White Adipose Tissue In Response Research Articles

Trauma-associated extracellular histones mediate inflammation via a MYD88-IRAK1-ERK signaling axis and induce lytic cell death in human adipocytes

Despite advances in the treatment and care of severe physical injuries, trauma remains one of the main reasons for disability-adjusted life years worldwide. Trauma patients often suffer from disturbances in energy utilization and metabolic dysfunction, including hyperglycemia and increased insulin resistance. White adipose tissue plays an essential role in the regulation of energy homeostasis and is frequently implicated in traumatic injury due to its ubiquitous body distribution but remains poorly studied. Initial triggers of the trauma response are mainly damage-associated molecular patterns (DAMPs) such as histones. We hypothesized that DAMP-induced adipose tissue inflammation contributes to metabolic dysfunction in trauma patients. Therefore, we investigated whether histone release during traumatic injury affects adipose tissue. Making use of a murine polytrauma model with hemorrhagic shock, we found increased serum levels of histones accompanied by an inflammatory response in white adipose tissue. In vitro, extracellular histones induced an inflammatory response in human adipocytes. On the molecular level, this inflammatory response was mediated via a MYD88-IRAK1-ERK signaling axis as demonstrated by pharmacological and genetic inhibition. Histones also induced lytic cell death executed independently of caspases and RIPK1 activity. Importantly, we detected increased histone levels in the bloodstream of patients after polytrauma. Such patients might benefit from a therapy consisting of activated protein C and the FDA-approved ERK inhibitor trametinib, as this combination effectively prevented histone-mediated effects on both, inflammatory gene activation and cell death in adipocytes. Preventing adipose tissue inflammation and adipocyte death in patients with polytrauma could help minimize posttraumatic metabolic dysfunction.

Activation of α7 nAChR improves white fat homeostasis and promotes beige adipogenesis and thermogenesis in obese mice

To investigate the effects of α7 nicotinic acetylcholine receptor (nAChR) agonist on β3-adrenoceptor agonist-induced impairment of white fat homeostasis and beige adipose formation and heat production in obese mice. Forty obese C57BL/6J mice were randomized into high-fat feeding group, β3-adrenoceptor agonist-treated model group, α7 nAChR agonist group, and α7 nAChR inhibitor group (n=10), with another 10 mice with normal feeding as the blank control group. White adipose tissue from the epididymis of the mice were sampled for HE staining of the adipocytes. The expression levels of TNF-α, IL-1β, IL-10 and TGF-β in the white adipose tissue were determined by ELISA, and the mRNA levels of iNOS, Arg1, UCP-1, PRDM-16 and PGC-1α were detected using RT-qPCR. Western blotting was performed to detect the expression levels of NF-κB P65, p-JAK2, p-STAT3 in the white adipose tissue. Compared with those in the blank control group, the mice with high-fat feeding showed significantly increased body weight, more fat vacuoles in the white adipose tissue, increased volume of lipid droplets in the adipocytes, upregulated iNOS mRNA expression and protein expression of TNF-α and IL-1β, and lowered expression of Arg-1 mRNA and IL-10 and TGF-β proteins (P < 0.01). Treatment with α7 nAChR significantly reduced mRNA levels of PRDM-16, PGC-1α and UCP-1, lowered TNF-α and IL-1β expressions, increased IL-10 and TGF-β expressions, and reduced M1/M2 macrophage ratio in the white adipose tissues (P < 0.05 or 0.01). Activation of α7 nAchR improves white adipose tissue homeostasis impairment induced by β3 agonist, promotes transformation of M1 to M2 macrophages, reduces inflammatory response in white adipose tissue, and promote beige adipogenesis and thermogenesis in obese mice.

Semi-Automated Isolation of the Stromal Vascular Fraction from Murine White Adipose Tissue Using a Tissue Dissociator.

The in vitro study of white, brown, and beige adipocyte differentiation enables the investigation of cell-autonomous functions of adipocytes and their mechanisms. Immortalized white preadipocyte cell lines are publicly available and widely used. However, the emergence of beige adipocytes in white adipose tissue in response to external cues is difficult to recapitulate to the full extent using publicly available white adipocyte cell lines. Isolation of the stromal vascular fraction (SVF) from murine adipose tissue is commonly executed to obtain primary preadipocytes and perform adipocyte differentiation. However, mincing and collagenase digestion of adipose tissue by hand can result in experimental variation and is prone to contamination. Here, we present a modified semi-automated protocol that utilizes a tissue dissociator for collagenase digestion to achieve easier isolation of the SVF, with the aim of reducing experimental variation, reducing contamination, and increasing reproducibility. The obtained preadipocytes and differentiated adipocytes can be used for functional and mechanistic analyses.

Circulating Asprosin Concentrations in Patients with Obesity and Carbohydrate Disturbances.

Asprosin is a fasting-induced glucogenic hormone, secreted by white adipose tissue in response to starvation. The aim of the current study was to determine the levels of asprosin in subjects from the entire spectrum of the carbohydrate metabolism. A total of 153 Causcasian subjects participated in this study: group 1, healthy volunteers; group 2, obese subjects without glycemic disturbances; group 3, subjects with prediabetes and group 4, patients with newly identified type 2 diabetes. Subject with body mass index≥30 kg/m2 and dysglycemia (prediabetes and diabetes) showed significantly high levels of asprosin (1.40 ng/ml [IQR=0.98-1.94]; 1.27 ng/ml [IQR=0.86-2.12]; 1.09 ng/ml [IQR=0.89-1.58]) compared to the control group (0.71 ng/ml [IQR=0.54-0.92]; p<0.001). Correlation analysis showed that serum asprosin also had significant positive associations with some anthropometric parameters, liver enzymes, fasting and post load glucose and insulin, LDL and triglycerides. Furthermore, we estimated a marked relationship between asprosin concentrations and intima media thickness of the common carotid artery as well as neuropathy disability and vibration sensitivity. The circulating asprosin levels for differentiating subjects with carbohydrate disturbances and those with obesity were determined by ROC analysis. The AUC for disturbances of the glucose metabolism was 0.672 (p<0.001; 95% CI=0.581-0.751) and for obesity AUC was 0.849 (p<0.001; 95% CI=0.785-0.919). Circulating asprosin could be used as a predictive factor for early carbohydrate disorders and might be a potential new therapeutic target for the treatment of dysglycemia and obesity. Further prospective studies are needed to confirm this observation.

Ponatinib modulates the metabolic profile of obese mice by inhibiting adipose tissue macrophage inflammation.

Obesity-induced metabolic syndrome is a rapidly growing conundrum, reaching epidemic proportions globally. Chronic inflammation in obese adipose tissue plays a key role in metabolic syndrome with a series of local and systemic effects such as inflammatory cell infiltration and inflammatory cytokine secretion. Adipose tissue macrophages (ATM), as one of the main regulators in this process, are particularly crucial for pharmacological studies on obesity-related metabolic syndrome. Ponatinib, a multi-targeted tyrosine kinase inhibitor originally used to treat leukemia, has recently been found to improve dyslipidemia and atherosclerosis, suggesting that it may have profound effect on metabolic syndrome, although the mechanisms underlying have not yet been revealed. Here we discovered that ponatinib significantly improved insulin sensitivity in leptin deficient obese mice. In addition to that, ponatinib treatment remarkably ameliorated high fat diet-induced hyperlipidemia and inhibited ectopic lipid deposition in the liver. Interestingly, although ponatinib did not reduce but increase the weight of white adipose tissue (WAT), it remarkably suppressed the inflammatory response in WAT and preserved its function. Mechanistically, we showed that ponatinib had no direct effect on hepatocyte or adipocyte but attenuated free fatty acid (FFA) induced macrophage transformation from pro-inflammatory to anti-inflammatory phenotype. Moreover, adipocytes co-cultured with FFA-treated macrophages exhibited insulin resistance, while pre-treat these macrophages with ponatinib can ameliorate this process. These results suggested that the beneficial effects of ponatinib on metabolic disorders are achieved by inhibiting the inflammatory phenotypic transformation of ATMs, thereby maintaining the physiological function of adipose tissue under excessive obesity. The data here not only revealed the novel therapeutic function of ponatinib, but also provided a theoretical basis for the application of multi-target tyrosine kinase inhibitors in metabolic diseases.

Knockout of ICAT in Adipose Tissue Alleviates Fibro-inflammation in Obese Mice.

TheE2 promoter binding factor 1 (E2F1) and the Wnt/β-catenin signaling are crucial in regulating metabolic homeostasis including obesity. Theβ-catenin interacting protein 1 (CTNNBIP1), also known as the inhibitor of β-catenin and TCF4 (ICAT), is required for E2F1 to inhibit the activity of β-catenin. However, the role of ICAT in E2F1 regulating obesity-related metabolic disorders remains unknown. In the present study, male adipose tissue-specific ICAT knockout (ICATadi-/-) C57BL/6J mice and control littermatesaged 6-8weeks were fed with high-fat diet (HFD) for 12weeks to explore the effect of ICAT on lipid metabolism and obesity-related disorders. Results showed that the adipose tissue-specific ICAT knockout had negligible effect on lipid metabolism, reflected by no difference in body weight, fat mass, and the expression of proteins involved in lipid metabolism in white adipose tissue (WAT) and the liver between the ICATadi-/- mice and the control littermate (ICATfl/fl) mice. However, the knockout of ICAT reduced inflammatory response in WAT and the liver. Additionally, Sirius red staining results showed that deletion of ICAT attenuated fibrosis and reduced mRNA levels of transforming growth factor β1(TGF-β1), matrix metallopeptidase 2 (Mmp2), Mmp3, and collagen, type V, alpha 1 (Col5a1) in WAT and the liver. These results suggested that knockout of ICAT improved the metabolic abnormalities of obese mice through attenuating adipose tissue and the liver inflammation as well as fibrosis. Our findings may provide a new insight to understand the role of ICAT in inflammation and fibrosis.

Time-of-day dependent effect of proanthocyanidins on adipose tissue metabolism in rats with diet-induced obesity.

Grape-seed proanthocyanidin extract (GSPE) improve white adipose tissue (WAT) expansion during diet-induced obesity. However, because adipose metabolism is synchronized by circadian rhythms, it is plausible to speculate that the bioactivity of dietary proanthocyanidins could be influenced by the time-of-day in which they are consumed. Therefore, the aim of the present study was to determine the interaction between zeitgeber time (ZT) and GSPE consumption on the functionality of WAT in rats with diet-induced obesity. Male Wistar rats were fed a cafeteria diet for 9 weeks. After 5 weeks, the animals were supplemented with 25 mg GSPE/kg for 4 weeks at the beginning of the light/rest phase (ZT0) or of the dark/active phase (ZT12). Body fat content was determined by nuclear magnetic resonance and histological analyses were performed in the epididymal (EWAT) and inguinal (IWAT) fat depots to determine adipocyte size and number. In addition, the expression of genes related to adipose metabolism and circadian clock function were analyzed by qPCR. GSPE consumption at ZT0 was associated with a potential antidiabetic effect without affecting adiposity and energy intake and downregulating the gene expression of inflammatory markers in EWAT. In contrast, GSPE consumption at ZT12 improved adipose tissue expansion decreasing adipocyte size in IWAT. In accordance with this adipogenic activity, the expression of genes involved in fatty acid metabolism were downregulated at ZT12 in IWAT. In turn, GSPE consumption at ZT12, but not at ZT0, repressed the expression of the clock gene Cry1 in IWAT. The interaction between ZT and GSPE consumption influenced the metabolic response of WAT in a tissue-specific manner. Understanding the impact of circadian clock on adipose metabolism and how this is regulated by polyphenols will provide new insights for the management of obesity.

Transcriptome profile of white, brown and beige adipose tissue

Adipose tissue dysregulation lies at the core metabolic syndrome. White adipose tissue (WAT) has the capacity to store energy, whereas brown adipose tissue (BAT) can dissipate energy through thermogenesis. Newly discovered beige adipocytes are brown like adipocytes recruited in WAT in response to b3‐adrenergic stimulation (CL316,243) or cold temperatures. Despite the important metabolic role, information regarding differential expression of molecular markers between different adipose tissue, especially beige and other depots is scarce. The objective of this study was to identify the shared and unique transcript signatures that characterize WAT, BAT and beige adipose tissue. We collected inguinal WAT (iWAT), gonadal WAT (gWAT) and BAT from male C57BL6/J mice fed a control diet (17% fat) for 13 weeks. Beige WAT (iWATCL) was isolated from mice treated with b3‐adrenergic agonist CL316, 243 for 7 days (1 mg/kg BW, ip). RNA sequencing was used to generate transcriptional profiles. Using gWAT as control, differentially expressed genes (DEGs) were determined with the DEseq2 and considered when the false discovery rate corrected P‐value was ≤ 0.05 and the fold change ≥ 4. Principal components analysis revealed that the transcriptional profiles differed across BAT, gWAT and iWAT and iWATCL. Across adipose tissues, 208 DEGs were shared. Between BAT and iWATCL, 223 DEGs were shared and were associated with muscle related pathways. Unique DEGs were 1130, 549, and 40 for BAT, iWAT, and iWATCL, respectively. Top upregulated unique genes included Lncbate10, Zic1, and Gm32468 in BAT; Dsc3, Cd22 and H2‐M2 in iWAT; and Cacng5, Mup11 and Mup9 in iWATCL. Among other processes, genes upregulated in BAT were associated with cellular respiration and fatty acid biosynthesis, in iWAT with immune responses and in iWATCL with sodium ion homeostasis and cellular response to lipids. Overall results show transcriptional heterogeneity across adipose tissues depots, especially the unique set of transcripts in CL316, 243 induced beige adipocytes that can serve specific markers.

IL-37 isoform D acts as an inhibitor of soluble ST2 to boost type 2 immune homeostasis in white adipose tissue

White adipose tissue (WAT) homeostasis substantiated by type 2 immunity is indispensable to counteract obesity and metabolic disorders. IL-33/suppression of tumorigenicity (ST) 2 signaling promotes type 2 response in WAT, while potential regulators remain to be discovered. We identified human IL-37 isoform D (IL-37D) as an effective trigger for ST2-mediated type 2 immune homeostasis in WAT. IL-37D transgene amplified ST2+ immune cells, promoted M2 macrophage polarization and type 2 cytokine secretion in WAT that mediate beiging and inflammation resolution, thereby increasing energy expenditure, reducing obesity and insulin resistance in high-fat diet (HFD)-fed mice. Mechanistically, either endogenous or exogenous IL-37D inhibited soluble ST2 (sST2) production from WAT challenged with HFD or TNF-α. Recombinant sST2 impaired the beneficial effects of IL-37D transgene in HFD-fed mice, characterized by damaged weight loss, insulin action, and type 2 cytokine secretion from WAT. In adipose-derived stem cells, IL-37D inhibited TNF-α-stimulated sST2 expression through IL-1 receptor 8 (IL-1R8)-dependent NF-κB inactivation. Collectively, human IL-37D suppresses sST2 to boost type 2 immune homeostasis in WAT, which may be a promising therapy target for obesity and metabolic disorders.

Lactobacillus rhamnosus Strain LRH05 Intervention Ameliorated Body Weight Gain and Adipose Inflammation via Modulating the Gut Microbiota in High-Fat Diet-Induced Obese Mice.

This study aims to investigate the underlying mechanism of a specific probiotic strain on suppression of adipogenesis and inflammatory response in white adipose tissue (WAT) of high-fat diet (HFD)-fed mice. Eight strains are screened in vitro for candidates of potential probiotics. Lactobacillus rhamnosus LRH05 (LRH05) and Lactobacillus reuteri LR47 (LR47) are screened out with lower triglyceride expression in vitro. The mice are fed a control diet (CD), HFD, or HFD supplemented with a dose of LRH05 or LR47 at 109 CFU per mouse per day for 10 weeks (n = 8), respectively. The results demonstrate that LRH05, but not LR47, significantly reduce body weight gain and the weight of WAT, as well as improve hepatic steatosis and glucose intolerance. LRH05 regulates the Mogat1, Igf-1, Mcp-1, and F4/80 mRNA expression and decreases macrophage infiltration in WAT. LRH05 shows an increase in butyric and propionic acid-producing bacteria, including Lachnoclostridium, Romboutsia, and Fusobacterium that is coincident with the increased fecal propionic acid and butyric acid levels. LRH05 shows a strain-specific effect on ameliorating the pro-inflammatory process by reducing inflammatory macrophage infiltration and the expression of inflammation-related genes in mice. Thus, LRH05 can be considered a potential probiotic strain to prevent obesity.

1208-P: Caloric Restriction Modified Factors Regulating Lipid Storage and Apoptosis in Inguinal but Not Epididymal Adipose Tissue of 24-Month-Old Male Rats

Calorie restriction (CR) and the resultant weight-loss are known to improve metabolic health, but the effects of CR on factors regulating white adipose tissue (WAT) metabolic function and remodeling remain unclear. The purpose of this study was to examine the effects of 8 weeks of CR on markers of lipid storage/release, endoplasmic reticulum (ER) stress, and apoptosis in inguinal and epididymal WAT collected from aged rats. We harvested inguinal and epididymal adipose tissue from 24-month-old male Fischer-344 x Brown Norway rats that were either fed an ad libitum chow diet (AL, n=6) or subjected to CR (consuming 35% below AL intake) of the same diet (n=6) for 8 weeks. Using standard immunoblotting methods, inguinal WAT from CR had a greater abundance of phosphorylated hormone sensitive lipase (pHSL) than AL (11.1±6.6 vs. 4.5±3.3 arbitrary units (AU); P=0.05) indicative of elevated lipase activity in CR, and a lower protein abundance of the esterification marker, diacylglycerol acyltransferase (DGAT; 1.0±0.2 vs. 2.4±0.6 AU; P&amp;lt;0.001). Interestingly, protein abundance of the ER stress marker, CCAAT-enhancer-binding protein homologous protein (CHOP) was also significantly lower in CR vs. AL (0.7±0.3 vs. 1.7±0.5 AU; P&amp;lt;0.01), and the protein abundance of Beclin, a marker of autophagy, tended to be lower in CR vs. AL (1.9±0.4 vs. 2.6±0.7 AU, for CR vs. AL; P=0.08). In contrast to these findings for inguinal WAT, there were no detectable differences in protein abundance for any of these markers in epididymal WAT from CR vs. AL. In summary, CR-mediated responses in WAT of aged rats appeared to be depot-specific and CR may contribute to important metabolic modifications and may impact cellular remodeling of inguinal adipose tissue. Disclosure P. Varshney: None. M. W. Schleh: None. C. Ahn: None. A. Zheng: None. E. B. Arias: None. G. D. Cartee: None. J. F. Horowitz: None. Funding National Institutes of Health (R01DK077966, P30DK089503, R01AG010026)

An immune‐neuron axis guides adipose tissue loss in cancer‐associated cachexia

Cancer-associated cachexia (CAC) is a hypermetabolic syndrome characterized by weight loss due to the atrophy of adipose tissue and skeletal muscle. CAC occurs in almost 80% of neoplastic diseases and causes death in more than 20% of cancer patients. The underlying mechanisms of the disease are poorly understood and no effective treatment for CAC is currently available. Recently, we and others demonstrated that a phenotypic switch from white- to beige adipocytes, a phenomenon called browning, accompanies CAC and contributes to increased energy dissipation. However, detailed mechanisms underlying this white to beige transition in CAC remained elusive. Here we show that cachexigenic tumors cause the activation of a type 2 immune response in white adipose tissue (WAT), which is functionally intertwined with neuron outgrowth, beta-adrenergic activation, and subsequent WAT browning. Immunohistochemical- and whole-mount immunofluorescence analyses revealed enhanced peripheral sympathetic neuron projections in cachectic WAT of mice and human patients. Using peripheral dopamine beta-hydroxylase (DBH) deficient mice, we demonstrate that local production of catecholamines by intra-adipose tissue neurons is responsible for increased lipolysis and initiation of the browning process. In vitro studies revealed that the increase in neuron arborization is promoted by alternatively activated macrophages which generate a neuroprotective environment covering the neurotoxic proinflammatory signals produced by cachexigenic tumors. In accordance with this concept, tumor-bearing mice that lack the interleukin-4 receptor exhibited a diminished type-2 immune response, attenuated neuronal outgrowth, and reduced browning of WAT compared to tumor-bearing wild type mice. Overall, our findings indicate that an immune–sympathetic neuron crosstalk increases local catecholamine production, lipolysis, and UCP-1 expression in WAT, thereby contributing to energy dissipation, and adipose tissue loss in CAC.

Curcumin improves insulin sensitivity and increases energy expenditure in high-fat-diet–induced obese mice associated with activation of FNDC5/irisin

ObjectiveCurcumin (Cur) has a beneficial role in preventing metabolic dysfunctions; however, the underlying mechanism are not yet fully understood. The aim of this study was to evaluate whether the beneficial metabolic effects of curcumin are associated with the regulation of energy metabolism and activation of fibronectin type 3 domain-containing protein 5 (FNDC5)/irisin. MethodsWe used cellular and molecular techniques to investigate the effects of Cur on C57 BL/6 mice that were fed either a control diet or a high-fat diet (HFD) with or without 0.2% Cur for 10 wk. Factors involved in energy metabolism, inflammatory responses, and insulin signaling, as well as the involvement of FNDC5/irisin pathway, were assessed. ResultsCur alleviated adiposity and suppressed inflammatory response in white adipose tissue (WAT) of HFD mice. Meanwhile, Cur administration increased plasma irisin concentration and improved insulin sensitivity of HFD mice. Cur increased the oxygen consumption and heat production and reduced respiratory exchange ratio (RES) in HFD mice, which were accompanied by the enhancement of metabolic activity in brown fat and inguinal WAT. Additionally, the improvement of basal metabolic rate by Cur may be partly regulated by the FNDC5/ p38 mitogen-activated protein kinase (p38 MAPK)/extracellular signal-related kinase (ERK) 1/2 pathway. ConclusionsThese findings demonstrated that dietary Cur alleviated diet-induced adiposity by improving insulin sensitivity and whole body energy metabolism via the FNDC5/p38 MAPK/ERK pathways.

Mitochondrial Unfolded Protein Responses in White Adipose Tissue: Lipoatrophy, Whole-Body Metabolism and Lifespan.

The mitochondrial unfolded protein response (UPRmt) is a stress response mediated by the expression of genes such as chaperones, proteases, and mitokines to maintain mitochondrial proteostasis. Certain genetically modified mice, which defect mitochondrial proteins specifically in adipocytes, developed atrophy of the white adipose tissue, resisted diet-induced obesity, and had altered whole-body metabolism. UPRmt, which has beneficial functions for living organisms, is termed “mitohormesis”, but its specific characteristics and detailed regulatory mechanism have not been elucidated to date. In this review, we discuss the function of UPRmt in adipose atrophy (lipoatrophy), whole-body metabolism, and lifespan based on the concept of mitohormesis.

Inhibition of cell death inducing DNA fragmentation factor-α-like effector c (CIDEC) by tumor necrosis factor-α induces lipolysis and inflammation in calf adipocytes

Dairy cows with ketosis exhibit signs of pronounced adipose tissue lipolysis and systemic inflammation, both of which exacerbate this metabolic disorder. In nonruminants, CIDEC plays a pivotal role in the formation of large unilocular lipid droplets. The present study aimed to ascertain the role of CIDEC in the lipolytic and inflammatory response of white adipose tissue (WAT) in vivo and in vitro. Subcutaneous adipose tissue and blood samples were collected from 15 healthy cows (blood β-hydroxybutyrate concentration < 1.2 mM) and 15 cows with clinical ketosis (blood β-hydroxybutyrate concentration > 3.0 mM) that had a similar number of lactations (median = 3, range = 2-4) and days in milk (median = 6 d, range = 3-9). Adipocytes isolated from 5 healthy Holstein calves (1 d old, female, 30-40 kg) were used for in vitro studies. Isolated adipocytes were treated with 0, 0.1, 1, or 10 ng/mL TNF-α for 3 h, transfected with CIDEC small interfering RNA for 48 h, or transfected with CIDEC overexpression adenovirus for 48 h followed by treatment with TNF-α (0.1 ng/mL) for 3 h. Serum concentrations of fatty acids were greater, and dry matter intake, milk yield, and serum glucose concentrations lower in cows with clinical ketosis. Protein and mRNA abundance of CIDEC were lesser in subcutaneous WAT of clinically ketotic versus healthy cows. Furthermore, the ratio of phosphorylated hormone sensitive lipase (p-LIPE) to LIPE, phosphorylated RELA (p-RELA) to RELA, and protein abundance of PNPLA2 and phosphorylated inhibitor of κBα (p-NFKBIA) were greater in dairy cows with clinical ketosis. The mRNA abundance of proinflammatory cytokines TNFA and IL1B were greater, and the anti-inflammatory cytokine IL10 was lower in WAT of dairy cows with clinical ketosis. In calf adipocytes, exogenous TNF-α (0.1, 1, or 10 ng/mL) decreased protein and mRNA abundance of CIDEC. In addition, exogenous TNF-α or knockdown of CIDEC reduced the secretion of the anti-inflammatory cytokine IL-10, but increased the ratio of p-LIPE to LIPE, p-RELA to RELA, protein abundance of PNPLA2 and p-NFKBIA, glycerol content, and the secretion of IL-1β in calf adipocytes. Overexpression of CIDEC in TNFα-treated adipocytes attenuated lipolysis and activation of the NF-κB signaling pathway. Overall, these data suggest that inhibition of lipid droplet-associated protein CIDEC by TNF-α contributes to the pronounced lipolysis and inflammation of calf adipocytes, and CIDEC is a relevant target in clinically ketotic cows.

The Ubiquitin Ligase SIAH2 Negatively Regulates Glucocorticoid Receptor Activity and Abundance.

Glucocorticoids are clinically essential drugs used routinely to control inflammation. However, a host of metabolic side effects manifests upon usage beyond a few days. In the present study, we tested the hypothesis that seven-in-absentia mammalian homolog-2 (SIAH2), a ubiquitin ligase that regulates adipogenesis, is important for controlling adipocyte size, inflammation, and the ability of adipose tissue to expand in response to a glucocorticoid challenge. Using mice with global deletion of SIAH2 exposed or not to corticosterone, we found that adipocytes are larger in response to glucocorticoids in the absence of SIAH2. In addition, SIAH2 regulates glucocorticoid receptor (GR) transcriptional activity and total GR protein abundance. Moreover, these studies reveal that there is an increased expression of genes involved in fibrosis and inflammatory signaling pathways found in white adipose tissue in response to glucocorticoids in the absence of SIAH2. In summary, this is the first study to identify a role for SIAH2 to regulate transcriptional activity and abundance of the GR, which leads to alterations in adipose tissue size and gene expression during in vivo exposure to glucocorticoids.

1939-P: Sex Dimorphism in Erythropoietin Regulation of Fat Mass

Erythropoietin (EPO), the cytokine required for erythropoiesis, contributes to metabolic regulation of fat mass and glycemic control. In male mice and ovariectomized female mice on high fat diet (HFD), EPO treatment increased hematocrit, improved glucose tolerance and attenuated body weight gain via reduced fat mass. Estradiol supplementation abrogated the EPO effect on body weight in ovariectomized female mice, providing evidence for estrogen related gender specific EPO action in metabolic regulation. To determine estrogen response in metabolic regulation with EPO treatment, we use the estrogen receptor α knockout (ERαKO) mice and mice with targeted deletion of estrogen receptor alpha in adipose tissue (ERαadipoKO). Male and female ERαKO mice on HFD exhibit increased fat mass and glucose intolerance. EPO treatment on HFD increased hematocrit in both wild type (WT) and ERαKO mice and reduced gain in fat mass in male mice as well as female ERαKO mice but did not affect fat mass in female WT mice. The improvement in glucose tolerance and insulin tolerance with EPO treatment during HFD was significantly greater in female ERaKO mice compared with female WT. We previously demonstrated that specific deletion of EPO receptor in adipose tissue in mice decreases glucose tolerance and insulin sensitivity and increases susceptibility to diet-induced obesity. Although no decrease in body weight was observed in female WT and ERαadipoKO with EPO treatment on HFD, female ERαadipoKO also showed significantly improved glycemic control with EPO treatment compared with WT control. EPO treatment decreased white fat associated gene expression, Psat1 and Wdnm-1like in subcutaneous fat pads from both female ERαKO mice and ERαadipoKO on HFD but not WT female mice. These results confirm the role of estrogen in the sex-differential EPO effect on fat mass regulation and suggest cross talk between EPO and estrogen signaling in metabolic homeostasis and fat mass regulation via estrogen response in white adipose tissue in female mice. Disclosure J. Lee: None. H. Rogers: None. C.T. Noguchi: None.

Differential Responses of White Adipose Tissue and Brown Adipose Tissue to Calorie Restriction During Aging.

Age-related adipose tissue dysfunction is potentially important in the development of insulin resistance and metabolic disorder. Caloric restriction (CR) is a robust intervention to reduce adiposity, improve metabolic health, and extend healthy life span. Both white adipose tissue (WAT) and brown adipose tissue (BAT) are involved in energy homeostasis. CR triggers the beiging of WAT in young mice; however, the effects of CR on beiging of WAT and function of BAT during aging are unclear. This study aimed to investigate how age and CR impact the beiging of WAT, the function of BAT, and metabolic health in mice. C57BL/6 mice were fed CR diet (40% less than the ad libitum [AL] diet) for 3 months initiated in young (3 months), middle-aged (12 months), and old (19 months) stage. We found age-related changes in different types of adipose tissue, including adipocyte enlargement, declined beiging of WAT, and declined thermogenic and β-oxidational function of BAT. Moreover, CR attenuated age-associated adipocyte enlargement and prevented the age-related decline in beiging potential of WAT. These protective effects on the beiging potential were significant in inguinal WAT at all three ages, which were significant in epididymal WAT at young and old age. In contrast, thermogenic and β-oxidational function of BAT further declined after CR in the young age group. In conclusion, our findings reveal the contribution of WAT beiging decline to age-related metabolic disorder and suggest nutritional intervention, specifically targeting WAT beiging, as an effective approach to metabolic health during aging.

Depot‐Specific Response of White Adipose Tissue to MicroRNA‐21 Ablation in Polycystic Ovary Syndrome

Background and aimPolycystic ovary syndrome (PCOS) is associated with obesity and insulin resistance. However, the molecular mechanisms that mediate this phenotype remain elusive. MicroRNA‐21 (miR‐21) is upregulated in PCOS and may be involved in the pathogenesis of the disease. We aim to study the role of miR‐21 in the deleterious metabolic effects associated with PCOS.MethodsThree week‐old miR‐21 knockout (miR21KO) or wild type (WT) female mice were treated with DHT (8 mg/silastic tube) or vehicle for 90 days (n=12/grp). Body composition was measured by EchoMRI. Energy expenditure (EE), oxygen consumption (VO2), and carbon dioxide production (VCO2) were measured by indirect calorimetry. Glucose homeostasis was measured by oral glucose tolerance test (OGTT). Gene expression was quantified by RT‐qPCR in subcutaneous (SC), retroperitoneal (RP) and mesenteric (M) white adipose tissue (WAT). Adipocyte size was evaluated by histological analysis.ResultsDHT increased body weight (25.07 ± 0.52 vs 21.79 ± 0.47 g, p&lt;0.05), fat mass (4.60 ± 0.46 vs 1.98 ± 0.12 g, p&lt;0.05) and adipocyte size (944 ±12 vs 513 ± 2 μm2, p&lt;0.05), impaired OGTT (29.70 ± 2.21 vs 22.33 ± 1.06 AUC, p&lt;0.05), and didn’t significantly change EE, VO2 or VCO2. Insulin signaling markers Irs1 and Slc2a4 expression was downregulated by DHT in all 3 WATs while Akt2 expression was upregulated. Adipogenesis markers Pparg and Adipoq were upregulated by DHT exclusively in the SC WAT. Inflammation marker Ccl2 was upregulated in all three WATs while Ikbkb was only upregulated in M WAT. Extracellular matrix (ECM) remodeling makers Mmp9 and Timp1 were upregulated by DHT only in RP and M WATs. Immune cells infiltration markers CD14, CD68, Itgam, Itgax, and CD86 were upregulated by DHT only in RP and M WATs.DHT‐treated miR21KO mice showed attenuated increase in body weight (23.84 ± 0.99 vs 25.07 ± 0.52 g, p&lt;0.05) with no change in fat mass or OGTT. Additionally, DHT‐treated miR21KO mice showed a trend to reduced EE, VO2 and VCO2 compared with DHT‐treated WT. Gene expression data showed lower expression of the insulin signaling marker Akt2, higher expression of ECM remodeling markers (Mmp9 and Timp1) and an exacerbated inflammatory and immune responses in M (Il‐18 and CD14) and SC (Il‐18 and CD86) WATs. Moreover, DHT‐treated miR21KO mice had exacerbated adipocyte hypertrophy compared with DHT‐treated WT controls (1,408 ± 16 vs 944 ±12 μm2, p&lt;0.05).Conclusion and significanceThese findings suggest that adipose tissue miR‐21 may have a protective role in PCOS and ameliorate some of the DHT‐mediated molecular changes observed in WAT in a fat depot‐specific manner. WAT‐specific modulation of miR‐21 levels could be a novel therapeutic approach for PCOS treatment.Support or Funding InformationSupported by National Institutes of Health grants P20 GM‐121334 (DGR and LLYC) and R21 DK‐113500 (DGR).

Lipokine 5-PAHSA Is Regulated by Adipose Triglyceride Lipase and Primes Adipocytes for De Novo Lipogenesis in Mice.

Branched esters of palmitic acid and hydroxystearic acid (PAHSA) are anti-inflammatory and antidiabetic lipokines that connect glucose and lipid metabolism. We aimed to characterize involvement of the 5-PAHSA regioisomer in the adaptive metabolic response of white adipose tissue (WAT) to cold exposure (CE) in mice, exploring the cross talk between glucose utilization and lipid metabolism. CE promoted local production of 5- and 9-PAHSAs in WAT. Metabolic labeling of de novo lipogenesis (DNL) using 2H2O revealed that 5-PAHSA potentiated the effects of CE and stimulated triacylglycerol (TAG)/fatty acid (FA) cycling in WAT through impacting lipogenesis and lipolysis. Adipocyte lipolytic products were altered by 5-PAHSA through selective FA re-esterification. The impaired lipolysis in global adipose triglyceride lipase (ATGL) knockout mice reduced free PAHSA levels and uncovered a metabolite reservoir of TAG-bound PAHSAs (TAG estolides) in WAT. Utilization of 13C isotope tracers and dynamic metabolomics documented that 5-PAHSA primes adipocytes for glucose metabolism in a different way from insulin, promoting DNL and impeding TAG synthesis. In summary, our data reveal new cellular and physiological mechanisms underlying the beneficial effects of 5-PAHSA and its relation to insulin action in adipocytes and independently confirm a PAHSA metabolite reservoir linked to ATGL-mediated lipolysis.