Regulation Of Adipocyte Function Research Articles

PAQR4 regulates adipocyte function and systemic metabolic health by mediating ceramide levels.

PAQR4 is an orphan receptor in the PAQR family with an unknown function in metabolism. Here, we identify a critical role of PAQR4 in maintaining adipose tissue function and whole-body metabolic health. We demonstrate that expression of Paqr4 specifically in adipocytes, in an inducible and reversible fashion, leads to partial lipodystrophy, hyperglycaemia and hyperinsulinaemia, which is ameliorated by wild-type adipose tissue transplants or leptin treatment. By contrast, deletion of Paqr4 in adipocytes improves healthy adipose remodelling and glucose homoeostasis in diet-induced obesity. Mechanistically, PAQR4 regulates ceramide levels by mediating the stability of ceramide synthases (CERS2 and CERS5) and, thus, their activities. Overactivation of the PQAR4-CERS axis causes ceramide accumulation and impairs adipose tissue function through suppressing adipogenesis and triggering adipocyte de-differentiation. Blocking de novo ceramide biosynthesis rescues PAQR4-induced metabolic defects. Collectively, our findings suggest a critical function of PAQR4 in regulating cellular ceramide homoeostasis and targeting PAQR4 offers an approach for the treatment of metabolic disorders.

Systems genetics analysis of human body fat distribution genes identifies adipocyte processes.

Excess abdominal fat is a sexually dimorphic risk factor for cardio-metabolic disease and is approximated by the waist-to-hip ratio adjusted for body mass index (WHRadjBMI). Whereas this trait is highly heritable, few causal genes are known. We aimed to identify novel drivers of WHRadjBMI using systems genetics. We used two independent cohorts of adipose tissue gene expression and constructed sex- and depot-specific Bayesian networks to model gene-gene interactions from 8,492 genes. Using key driver analysis, we identified genes that, in silico and putatively in vitro, regulate many others. 51-119 key drivers in each network were replicated in both cohorts. In other cell types, 23 of these genes are found in crucial adipocyte pathways: Wnt signaling or mitochondrial function. We overexpressed or down-regulated seven key driver genes in human subcutaneous pre-adipocytes. Key driver genes ANAPC2 and RSPO1 inhibited adipogenesis, whereas PSME3 increased adipogenesis. RSPO1 increased Wnt signaling activity. In differentiated adipocytes, MIGA1 and UBR1 down-regulation led to mitochondrial dysfunction. These five genes regulate adipocyte function, and we hypothesize that they regulate fat distribution.

Deletion of CD44 promotes adipogenesis by regulating PPARγ and cell cycle-related pathways.

CD44, a cell surface adhesion receptor and stem cell biomarker, is recently implicated in chronic metabolic diseases. Ablation of CD44 ameliorates adipose tissue inflammation and insulin resistance in obesity. Here, we investigated cell type-specific CD44 expression in human and mouse adipose tissue and further studied how CD44 in preadipocytes regulates adipocyte function. Using Crispr Cas9-mdediated gene deletion and lentivirus-mediated gene re-expression, we discovered that deletion of CD44 promotes adipocyte differentiation and adipogenesis, whereas re-expression of CD44 abolishes this effect and decreases insulin responsiveness and adiponectin secretion in 3T3-L1 cells. Mechanistically, CD44 does so via suppressing Pparg expression. Using quantitative proteomics analysis, we further discovered that cell cycle-regulated pathways were mostly decreased by deletion of CD44. Indeed, re-expression of CD44 moderately restored expression of proteins involved in all phases of the cell cycle. These data were further supported by increased preadipocyte proliferation rates in CD44-deficient cells and re-expression of CD44 diminished this effect. Our data suggest that CD44 plays a crucial role in regulating adipogenesis and adipocyte function possibly through regulating PPARγ and cell cycle-related pathways. This study provides evidence for the first time that CD44 expressed in preadipocytes plays key roles in regulating adipocyte function outside immune cells where CD44 is primarily expressed. Therefore, targeting CD44 in (pre)adipocytes may provide therapeutic potential to treat obesity-associated metabolic complications.

The Transcription Factor HOXA5: Novel Insights into Metabolic Diseases and Adipose Tissue Dysfunction.

The transcription factor HOXA5, from the HOX gene family, has long been studied due to its critical role in physiological activities in normal cells, such as organ development and body patterning, and pathological activities in cancer cells. Nonetheless, recent evidence supports the hypothesis of a role for HOXA5 in metabolic diseases, particularly in obesity and type 2 diabetes (T2D). In line with the current opinion that adipocyte and adipose tissue (AT) dysfunction belong to the group of primary defects in obesity, linking this condition to an increased risk of insulin resistance (IR) and T2D, the HOXA5 gene has been shown to regulate adipocyte function and AT remodeling both in humans and mice. Epigenetics adds complexity to HOXA5 gene regulation in metabolic diseases. Indeed, epigenetic mechanisms, specifically DNA methylation, influence the dynamic HOXA5 expression profile. In human AT, the DNA methylation profile at the HOXA5 gene is associated with hypertrophic obesity and an increased risk of developing T2D. Thus, an inappropriate HOXA5 gene expression may be a mechanism causing or maintaining an impaired AT function in obesity and potentially linking obesity to its associated disorders. In this review, we integrate the current evidence about the involvement of HOXA5 in regulating AT function, as well as its association with the pathogenesis of obesity and T2D. We also summarize the current knowledge on the role of DNA methylation in controlling HOXA5 expression. Moreover, considering the susceptibility of epigenetic changes to reversal through targeted interventions, we discuss the potential therapeutic value of targeting HOXA5 DNA methylation changes in the treatment of metabolic diseases.

1647-P: Novel Functions of CXCL12 as a Brown Adipocyte–Derived Secreted Factor

The capacity of brown adipose tissue (BAT) to regulate metabolism through cold-induced thermogenesis and secretion of molecules appeals as a potential therapeutic against metabolic disease. BAT secreted factors have gained interest as they may coordinate the adaptive response to cold and tissue remodeling. Yet, BAT secretory functions need deeper investigation before identifying druggable candidates. Thus, our group aimed to study secreted factors from active brown adipocytes that may sway adipocyte function and/or may orchestrate inter-cellular communications. For this, mRNA levels in mature adipocytes of brown, beige and white adipose depots from mice exposed to 21 days of cold were evaluated using RNA sequencing, and bioinformatic analysis was used to filter for potentially secreted factors. Cxcl12 was found to be the most cold-induced chemokine specifically in BAT. mRNA expression analysis of Cxcl12 by qPCR and fluorescence in-situ hybridization revealed its enrichment in brown adipocytes upon cold. Cold also increased CXCL12 levels in BAT media but not plasma, indicating a potential local action. Cxcl12 knockdown in active brown adipocytes impaired thermogenesis, estimated by Ucp1 gene expression, norepinephrine-induced lipolysis and mitochondrial respiration, despite intact β-adrenergic signaling, as estimated by immunoblotting of phosphorylated proteins. Together, these data suggest that Cxcl12 regulates adipocyte function independently from the β-adrenergic pathway. Additionally, CXCL12 exerted inter-cellular crosstalks by promoting macrophage chemotaxis and neurite outgrowth in vitro. Here we present CXCL12 as a novel brown adipocyte, cold-induced secreted factor involved in adipocyte function and inter-cellular crosstalk within BAT. Further characterization of the role of CXCL12 and other secreted factors in BAT will provide valuable knowledge on the adaptive mechanisms of BAT to cold and may open new avenues to leverage BAT functions for the treatment of metabolic disease. Disclosure M.Agueda oyarzabal: None. M.Tozzi: None. T.W.Schwartz: None. C.C.Schéele: Research Support; Novo Nordisk A/S. B.Emanuelli: None. K.Plucinska: None. M.Isidor: None. L.R.Ingerslev: None. P.Petersen: None. O.Dmytriyeva: None. J.Henningsen: None. E.Brown: None. K.Rupar: None. Funding Novo Nordisk Foundation (18CC0034900, 19SA0035436); Copenhagen Bioscience PhD Program

The role of somatosensory innervation of adipose tissues

Adipose tissues communicate with the central nervous system to maintain whole-body energy homeostasis. The mainstream view is that circulating hormones secreted by the fat convey the metabolic state to the brain, which integrates peripheral information and regulates adipocyte function through noradrenergic sympathetic output1. Moreover, somatosensory neurons of the dorsal root ganglia innervate adipose tissue2. However, the lack of genetic tools to selectively target these neurons has limited understanding of their physiological importance. Here we developed viral, genetic and imaging strategies to manipulate sensory nerves in an organ-specific manner in mice. This enabled us to visualize the entire axonal projection of dorsal root ganglia from the soma to subcutaneous adipocytes, establishing the anatomical underpinnings of adipose sensory innervation. Functionally, selective sensory ablation in adipose tissue enhanced the lipogenic and thermogenetic transcriptional programs, resulting in an enlarged fat pad, enrichment of beige adipocytes and elevated body temperature under thermoneutral conditions. The sensory-ablation-induced phenotypes required intact sympathetic function. We postulate that beige-fat-innervating sensory neurons modulate adipocyte function by acting as a brake on the sympathetic system. These results reveal an important role of the innervation by dorsal root ganglia of adipose tissues, and could enable future studies to examine the role of sensory innervation of disparate interoceptive systems.

Toll-like Receptor 7 (TLR7) Is Expressed in Adipocytes and the Pharmacological TLR7 Agonist Imiquimod and Adipocyte-Derived Cell-Free Nucleic Acids (cfDNA) Regulate Adipocyte Function.

Endosome-localized Toll-like receptors (TLRs) 3 and 9 are expressed and functionally active in adipocytes. The functionality and role of TLR7 in adipocyte biology and innate immunity of adipose tissue (AT) is poorly characterized. We analyzed TLR7 mRNA and protein expression in murine 3T3-L1 and primary adipocytes, in co-cultures of 3T3-L1 adipocytes with murine J774A.1 monocytes and in human AT. The effects of TLR7 agonists imiquimod (IMQ) and cell-free nucleic acids (cfDNA) on adipokine concentration in cell-culture supernatants and gene expression profile were investigated. We found that TLR7 expression is strongly induced during adipocyte differentiation. TLR7 gene expression in adipocytes and AT stroma-vascular cells (SVC) seems to be independent of TLR9. IMQ downregulates resistin concentration in adipocyte cell-culture supernatants and modulates gene expression of glucose transporter Glut4. Adipocyte-derived cfDNA reduces adiponectin and resistin in cell-culture supernatants and potentially inhibits Glut4 gene expression. The responsiveness of 3T3-L1 adipocytes to imiquimod is preserved in co-culture with J774A.1 monocytes. Obesity-related, adipocyte-derived cfDNA engages adipocytic pattern recognition receptors (PRRs), modulating AT immune and metabolic homeostasis during adipose inflammation.

IL-25 directly modulates adipocyte function and inflammation through the regulation of adiponectin.

This study aimed to investigate the direct role of IL-25 in modulating adipocyte function during homeostasis and low-grade inflammation induced by lipopolysaccharide (LPS). The 3T3-L1 preadipocyte cell lines and primary cultures of adipose-derived stromal vascular precursor cells of wild-type and IL-17RB-deficient mice were used to determine the direct function of IL-25. The expression of IL-17RB in differentiating adipocyte was determined using real-time PCR and flow cytometry analysis. The effect of IL-25 on lipid accumulation, triglyceride content, lipolysis, glucose uptake, and adipokine expression in the mature adipocytes was evaluated. IL-25 modulating the expression of inflammatory cytokines in adipocytes induced by low dose LPS was determined using real-time PCR and ELISA. The receptor for IL-25 was up-regulated during adipocyte differentiation and IL-25 directly modulated adipocyte function by reducing lipid accumulation and triglyceride concentration and enhancing lipolysis without affecting an insulin-stimulated glucose uptake. Interestingly, IL-25 induced adiponectin secretion through the PI3K/AKT signaling pathway. In 3T3-L1 adipocytes under low-grade inflammation, IL-25 attenuated the expression of IL-6 and CCL5 through the induction of adiponectin. Our studies suggest that IL-25 directly regulates adipocyte function by maintaining the adiponectin level during homeostasis and by alleviating inflammatory response through the regulation of adiponectin during low-grade inflammation in adipocytes.

Baicalin attenuates diet-induced obesity partially through promoting thermogenesis in adipose tissue

BackgroundAdipose tissues have essential roles on energy homeostasis and the development of metabolic syndrome and obesity, they have become critical targets for treating obesity and metabolic disorders. Baicalin is a flavonoid that derived from the root of Scutellaria baicalensis, and it has been reported to take part in the regulation of adipocyte function. All these highlighted the potential of baicalin in the regulation of fat accumulation and obesity. Yet the impact of baicalin on thermogenic function of adipocytes remains to be deciphered. ObjectiveThis study aims to explore the anti-obesity effects of baicalin. Materials & MethodsThe level of mRNA was detected by qRT-PCR and the protein expression level was examined by western blot. H&E staining was used for the observation of the structure of adipose tissue. Serum triglyceride and insulin levels were detected by commercial test kits. ResultsOur data demonstrated that baicalin up-regulates the expression of UCP1 and PGC1a in a dose-dependent manner in vitro. Baicalin also increases ERK phosphorylation, and the increased expression of UCP1 and PGC1a in adipocytes could be inhibited by an ERK inhibitor, U0126. Moreover, dietary baicalin ameliorates high fat diet (HFD)-induced obesity without affecting food intake. In addition, dietary baicalin inhibits adipocyte hypertrophy and enhances thermogenic gene program in sWAT and intrascapular brown adipose tissue (iBAT) in vivo. Discussion & ConclusionBaicalin prevents HFD-induced obesity partially through promoting adipocyte thermogenesis. Baicalin may be a promising compound against human obesity and related metabolic diseases.

Adipocyte CAMK2 deficiency improves obesity-associated glucose intolerance

ObjectiveObesity-related adipose tissue dysfunction has been linked to the development of insulin resistance, type 2 diabetes, and cardiovascular disease. Impaired calcium homeostasis is associated with altered adipose tissue metabolism; however, the molecular mechanisms that link disrupted calcium signaling to metabolic regulation are largely unknown. Here, we investigated the contribution of a calcium-sensing enzyme, calcium/calmodulin-dependent protein kinase II (CAMK2), to adipocyte function, obesity-associated insulin resistance, and glucose intolerance. MethodsTo determine the impact of adipocyte CAMK2 deficiency on metabolic regulation, we generated a conditional knockout mouse model and acutely deleted CAMK2 in mature adipocytes. We further used in vitro differentiated adipocytes to dissect the mechanisms by which CAMK2 regulates adipocyte function. ResultsCAMK2 activity was increased in obese adipose tissue, and depletion of adipocyte CAMK2 in adult mice improved glucose intolerance and insulin resistance without an effect on body weight. Mechanistically, we found that activation of CAMK2 disrupted adipocyte insulin signaling and lowered the amount of insulin receptor. Further, our results revealed that CAMK2 contributed to adipocyte lipolysis, tumor necrosis factor alpha (TNFα)–induced inflammation, and insulin resistance. ConclusionsThese results identify a new link between adipocyte CAMK2 activity, metabolic regulation, and whole-body glucose homeostasis.

Voluntary exercise augments gene transcription associated with futile cycling in white adipocytes from lean and obese mice

Adipose tissue (AT) is a major node that links obesity and cardiometabolic disorders and thus is an important area of focus for studying obesity and related diseases. While a well-balanced diet and regular physical activity are known to combat negative effects of obesity and improve AT function, little is known about how exercise influences adipocytesgene expression. We determined how diet and exercise regulate the adipocyte transcriptome in mice. Eight-week-old male mice were given either a high-fat diet (HF) or low-fat diet (LF), and within each group, either remained sedentary (Sed) or were given access to a running wheel (Ex). One group of mice was fed a HF, but their caloric intake was restricted so that they were weight-matched (WM) to the HF Ex group of mice to determine the effects of physical activity while eliminating differences in body weight. Following eight weeks of interventions, mice were euthanized, and adipocytes were isolated from epididymal AT (eAT). Next Generation RNA transcriptomics were conducted on adipocytes from eAT. Physically active mice had lower body weight and fat mass relative to Sed ad libitum fed animals, regardless of diet. By design, body mass was not different between HF Ex and HF WM animals, however fat mass was 36% lower in HF Ex compared to HF WM mice. KEGG pathway analysis revealed that genes linked with lysosomal cation transport and protease/lipase activity (Atp6v0d2, Atp6v1b2, Ctss, Ctsk, Lipa) were most highly upregulated by HF-induced obesity vs lean LF-fed mice; whereas HF Ex mice downregulated genes in this pathway. Physically active mice upregulated genes associated with oxidative phosphorylation (Atp8, Nd4l, Nd3, Cytb, Co1, Co3), fatty acid degradation (Ehhadh, Hadh, Echs1), fatty acid biosynthesis (Fasn, Me1, Acaca, Acly) and fatty acid esterification/glycerol metabolism (Pcx, Pck1, Gpd1, Mogat1, Dgat1) compared with sedentary animals, irrespective of diet. Compared to HF Sed mice, HF WM animals upregulated genes pertinent to oxidative phosphorylation and fatty acid degradation; whereas, relative to HF Ex, genes involved in lipogenesis and fatty acid esterification were lower in HFD WM animals. Collectively, these data reveal transcriptional changes upregulated by chronic exercise in isolated adipocytes that reflect a phenotype consistent with lipid turnover and oxidative metabolism, which is hypothesized to decrease stagnant lipid pools associated with physical inactivity in both lean and obese states. These preliminary findings provide a framework for future hypothesis-driven experiments to interrogate the mechanisms by which exercise regulates adipocyte function in vivo.

Type VI collagen and its cleavage product, endotrophin, cooperatively regulate the adipogenic and lipolytic capacity of adipocytes

ObjectiveObesity-induced adipose tissue remodeling is closely associated with systemic insulin resistance. However, the mechanistic involvement of adipocyte-derived extracellular matrix proteins under pathophysiological conditions remains unclear. Our aim was to investigate the distinctive contributions of each chain of type VI collagens (Col6) and its cleavage protein endotrophin to adipocyte functions and insulin sensitivity. MethodsCol6 comprises three alpha chains: Col6a1, Col6a2, and Col6a3. We generated Col6a1-, Col6a2-, and Col6a3-deficient 3T3-L1 adipocytes using the CRISPR-Cas9 system as well as a novel Col6a3-deficient (Col6a3KO) mouse model for loss-of-function studies. Adenoviral-endotrophin and adipocyte-specific doxycycline-inducible endotrophin transgenic mice were utilized for the gain-of-function analysis. ResultsThe holo-Col6 fibrils were found to be required for mature adipocyte differentiation. Only Col6a3-deficient 3T3-L1 adipocytes showed decreased inflammation and basal adipocyte lipolysis and prevented ER-stress-induced insulin resistance. Consistently, Col6a3KO mice showed decreased adipocyte size and fat mass of epididymal adipose tissues due to a defect in adipogenic and lipolytic capacity of adipocytes. Beyond the structural role of Col6a3, overexpression of endotrophin in obese mice further augmented insulin resistance, which was tightly associated with a significant increase in lipolysis, inflammation, and cellular apoptosis in adipose tissues, whereas this showed a limited effect on adipogenesis. ConclusionsThese novel findings corroborate our previous observations suggesting that adipose tissue extracellular matrix regulates adipocyte function and insulin sensitivity in pathophysiological conditions. Mechanistically, holo-Col6 fibrils and their signaling derivative endotrophin govern adipocyte function independently of their role as structural supports via MAPK signaling pathways, and the latter could be an important metabolic effector in obesity-related metabolic diseases.

1694-P: Beta-Arrestin 1 Suppresses Myogenic Reprogramming of Brown Fat to Maintain Euglycemia

Background: Adipocyte dysfunction links obesity to type 2 diabetes (T2D). To develop novel antidiabetic drugs, it is essential to delineate the signaling pathways that regulate adipocyte function under physiological and pathophysiological conditions. Recent studies have shown that β-arrestin-1 and -2 (barr1 and barr2, respectively), two intracellular signaling proteins known for their ability to modulate signaling via G protein-coupled receptors (GPCRs), regulate many important metabolic functions, sometimes even in a GPCR-independent fashion. Objective: To understand the role of barr1 in adipocyte function, obesity, and whole body glucose and energy homeostasis. Methods: To assess the role of barr1 in regulating adipocyte function, we used Cre-lox technology to generate mutant mice that lacked barr1 selectively in adipocytes(adipo-barr1-KO). These mutant mice, together with their control littermates, were subjected to a series of metabolic tests. Moreover, we used various molecular and biochemical techniques to probe the molecular mechanism through which barr1 modulates adipocyte function. Results: We found that adipo-barr1-KO mice showed greatly impaired glucose tolerance and insulin sensitivity when consuming an obesogenic diet. In contrast, transgenic mice that overexpressed barr1 in adipocytes were protected against the metabolic deficits caused by a high-calorie diet. At the cellular level, barr1 deficiency led to a myogenic re-programming of brown adipose tissue (BAT), causing elevated plasma myostatin levels, which in turn led to impaired insulin signaling in peripheral tissues. Additional in vivo studies indicated that barr1-mediated suppression of myostatin expression by BAT is required for maintaining euglycemia. Conclusion: Our data suggest that strategies aimed at enhancing barr1 expression or activity in BAT may prove beneficial for the treatment of T2D. Disclosure S. Pydi: None. S. Jain: None. L.F. Barella: None. J. Meister: None. L. Wang: None. Y. Cui: None. O. Gavrilova: None. J. Wess: None. Funding National Institute of Diabetes and Digestive and Kidney Diseases

RNAseq in fat pads from Alstrom syndrome 1 (ALMS1) Knockout rats support a role for ALMS1 in leptin release and fat metabolism in adipocytes.

Inactivating mutations in the ALMS1 gene in humans cause early onset obesity and metabolic syndrome. Deletion of ALMS1 in rodents induces early onset obesity and insulin resistance on normal chow. The mechanisms by which ALMS1 cause obesity are not clear but likely involve the development of leptin resistance since adult ALMS1 KO rats are hyperphagic and hyperleptinemic. It is not known whether ALMS1 controls leptin release and expression in adipocytes prior to development of obesity. ALMS1 has been associated to ciliary function which may play a role in adipogenesis. We hypothesized that deletion of ALMS1 would alter gene pathways involved in fat metabolism and cilia formation in young ALMS1 KO rats, which should contribute to hyperleptinemia. To study this, we obtained abdominal fat pads from young ALMS1 KO rats shortly after weaning (7 weeks) and prior to changes in body weight. We found that plasma leptin levels were higher in young ALMS1 KO rats (ALMS1: 3530±300 vs. WT: 172±16 pg/ml, p<0.05) despite no differences in body weight or baseline glucose. Leptin release (normalized to fat weight) was 120% higher (wild type: 30±8, ALMS1‐KO: 56± 15 pg/mg tissue/2h) in abdominal fat pads from ALMS1 KO rats. To study the genes involved in higher leptin release we performed RNAseq in freshly isolated abdominal fat pads. 12,860 genes were identified and quantified. In fat pads from ALMS1 KO there were 163 upregulated and 237 significantly downregulated genes. Leptin mRNA was significantly increased (1.68 Log2 fold, p<0.001, n=4) as were several genes involved in lipogenesis (Elovl6, Scd1, Scd2, Mogat2, Acly, Lpl, Dgat2, Fasn). Ingenuity pathway analysis indicated that the top canonical pathways impacted were: oleate biosynthesis, glycolysis I, hepatic fibrosis, and clathrin‐mediated endocytosis signaling. Importantly, at least 10 ciliary genes were significantly decreased (Traf3ip1, Pard6g, Cngb1, Ak7, Ttll1, Mlf1, Ccdc162, Dnah6, Ccdc187, Foxj1) suggesting a role for ALMS1 in ciliary function. To examine this, we immunolabeled cilia in abdominal adipocytes with gamma acetylated tubulin antibodies, and examined sections by confocal microscopy. Short cilia (1–2 μm) were observed in both WT and ALMS1 KO adipocytes. We concluded that ALMS1 is likely involved in leptin release from adipocytes. It is not clear whether increased leptin expression and release is secondary to neuroendocrine alterations or alterations in cilia‐induced signaling. Our data also point to an important role for ALMS1 in regulating adipocyte function and leptin physiology.Support or Funding InformationHenry Ford Hospital Research Fund

Regulation of Adipsin Expression by Endoplasmic Reticulum Stress in Adipocytes.

Adpsin is an adipokine that stimulates insulin secretion from β-cells and improves glucose tolerance. Its expression has been found to be markedly reduced in obese animals. However, it remains unclear what factors lead to downregulation of adipsin in the context of obesity. Endoplasmic reticulum (ER) stress response is activated in various tissues under obesity-related conditions and can induce transcriptional reprogramming. Therefore, we aimed to investigate the relationship between adipsin expression and ER stress in adipose tissues during obesity. We observed that obese mice exhibited decreased levels of adipsin in adipose tissues and serum and increased ER stress markers in adipose tissues compared to lean mice. We also found that ER stress suppressed adipsin expression via adipocytes-intrinsic mechanisms. Moreover, the ER stress-mediated downregulation of adipsin was at least partially attributed to decreased expression of peroxisome proliferator-activated receptor γ (PPARγ), a key transcription factor in the regulation of adipocyte function. Finally, treatment with chemical chaperones recovered the ER stress-mediated downregulation of adipsin and PPARγ in vivo and in vitro. Our findings suggest that activated ER stress in adipose tissues is an important cause of the suppression of adipsin expression in the context of obesity.

The role of ADP-ribose metabolism in metabolic regulation, adipose tissue differentiation, and metabolism.

Poly(ADP-ribose) polymerases (PARPs or ARTDs), originally described as DNA repair factors, have metabolic regulatory roles. PARP1, PARP2, PARP7, PARP10, and PARP14 regulate central and peripheral carbohydrate and lipid metabolism and often channel pathological disruptive metabolic signals. PARP1 and PARP2 are crucial for adipocyte differentiation, including the commitment toward white, brown, or beige adipose tissue lineages, as well as the regulation of lipid accumulation. Through regulating adipocyte function and organismal energy balance, PARPs play a role in obesity and the consequences of obesity. These findings can be translated into humans, as evidenced by studies on identical twins and SNPs affecting PARP activity.

The Rho GTPase RND3 regulates adipocyte lipolysis

BackgroundAdipose tissue plays a crucial role in diet- and obesity-related insulin resistance, with implications for several metabolic diseases. Identification of novel target genes and mechanisms that regulate adipocyte function could lead to improved treatment strategies. RND3 (RhoE/Rho8), a Rho-related GTP-binding protein that inhibits Rho kinase (ROCK) signaling, has been linked to diverse diseases such as apoptotic cardiomyopathy, heart failure, cancer and type 2 diabetes, in part by regulating cytoskeleton dynamics and insulin-mediated glucose uptake. ResultsWe here investigated the expression of RND3 in adipose tissue in human obesity, and discovered a role for RND3 in regulating adipocyte metabolism. In cross-sectional and prospective studies, we observed 5-fold increased adipocyte levels of RND3 mRNA in obesity, reduced levels after surgery-induced weight loss, and positive correlations of RND3 mRNA with adipocyte size and surrogate measures of insulin resistance (HOMA2-IR and circulating triglyceride/high-density lipoprotein cholesterol (TAG/HDL-C) ratio). By screening for RND3-dependent gene expression following siRNA-mediated RND3 knockdown in differentiating human adipocytes, we found downregulation of inflammatory genes and upregulation of genes related to adipocyte ipolysis and insulin signaling. Treatment of adipocytes with tumor necrosis factor alpha (TNFα), lipopolysaccharide (LPS), hypoxia or cAMP analogs increased RND3 mRNA levels 1.5–2-fold. Functional assays in primary human adipocytes confirmed that RND3 knockdown reduces cAMP- and isoproterenol-induced lipolysis, which were mimicked by treating cells with ROCK inhibitor. This effect could partly be explained by reduced protein expression of adipose triglyceride lipase (ATGL) and phosphorylated hormone-sensitive lipase (HSL). ConclusionWe here uncovered a novel differential expression of adipose RND3 in obesity and insulin resistance, which may at least partly depend on a causal effect of RND3 on adipocyte lipolysis.

Adipocyte \u03b2-arrestin-2 is essential for maintaining whole body glucose and energy homeostasis

β-Arrestins are major regulators of G protein-coupled receptor-mediated signaling processes. Their potential roles in regulating adipocyte function in vivo remain unexplored. Here we report the novel finding that mice lacking β-arrestin-2 (barr2) selectively in adipocytes show significantly reduced adiposity and striking metabolic improvements when consuming excess calories. We demonstrate that these beneficial metabolic effects are due to enhanced signaling through adipocyte β3-adrenergic receptors (β3-ARs), indicating that barr2 represents a potent negative regulator of adipocyte β3-AR activity in vivo. Interestingly, essentially all beneficial metabolic effects caused by adipocyte barr2 deficiency are absent in adipocyte barr2-PRDM16 double KO mice, indicating that the metabolic improvements caused by the lack of barr2 in adipocytes are mediated by the browning/beiging of white adipose tissue. Our data support the novel concept that ‘G protein-biased’ β3-AR agonists that do not promote β3-AR/barr2 interactions may prove useful for the treatment of obesity and related metabolic disorders.

1781-P: Adipocyte GI Signaling Regulates Whole-Body Glucose Homeostasis and Insulin Sensitivity

Adipocyte dysfunction links obesity to insulin resistance and type 2 diabetes (T2D). Receptor-mediated activation of heterotrimeric G proteins plays a pivotal role in regulating adipocyte function. Little is known about the potential in vivo metabolic roles of Gi-type G proteins expressed by adipocytes, primarily due to the lack of suitable animal models. To address this issue, we generated adipocre-ROSA 26PTXmice in which Gisignaling was selectively disrupted in adipocytes ('adipo-Gi KO mice'). Compared to control mice, adipo-Gi KO mice displayed increased lipolysis and decreased fat mass when maintained on regular chow or a high fat diet (HFD). Moreover, HFD adipo-Gi KO mice showed impaired glucose tolerance and reduced insulin sensitivity. In vitro studies with primary adipocytes prepared from adipo-Gi KO mice indicated that the lack of Gisignaling led to impaired insulin signaling and reduced insulin-stimulated glucose uptake. In parallel, we also used a chemo-genetic approach to generate a mouse line selectively expressing a CNO-sensitive, Gi-coupled DREADD (designer receptor exclusively activated by a designer drug) in adipocytes (adipo-GiD mice). CNO-induced activation of adipocyte Gi signaling in adipo-GiD mice significantly improved glucose tolerance and insulin sensitivity independent of the diet that the mice consumed (regular chow or HFD). In vivo 14C-2-deoxy-glucose uptake assays demonstrated that activation of adipocyte Gisignaling enhanced insulin-induced glucose uptake in adipose tissues. Stimulation of adipocyte Gisignaling activation also further enhanced insulin signaling in adipose tissues. Taken together, our data clearly demonstrate that adipocyte Gisignaling is a critical regulator of insulin signaling in adipocytes and is critical for the maintenance of glucose homeostasis. A better understanding of the mechanisms through which Gipromotes adipocyte insulin signaling should stimulate the development of novel antidiabetic drugs. Disclosure L. Wang: None. S. Pydi: None. L. Zhu: None. S. Jain: None. Y. Cui: None. O. Gavrilova: None. J. Wess: None. Funding National Institutes of Health

Lrp4 expression by adipocytes and osteoblasts differentially impacts sclerostin’s endocrine effects on body composition and glucose metabolism

Sclerostin exerts profound local control over bone acquisition and also mediates endocrine communication between fat and bone. In bone, sclerostin's anti-osteoanabolic activity is enhanced by low-density lipoprotein receptor-related protein 4 (Lrp4), which facilitates its interaction with the Lrp5 and Lrp6 Wnt co-receptors. To determine whether Lrp4 similarly affects sclerostin's endocrine function, we examined body composition as well as glucose and fatty acid metabolism in mice rendered deficient of Lrp4 in the adipocyte (AdΔLrp4) or the osteoblast (ObΔLrp4). AdΔLrp4 mice exhibit a reduction in adipocyte hypertrophy and improved glucose and lipid homeostasis, marked by increased glucose and insulin tolerance and reduced serum fatty acids, and mirror the effect of sclerostin deficiency on whole-body metabolism. Indeed, epistasis studies place adipocyte-expressed Lrp4 and sclerostin in the same genetic cascade that regulates adipocyte function. Intriguingly, ObΔLrp4 mice, which exhibit dramatic increases in serum sclerostin, accumulate body fat and develop impairments in glucose tolerance and insulin sensitivity despite development of a high bone mass phenotype. These data indicate that expression of Lrp4 by both the adipocyte and osteoblast is required for normal sclerostin endocrine function and that the impact of sclerostin deficiency on adipocyte physiology is distinct from the effect on osteoblast function.