Obese Visceral Adipose Tissue Research Articles

Obese visceral adipose dendritic cells downregulate regulatory T cell development through IL-33.

Regulatory T cells (Tregs) residing in visceral adipose tissue (VAT) play a pivotal role in regulating tissue inflammation and metabolic dysfunction associated with obesity. However, the specific phenotypic and functional characteristics of Tregs in obese VAT, as well as the regulatory mechanisms shaping them, remain elusive. This study demonstrates that obesity selectively reduces Tregs in VAT, characterized by restrained proliferation, heightened PD-1 expression, and diminished ST2 expression. Additionally, obese VAT displays distinctive maturation of dendritic cells (DCs), marked by elevated expressions of MHC-II, CD86, and PD-L1, which are inversely correlated with VAT Tregs. In an in vitro co-culture experiment, only obese VAT DCs, not macrophages or DCs from subcutaneous adipose tissue (SAT) and spleen, result in decreased Treg differentiation and proliferation. Furthermore, Tregs differentiated by obese VAT DCs exhibit distinct characteristics resembling those of Tregs in obese VAT, such as reduced ST2 and IL-10 expression. Mechanistically, obesity lowers IL-33 production in VAT DCs, contributing to the diminished Treg differentiation. These findings collectively underscore the critical role of VAT DCs in modulating Treg generation and shaping Treg phenotype and function during obesity, potentially contributing to the regulation of VAT Treg populations.

Unraveling adipose tissue proteomic landscapes in severe obesity: insights into metabolic complications and potential biomarkers.

In this study, we aimed to comprehensively characterize the proteomic landscapes of subcutaneous adipose tissue (SAT) and visceral adipose tissue (VAT) in patients with severe obesity, to establish their associations with clinical characteristics, and to identify potential serum protein biomarkers indicative of tissue-specific alterations or metabolic states. We conducted a cross-sectional analysis of 32 patients with severe obesity (16 males and 16 females) of Central European descent who underwent bariatric surgery. Clinical parameters and body composition were assessed using dual-energy X-ray absorptiometry (DXA) and bioelectrical impedance, with 15 patients diagnosed with type 2 diabetes (T2D) and 17 with hypertension. Paired SAT and VAT samples, along with serum samples, were subjected to state-of-the-art proteomics liquid chromatography-mass spectrometry (LC-MS). Our analysis identified 7,284 proteins across SAT and VAT, with 1,249 differentially expressed proteins between the tissues and 1,206 proteins identified in serum. Correlation analyses between differential protein expression and clinical traits suggest a significant role of SAT in the pathogenesis of obesity and related metabolic complications. Specifically, the SAT proteomic profile revealed marked alterations in metabolic pathways and processes contributing to tissue fibrosis and inflammation. Although we do not establish a definitive causal relationship, it appears that VAT might respond to SAT metabolic dysfunction by potentially enhancing mitochondrial activity and expanding its capacity. However, when this adaptive response is exceeded, it could possibly contribute to insulin resistance (IR) and in some cases, it may be associated with the progression to T2D. Our findings provide critical insights into the molecular foundations of SAT and VAT in obesity and may inform the development of targeted therapeutic strategies.NEW & NOTEWORTHY This study provides insights into distinct proteomic profiles of subcutaneous adipose tissue (SAT), visceral adipose tissue (VAT), and serum in patients with severe obesity and their associations with clinical traits and body composition. It underscores SAT's crucial role in obesity development and related complications, such as insulin resistance (IR) and type 2 diabetes (T2D). Our findings emphasize the importance of understanding the SAT and VAT balance in energy homeostasis, proteostasis, and the potential role of SAT capacity in the development of metabolic disorders.

Identification of major hub genes involved in high-fat diet-induced obese visceral adipose tissue based on bioinformatics approach

Identification of major hub genes involved in high-fat diet-induced obese visceral adipose tissue based on bioinformatics approach

Mitochondrial sourcing of interferogenic ligands and an autoantigen in human obesity-associated metaflammation.

Visceral adipose tissue (VAT) inflammation contributes to metabolic dysregulation in obesity. VAT recruitment and activation of plasmacytoid dendritic cells (pDCs) through toll-like receptor 9 (TLR9) recognition of self-DNA, leading to induction of type I interferons, arecrucial innate triggers for this VAT inflammation. It was hypothesized that mitochondrial DNA (mtDNA) can contribute to TLR9 activation in VAT-recruited pDCs in obesity, and this study aimed to identify the carrier protein for ligand access to TLR9 and to explorewhether this also provides for a source of autoantigensin this context. VAT samples, used for gene expression studies as well as adipose explant cultures, were collected from patients with obesity (n = 54) and lean patients (n = 10). Supernatants from human pDC cultures, treated with adipose explant culture supernatants, were used for interferon α ELISA. Venous plasma, from patients with (n = 114) and without (n = 45) obesity, was used for an ELISA for autoantibodies. MtDNA from VAT in obesity, in complex with mitochondrial transcription factor A protein (TFAM), acts as interferogenic ligands for pDCs. Humoral autoreactivity against TFAM is also induced in obesity. Interferogenic ligands and an autoantigen can be sourced from dysfunctional mitochondria in VAT of humans with obesity. Further therapeutic and prognostic potential for this immune mechanism in obesity warrants exploration.

Expression analysis of MMP14: Key enzyme action in modulating visceral adipose tissue plasticity in patients with obesity.

Compromised adipose tissue plasticity is a hallmark finding of obesity orchestrated by the intricate interplay between various extracellular matrix components. Collagen6 (COL6) is well characterized in obese visceral adipose tissue (VAT), not much is known about MMP14 which is hypothesized to be the key player in matrix reorganization. Subjects with obesity (BMI ≥40; n = 50) aged 18-60 years undergoing bariatric surgery and their age-matched controls (BMI < 25; n = 30) were included. MMP14, Col6A3 and Tissue inhibitor of metalloproteinase 2 (TIMP2) mRNA expression was assessed in VAT and their serum levels along with endotrophin were estimated in both groups preoperatively and post-operatively in the obese group. The results were analysed statistically and correlated with anthropometric and glycaemic parameters, namely fasting glucose and insulin, HbA1c, HOMA-IR, HOMA-β and QUICKI. Circulating levels as well as mRNA expression profiling revealed significant differences between the individuals with and without obesity (p < .05), more so in individuals with diabetes and obesity (p < .05). Follow-up serum analysis revealed significantly raised MMP14 (p < .001), with decreased Col6A3, endotrophin and TIMP2 levels (p < .01, p < .001 and p < .01, respectively). A rise in serum MMP14 protein, simultaneous with post-surgical weight loss and decreased serum levels of associated extracellular matrix (ECM) remodellers, suggests its crucial role in modulating obesity-associated ECM fibrosis and pliability of VAT.

Molecular Mechanisms for the Vicious Cycle between Insulin Resistance and the Inflammatory Response in Obesity.

The comprehensive anabolic effects of insulin throughout the body, in addition to the control of glycemia, include ensuring lipid homeostasis and anti-inflammatory modulation, especially in adipose tissue (AT). The prevalence of obesity, defined as a body mass index (BMI) ≥ 30 kg/m2, has been increasing worldwide on a pandemic scale with accompanying syndemic health problems, including glucose intolerance, insulin resistance (IR), and diabetes. Impaired tissue sensitivity to insulin or IR paradoxically leads to diseases with an inflammatory component despite hyperinsulinemia. Therefore, an excess of visceral AT in obesity initiates chronic low-grade inflammatory conditions that interfere with insulin signaling via insulin receptors (INSRs). Moreover, in response to IR, hyperglycemia itself stimulates a primarily defensive inflammatory response associated with the subsequent release of numerous inflammatory cytokines and a real threat of organ function deterioration. In this review, all components of this vicious cycle are characterized with particular emphasis on the interplay between insulin signaling and both the innate and adaptive immune responses related to obesity. Increased visceral AT accumulation in obesity should be considered the main environmental factor responsible for the disruption in the epigenetic regulatory mechanisms in the immune system, resulting in autoimmunity and inflammation.

Identification of major hub genes involved in high-fat diet-induced obese visceral adipose tissue based on bioinformatics approach

ABSTRACT High-fat diet (HFD) can cause obesity, inducing dysregulation of the visceral adipose tissue (VAT). This study aimed to explore potential biological pathways and hub genes involved in obese VAT, and for that, bioinformatic analysis of multiple datasets was performed. The expression profiles (GSE30247, GSE167311 and GSE79434) were downloaded from Gene Expression Omnibus. Overlapping differentially expressed genes (ODEGs) between normal diet and HFD groups in GSE30247 and GSE167311 were selected to run protein–protein interaction network, GO and KEGG analysis. The hub genes in ODEGs were screened by Cytoscape software and further verified in GSE79434 and obese mouse model. A total of 747 ODEGs (599 up-regulated and 148 down-regulated) were screened, and the GO and KEGG analysis showed that the up-regulated ODEGs were significantly enriched in inflammatory response and extracellular matrix receptor interaction pathways. On the other hand, the down-regulated ODEGs were involved in metabolic pathways; however, there were no significant KEGG pathways. Furthermore, six hub genes, Mki67, Rac2, Itgb2, Emr1, Tyrobp and Csf1r were acquired. These pathways and genes were verified in GSE79434 and VAT of obese mice. This study revealed that HFD induced VAT expansion, inflammation and fibrosis, and the hub genes could be used as therapeutic biomarkers in obesity.

Drastic transformation of visceral adipose tissue and peripheral CD4 T cells in obesity.

Obesity has a pronounced effect on the immune response in systemic organs that results in not only insulin resistance but also altered immune responses to infectious diseases and malignant tumors. Obesity-associated microenvironmental changes alter transcriptional expression and metabolism in T cells, leading to alterations in T-cell differentiation, proliferation, function, and survival. Adipokines, cytokines, and lipids derived from obese visceral adipose tissue (VAT) may also contribute to the systemic T-cell phenotype, resulting in obesity-specific pathogenesis. VAT T cells, which have multiple roles in regulating homeostasis and energy utilization and defending against pathogens, are most susceptible to obesity. In particular, many studies have shown that CD4 T cells are deeply involved in the homeostasis of VAT endocrine and metabolic functions and in obesity-related chronic inflammation. In obesity, macrophages and adipocytes in VAT function as antigen-presenting cells and contribute to the obesity-specific CD4 T-cell response by inducing CD4 T-cell proliferation and differentiation into inflammatory effectors via interactions between major histocompatibility complex class II and T-cell receptors. When obesity persists, prolonged stimulation by leptin and circulating free fatty acids, repetitive antigen stimulation, activating stress responses, and hypoxia induce exhaustion of CD4 T cells in VAT. T-cell exhaustion is characterized by restricted effector function, persistent expression of inhibitory receptors, and a transcriptional state distinct from functional effector and memory T cells. Moreover, obesity causes thymic regression, which may result in homeostatic proliferation of obesity-specific T-cell subsets due to changes in T-cell metabolism and gene expression in VAT. In addition to causing T-cell exhaustion, obesity also accelerates cellular senescence of CD4 T cells. Senescent CD4 T cells secrete osteopontin, which causes further VAT inflammation. The obesity-associated transformation of CD4 T cells remains a negative legacy even after weight loss, causing treatment resistance of obesity-related conditions. This review discusses the marked transformation of CD4 T cells in VAT and systemic organs as a consequence of obesity-related microenvironmental changes.

Visceral Adipose Tissue E2F1-miRNA206/210 Pathway Associates with Type 2 Diabetes in Humans with Extreme Obesity.

Objective: Up-regulated expression of transcription-factor E2F1 in human visceral adipose tissue (VAT) characterizes a dysmetabolic obesity sub-phenotype. An E2F1-miRNA network has been described in multiple cancers. Here we investigated whether elevated VAT-E2F1 in obesity is associated with VAT-miRNA alterations similar to, or distinct from, those described in cancer. Furthermore, we assessed if E2F1-associated miRNA changes may contribute to the link between high- VAT-E2F1 and a dysmetabolic obesity phenotype. Methods: We assembled a cohort of patients with obesity and high-VAT-E2F1, matched by age, sex, ±BMI to patients with low-VAT-E2F1, with and without obesity (8 patients/groupX3 groups). We performed Nanostring©-based miRNA profiling of VAT samples from all 24 patients. Candidate E2F1-related miRNAs were validated by qPCR in an independent cohort of patients with extreme obesity, with or without type-2-diabetes (T2DM) (n = 20). Bioinformatic tools and manipulation of E2F1 expression in cells were used to establish the plausibility of the functional VAT-E2F1-miRNA network in obesity. Results: Among n = 798 identified miRNAs, 17 were differentially expressed in relation to E2F1 and not to obesity itself. No evidence for the cancer-related E2F1-miRNA network was identified in human VAT in obesity. In HEK293-cells, overexpression/downregulation of E2F1 correspondingly altered the expression of miRNA-206 and miRNA-210-5p, two miRNAs with reported metabolic functions consistent with those of E2F1. In VAT from both cohorts, the expression of both miRNA-206 and 210-5p intercorrelated, and correlated with the expression of E2F1. In cohort 1 we did not detect significant associations with biochemical parameters. In cohort 2 of patients with extreme obesity, all those with high VAT-E2F1 showed a diabetes-complicated obesity phenotype and higher expression of miRNA-206 and miRNA-210-5p, which also correlated with fasting glucose levels (both miRNAs) and fasting insulin (miRNA-210-5p). Conclusions: Whilst the previously described cancer-related E2F1-miRNA network does not appear to operate in VAT in obesity, miRNAs-206 and 210-5p may link high-E2F1 expression in VAT with diabetes-complicated extreme obesity phenotype.

1417-P: Gut Microbiome Translocation to Visceral Adipose Tissue Promotes Recruitment of Adipose-Specific Neutrophils in Human Obesity

Neutrophils protect against invading bacteria, and their unique role in adipose tissue inflammation and insulin resistance (IR) in obesity is increasingly recognized. We found that visceral adipose tissue (VAT) of obese (n=83) compared to lean (n=13) individuals had higher neutrophil abundance, correlating with IR. The obese subjects had elevated circulating plasma zonulin, a marker of intestinal permeability, and lipopolysaccharide-binding protein (LPSBP) , a marker of bacterial presence, consistent with evidence suggesting that changes in the microbiome and gut permeability in obesity permits the entry of gut bacteria and bacterial products into the circulation. Therefore, we tested for bacterial presence in VAT under sterile conditions. 16s rRNA analyses revealed that the family Streptococcaceae and an unnamed genus in the family Ruminococaceae were enriched in obese VAT contrary to order Bacilliales and genus Marvinobryantia in lean VAT, majorly originating from the GI tract. To confirm whether higher bacterial load is causal for VAT neutrophil abundance in obesity, we gavaged high-fat diet fed mice with fecal material from an obese human, which increased neutrophil, but not macrophage recruitment to VAT. As VAT is highly vascularized, we used RNA-seq to compare the gene expression signature of VAT neutrophils to peripheral blood neutrophils to eliminate the possibility that VAT neutrophils were circulating cells. Evidently, VAT neutrophils had a differential activation state compared to the circulating neutrophils, with a distinct gene expression pattern representing upregulated inflammatory, chemotactic, extracellular matrix, and ROS generation pathways. Overall, we find that that increased gut bacterial translocation enhances the recruitment of VAT specific neutrophils in obesity offering novel targets for managing VAT inflammation and comorbidities of obesity. Disclosure D.Shantaram: None. N.F.Williams: None. D.Spakowicz: None. W.Hsueh: None. A.M.Blaszczak: None. R.Hoyd: None. A.Smith: None. L.Antwi: None. J.Z.Liu: None. V.P.Wright: None. D.Bradley: None. Y.Liu: None.

Effects of systemic inflammation due to hepatic ischemia-reperfusion injury upon lean or obese visceral adipose tissue.

ABSTRACTPurpose:To evaluate how the induction of liver damage by ischemia and reperfusion affects the adipose tissue of lean and obese mice.Methods:Lean and diet-induced obese mice were subjected to liver ischemia (30 min) followed by 6 h of reperfusion. The vascular stromal fraction of visceral adipose tissue was analyzed by cytometry, and gene expression was evaluated by an Array assay and by RT-qPCR. Intestinal permeability was assessed by oral administration of fluorescein isothiocyanate (FITC)-dextran and endotoxemia by serum endotoxin measurements using a limulus amebocyte lysate assay.Results:It was found that, after liver ischemia and reperfusion, there is an infiltration of neutrophils, monocytes, and lymphocytes, as well as an increase in the gene expression that encode cytokines, chemokines and their receptors in the visceral adipose tissue of lean mice. This inflammatory response was associated with the presence of endotoxemia in lean mice. However, these changes were not observed in the visceral adipose tissue of obese mice.Conclusions:Liver ischemia and reperfusion induce an acute inflammatory response in adipose tissue of lean mice characterized by an intense chemokine induction and leukocyte infiltration; however, inflammatory alterations are already present at baseline in the obese adipose tissue and liver ischemia and reperfusion do not injure further.

Metabolic link between obesity and autoimmune diseases.

The abnormal accumulation of visceral adipose tissue in obesity is associated with metabolic changes that include altered glucose tolerance, insulin resistance, hyperlipidemia, and metabolic syndrome. Obesity also coincides with increased incidence of autoimmune diseases. Accumulating evidence suggest that prolonged metabolic overload related to overnutrition, influenced by genetic and epigenetic factors, might affect immunologic self-tolerance through changes in the energy metabolism of immune cells, particularly regulatory T (Treg) cells. A strong activation of nutrient-energy signaling pathways blocks the induction of the transcription factor forkhead P3 (FOXP3), a master regulator of Treg cells, consequently inhibiting their generation and proliferation, thereby promoting proinflammatory response. Expanding our knowledge on the topic, particularly on metabolic T cell flexibility in vivo will provide new insights that can be used to develop therapeutic strategies for various inflammatory diseases, including obesity and autoimmune diseases. Targeting specific metabolic pathways is emerging as an important approach to control immune response and maintain immunological homeostasis.

Expression of LONP1 Is High in Visceral Adipose Tissue in Obesity, and Is Associated with Glucose and Lipid Metabolism.

BackgroundThe nature and role of the mitochondrial stress response in adipose tissue in relation to obesity are not yet known. To determine whether the mitochondrial unfolded protein response (UPRmt) in adipose tissue is associated with obesity in humans and rodents.MethodsVisceral adipose tissue (VAT) was obtained from 48 normoglycemic women who underwent surgery. Expression levels of mRNA and proteins were measured for mitochondrial chaperones, intrinsic proteases, and components of electron-transport chains. Furthermore, we systematically analyzed metabolic phenotypes with a large panel of isogenic BXD inbred mouse strains and Genotype-Tissue Expression (GTEx) data.ResultsIn VAT, expression of mitochondrial chaperones and intrinsic proteases localized in inner and outer mitochondrial membranes was not associated with body mass index (BMI), except for the Lon protease homolog, mitochondrial, and the corresponding gene LONP1, which showed high-level expression in the VAT of overweight or obese individuals. Expression of LONP1 in VAT positively correlated with BMI. Analysis of the GTEx database revealed that elevation of LONP1 expression is associated with enhancement of genes involved in glucose and lipid metabolism in VAT. Mice with higher Lonp1 expression in adipose tissue had better systemic glucose metabolism than mice with lower Lonp1 expression.ConclusionExpression of mitochondrial LONP1, which is involved in the mitochondrial quality control stress response, was elevated in the VAT of obese individuals. In a bioinformatics analysis, high LONP1 expression in VAT was associated with enhanced glucose and lipid metabolism.

1973-P: Single-Nuclei Transcriptomics of Human Adipose Tissue Identify Distinct Adipocyte Progenitor Subpopulations in Type 2 Diabetes

Altered adipocyte function underlies type 2 diabetes (DM) pathogenesis. We hypothesized that adipocyte dysfunction in visceral adipose tissue (VAT) in DM originates from distinct adipocyte progenitor cell (APC) subpopulations. VAT was collected from bariatric surgery patients with (HbA1c&amp;gt;6.5) and without DM (NDM, HbA1c&amp;lt;5.7). Single-nuclei RNA sequencing was performed in VAT nuclei (n= 1 NDM, 2 DM) using 10X Genomics. Transcriptionally distinct cell subpopulations were identified by DEG: mature adipocytes (ADIPOQ, LEP), endothelial cells (VWR), immune cells (F13A1, MSR1), and two major APC subpopulations (S1: FABP4, ADIRF, ITLN1 and MSLN; S2: APOD, CFD and WISP2). S2 was uniquely identified by the upregulation of extracellular matrix accumulation and fibrosis markers: LUM, DCN, FBN1, and COL1A1, 3A1, 6A1 and 6A3. S2 had higher expression of APC and committed preadipocyte markers, such as PLIN1, GNAI1, CXCL14, CP, and CRISPLD2, while S1 had higher mesothelial-like cell markers GPM6A, PKHD1L1, LRRN4, CXADR, SLPI, and UPK3B. We FACS-isolated S1 and S2 from VAT CD31-CD45- cells (n= 3 NDM, 5 DM) using the surface marker TM4SF1, which was abundantly expressed in S1, but not S2. TM4SF1 expression (qPCR) was 50% higher in S1 (TM4SF1+) than in S2 (TM4SF1-). TM4SF1 immunostaining of human SVF confirmed distinct APC subpopulations. S1 corresponded to 15 ± 3% and S2 to 69 ± 2% of CD31-CD45- cells. In DM compared to NDM, S2 fraction was increased by 12% while S1 was decreased by 45%. We tested the metabolic function of S1 and S2 sorted from 5 DM VAT. Both subpopulations differentiated into lipid-filled adipocytes upon adipogenic induction in vitro. However, S2 compared to S1, manifested decreased proliferation and increased intracellular lipid accumulation and lipolytic response, consistent with a pro-hypertrophic, lipolytic adipocyte phenotype in DM. Our data suggest that distinct APC subpopulations in human obese VAT contribute to adipose tissue dysfunction in DM. Disclosure C. Strieder-Barboza: None. C.G. Flesher: None. L.M. Geletka: None. R.W. ORourke: None. C.N. Lumeng: None. Funding National Institute of Diabetes and Digestive and Kidney Diseases (R01DK115190, DK090262)

Enhanced glycolysis and HIF-1\u03b1 activation in adipose tissue macrophages sustains local and systemic interleukin-1\u03b2 production in obesity

During obesity, macrophages infiltrate the visceral adipose tissue and promote inflammation that contributes to type II diabetes. Evidence suggests that the rewiring of cellular metabolism can regulate macrophage function. However, the metabolic programs that characterize adipose tissue macrophages (ATM) in obesity are poorly defined. Here, we demonstrate that ATM from obese mice exhibit metabolic profiles characterized by elevated glycolysis and oxidative phosphorylation, distinct from ATM from lean mice. Increased activation of HIF-1α in ATM of obese visceral adipose tissue resulted in induction of IL-1β and genes in the glycolytic pathway. Using a hypoxia-tracer, we show that HIF-1α nuclear translocation occurred both in hypoxic and non-hypoxic ATM suggesting that both hypoxic and pseudohypoxic stimuli activate HIF-1α and its target genes in ATM during diet-induced obesity. Exposure of macrophages to the saturated fatty acid palmitate increased glycolysis and HIF-1α expression, which culminated in IL-1β induction thereby simulating pseudohypoxia. Using mice with macrophage-specific targeted deletion of HIF-1α, we demonstrate the critical role of HIF-1α-derived from macrophages in regulating ATM accumulation, and local and systemic IL-1β production, but not in modulating systemic metabolic responses. Collectively, our data identify enhanced glycolysis and HIF-1α activation as drivers of low-grade inflammation in obesity.

Visceral and subcutaneous adipose tissue as markers of local and systemic inflammation: a comparison between celiac and obese patients using MRI.

Celiac disease (CD) is a systemic inflammatory disease, which primarily affects the gastrointestinal tract. It has been recently demonstrated that adipose-tissue infiltration by proinflammatory immune cells causes a chronic low-grade inflammation in obese patients. Magnetic resonance imaging (MRI) has already proved to be useful in evaluation of inflammatory states. The aim of the present study was to determine whether alterations of visceral and subcutaneous adipose tissue, identified with MRI, could serve as markers of local and systemic inflammation in patients with CD. A pilot study was conducted comparing alterations in visceral adipose tissue (VAT) and subcutaneous adipose tissue (SAT) in CDpatients vsobese patients and healthy controls. Fifty patients were enrolled and assigned to one of the following groups: Group A: 11 active CD patients; Group B: 11 CD patients in remission; Group C: 16 obese patients; Group D: 12 healthy controls. A 3-T MRI unit was used and T2-weighted TSE images of VAT and SAT were obtained in specific regions of interest. Serum cytokine concentrations (TNF-α, IL-6, adiponectin, leptin, IL-2, IFN-γ) were determined. There was a significant difference in VAT T2 relaxation time between Group A and B (p < 0.001), A and D (p < 0.01), B and C (p < 0.001). There was a statistically significant difference in SAT T2 relaxation time between Group A and B (p < 0.001), A and C (p < 0.05), A and D (p < 0.001), B and C (p < 0.01). In addition, VAT/SAT T2 relaxation time ratio showed a statistically significant difference between Group A and C (p < 0.05) and between Group B and C (p < 0.01). Only TNF-α and IL-6 significantly correlated with both VAT and VAT/SAT ratio in active CD. MRI showed similar increased visceral inflammatory signals in patients with active CD and obese patients. However, subcutaneous inflammatory signals werehigherin active CD than in all the other groups. These data show that there is a systemic inflammatory state in active CD, whereas chronic inflammation appears confined to VAT in obesity. These data were only partially confirmed by serological cytokine profiles, which showed less specificity than MRI.

Refining the adipose progenitor cell landscape in healthy and obese visceral adipose tissue using single-cell gene expression profiling.

Obesity is a serious health concern and is associated with a reduced quality of life and a number of chronic diseases, including diabetes, heart disease, stroke, and cancer. With obesity rates on the rise worldwide, adipose tissue biology has become a top biomedical research priority. Despite steady growth in obesity-related research, more investigation into the basic biology of adipose tissue is needed to drive innovative solutions aiming to curtail the obesity epidemic. Adipose progenitor cells (APCs) play a central role in adipose tissue homeostasis and coordinate adipose tissue expansion and remodeling. Although APCs are well studied, defining and characterizing APC subsets remains ambiguous because of ill-defined cellular heterogeneity within this cellular compartment. In this study, we used single-cell RNA sequencing to create a cellular atlas of APC heterogeneity in mouse visceral adipose tissue. Our analysis identified two distinct populations of adipose tissue-derived stem cells (ASCs) and three distinct populations of preadipocytes (PAs). We identified novel cell surface markers that, when used in combination with traditional ASC and preadipocyte markers, could discriminate between these APC subpopulations by flow cytometry. Prospective isolation and molecular characterization of these APC subpopulations confirmed single-cell RNA sequencing gene expression signatures, and ex vivo culture revealed differential expansion/differentiation capabilities. Obese visceral adipose tissue featured relative expansion of less mature ASC and PA subpopulations, and expression analyses revealed major obesity-associated signaling alterations within each APC subpopulation. Taken together, our study highlights cellular and transcriptional heterogeneity within the APC pool, provides new tools to prospectively isolate and study these novel subpopulations, and underscores the importance of considering APC diversity when studying the etiology of obesity.

CD8+ T cell/adipocyte inflammatory cross talk and ensuing M1 macrophage polarization are reduced by fish-oil-derived n-3 polyunsaturated fatty acids, in part by a TNF-α-dependent mechanism

Obese visceral adipose tissue (AT) inflammation is driven by adipokine-mediated cross talk between CD8+ T cells and adipocytes, a process mitigated by long-chain (LC) n-3 polyunsaturated fatty acids (PUFA) but underlying mechanisms and ensuing effects on macrophage polarization status are unknown. Using an in vitro co-culture model that recapitulates the degree of CD8+ T cell infiltration reported in obese AT, 3T3-L1 adipocytes were co-cultured for 24 h with purified splenic CD8+ T cells from C57Bl/6 mice consuming either a 10% w/w safflower oil (control, CON) or 7% w/w safflower oil + 3% w/w fish oil (FO) diet for 4 weeks (n=8–10/diet). Co-cultured cells were in direct contact or in a contact-independent condition separated by a Transwell permeable membrane and stimulated with lipopolysaccharide (10 ng/ml) to mimic in vivo obese endotoxin levels. In contact-dependent co-cultures, FO reduced inflammatory (IL-6, TNFα, IFN-γ) and macrophage chemotactic (CCL2, CCL7, CCL3) mRNA expression and/or secreted protein, NF-κB p65 activation, ROS accumulation, NLRP3 inflammasome priming (Nlrp3, Il1β mRNA) and activation (caspase-1 activity) compared to CON (P<.05). The anti-inflammatory action of FO was reproduced by the addition of a TNF-α neutralizing antibody (1 μg/ml) to CON co-cultures (CON/anti-TNF-α), albeit to a lesser degree. Conditioned media from FO and CON/anti-TNF-α co-cultures, in turn, reduced RAW 264.7 macrophage mRNA expression of M1 polarization markers (iNos, Cd11c, Ccr2) and associated inflammatory cytokines (Il6, Tnfα, Il1β) compared to CON. These data suggest that inflammatory CD8+ T cell/adipocyte cross talk is partially attributable to TNF-α signaling, which can be mitigated by LC n-3 PUFA.

Adaptive adipose tissue stromal plasticity in response to cold stress and antibody-based metabolic therapy

In response to environmental and nutrient stress, adipose tissues must establish a new homeostatic state. Here we show that cold exposure of obese mice triggers an adaptive tissue remodeling in visceral adipose tissue (VAT) that involves extracellular matrix deposition, angiogenesis, sympathetic innervation, and adipose tissue browning. Obese VAT is predominated by pro-inflammatory M1 macrophages; cold exposure induces an M1-to-M2 shift in macrophage composition and dramatic changes in macrophage gene expression in both M1 and M2 macrophages. Antibody-mediated CSF1R blocking prevented the cold-induced recruitment of adipose tissue M2 macrophages, suggesting the role of CSF1R signaling in the process. These cold-induced effects in obese VAT are phenocopied by an administration of the FGF21-mimetic antibody, consistent with its action to stimulate sympathetic nerves. Collectively, these studies illuminate adaptive visceral adipose tissue plasticity in obese mice in response to cold stress and antibody-based metabolic therapy.

Netrin-1 Alters Adipose Tissue Macrophage Fate and Function in Obesity.

Macrophages accumulate prominently in the visceral adipose tissue (VAT) of obese humans and high fat diet (HFD) fed mice, and this is linked to insulin resistance and type II diabetes. While the mechanisms regulating macrophage recruitment in obesity have been delineated, the signals directing macrophage persistence in VAT are poorly understood. We previously showed that the neuroimmune guidance cue netrin-1 is expressed in the VAT of obese mice and humans, where it promotes macrophage accumulation. To better understand the source of netrin-1 and its effects on adipose tissue macrophage (ATM) fate and function in obesity, we generated mice with myeloid-specific deletion of netrin-1 (Ntn1fl/fl LysMCre+/–; Ntn1Δmac). Interestingly, Ntn1Δmac mice showed a modest decrease in HFD-induced adiposity and adipocyte size, in the absence of changes in food intake or leptin, that was accompanied by an increase in markers of adipocyte beiging (Prdm16, UCP-1). Using single cell RNA-seq, combined with conventional histological and flow cytometry techniques, we show that myeloid-specific deletion of netrin-1 caused a 50% attrition of ATMs in HFD-fed mice, particularly of the resident macrophage subset, and altered the phenotype of residual ATMs to enhance lipid handling. Pseudotime analysis of single cell transcriptomes showed that in the absence of netrin-1, macrophages in the obese VAT underwent a phenotypic switch with the majority of ATMs activating a program of genes specialized in lipid handling, including fatty acid uptake and intracellular transport, lipid droplet formation and lipolysis, and regulation of lipid localization. Furthermore, Ntn1Δmac macrophages had reduced expression of genes involved in arachidonic acid metabolism, and targeted LCMS/MS metabololipidomics analysis revealed decreases in proinflammatory eicosanoids (5-HETE, 6-trans LTB4, TXB2, PGD2) in the obese VAT. Collectively, our data show that targeted deletion of netrin-1 in macrophages reprograms the ATM phenotype in obesity, leading to reduced adipose inflammation, and improved lipid handling and metabolic function.