, CBS, and MPST, in the physiology of adipose tissue and the differentiation of preadipocyte into adipocytes. Here, our objective was to investigate the combined role of CTH, CBS, and MPST in the preservation of adipocyte protein persulfidation and adipogenesis. Combined partial CTH, CBS, and MPST gene knockdown was achieved treating fully human adipocytes with siRNAs against these transcripts (siRNA_MIX). Adipocyte protein persulfidation was analyzed using label-free quantitative mass spectrometry coupled with a dimedone-switch method for protein labeling and purification. Proteomic analysis quantified 216 proteins with statistically different levels of persulfidation in KD cells compared to control adipocytes. In fully differentiated adipocytes, CBS and MPST mRNA and protein levels were abundant, while CTH expression was very low. It is noteworthy that siRNA_MIX administration resulted in a significant decrease in CBS and MPST expression, without impacting on CTH. The combined partial knockdown of the CBS and MPST genes resulted in reduced cellular sulfide levels in parallel to decreased expression of relevant genes for adipocyte biology, including adipogenesis, mitochondrial biogenesis, and lipogenesis, but increased proinflammatory- and senescence-related genes. It should be noted that the combined partial knockdown of CBS and MPST genes also led to a significant disruption in the persulfidation pattern of the adipocyte proteins. Although among the less persulfidated proteins, we identified several relevant proteins for adipocyte adipogenesis and function, among the most persulfidated, key mediators of adipocyte inflammation and dysfunction as well as some proteins that might play a positive role in adipogenesis were found. In conclusion, the current study indicates that the combined partial elimination of CBS and MPST (but not CTH) in adipocytes affects the expression of genes related to the maintenance of adipocyte function and promotes inflammation, possibly by altering the pattern of protein persulfidation in these cells, suggesting that these enzymes were required for the functional maintenance of adipocytes.

New treatments for obesity and associated metabolic disease are increasingly warranted with the growth of the obesity pandemic. Brown adipose tissue (BAT) may represent a promising therapeutic target to treat obesity, as this tissue has been shown to regulate energy expenditure through non-shivering thermogenesis. Three different strategies could be employed for therapeutic targeting of human thermogenic adipocytes: increasing BAT mass through stimulation of BAT progenitors, increasing BAT function through regulatory pathways, and increasing WAT browning through promotion of beige adipocyte formation. However, these strategies require deeper understanding of human brown and beige adipocytes. While murine studies have greatly increased our understanding of BAT, it is becoming clear that human BAT does not exactly resemble that of the mouse, highlighting the need for human in vitro models of brown adipocytes. Several different human brown adipocyte models will be discussed here, along with the potential to improve brown adipocyte culture through recreation of the BAT microenvironment. [ABSTRACT FROM AUTHOR]

Context: Dysfunctional endoplasmic reticulum (ER) and mitochondria are known to contribute to the pathology of metabolic disease. This damage may occur, in part, as a consequence of ER-mitochondria cross-talk in conditions of nutrient excess such as obesity. To date, insight into this dynamic relationship has not been characterized in adipose tissue. Therefore, this study investigated whether ER stress contributes to the development of mitochondrial inefficiency in human adipocytes from lean and obese participants.Methods: Human differentiated adipocytes from Chub-S7 cell line and primary abdominal subcutaneous adipocytes from lean and obese participants were treated with tunicamycin to induce ER stress. Key parameters of mitochondrial function were assessed, including mitochondrial respiration, membrane potential (MMP), and dynamics.Results: ER stress led to increased respiratory capacity in a model adipocyte system (Chub-S7 adipocytes) in a concentration and time dependent manner (24 h: 23%↑; 48 h: 68%↑, P < 0.001; 72 h: 136%↑, P < 0.001). This corresponded with mitochondrial inefficiency and diminished MMP, highlighting the formation of dysfunctional mitochondria. Morphological analysis revealed reorganization of mitochondrial network, specifically mitochondrial fragmentation. Furthermore, p-DRP1, a key protein in fission, significantly increased (P < 0.001). Additionally, adipocytes from obese subjects displayed lower basal respiration (49%↓, P < 0.01) and were unresponsive to tunicamycin in contrast to their lean counterparts, demonstrating inefficient mitochondrial oxidative capacity.Conclusion: These human data suggest that adipocyte mitochondrial inefficiency is driven by ER stress and exacerbated in obesity. Nutrient excess-induced ER stress leads to mitochondrial dysfunction that may therefore shift lipid deposition ectopically and thus have further implications on the development of related metabolic disorders. [ABSTRACT FROM AUTHOR]

The striking rise of obesity-related metabolic disorders has focused attention on adipocytes as critical mediators of disease phenotypes. To better understand the role played by excess adipose in metabolic dysfunction it is crucial to decipher the transcriptional underpinnings of the low-grade adipose inflammation characteristic of diseases such as type 2 diabetes. Through employing a comparative transcriptomics approach, we identified IRF1 as differentially regulated between primary and in vitro-derived genetically matched adipocytes. This suggests a role as a mediator of adipocyte inflammatory phenotypes, similar to its function in other tissues. Utilizing adipose-derived mesenchymal progenitors we subsequently demonstrated that expression of IRF1 in adipocytes indeed contributes to upregulation of inflammatory processes, both in vitro and in vivo. This highlights IRF1's relevance to obesity-related inflammation and the resultant metabolic dysregulation.

We have previously demonstrated in Caco‐2 cells that tumor necrosis factor‐α (TNF‐α) inhibits sugar uptake, acting from the apical membrane, by decreasing the expression of the Na+‐glucose cotransporter SGLT1 in the brush border membrane. The goal was to investigate the hypothesis that TNF‐α from abdominal adipose tissue (adipocytes and macrophages) would decrease sugar and amino acid transport acting from the basolateral membrane of the enterocytes. TNF‐α placed in the basal compartment of Caco‐2 cells decreased α‐methyl‐ d‐glucose (αMG) and glutamine uptake. The apical medium derived from these Caco‐2 cells apically placed in another set of cells, also reduced sugar and glutamine transport. Reverse‐transcription polymerase chain reaction analysis demonstrated upregulation of TNF‐α, IL‐1β, and MCP1 expression in Caco‐2 cells exposed to basal TNF‐α. Similarly, αMG uptake was inhibited after Caco‐2 cells were incubated, in the basal compartment, with medium from visceral human mesenchymal stem cells‐derived adipocytes of overweight individuals. The apical medium collected from those Caco‐2 cells, and placed in the upper side of other set of cells, also decreased sugar uptake. Basal presence of medium derived from lipopolysaccharide‐activated macrophages and nonactivated macrophages decreased αMG uptake as well. Diet‐induced obese mice showed an increase in the visceral adipose tissue surrounding the intestine. In this physiological condition, there was a reduction on αMG uptake in jejunal everted rings. Altogether, these results suggest that basolateral TNF‐α, which can be produced by adipocytes and macrophages during obesity, would be able to activate TNF‐α and other proinflammatory proteins expression in the small intestine and diminish intestinal sugar and amino acids transport. Altogether, the present results support our hypothesis (Figure 8). Thus, obesity would induce the growth of visceral adipose tissue next to the intestine. TNF‐α coming from adipocytes would bind to the basolateral receptor TNFR1 (Sonnier et al., 2010) and stimulate TNF‐α apical secretion by the enterocytes which, in turn, would bind to apical TNFR1 triggering nutrients transporters internalization and thus, inhibiting nutrients uptake (Barrenetxe et al., 2013). TNF‐α from activated macrophages would also decrease nutrients transport by directly signaling from the basolateral membrane. [ABSTRACT FROM AUTHOR]

Abstract Aims/hypothesis Salt-inducible kinase 2 (SIK2) is downregulated in adipose tissue from obese or insulin-resistant individuals and inhibition of SIK isoforms results in reduced glucose uptake and insulin signalling in adipocytes. However, the regulation of SIK2 itself in response to insulin in adipocytes has not been studied in detail. The aim of our work was to investigate effects of insulin on various aspects of SIK2 function in adipocytes. Methods Primary adipocytes were isolated from human subcutaneous and rat epididymal adipose tissue. Insulin-induced phosphorylation of SIK2 and HDAC4 was analyzed using phosphospecific antibodies and changes in the catalytic activity of SIK2 with in vitro kinase assay. SIK2 protein levels were analyzed in primary adipocytes treated with the proteasome inhibitor MG132. Results We have identified a novel regulatory pathway of SIK2 in adipocytes, which involves insulin-induced phosphorylation at Thr484. This phosphorylation is impaired in individuals with a reduced insulin action. Insulin stimulation does not affect SIK2 catalytic activity or cellular activity towards HDAC4, but is associated with increased SIK2 protein levels in adipocytes. Conclusion/interpretation Our data suggest that downregulation of SIK2 in the adipose tissue of insulin-resistant individuals can partially be caused by impaired insulin signalling, which might result in defects in SIK2 expression and function. Graphical abstract Unlabelled Image Highlights • SIK2 is phosphorylated at Thr484 in response to insulin in adipocytes. • Phosphorylation at Thr484 is impaired in humans with reduced insulin action. • Insulin stimulation does not alter the catalytic activity of SIK2. • SIK2 protein levels are increased in response to insulin in adipocytes. [ABSTRACT FROM AUTHOR]

Human lifetime exposure to arsenic through drinking water, food supply or industrial pollution leads to its accumulation in many organs such as liver, kidneys, lungs or pancreas but also adipose tissue. Recently, population-based studies revealed the association between arsenic exposure and the development of metabolic diseases such as obesity and type 2 diabetes. To shed light on the molecular bases of such association, we determined the concentration that inhibited 17% of cell viability and investigated the effects of arsenic acute exposure on adipose-derived human mesenchymal stem cells differentiated in vitro into mature adipocytes and treated with sodium arsenite (NaAsO 2 , 10 nM to 10 µM). Untargeted metabolomics and gene expression analyses revealed a strong dose-dependent inhibition of lipogenesis and lipolysis induction, reducing the cellular ability to store lipids. These dysregulations were emphasized by the inhibition of the cellular response to insulin, as shown by the perturbation of several genes and metabolites involved in the mentioned biological pathways. Our study highlighted the activation of an adaptive oxidative stress response with the strong induction of metallothioneins and increased glutathione levels in response to arsenic accumulation that could exacerbate the decreased insulin sensitivity of the adipocytes. Arsenic exposure strongly affected the expression of arsenic transporters, responsible for arsenic influx and efflux, and induced a pro-inflammatory state in adipocytes by enhancing the expression of the inflammatory interleukin 6 (IL6). Collectively, our data showed that an acute exposure to low levels of arsenic concentrations alters key adipocyte functions, highlighting its contribution to the development of insulin resistance and the pathogenesis of metabolic disorders. • As impacts adipocyte functions at low levels of exposure. • As induces insulin resistance and perturbs lipid storage in human adipocytes. • As activates cell defense mechanisms and inflammation in human adipocytes. • As induces an adaptive oxidative stress response in human adipocytes. • As effects in human adipocytes are associated with the inhibition of As transporters. [ABSTRACT FROM AUTHOR]

Increased consumption of sugar and fructose as sweeteners has resulted in the utilization of fructose as an alternative metabolic fuel that may compete with glucose and alter its metabolism. To explore this, human Simpson-Golabi-Behmel Syndrome (SGBS) preadipocytes were differentiated to adipocytes in the presence of 0, 1, 2.5, 5 or 10 mM of fructose added to a medium containing 5 mM of glucose representing the normal blood glucose concentration. Targeted tracer [1,2-13C2]-d-glucose fate association approach was employed to examine the influence of fructose on the intermediary metabolism of glucose. Increasing concentrations of fructose robustly increased the oxidation of [1,2-13C2]-d-glucose to 13CO2 (p < 0.000001). However, glucose-derived 13CO2 negatively correlated with 13C labeled glutamate, 13C palmitate, and M+1 labeled lactate. These are strong markers of limited tricarboxylic acid (TCA) cycle, fatty acid synthesis, pentose cycle fluxes, substrate turnover and NAD+/NADP+ or ATP production from glucose via complete oxidation, indicating diminished mitochondrial energy metabolism. Contrarily, a positive correlation was observed between glucose-derived 13CO2 formed and 13C oleate and doses of fructose which indicate the elongation and desaturation of palmitate to oleate for storage. Collectively, these results suggest that fructose preferentially drives glucose through serine oxidation glycine cleavage (SOGC pathway) one-carbon cycle for NAD+/NADP+ production that is utilized in fructose-induced lipogenesis and storage in adipocytes.

Trifuhalol A, a fucol-type phlorotannin, was extracted and identified from the brown algae Agarum cribrosum . The total yield and purity of trifuhalol A from A. cribrosum were 0.98% and 86%, respectively. Trifuhalol A at 22 and 44 μM inhibited lipid accumulation in human primary adipocytes. Consistently trifuhalol A suppressed the expression of adipogenesis-related genes, such as proliferator-activated receptor-gamma (PPAR-γ), CCAAT/enhancer-binding protein-alpha (C/EBP-α), fatty acid synthase (FAS), and sterol regulatory element-binding protein-1 (SREBP-1), in a dose-dependent manner. Trifuhalol A increased the level of proteins such as wingless/integrated (Wnt)10b, nuclear-β-catenin, total-β-catenin, phospho-AMP-activated protein kinase ( p AMPK), and phospho-liver kinase B1 ( p LKB1) as well as the expression of genes such as Wnt10b, Frizzled 1, and low-density lipoprotein receptor-related protein 6 (LRP6). Additionally, trifuhalol A decreased the expression of the glycogen synthase kinase-3beta (GSK3β) gene. These results suggest that trifuhalol A reduces fat accumulation in human adipocytes via the Wnt/β-catenin- and AMPK-dependent pathways., Competing Interests: The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Given her role as the Associate Editor at the time of submission, Dr. Yeonhwa Park was not involved in the peer review of this article and had no access to information regarding its peer review. Full responsibility for the editorial process for this article was delegated to another editor, as per the Journal guidelines. The authors declare that they have no other known competing financial interests that could have appeared to influence the work reported in this paper., (© 2023 The Authors.)

Activation of AMP-activated protein kinase (AMPK) is considered an attractive strategy for the treatment of type 2 diabetes. Favorable metabolic effects of AMPK activation are mainly observed in skeletal muscle and liver tissue, whereas the effects in human adipose tissue are only poorly understood. Previous studies, which largely employed the AMPK activator 5-aminoimidazole-4-carboxamide-1-β-d-ribofuranoside (AICAR), suggest an antilipolytic role of AMPK in adipocytes. The aim of this work was to reinvestigate the role of AMPK in the regulation of lipolysis, using the novel allosteric small-molecule AMPK activators A-769662 and 991, with a focus on human adipocytes. For this purpose, human primary subcutaneous adipocytes were treated with A-769662, 991, or AICAR, as a control, before being stimulated with isoproterenol. AMPK activity status, glycerol release, and the phosphorylation of hormone-sensitive lipase (HSL), a key regulator of lipolysis, were then monitored. Our results show that both A-769662 and 991 activated AMPK to a level that was similar to, or greater than, that induced by AICAR. In contrast to AICAR, which as expected was antilipolytic, neither A-769662 nor 991 affected lipolysis in human adipocytes, although 991 treatment led to altered HSL phosphorylation. Furthermore, we suggest that HSL Ser660 is an important regulator of lipolytic activity in human adipocytes. These data suggest that the antilipolytic effect observed with AICAR in previous studies is, at least to some extent, AMPK independent. [ABSTRACT FROM AUTHOR]

Abstract Spexin (SPX, NPQ) is a novel peptide involved in the regulation of energy metabolism. SPX inhibits food intake and reduces body weight. In obese humans, SPX is the most down-regulated gene in fat. Therefore, SPX might be involved in the regulation of lipid metabolism. Here, we study the effects of SPX on lipolysis, lipogenesis, glucose uptake, adipogenesis, cell proliferation and survival in isolated human adipocytes or murine 3T3-L1 cells. SPX and its receptors, GALR2 and GALR3, are present at mRNA and protein levels in murine 3T3-L1 cells and human adipocytes. SPX inhibits adipogenesis and down-regulates mRNA expression of proadipogenic genes such as Pparγ, C/ebpα, C/ebpβ and Fabp4. SPX stimulates lipolysis by increasing the phosphorylation of hormone sensitive lipase (HSL). Simultaneously, SPX inhibits lipogenesis and glucose uptake in human adipocytes and murine 3T3-L1 cells. SPX has no effect on murine 3T3-L1 cell proliferation and viability. Moreover, our research showed that the SPX effect on adipocytes metabolism is mediated via GALR2 and GALR3 receptors. SPX is a novel regulator of lipid metabolism in murine 3T3-L1 and human adipocytes. Highlights • Spexin is expressed in murine 3T3-L1 adipocytes and isolated human adipocytes. • Spexin inhibits adipogenesis. • Spexin stimulates lipolysis in human and murine 3T3-L1 adipocytes. • Spexin inhibits lipogenesis and glucose uptake in human and murine 3T3-L1 adipocytes. • Spexin effect on adipocytes metabolism is mediated via GALR2 and GALR3 receptors. [ABSTRACT FROM AUTHOR]

Brown/beige adipocytes express uncoupling protein-1 (UCP1) that enables them to dissipate energy as heat. Systematic activation of this process can alleviate obesity. Human brown adipose tissues are interspersed in distinct anatomical regions including deep neck. We found that UCP1 enriched adipocytes differentiated from precursors of this depot highly expressed ThTr2 transporter of thiamine and consumed thiamine during thermogenic activation of these adipocytes by cAMP which mimics adrenergic stimulation. Inhibition of ThTr2 led to lower thiamine consumption with decreased proton leak respiration reflecting reduced uncoupling. In the absence of thiamine, cAMP-induced uncoupling was diminished but restored by thiamine addition reaching the highest levels at thiamine concentrations larger than present in human blood plasma. Thiamine is converted to thiamine pyrophosphate (TPP) in cells; the addition of TPP to permeabilized adipocytes increased uncoupling fueled by TPP-dependent pyruvate dehydrogenase. ThTr2 inhibition also hampered cAMP-dependent induction of UCP1, PGC1a, and other browning marker genes, and thermogenic induction of these genes was potentiated by thiamine in a concentration-dependent manner. Our study reveals the importance of amply supplied thiamine during thermogenic activation in human adipocytes which provides TPP for TPP-dependent enzymes not fully saturated with this cofactor and by potentiating the induction of thermogenic genes. [Display omitted] [ABSTRACT FROM AUTHOR]

The anti-obesity thyroid hormone, triiodothyronine (T3), and irisin, an exercise- and/or cold-induced myokine, stimulate thermogenesis and energy consumption while decreasing lipid accumulation. The involvement of ATP signaling in adipocyte cell function and obesity has attracted increasing attention, but the crosstalk between the purinergic signaling cascade and anti-obesity hormones lacks experimental evidence. In this study, we investigated the effects of T3 and irisin in the transcriptomics of membrane-bound purinoceptors, ectonucleotidase enzymes and nucleoside transporters participating in the purinergic signaling in cultured human adipocytes. The RNA-seq analysis revealed that differentiated adipocytes express high amounts of ADORA1, P2RY11, P2RY12, and P2RX6 gene transcripts, along with abundant levels of transcriptional products encoding to purine metabolizing enzymes (ENPP2, ENPP1, NT5E, ADA and ADK) and transporters (SLC29A1, SCL29A2). The transcriptomics of purinergic signaling markers changed in parallel to the upsurge of "browning" adipocyte markers, like UCP1 and P2RX5, after treatment with T3 and irisin. Upregulation of ADORA1, ADORA2A and P2RX4 gene transcription was obtained with irisin, whereas T3 preferentially upregulated NT5E, SLC29A2 and P2RY11 genes. Irisin was more powerful than T3 towards inhibition of the leptin gene transcription, the SCL29A1 gene encoding for the ENT1 transporter, the E-NPP2 (autotaxin) gene, and genes that encode for two ADP-sensitive P2Y receptors, P2RY1 and P2RY12. These findings indicate that anti-obesity irisin and T3 hormones differentially affect the purinergic signaling transcriptomics, which might point towards new directions for the treatment of obesity and related metabolic disorders that are worth to be pursued in future functional studies. • T3 and the myokine, irisin, promote transdifferentiation of human WAT into BAT. • These anti-obesity hormones differently affect purinergic signal transcriptomics. • Upregulation of ADORA1, ADORA2A and P2RX4 gene expression is preferential by irisin. • T3 preferentially upregulates NT5E, SLC29A2 and P2RY11 genes transcription. • Irisin (and T3) inhibits SCL29A1/ENT1, E-NPP2 and P2RY1/P2RY12 genes transcription. [ABSTRACT FROM AUTHOR]

Brown adipose tissue is a promising therapeutic target for opposing obesity, glucose intolerance and insulin resistance. The ability to modulate gene expression in mature brown adipocytes is important to understand brown adipocyte function and delineate novel regulatory mechanisms of non-shivering thermogenesis. The aim of this study was to optimize a lipofection-based small interfering RNA (siRNA) transfection protocol for efficient silencing of gene expression in mature brown adipocytes. We determined that a critical parameter was to deliver the siRNA to mature adipocytes by reverse transfection, i.e. transfection of non-adherent cells. Using this protocol, we effectively knocked down both high- and low-abundance transcripts in a model of mature brown adipocytes (WT-1) as well as in primary mature mouse brown adipocytes. A functional consequence of the knockdown was confirmed by an attenuated increase in uncoupled respiration (thermogenesis) in response to β-adrenergic stimulation of mature WT-1 brown adipocytes transfected with uncoupling protein 1 siRNA. Efficient gene silencing was also obtained in various mouse and human white adipocyte models (3T3-L1, primary mouse white adipocytes, hMADS) with the ability to undergo "browning." In summary, we report an easy and versatile reverse siRNA transfection protocol to achieve specific silencing of gene expression in various models of mature brown and browning-competent white adipocytes, including primary cells. [ABSTRACT FROM AUTHOR]

Second generation antipsychotics are useful for the treatment of schizophrenia, but concerns have been raised about the side effects of diabetes mellitus and obesity. Olanzapine, especially, is associated with more weight gain than the others. It has been reported that olanzapine promotes adipocyte-differentiation in rodents both in vivo and in vitro . In this study the effects of antipsychotics on human adipocytes were investigated by using human mesenchymal stem cells (hMSCs). When hMSCs were differentiated and treated with various antipsychotics, olanzapine and clozapine increased intracellular lipids. Olanzapine induced lipid accumulation in a dose-dependent manner. Proteomic analysis revealed that PLIN4 and several enzymes for lipid metabolism were increased in the hMSCs after olanzapine treatment. During adipocyte differentiation, olanzapine increased the protein expression of PLIN1, PLIN2 and PLIN4. These proteins are known to be associated with the initial stage of lipid droplet formation. Immunocytochemistry showed that olanzapine increased and enlarged the lipid droplets coated with PLIN1 and PLIN2 while PLIN4 was largely distributed in the cytosol. mRNA expression of PLIN2, but not PLIN1 or PLIN4, was increased by olanzapine. On the other hand, olanzapine did not alter the mRNA level of transcription regulators involved in adipocyte-differentiation or adipokines. The present study shows that olanzapine induced transient PLIN2 expression in hMSCs that could result in an accumulation of lipid droplets and overexpression of PLIN1 and PLIN4, providing information of possible interest for olanzapine-induced weight gain. [ABSTRACT FROM AUTHOR]

The development of obesity is becoming an international problem and the role of fructose is unclear. Studies using liver tissue and hepatocytes have contributed to the understanding of fructose metabolism. Excess fructose consumption also affects extra hepatic tissues including adipose tissue. The effects of fructose on human adipocytes are not yet fully characterized, although in vivo studies have noted increased adiposity and weight gain in response to fructose sweetened-beverages. In order to understand and predict the metabolic responses of adipocytes to fructose, this study examined differentiating and differentiated human adipocytes in culture, exposed to a range of fructose concentrations equivalent to that reported in blood after consuming fructose. A stable isotope based dynamic profiling method using [U-C]- d-fructose tracer was used to examine the metabolism and fate of fructose. A targeted stable isotope tracer fate association method was used to analyze metabolic fluxes and flux surrogates with exposure to escalating fructose concentration. This study demonstrated that fructose stimulates anabolic processes in adipocytes robustly, including glutamate and de novo fatty acid synthesis. Furthermore, fructose also augments the release of free palmitate from fully differentiated adipocytes. These results imply that in the presence of fructose, the metabolic response of adipocytes in culture is altered in a dose dependent manner, particularly favoring increased glutamate and fatty acid synthesis and release, warranting further in vivo studies. [ABSTRACT FROM AUTHOR]

Background: Functional brown adipose tissue (BAT), involved in energy expenditure, has recently been detected in substantial amounts in adults. Formerly overlooked BAT has now become an attractive anti-obesity target. Methods and results: Molecular characterization of human brown and white adipocytes, using a myriad of techniques including high-throughput RNA sequencing and functional assays, showed that PAZ6 and SW872 cells exhibit classical molecular and phenotypic markers of brown and white adipocytes, respectively. However, the pre-adipocyte cell line SGBS presents a versatile phenotype. A transit expression of classical brown markers such as UCP1 and PPARγ peaked and declined at day 28 post-differentiation initiation. Conversely, white adipocyte markers, including Tcf21, showed reciprocal behavior. Interestingly, leptin levels peaked at day 28 whereas the highest adiponectin mRNA levels were detected at day 14 of differentiation. Phenotypic analysis of the abundance and shape of lipid droplets were consistent with the molecular patterns. Accordingly, the oxidative capacity of SGBS adipocytes peaked on differentiation day 14 and declined progressively towards differentiation day 28. Conclusions: Our studies have unveiled a new phenotype of human adipocytes, providing a tool to identify molecular gene expression patterns and pathways involved in the conversion between white and brown adipocytes. [ABSTRACT FROM AUTHOR]

Objective Insulin-like-growth factor binding protein 2 (IGFBP-2) is thought to be a marker for the phosphatase and tensin homolog (PTEN) status and activity of the phosphatidylinositol 3-kinase (PI3K)/AKT/mammalian target of rapamycin (mTOR) pathway. We aimed to evaluate whether or not lipoma cells of a patient with a heterozygous deletion in the PTEN gene would produce more IGFBP-2 than PTEN non deficient control cells. Moreover, we analysed the influence of pharmacological inhibitors of the PI3K/AKT/mTOR pathway on IGFBP-2 production. Design PTEN deficient preadipocytes and control PTEN non deficient preadipocytes were differentiated in vitro and treated with the respective inhibitors. PTEN was transiently down regulated by siRNA in human preadipocytes. IGFBP-2 mRNA and protein expression and IGFBP-2 in culture supernatant were measured. Results PTEN deficient lipoma cells were found to produce IGFBP-2 during in vitro differentiation in comparable amounts to PTEN non deficient cells. In contrast, acute down regulation of PTEN in preadipocytes resulted in enhanced production of IGFBP-2. Incubation with the PI3K inhibitors LY294002 and wortmannin decreased IGFBP-2 mRNA and protein. Neither the mTOR complex 1 inhibitor rapamycin nor PD98059, an inhibitor of MEK (mitogen-activated protein kinase kinase), showed a significant effect on IGFBP-2 production. Conclusion IGFBP-2 production in PTEN deficient preadipocytes was not influenced by PTEN deficiency or by inhibition of mTORC1 and MAPK. In contrast, inhibition of PI3K decreased IGFBP-2 expression and secretion. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs) are small non-coding RNAs involved in the regulation of gene expression. MiR-1908 is a recently identified miRNA that is highly expressed in human adipocytes. However, it is not known what role of miR-1908 is involved in the regulation of human adipocytes. In this study, we demonstrate that the level of miR-1908 increases during the adipogenesis of human multipotent adipose-derived stem (hMADS) cells and human preadipocytes-visceral. Overexpression of miR-1908 in hMADS cells inhibited adipogenic differentiation and increased cell proliferation, suggesting that miR-1908 is involved in the regulation of adipocyte cell differentiation and metabolism, and, thus, may have an effect on human obesity. [ABSTRACT FROM AUTHOR]

Overweight (OW) and obese individuals are considered to be graded parts of the scale having increasing weight as a common feature. They may not, however, be part of the same continuum and may differ metabolically. In this study we applied an untargeted proteomic approach to compare protein abundances in mature adipocytes derived from the subcutaneous adipose tissue of overweight and morbidly obese female subjects to those of lean age matched controls. Mature adipocytes were isolated from liposuction samples of abdominal subcutaneous adipose tissue collected from both lean (L; n = 7, 23.3 ± 0.4 kg/m 2 ; mean BMI ± SD), overweight (OW; n = 8, 27.9 ± 0.6 kg/m 2 ; mean BMI ± SD) and morbidly obese (MOB; n = 7, 44.8 ± 3.8 kg/m 2 ; mean BMI ± SD) individuals. Total protein extracts were then compared by two-dimensional difference in gel electrophoresis (2D DIGE). One hundred and ten differentially expressed protein spots (i.e., fitting the statistical criteria ANOVA test, p < 0.05; fold-change ≥1.5) were detected, and of these, 89 were identified by MALDI-TOF mass spectrometry. Of these, 66 protein spots were common to both groups whereas 23 were unique to the MOB group. Significant differences were evident in the abundances of key proteins involved in glucose and lipid metabolism, energy regulation, cytoskeletal structure and redox control signaling pathways. Differences in the abundance of some chaperones were also evident. The differentially abundant proteins were investigated using Ingenuity Pathway Analysis (IPA) to establish their associations with known biological functions. The network identified in the OW group with the highest score relates to-: cell-to-cell signaling and interaction; in contrast, in the MOB group the major interacting pathways are associated with lipid metabolism, small molecule biochemistry and cancer. The differences in abundance of the differentially regulated proteins were validated by immunoblotting. These findings provide insights into metabolic differences in OW and MOB individuals. [ABSTRACT FROM AUTHOR]

Obesity is characterized by dysfunctional white adipose tissue (WAT) that ultimately may lead to metabolic diseases. Calorie restriction (CR) reduces the risk for age and obesity-associated complications. The impact of CR on obesity has been examined with human intervention studies, which showed alterations in circulating adipokines. However, a direct effect of CR on the human adipocyte secretome remains elusive. Therefore, the effect of a 96h low glucose CR on the secretion profile of in vitro cultured mature human SGBS adipocytes was investigated by using proteomics technology. Low-glucose CR decreased the adipocyte triglyceride contents and resulted in an altered secretion profile. Changes in the secretome indicated an improved inflammatory phenotype. In addition, several adipocyte-secreted proteins related to insulin resistance showed a reversed expression after low-glucose CR. Furthermore, 6 novel CR-regulated adipocyte-secreted proteins were identified. Since resveratrol (RSV) mimics CR we compared results from this study with data from our previous RSV study on the SGBS adipocyte secretome. The CR and RSV adipocyte secretomes partly differed from each other, although both treatment strategies lead to secretome changes indicating a less inflammatory phenotype. Furthermore, both treatments induced SIRT1 expression and resulted in a reversed expression of detrimental adipokines associated with metabolic complications. [ABSTRACT FROM AUTHOR]

Adipose tissue expansion during obesity is associated with increased macrophage infiltration. Macrophage- derived factors significantly alter adipocyte function, inducing inflammatory responses and decreasing insulin sensitivity. Identification of the major factors that mediate detrimental effects of macrophages on adipocytes may offer potential therapeutic targets. IL-1β, a proinflammatory cytokine, is suggested to be involved in the development of insulin resistance. This study investigated the role of IL-1β in macrophage-adipocyte cross-talk, which affects insulin signaling in human adipocytes. Using macrophage-conditioned (MC) medium and human primary adipocytes, we examined the effect of IL-1β antagonism on the insulin signaling pathway. Gene expression profile and protein abundance of insulin signaling molecules were determined, as was the production of proinflammatory cytokine/chemokines. We also examined whether IL-1β mediates MC medium-induced alteration in adipocyte lipid storage. MC medium and IL-1β significantly reduced gene expression and protein abundance of insulin signaling molecules, including insulin receptor substrate-1, phosphoinositide 3-kinase p85α, and glucose transporter 4 and phosphorylation of Akt. In contrast, the expression and release of the proinflammatory markers, including IL-6, IL-8, monocyte chemotactic protein-1, and chemokine (C-C motif) ligand 5 by adipocytes were markedly increased. These changes were significantly reduced by blocking IL-1β activity, its receptor binding, or its production by macrophages. MC mediuminhibited expression of the adipogenic factors and -stimulated lipolysis was also blunted with IL-1β neutralization. We conclude that IL-1β mediates, at least in part, the effect of macrophages on insulin signaling and proinflammatory response in human adipocytes. Blocking IL-1β could be beneficial for preventing obesity-associated insulin resistance and inflammation in human adipose tissue. [ABSTRACT FROM AUTHOR]

Abdominal obesity associates with cardiometabolic disease and an accumulation of lipids in the visceral adipose depot, whereas lipid accumulation in the subcutaneous depot is more benign. We aimed to further investigate whether the adipogenic properties where cell-intrinsic, or dependent on a depot-specific or obesity-produced microenvironment. We obtained visceral and subcutaneous biopsies from non-obese women ( n = 14) or women living with morbid obesity ( n = 14) and isolated adipose stem and progenitor cells (ASPCs) from the stromal vascular fraction of non-obese ( n = 13) and obese ( n = 13). Following in vitro differentiation into mature adipocytes, we observed a contrasting pattern with a lower gene expression of adipogenic markers and a higher gene expression of immunogenic markers in the visceral compared to the subcutaneous adipocytes. We identified the immunogenic factor BST2 as a marker for visceral ASPCs. The effect of obesity and insulin resistance on adipogenic and immunogenic markers in the in vitro differentiated cells was minor. In contrast, differentiation with exogenous Tumor necrosis factor resulted in increased immunogenic signatures, including increased expression of BST2 , and decreased adipogenic signatures in cells from both depots. Our data, from 26 women, underscore the intrinsic differences between human visceral and subcutaneous adipose stem and progenitor cells, suggest that dysregulation of adipocytes in obesity mainly occurs at a post-progenitor stage, and highlight an inflammatory microenvironment as a major constraint of human adipogenesis., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Mathur, Severinsen, Jensen, Naver, Schrölkamp, Laye, Watt, Nielsen, Krogh-Madsen, Pedersen and Scheele.)