Obese Adipose Tissue Research Articles

Adipose Tissue Vasculature Facilitates Immune Activation of Macrophages Via the CD200-CD200R Axis

The activation of adipose tissue macrophages (ATMs) in obesity is a key mediator of metabolic complications. Therefore, deciphering the cellular and molecular mechanisms initiating ATM activation could reveal regulatory pathways important to prevent obesity‐associated metabolic disorders. While stressed or dying adipocytes have been described as the source of factors activating ATMs, herein we discovered that endothelial cells directly communicate with a distinct population of ATMs to induce their transcriptional reprogramming in response to obesity in mice and humans. CD200R expressed by these perivascular ATMs regulated the acquisition of unique properties shaped by the obese adipose tissue vasculature. Systems biology and ATM-specific silencing in obese mice revealed a unique downstream signaling whereby CD200R act as a rheostat controlling multiple functions of ATMs. Together, these studies provide evidence for a CD200-CD200R axis triggering the reprogramming of ATMs by endothelial cells in obesity.

Механизмы адипотропного действия цинка и их роль в патогенезе ожирения

Клинические и экспериментальные работы демонстрируют взаимосвязь между ожирением и нарушением метаболизма цинка, однако локальные эффекты цинка в жировой ткани изучены недостаточно. В связи с этим, целью обзора явился анализ данных о механизмах адипотропного действия цинка и их роли в патогенезе ожирения. Экспериментальные работы показали, как снижение уровня цинка в жировой ткани при ожирении, так и влияние цинка на дифференцировку и функционирование адипоцитов. Таким образом, цинк может рассматриваться в качестве одного из регуляторов адипогенеза, в связи с чем нарушение обмена цинка может сопровождаться дисфункцией жировой ткани. Модуляция активности транспортеров цинка обусловливает изменение уровня цинка в клетках, в то же время дальнейшие процессы реализации адипотропных функций цинка связаны с функционированием эффекторных молекул. Среди транспортеров цинка, функционирование которых тесно связано с регуляцией адипогенеза, выделяются ZIP14 и ZNT7, активно экспрессирующиеся в процессе созревания адипоцитов. Цинк-α2-гликопротеин является адипокином, действующим как аутокринный и паракринный регулятор метаболизма адипоцитов, а также как фактор активации липолиза. Влияние цинка на дифференцировку адипоцитов также может обусловливаться функционированием ряда белков цинковых пальцев, в первую очередь Zfp423 и Zfp521. Zfp423 является фактором дифференцировки адипоцитов, оказывающим стимулирующее влияние на экспрессию PPARγ, тогда как тормозное влияние Zfp521 достигается, в том числе, за счет подавления активности Zfp423. Несмотря на роль цинксодержащих металлопротеинов, значительная часть эффектов может быть обусловлена действием низкомолекулярных соединений цинка или непосредственно катионом Zn2+. Таким образом, коррекция обмена цинка в организме в целом и в жировой ткани в частности может рассматриваться в качестве одного из способов нормализации метаболизма жировой ткани при ожирении. Experimental and clinical studies have demonstrated an association between obesity and impaired zinc metabolism, although local effects of zinc in adipose tissue are insufficiently studied. The objective of this study was to review current data on adipotropic effects of zinc and its role in the pathogenesis of obesity. Experimental studies have shown both decreased content of zinc in the adipose tissue in obesity and effects of zinc on adipocyte differentiation and functioning. Therefore, zinc can be considered as a regulator of adipogenesis, and, thus, impaired zinc metabolism may be associated with adipose tissue dysfunction. Modulation of zinc transporters underlies alteration of intracellular zinc levels, whereas further effects of zinc are related with functioning of zinc-containing effector molecules. The zinc transporters known to play a significant role in regulation of adipogenesis include ZIP14 and ZNT7 that are actively expressed during adipocyte maturation. Zinc-α2-glycoprotein is an adipokine acting as an autocrine and paracrine regulator of adipocyte metabolism as well as a lipolytic factor. The effect of zinc on adipose tissue differentiation may be also mediated by zinc-finger proteins, primarily Zfp423 and Zfp521. Zfp423 is an adipocyte differentiation factor stimulating the PPARγ expression whereas the antiadipogenic effect of Zfp521 is mediated by Zfp423 down-regulation. Despite a clearly demonstrated effect of Zn-containing metalloproteins a significant part of the effects of zinc may be mediated by biological activity of low-molecular weight zinc compounds or directly by Zn2+ cation. Therefore, improvement of zinc metabolism in the body in general and the adipose tissue in particular can be considered as a strategy for improvement of adipose tissue metabolism in obesity.

1896-P: Adipocyte-Derived Fibroblast Growth Factor 21 (FGF21) Contributes to Circulating FGF21 Pools in Diet-Induced Obese Mice

Fibroblast growth factor 21 (FGF-21) is a hormone derived from hepatocytes and adipocytes that plays a role in energy homeostasis, and promotes weight loss when given pharmacologically. Nutritional stress such as fasting increases liver-derived FGF-21 expression, whereas over-nutrition (i.e., obesity) promotes adipocyte-derived FGF-21 (Ad-FGF-21) production. Despite the current dogma that Ad-FGF-21 acts only in a paracrine manner, obesity is associated with elevated circulating FGF-21 levels which correlate with adiposity and adipose tissue FGF-21 mRNA expression. To date, a concise role for Ad-FGF-21 in obesity-associated energy balance has not been identified. To determine whether Ad-FGF-21 plays a pathological role in obesity, and whether it contributes to circulating FGF-21 levels, we generated mice with Ad-FGF-21 deficiency (Ad-FGF-21 KO) and placed them on an obesogenic high fat high sucrose diet for 18 weeks. Body composition, energy expenditure, glucose and insulin tolerance were assessed. Male and female Ad-FGF-21 KO mice were protected from obesity, gained less body fat, and exhibited improved glucose metabolism over wild type (WT) controls. Circulating FGF-21 levels were higher in obese WT mice than age-matched chow-fed WT or obese Ad-FGF-21 KO mice with no changes in hepatic FGF-21, suggesting that adipocytes are a source of FGF-21 in obesity. Moreover, adipose tissue explants from obese WT mice secreted higher levels of FGF-21 than explants from chow-fed mice, an effect that was blunted in Ad-FGF-21 KO mice. Finally, expression of fibroblast activation protein (FAP), an enzyme that cleaves FGF-21 to blunt its activity, was reduced from obese adipose tissue, providing a potential mechanism by which adipocyte-derived FGF-21 circulates in obesity. In summary, adipocyte-derived FGF-21 may contribute to obesity-associated increases in circulating FGF-21, with the potential to influence the pathology of obesity. Disclosure S. Wang: None. L. Goodspeed: None. K. Ogimoto: None. L. den Hartigh: None. Funding National Institutes of Health (HL092969)

2010-P: Adipocyte Clusterin Is a Link between Cardiometabolism and Alzheimer’s Disease

Background: Obesity heightens the risk for both cardiometabolic complications and Alzheimer’s disease (AD). Although the reasons have yet to be fully elucidated, weight loss improves cognition. Given this link, factors overproduced by obese adipose tissue are ideal candidates for novel investigation into AD pathogenesis. We have recently shown that clusterin from the adipocyte is related to multiple factors of the metabolic syndrome (insulin resistance, dyslipidemia, NAFLD, and atherosclerosis). Clusterin is also involved in CNS lipid trafficking and has been proposed as an AD biomarker. Rationale: Our goals were to characterize the effect of high-fat diet (HFD) feeding on CNS clusterin and neuronal insulin signaling in an AD mouse model (5XFAD); and furthermore, determine the relationship of adipocyte clusterin with neurocognition and insulin sensitivity in obese humans at baseline and following bariatric surgery. Methods: We treated 5XFAD and WT mice with normal chow and HFD for 4 mo., measured CNS clusterin, and determined its effect on neuronal insulin signaling. We next measured subcutaneous adipocyte (SAd) clusterin expression, and performed neuropsychological evaluation and a hyperinsulinemic-euglycemic clamp procedure in obese human subjects (n=11) before and after sleeve gastrectomy. Results: In the 5XFAD mice, HFD increases brain clusterin levels compared to WT, and in primary cortical neurons clusterin attenuates insulin signaling in a dose-dependent manner. In humans, SAd clusterin is related to hepatic insulin resistance, higher fasting glucose, and reduced working memory/attention. After sleeve gastrectomy SAd clusterin expression is reduced, with a significant inverse association between clusterin and working memory/attention and hepatic insulin sensitivity. Conclusions: Our data suggests that SAd clusterin is a potential player in AD pathology, providing a critical link between excess adiposity, T2D, insulin resistance, and heightened AD risk. Disclosure D. Bradley: None. J.Z. Liu: None. A.M. Blaszczak: None. V.P. Wright: None. R. Tarawneh: None. Y. Liu: None. J. Ryu: None. S. Yoon: None. W. Hsueh: None. Funding American Diabetes Association (1-16-ICTS-049 to W.H.); National Institutes of Health (KL2TR001068, R01HL135622)

208-OR: Emc10emc10, a Novel Circulating Factor for Systemic Energy and Metabolic Homeostasis

Proteins secreted from metabolic organs play critical roles in fine-tuning and maintenance of systemic metabolic and energy homeostasis. In this study, we identified a novel obesity-regulated circulating protein, scEmc10 (secreted-isoform of endoplasmic reticulum membrane complex subunit 10), that regulates systemic energy homeostasis. We initially observed that scEmc10 is modulated in adipose tissues in diet-induced obesity (DIO) models and subsequently in obese patients. Using an in-house-developed chemiluminescent immunoassay, we found that circulating Emc10 is significantly upregulated in overweight and obese patients and can be normalized by bariatric surgery-induced weight loss. Subsequently, we observed that ablation of Emc10 protects mice from DIO and associated metabolic dysregulation. Conversely, enhanced circulating Emc10 by expression of secretable AAV-hscEmc10 in the liver promotes obesity and associated metabolic dysfunctions in mice fed with either chow or high fat diet. Subsequent metabolic analysis revealed that Emc10 KO mice on HFD have elevated whole body energy expenditure compared to WT controls via an increase in oxygen consumption in both the brown and inguinal subcutaneous fat. Our mechanistic analysis demonstrated that ablation of scEmc10 promotes adaptive thermogenesis in adipocytes via activation of the PKA signaling pathway and its downstream targets, particularly CREB and p38MAPK. Finally, we also showed that neutralization of scEmc10 in the circulation with a monoclonal antibody reduced body weight and improved diet-induced metabolic dysfunction in obese mice. Our findings implicate Emc10 as a novel therapeutic target for counteracting the development of obesity and obesity-induced metabolic dysfunction. Disclosure Y. Li: None. X. Wang: None. G. Qiang: None. M.A. McCann: None. V. Gil: None. M. Blüher: Advisory Panel; Self; AstraZeneca, Boehringer Ingelheim Pharmaceuticals, Inc., Lilly Diabetes, Novo Nordisk A/S, Sanofi. Speaker’s Bureau; Self; Amgen, Novartis AG. C. Liew: None. Funding National Institutes of Health (R01DK090210, R01DK109015); National Natural Science Foundation of China (81070647, 81370936, 81873645, 81873853)

Impact of osteopontin on the development of non-alcoholic liver disease and related hepatocellular carcinoma.

Background & aimsOsteopontin, a multifunctional protein and inflammatory cytokine, is overexpressed in adipose tissue and liver in obesity and contributes to the induction of adipose tissue inflammation and non‐alcoholic fatty liver (NAFL). Studies performed in both mice and humans also point to a potential role for OPN in malignant transformation and tumour growth. To fully understand the role of OPN on the development of NAFL‐derived hepatocellular carcinoma (HCC), we applied a non‐alcoholic steatohepatitis (NASH)‐HCC mouse model on osteopontin‐deficient (Spp1−/−) mice analysing time points of NASH, fibrosis and HCC compared to wild‐type mice.MethodsTwo‐day‐old wild‐type and Spp1−/− mice received a low‐dose streptozotocin injection in order to induce diabetes, and were fed a high‐fat diet starting from week 4. Different cohorts of mice of both genotypes were sacrificed at 8, 12 and 19 weeks of age to evaluate the NASH, fibrosis and HCC phenotypes respectively.ResultsSpp1−/− animals showed enhanced hepatic lipid accumulation and aggravated NASH, as also increased hepatocellular apoptosis and accelerated fibrosis. The worse steatotic and fibrotic phenotypes observed in Spp1−/− mice might be driven by enhanced hepatic fatty acid influx through CD36 overexpression and by a pathological accumulation of specific diacylglycerol species during NAFL. Lack of osteopontin lowered systemic inflammation, prevented HCC progression to less differentiated tumours and improved overall survival.ConclusionsLack of osteopontin dissociates NASH‐fibrosis severity from overall survival and HCC malignant transformation in NAFLD, and is therefore a putative therapeutic target only for advanced chronic liver disease.

Examination of ex-vivo viability of human adipose tissue slice culture.

Obesity is associated with significantly higher mortality rates, and excess adipose tissue is involved in respective pathologies. Here we established a human adipose tissue slice cultures (HATSC) model ex vivo. HATSC match the in vivo cell composition of human adipose tissue with, among others, mature adipocytes, mesenchymal stem cells as well as stroma tissue and immune cells. This is a new method, optimized for live imaging, to study adipose tissue and cell-based mechanisms of obesity in particular. HATSC survival was tested by means of conventional and immunofluorescence histological techniques, functional analyses and live imaging. Surgery-derived tissue was cut with a tissue chopper in 500 μm sections and transferred onto membranes building an air-liquid interface. HATSC were cultured in six-well plates filled with Dulbecco’s Modified Eagle’s Medium (DMEM), insulin, transferrin, and selenium, both with and without serum. After 0, 1, 7 and 14 days in vitro, slices were fixated and analyzed by morphology and Perilipin A for tissue viability. Immunofluorescent staining against IBA1, CD68 and Ki67 was performed to determine macrophage survival and proliferation. These experiments showed preservation of adipose tissue as well as survival and proliferation of monocytes and stroma tissue for at least 14 days in vitro even in the absence of serum. The physiological capabilities of adipocytes were functionally tested by insulin stimulation and measurement of Phospho-Akt on day 7 and 14 in vitro. Viability was further confirmed by live imaging using Calcein-AM (viable cells) and propidium iodide (apoptosis/necrosis). In conclusion, HATSC have been successfully established by preserving the monovacuolar form of adipocytes and surrounding macrophages and connective tissue. This model allows further analysis of mature human adipose tissue biology ex vivo.

Low-Grade Inflammation Is Not Present in Former Obese Males but Adipose Tissue Macrophage Infiltration Persists.

Macrophage infiltration in two subcutaneous adipose tissue depots and systemic low-grade inflammation were studied in post-obese (PO), obese (O), and control (C) subjects. Young males were recruited into PO: (n = 10, weight-loss avg. 26%, BMI: 26.6 ± 0.7, mean ±SEM kg/m2), O: (n = 10, BMI: 33.8 ± 1.0kg/m2) and C: (n = 10, BMI: 26.6 ± 0.6 kg/m2). PO and C were matched by BMI. Blood and abdominal and gluteal subcutaneous adipose tissue were obtained in the overnight fasted state. Plasma concentrations of IL-6 and CRP were higher (p < 0.05) in O than in PO and C, TNF-α was higher (p < 0.05) only in O compared to PO and IL-18 was similar between groups. The number of CD68+ macrophages was higher (p < 0.05) in the gluteal than the abdominal depot, and higher (p < 0.05) in O and PO compared to C in both depots. The content of CD163+ macrophages was similar between depots but was higher (p < 0.05) in PO compared to C and O in the gluteal depot. In post obese men with a long-term sustained weight loss, systemic low-grade inflammation was similar to non-obese controls despite a higher subcutaneous adipose tissue CD68+ macrophage content. Interestingly, the anti-inflammatory CD163+ macrophage adipose tissue content was consistently higher in post obese than obese and controls.

SUN-544 Immunologic Effects of GLP-1 Activation in Obese Adipose Tissue

Background: Obesity is associated with systemic inflammation which is thought to stem, in part, from adipose tissue (AT). Obese AT is characterized by infiltration of pro-inflammatory T cells that promote macrophage activation and inflammation. GLP-1 has been shown to have anti-inflammatory effects in previous studies. We hypothesized that promotion of GLP-1 signaling with liraglutide or sitagliptin would reduce inflammation in association with an increase in the number of anti-inflammatory invariant natural killer T cells (iNKTs), group 2 innate lymphoid cell (ILC2s) and regulatory T cells (Treg) in blood and AT.Methods: Obese adults with pre-diabetes were randomized to pharmacologic treatment resulting in increased GLP-1 signaling (liraglutide or sitagliptin, N=8), or hypocaloric diet (N=3). This ongoing study is blinded, so the effects of liraglutide and sitagliptin are combined in analyses and referred to as “drug”. Subcutaneous abdominal AT and peripheral blood mononuclear cells (PBMCs) were collected at baseline (“pre”) and after 12 weeks of therapy (“post”). Phenotypic marker expression of blood and AT T cells were characterized by flow cytometry. Whole AT inflammatory gene expression in the pre and post groups was assessed by Nanostring.Results: Using the Nanostring inflammation panel, we found that a number of pro-inflammatory genes were significantly downregulated in whole AT after treatment with drug, including CD163, CD86, CCR1, MCP-2, and MCP-4.Blood ILC2s were significantly decreased with drug treatment (pre 3.95%±3.05, post 1.71% ±1.65, p=0.01), but not diet (pre 2.17% ±1.91, post 1.46% ±1.68, p=0.18). We did not detect a change in Treg numbers after treatment with either diet (pre 5.78% ±1.91, post 6.09% ±1.50, p=0.29) or drug (pre 6.49% ±2.29, post 5.94% ±2.15, p=0.12). Similarly, no difference in blood iNKT numbers was detected after diet (pre 0.063% ±0.044, post 0.082% ±0.049, p=0.11) or drug (pre 0.077% ±0.106, post 0.091% ±0.130, p=0.67).As observed in the PBMCs, adipose ILC2s were decreased after drug (pre 2.04% ±1.67, post 1.32% ±1.58, p=0.07, N=4). We did not detect a change in AT Treg and iNKT numbers (Treg pre 9.24% ±5.36, post 6.23% ±1.55, p=0.14; iNKT pre 0.12% ±0.08, post 0.09% ±0.05, p=0.47, N=4).Conclusions: In a small pilot study of obese pre-diabetic patients treated with drugs that activate GLP-1 signaling (liraglutide or sitagliptin) or hypocaloric diet, global transcriptional analysis of whole AT suggested decreased inflammation with drug therapy. However, we found decreased percentages of ILC2 cells (considered anti-inflammatory in adipose) in both blood and AT after drug treatment. Future experiments will further characterize the function of these cell types, and evaluate other immune subsets in PBMC and AT that may be responsible for decreasing inflammation.

SAT-126 Breast Adipose Tissue Extracellular Vesicles from Obese Women Increase Breast Cancer Aggressiveness - a Novel Mechanism for the Obesity-Breast Cancer Link

Background and Objectives: Breast cancer is among the most common cancer in women with 2.1 million new cases detected each year. Numerous studies have demonstrated a connection between body mass index (BMI) and cancer incidence, with obesity (BMI ≥ 30) being responsible for the development of at least 13 types of cancer, and 15% to 20% of total cancer-related mortality. The effects of extracellular vesicles (EVs) derived from the obese adipose tissue microenvironment on breast cancer have not yet been clearly elucidated. Methods: EVs were obtained from media conditioned with human breast adipose tissue from reduction mammoplasty (n=31). Women were healthy at the time of surgery and had no history of breast cancer. Patient samples were stratified based on their body mass index (BMI), with a BMI < 25 considered healthy and a BMI ≥ 25 considered overweight/obese. Breast adipose tissue-derived EVs (AT-EVs) were characterized (Quantitative Mass Spectrometry) and used to treat human breast cancer cell lines, including the ER+ MCF7 and triple negative breast cancer (TNBC) MDA-MB-231. Effects on cell proliferation and migration in vitro, and on tumor growth in a mouse xenograft model, were examined after long-term education with EVs. RNA sequencing was performed to investigate potential reprogramming induced by AT-EVs. Results: We found a positive correlation between protein amount per AT-EV and BMI. Quantitative proteomics of AT-EVs revealed 46 proteins that were significantly higher and 54 proteins that were significantly lower in specimens from women with a BMI ≥ 25 compared to women with a BMI < 25. AT-EVs from patients with a BMI ≥ 25 induced proliferation of MCF7 cells compared to AT-EVs from patients with a BMI < 25. Obese EVs induced a more aggressive phenotype in MDA-MB-231 cells, increasing their invasiveness in vitro. Obese EVs also increased the growth of MCF7 and MDA-MB-231 cells in vivo. Ingenuity pathway analysis of RNA-Seq data identified significant differences in mTOR signaling and canonical pathways associated with altered mitochondrial function. Conclusion: Our studies identify a novel mechanism to explain the obesity-breast cancer link in older women. Namely, that in obesity, the breast microenvironment produces EVs capable of reprogramming breast cancer cells to grow faster and be more aggressive. Identifying which cargo in breast AT-EV mediates these effects may provide new targets for intervention.

Evaluating the effects of honey bee (Apis mellifera L.) venom on the expression of insulin sensitivity and inflammation‐related genes in co‐culture of adipocytes and macrophages

AbstractObese adipose tissue is characterized by adipocyte hypertrophy and a massive macrophage infiltration. The interaction between macrophages with mature adipocytes releases pro‐inflammatory cytokines. This chronic inflammatory state can contribute to obesity‐related complications, such as insulin resistance, type 2 diabetes and cardiovascular disease. Therefore, we can attempt to prevent and treat obesity‐related diseases by inhibiting macrophage infiltration and blocking their interaction with adipocytes. Honey bee (Apis mellifera) venom (BV) has been reported to have anti‐inflammatory effects. Although BV is used to treat chronic inflammatory diseases, few studies have addressed its use in obesity‐associated inflammation. This study examines the inhibitory effects of BV on lipid accumulation in differentiating preadipocytes, inflammation, and insulin resistance in macrophages and adipocyte‐macrophage co‐culture system. We treated 3 T3‐L1 preadipocytes with BV during differentiation. We later measured lipid accumulation and gene expression of master adipogenic transcription factors. After RAW264.7 and 3 T3‐L1 cells were pretreated with BV, RAW264.7 cells were activated with LPS or co‐cultured with pretreated 3 T3‐L1 cells. Gene expression of pro‐inflammatory cytokines and insulin sensitizing genes was measured in these cells. BV inhibited lipid accumulation and C/EBPα and PPARγ gene expression during intermediate and late 3 T3‐L1 cell differentiation. BV also suppressed gene expression of pro‐inflammatory cytokines (COX‐2, iNOS, MCP‐1, TNF‐α, IL‐1β and IL‐6) in LPS‐stimulated macrophages, and in co‐culture of 3 T3‐L1 adipocytes and RAW264.7 macrophages. However, adiponectin and GLUT‐4 expression were both significantly increased by BV in co‐culture. These findings demonstrate that BV attenuates adipocyte hypertrophy and improves obesity‐related inflammation and insulin resistance in obese adipose tissue.

Identification and characterization of adipose tissue-derived human antibodies with “anti-self” specificity

Abstract We have previously shown that the human obese adipose tissue (AT) contributes to increased secretion of adipocyte-specific IgG antibodies in individuals with obesity. We have also shown that this occurs without any exogenous stimulation, because the ongoing process of cell death in the obese AT leads to the release of “self” antigens able to induce chronic stimulation of B cells. Here, we confirm and extend our initial observation on a different cohort of individuals, and we show that also the plasma of obese individuals is enriched in IgG antibodies with specificities for adipocyte-derived antigens. Adipocyte-specific IgG secreted in the obese AT are significantly correlated with those present in plasma. Using immunoprecipitation and mass spectrometry, we have identified these antigenic specificities. The antigens are almost exclusively intracellular or cell-associated, usually not “self” antigens, but they are released by cells dying in the AT. These antigens will be used in protein arrays to screen plasma from individuals with obesity and also with autoimmune diseases. We have indeed preliminary evidence that the plasma of Rheumatoid Arthritis patients is enriched in adipocyte-specific IgG antibodies, suggesting that obesity may contribute to pathogenicity in autoimmune patients. Finally, we show for the first time that not only macrophages but also adipocytes in the obese AT are efficient antigen-presenting cells and stimulate the secretion of IgG autoimmune antibodies. They do so because they express the antigen-presenting molecules CD1d and, to a lesser extent, MHC class II, as well as the co-stimulatory molecule CD86. These results may lead to the development of novel therapeutic strategies to controlautoimmunity.

Blimp-1 in adipose resident Tregs controls adipocyte beiging and obesity

Abstract Visceral adipose tissue regulatory T cells (VAT Tregs) protect against systemic inflammation and metabolic disease by limiting expansion of pro-inflammatory Th1 cells and M1 macrophages, and by preserving insulin sensitivity and glucose tolerance. Although their basic markers and roles have been studied, less is known about the transcriptional machinery regulating their differentiation and function. B lymphocyte-induced maturation protein-1 (Blimp-1) is a transcriptional regulator known to be involved in the development, polarization, and maintenance of various immune cells including CD4+ T cells. Using Blimp-1 reporter mice, we discovered that Blimp-1 is constitutively expressed in a subset of VAT Tregs compared to lymphoid Tregs, and that Blimp-1+ VAT Tregs are phenotypically distinct from their Blimp-1− counterparts. Blimp-1 is not required for VAT Treg development, however Treg-specific deletion of Blimp-1 led to unique changes in VAT Treg markers in lean versus obese adipose tissue. In addition, Blimp-1 knockout mice fed high fat diet had fewer adipose-resident NK cells and increased CD8 T cells. Surprisingly, loss of Blimp-1+ Tregs led to less adipose tissue IL-10, increased expression of thermogenic genes, reduced body fat, decreased weight, and improved insulin sensitivity. Based on these data, we hypothesize that Blimp-1+ Treg dependent IL-10 production suppresses adipocyte beiging, and that loss of these cells results in increased thermogenesis, greater weight loss and improved insulin sensitivity in obese mice.

Macrophage Subsets in Obesity, Aligning the Liver and Adipose Tissue.

The increasing prevalence of obesity is accompanied by a rising incidence in metabolic syndrome and related pathologies such as non-alcoholic fatty liver disease. Macrophages are hypothesized to play central roles in these diseases, through their role as inflammatory mediators and as such are thought to be potential targets for future therapies. Recently, single cell technologies have revealed significant heterogeneity within the macrophage pool in both liver and adipose tissue in obesity. Thus current efforts are focused on dissecting this heterogeneity and understanding the distinct functions of the individual subsets. In this review, we discuss the current knowledge regarding macrophage heterogeneity, ontogeny and functions in the context of obese adipose tissue and fatty liver disease and attempt to align the distinct populations described to date.

Fat Quality Matters: Distinct Proteomic Signatures Between Lean and Obese Cardiac Visceral Adipose Tissue Underlie its Differential Myocardial Impact.

Heart failure with preserved ejection fraction (HFpEF) is recognised as an important cause of cardiovascular mortality and morbidity, accounting for approximately 50% of heart failure cases. Metabolic-related complications, such as obesity, have been associated with the pathophysiology of this complex syndrome. The anatomic proximity between cardiac visceral adipose tissue (CVAT) and the myocardium has been drawing attention due to its potential pathogenic role in cardiac diseases. Thus, we aimed to characterise the phenotypic and proteomic differences between CVAT from ZSF1 lean (control) and ZSF1 obese (HFpEF) rats as well as to evaluate the myocardial impact of conditioned media derived from CVAT of these 2 groups. CVAT of 20-weeks-old lean and obese ZSF1 rats was collected for: 1) 24h DMEM incubation to obtain conditioned media, 2) separation of proteins to mass spectrometry identification, 3) adipokines' expression, 4) adipocytes cross-sectional area assessment. Organotypic cultures were prepared from 7 days-old Wistar Han cardiac explants and incubated for 24h with the conditioned media. After incubation, cross-section area of cardiomyocytes and fibrosis were evaluated. Cardiomyocytes were isolated from Wistar Han and incubated with conditioned media for viability studies. CVAT from lean rats presented a higher expression of uncoupling protein-1 (UCP-1) protein, associated with a multilocular appearance and an increased expression of brown adipose tissue markers. Contrarily, CVAT from obese rats revealed a white adipose tissue-like phenotype accompanied by hypertrophy of adipocytes. The analysis of the CVAT proteome reinforced the phenotypic differences between lean and obese CVAT, showing enrichment of proteins involved in triglyceride metabolic processes in obese CVAT. In contrast, mitochondrial proteins were prominent in lean CVAT, further suggesting a brown adipose tissue-like phenotype. The twenty-four hours-long incubation of myocardial organo-cultures with conditioned media obtained from CVAT obese (CM-obese) rats significantly reduced cell viability, induced cardiomyocytes hypertrophy and fibrosis, in stark contrast with the incubation with the conditioned media from lean rats CVAT (CM-lean). Furthermore, the deleterious effect imposed by CM-obese was associated with a pro-inflammatory profile, characterised by an increased expression of several pro-inflammatory adipokines. Obesity promotes alterations in CVAT proteome signature, structure, composition and secretome, translating into dramatic myocardial consequences.

Immunometabolism: From basic mechanisms to translation.

Immunometabolism has emerged as a major mechanism central to adaptive and innate immune regulation. From early observations that inflammatory cytokines were induced in obese adipose tissue and that these cytokines contributed to metabolic disease, it was clear that metabolism and the immunological state are inextricably linked. With a second research wave arising from studies in cancer metabolism to also study the intrinsic metabolic pathways of immune cells themselves and how those pathways influence cell fate and function, immunometabolism is a rapidly maturing area of research. Several key themes and goals drive the field. There is abundant evidence that metabolic pathways are closely tied to cell signaling and differentiation which leads different subsets of immune cells to adopt unique metabolic programs specific to their state and environment. In this way, metabolic signaling drives cell fate. It is also apparent that microenvironment greatly influences cell metabolism. Immune cells adopt programs specific for the tissues where they infiltrate and reside. Ultimately, a central goal of the field is to apply immunometabolism findings to the discovery of novel therapeutic strategies in a wide range of diseases, including cancer, autoimmunity, and metabolic syndrome. This review summarizes these facets of immunometabolism and highlights opportunities for clinical translation.

The influence of equine body weight gain on inflammatory cytokine expressions of adipose tissue in response to endotoxin challenge

BackgroundHuman obesity is linked with systemic inflammation. However, it is still controversial if equines produce more inflammatory cytokines with increasing body weight and if the production of those show breed type specific patterns. The main objective of this study was to determine if diet induced obesity is associated with increased inflammatory signatures in adipose tissue of equines and if a breed predisposition exists between ponies and horses. Additionally, we aimed to identify adipose tissue depot differences in inflammatory cytokine expression. Nineteen healthy, non-overweight and metabolically healthy equines received a hypercaloric diet for 2 years. Body weight, body condition score and cresty neck score were assessed weekly throughout the study. At three time points, insulin sensitivity was determined by a combined glucose-insulin test. Adipose tissue samples were collected from two intra-abdominal and two subcutaneous depots under general anesthesia at each time point after an endotoxin trigger. In the adipose tissue samples levels of CD68 mRNA (a marker of macrophage infiltration) and pro-inflammatory cytokine mRNA (IL-1β, IL-6 and TNFα) were analyzed with RT-qPCR. As markers of lipid metabolism mRNA levels of lipoprotein lipase (LPL) and fatty acid binding protein 4 (FABP4) were determined with RT-qPCR.ResultsCD68 mRNA levels increased with body weight gain in several adipose tissue (AT) depots (Wilcoxon signed rank test with Bonferroni correction; retroperitoneal AT horses: P = 0.023, mesocolonial AT horses: P = 0.023, subcutaneous tail head AT ponies: P = 0.015). In both abdominal depots CD68 mRNA levels were higher than in subcutaneous adipose tissue depots (Kruskal–Wallis-ANOVA with Bonferroni correction: P < 0.05). No breed related differences were found. Pro-inflammatory cytokine mRNA IL-1β, IL-6 and TNFα levels were higher in subcutaneous depots compared to abdominal depots after body weight gain. IL-1β, IL-6 and TNFα mRNA levels of mesocolon adipose tissue were higher in obese horses compared to obese ponies (Mann–Whitney-U test; IL-1β: P = 0.006; IL-6: P = 0.003; TNFα: P = 0.049). In general, horses had higher FABP4 and LPL mRNA levels compared to ponies in neck AT and tail AT at all time points.ConclusionOur findings suggest an increased invasion of macrophages in intra-abdominal adipose tissue with increasing body weight gain in equines in combination with a low dose endotoxin stimulus. This might predispose equines to obesity related comorbidities. In obese horses mesocolon adipose tissue showed higher inflammatory cytokine expression compared to obese ponies. Additionally, subcutaneous adipose tissue expressed more pro-inflammatory cytokines compared to intra-abdominal adipose tissue. Horses had higher FABP4 and LPL mRNA levels in selected AT depots which may indicate a higher fat storage capacity than in ponies. The differences in lipid storage might be associated with a higher susceptibility to obesity-related comorbidities in ponies in comparison to horses.

Hypoxia‑inducible factor‑1α regulates Lipin1 differently in pre‑adipocytes and mature adipocytes.

Hypoxia-inducible factor (HIF)-1α is a transcription factor that is activated in low oxygen conditions. Adipose tissues are poorly oxygenated in patients with obesity. The low oxygen conditions in obese adipose tissues induce HIF‑1α in adipocytes. Previous studies using genetically modified mice suggest that HIF‑1α contributes to dysfunction in adipocytes. Lipin1 is a bifunctional protein that works as a phosphatidate phosphatase and transcriptional coactivator, which regulates lipid metabolism and adipogenesis, respectively. HIF‑1α directly regulates Lipin1 in hepatocytes. However, the regulation of Lipin1 by HIF‑1α in adipocytes is not well determined. Therefore, the present study investigated the regulation of Lipin1 by HIF‑1α in adipocytes. Expression levels of Lipin1 were reduced in epididymal adipose tissues of adipocyte‑specific HIF‑1α knockout mice, indicating that HIF‑1α regulates Lipin1 in adipocytes. In differentiated mature adipocytes, a HIF‑1α activator, dimethyloxallyl glycine(DMOG), was demonstrated to increase Lipin1, and a HIF‑1α inhibitor, 3‑(5'‑hydroxymethyl‑2'‑furyl)-1‑benzylindazole(YC‑1), reversed this increase, indicating that HIF‑1α regulates Lipin1 in differentiated adipocytes. However, during differentiation of pre‑adipocytes into adipocytes, YC‑1 increased Lipin1 even though HIF‑1α was decreased. The differentiation efficiency increased with YC‑1 treatment. In addition, DMOG reduced Lipin1 expression levels during differentiation despite increased HIF‑1α. Under these conditions, differentiation efficiency was reduced. These results suggest that Lipin1 is negatively regulated by HIF‑1α in pre‑adipocytes. Our results show that regulation of Lipin1 by HIF‑1α is different in adipocytes and pre‑adipocytes.

ASK1 inhibits browning of white adipose tissue in obesity

Increasing energy expenditure via induction of adipose tissue browning has become an appealing strategy to treat obesity and associated metabolic complications. Herein, we identify adipocyte-expressed apoptosis signal-regulating kinase 1 (ASK1) as regulator of adipose tissue browning. High fat diet-fed adipocyte-specific ASK1 knockout mice reveal increased UCP1 protein levels in inguinal adipose tissue concomitant with elevated energy expenditure, reduced obesity and ameliorated glucose tolerance compared to control littermates. In addition, ASK1-depletion blunts LPS-mediated downregulation of isoproterenol-induced UCP1 in subcutaneous fat both in vitro and in vivo. Conversely, adipocyte-specific ASK1 overexpression in chow-fed mice attenuates cold-induced UCP1 protein levels in inguinal fat. Mechanistically, ASK1 phosphorylates interferon regulatory factor 3 (IRF3) resulting in reduced Ucp1 expression. Taken together, our studies unravel a role of ASK1 in mediating the inhibitory effect of caloric surplus or LPS-treatment on adipose tissue browning. Adipocyte ASK1 might be a pharmacological target to combat obesity and associated morbidities.

Myeloid mineralocorticoid receptor activation alters metabolism in HFD‐salt fed mice

Context and ObjectivesIn obese mice and patients, mineralocorticoid receptor (MR) activation has deleterious consequences on metabolic and cardiovascular parameters. In mice, adipose tissue MR triggers mitochondrial and vascular dysfunction. Adipocyte MR activation induces metabolic and vascular dysfunction in a context of obesity, however, obese adipose tissue is also infiltrated with macrophages, which are capable of expressing MR. Thus, we hypothesised that macrophage MR could also be responsible for metabolic and cardiovascular complications in high‐fat high‐salt diet‐fed mice.MethodsWe fed myeloid cell‐specific MR knock‐out male and female mice (My‐MRKO) with a 6‐week high fat – high salt diet (60% kcal from fat, 1.2% salt in drinking water) and studied weight gain, whole body and specific organs composition by nuclear magnetic resonance and insulin sensitivity to explore the metabolic phenotype. Blood pressure measurements by plethysmography and vascular reactivity studies by wire myography were also performed to characterise the cardiovascular phenotype.ResultsMy‐MRKO male mice under high fat – high salt diet displayed reduced weight (37.8 g [interquartile range 34.1–40.8] vs 42.5 g [39.5–45.2]; P = 0.0128), preserved muscle mass (61.5 % [55.8–63.3] vs 56.8 % [54.1–58.5]; P = 0.0337) and a strong tendency to decreased fat mass (26.1 % [23.7–33.9] vs 31.7 % [27.1–35.4]; P = 0.0728). Moreover, liver fat content at sacrifice was decreased (5.8 % [2.1–9.1] vs 9.3 % [5.9–13.4]; P = 0.0491). Those effects were not observed in female mice. Fasting glycaemia was decreased in male My‐MRKO mice (9.2 mmol/L [6.4–12.8] vs 13.2 mmol/L [10.4–16.1]; P = 0.0005) and insulin resistance was decreased in both gender (area under curve 9915 [9398–10043] vs 11550 [9795–12848]; P = 0.0458 in males, 8291 [7812–8884] vs 9544 [8918–9735]; P = 0.0177 in females), as assessed by an insulin tolerance test. A clear effect was observed on blood pressure only in female mice (blood pressure 96 mmHg [94–97] vs 106 mmHg [104–109]; P = 0.0004), whereas there was no modification in mesenteric arteries vascular reactivity in both gender.ConclusionsThus, our findings show that myeloid cell‐MR regulates systemic metabolic parameters, highlighting a new role in obesity and its associated complications for MR expressed in myeloid cells.Support or Funding InformationSorbonne Université, Inserm, CARMMA‐RHU