Macrophage and T cell networks in adipose tissue.

The signals and structure of the tissues in which leukocytes reside critically mould leukocyte function and development and have challenged our fundamental understanding of how to define and categorize tissue-resident immune cells. One specialized tissue niche that has a powerful effect on immune cell function is adipose tissue. The field of adipose tissue leukocyte biology has expanded dramatically and has revealed how tissue niches can shape immune cell function and reshape them in a setting of metabolic stress, such as obesity. Most notably, adipose tissue macrophages and T cells are under intense investigation due to their contributions to adipose tissue in the lean and obese states. Both adipose tissue macrophages and T cells have features associated with the metabolic function of adipose tissue that are distinct from features of macrophages and T cells that are classically characterized in other tissues. This Review provides state-of-the-art understanding of adipose tissue macrophages and T cells and discusses how their unique niche can help us to better understand diversity in leukocyte responses.

TBK1-mTOR Signaling Attenuates Obesity-Linked Hyperglycemia and Insulin Resistance.

The innate immune kinase TBK1 (TANK-binding kinase 1) responds to microbial-derived signals to initiate responses against viral and bacterial pathogens. More recent work implicates TBK1 in metabolism and tumorigenesis. The kinase mTOR (mechanistic target of rapamycin) integrates diverse environmental cues to control fundamental cellular processes. Our prior work demonstrated in cells that TBK1 phosphorylates mTOR (on S2159) to increase mTORC1 and mTORC2 catalytic activity and signaling. Here we investigate a role for TBK1-mTOR signaling in control of glucose metabolism in vivo. We find that mice with diet-induced obesity (DIO) but not lean mice bearing a whole-body "TBK1-resistant" Mtor S2159A knock-in allele (MtorA/A) display exacerbated hyperglycemia and systemic insulin resistance with no change in energy balance. Mechanistically, Mtor S2159A knock-in in DIO mice reduces mTORC1 and mTORC2 signaling in response to insulin and innate immune agonists, reduces anti-inflammatory gene expression in adipose tissue, and blunts anti-inflammatory macrophage M2 polarization, phenotypes shared by mice with tissue-specific inactivation of TBK1 or mTOR complexes. Tissues from DIO mice display elevated TBK1 activity and mTOR S2159 phosphorylation relative to lean mice. We propose a model whereby obesity-associated signals increase TBK1 activity and mTOR phosphorylation, which boost mTORC1 and mTORC2 signaling in parallel to the insulin pathway, thereby attenuating insulin resistance to improve glycemic control during diet-induced obesity.

Neuregulin 4 suppresses NASH-HCC development by restraining tumor-prone liver microenvironment.

The mammalian liver comprises heterogeneous cell types within its tissue microenvironment that undergo pathophysiological reprogramming in disease states, such as non-alcoholic steatohepatitis (NASH). Patients with NASH are at an increased risk for the development of hepatocellular carcinoma (HCC). However, the molecular and cellular nature of liver microenvironment remodeling that links NASH to liver carcinogenesis remains obscure. Here, we show that diet-induced NASH is characterized by the induction of tumor-associated macrophage (TAM)-like macrophages and exhaustion of cytotoxic CD8+ T cells in the liver. The adipocyte-derived endocrine factor Neuregulin 4 (NRG4) serves as a hormonal checkpoint that restrains this pathological reprogramming during NASH. NRG4 deficiency exacerbated the induction of tumor-prone liver immune microenvironment and NASH-related HCC, whereas transgenic NRG4 overexpression elicited protective effects in mice. In a therapeutic setting, recombinant NRG4-Fc fusion protein exhibited remarkable potency in suppressing HCC and prolonged survival in the treated mice. These findings pave the way for therapeutic intervention of liver cancer by targeting the NRG4 hormonal checkpoint.

Human CD206+ macrophages associate with diabetes and adipose tissue lymphoid clusters.

Increased adipose tissue macrophages (ATMs) correlate with metabolic dysfunction in humans and are causal in development of insulin resistance in mice. Recent bulk and single-cell transcriptomics studies reveal a wide spectrum of gene expression signatures possible for macrophages that depends on context, but the signatures of human ATM subtypes are not well defined in obesity and diabetes. We profiled 3 prominent ATM subtypes from human adipose tissue in obesity and determined their relationship to type 2 diabetes. Visceral adipose tissue (VAT) and s.c. adipose tissue (SAT) samples were collected from diabetic and nondiabetic obese participants to evaluate cellular content and gene expression. VAT CD206+CD11c- ATMs were increased in diabetic participants, were scavenger receptor-rich with low intracellular lipids, secreted proinflammatory cytokines, and diverged significantly from 2 CD11c+ ATM subtypes, which were lipid-laden, were lipid antigen presenting, and overlapped with monocyte signatures. Furthermore, diabetic VAT was enriched for CD206+CD11c- ATM and inflammatory signatures, scavenger receptors, and MHC II antigen presentation genes. VAT immunostaining found CD206+CD11c- ATMs concentrated in vascularized lymphoid clusters adjacent to CD206-CD11c+ ATMs, while CD206+CD11c+ were distributed between adipocytes. Our results show ATM subtype-specific profiles that uniquely contribute to the phenotypic variation in obesity.

The human type 2 diabetes-specific visceral adipose tissue proteome and transcriptome in obesity.

Dysfunctional visceral adipose tissue (VAT) in obesity is associated with type 2 diabetes (DM) but underlying mechanisms remain unclear. Our objective in this discovery analysis was to identify genes and proteins regulated by DM to elucidate aberrant cellular metabolic and signaling mediators. We performed label-free proteomics and RNA-sequencing analysis of VAT from female bariatric surgery subjects with DM and without DM (NDM). We quantified 1965 protein groups, 23 proteins, and 372 genes that were differently abundant in DM vs. NDM VAT. Proteins downregulated in DM were related to fatty acid synthesis and mitochondrial function (fatty acid synthase, FASN; dihydrolipoyl dehydrogenase, mitochondrial, E3 component, DLD; succinate dehydrogenase-α, SDHA) while proteins upregulated in DM were associated with innate immunity and transcriptional regulation (vitronectin, VTN; endothelial protein C receptor, EPCR; signal transducer and activator of transcription 5B, STAT5B). Transcriptome indicated defects in innate inflammation, lipid metabolism, and extracellular matrix (ECM) function, and components of complement classical and alternative cascades. The VAT proteome and transcriptome shared 13 biological processes impacted by DM, related to complement activation, cell proliferation and migration, ECM organization, lipid metabolism, and gluconeogenesis. Our data revealed a marked effect of DM in downregulating FASN. We also demonstrate enrichment of complement factor B (CFB), coagulation factor XIII A chain (F13A1), thrombospondin 1 (THBS1), and integrins at mRNA and protein levels, albeit with lower q-values and lack of Western blot or PCR confirmation. Our findings suggest putative mechanisms of VAT dysfunction in DM.

Obesity results in adipose tissue T cell exhaustion.

Despite studies implicating adipose tissue T cells (ATT) in the initiation and persistence of adipose tissue inflammation, fundamental gaps in knowledge regarding ATT function impedes progress toward understanding how obesity influences adaptive immunity. We hypothesized that ATT activation and function would have tissue-resident-specific properties and that obesity would potentiate their inflammatory properties. We assessed ATT activation and inflammatory potential within mouse and human stromal vascular fraction (SVF). Surprisingly, murine and human ATTs from obese visceral white adipose tissue exhibited impaired inflammatory characteristics upon stimulation. Both environmental and cell-intrinsic factors are implicated in ATT dysfunction. Soluble factors from obese SVF inhibit ATT activation. Additionally, chronic signaling from macrophage major histocompatibility complex II (MHCII) is necessary for ATT impairment in obese adipose tissue but is independent of increased PD1 expression. To assess intracellular signaling mechanisms responsible for ATT inflammation impairments, single-cell RNA sequencing of ATTs was performed. ATTs in obese adipose tissue exhibit enrichment of genes characteristic of T cell exhaustion and increased expression of coinhibitory receptor Btla. In sum, this work suggests that obesity-induced ATTs have functional characteristics and gene expression resembling T cell exhaustion induced by local soluble factors and cell-to-cell interactions in adipose tissue.

Myeloid interleukin-4 receptor α is essential in postmyocardial infarction healing by regulating inflammation and fibrotic remodeling.

Interleukin-4 receptor α (IL4Rα) signaling plays an important role in cardiac remodeling during myocardial infarction (MI). However, the target cell type(s) of IL4Rα signaling during this remodeling remains unclear. Here, we investigated the contribution of endogenous myeloid-specific IL4Rα signaling in cardiac remodeling post-MI. We established a murine myeloid-specific IL4Rα knockout (MyIL4RαKO) model with LysM promoter-driven Cre recombination. Macrophages from MyIL4RαKO mice showed significant downregulation of alternatively activated macrophage markers but an upregulation of classical activated macrophage markers both in vitro and in vivo, indicating the successful inactivation of IL4Rα signaling in macrophages. To examine the role of myeloid IL4Rα during MI, we subjected MyIL4RαKO and littermate floxed control (FC) mice to MI. We found that cardiac function was significantly impaired as a result of myeloid-specific IL4Rα deficiency. This deficiency resulted in a dysregulated inflammatory response consisting of decreased production of anti-inflammatory cytokines. Myeloid IL4Rα deficiency also led to reduced collagen 1 deposition and an imbalance of matrix metalloproteinases (MMPs)/tissue inhibitors of metalloproteinases (TIMPs), with upregulated MMPs and downregulated TIMPs, which resulted in insufficient fibrotic remodeling. In conclusion, this study identifies that myeloid-specific IL4Rα signaling regulates inflammation and fibrotic remodeling during MI. Therefore, myeloid-specific activation of IL4Rα signaling could offer protective benefits after MI.NEW & NOTEWORTHY This study showed, for the first time, the role of endogenous IL4Rα signaling in myeloid cells during cardiac remodeling and the underlying mechanisms. We identified myeloid cells are the critical target cell types of IL4Rα signaling during cardiac remodeling post-MI. Deficiency of myeloid IL4Rα signaling causes deteriorated cardiac function post-MI, due to dysregulated inflammation and insufficient fibrotic remodeling. This study sheds light on the potential of activating myeloid-specific IL4Rα signaling to modify remodeling post-MI. This brings hope to patients with MI and diminishes side effects by cell type-specific instead of whole body treatment.

Wnt/ß-catenin signaling regulates adipose tissue lipogenesis and adipocyte-specific loss is rigorously defended by neighboring stromal-vascular cells.

OBJECTIVE: Canonical Wnt/ß-catenin signaling is a well-studied endogenous regulator of mesenchymal cell fate determination, promoting osteoblastogenesis and inhibiting adipogenesis. However, emerging genetic evidence in humans links a number of Wnt pathway members to body fat distribution, obesity, and metabolic dysfunction, suggesting that this pathway also functions in adipocytes. Recent studies in mice have uncovered compelling evidence that the Wnt signaling pathway plays important roles in adipocyte metabolism, particularly under obesogenic conditions. However, complexities in Wnt signaling and differences in experimental models and approaches have thus far limited our understanding of its specific roles in this context. METHODS: To investigate roles of the canonical Wnt pathway in the regulation of adipocyte metabolism, we generated adipocyte-specific ß-catenin (ß-cat) knockout mouse and cultured cell models. We used RNA sequencing, ChIP sequencing, and molecular approaches to assess expression of Wnt targets and lipogenic genes. We then used functional assays to evaluate effects of ß-catenin deficiency on adipocyte metabolism, including lipid and carbohydrate handling. In mice maintained on normal chow and high-fat diets, we assessed the cellular and functional consequences of adipocyte-specific ß-catenin deletion on adipose tissues and systemic metabolism. RESULTS: We report that in adipocytes, the canonical Wnt/ß-catenin pathway regulates de novo lipogenesis (DNL) and fatty acid monounsaturation. Further, ß-catenin mediates effects of Wnt signaling on lipid metabolism in part by transcriptional regulation of Mlxipl and Srebf1. Intriguingly, adipocyte-specific loss of ß-catenin is sensed and defended by CD45-/CD31- stromal cells to maintain tissue-wide Wnt signaling homeostasis in chow-fed mice. With long-term high-fat diet, this compensatory mechanism is overridden, revealing that ß-catenin deletion promotes resistance to diet-induced obesity and adipocyte hypertrophy and subsequent protection from metabolic dysfunction. CONCLUSIONS: Taken together, our studies demonstrate that Wnt signaling in adipocytes is required for lipogenic gene expression, de novo lipogenesis, and lipid desaturation. In addition, adipose tissues rigorously defend Wnt signaling homeostasis under standard nutritional conditions, such that stromal-vascular cells sense and compensate for adipocyte-specific loss. These findings underscore the critical importance of this pathway in adipocyte lipid metabolism and adipose tissue function.

Cholesterol 25-hydroxylase (CH25H) as a promoter of adipose tissue inflammation in obesity and diabetes.

OBJECTIVE: Expansion of visceral adipose tissue (VAT) and metabolic inflammation are consequences of obesity and associated with type 2 diabetes (T2DM). Metabolically activated adipose tissue macrophages (ATMs) undergo qualitative and quantitative changes that influence their inflammatory responses. How these cells contribute to insulin resistance (IR) in humans is not well understood. Cholesterol 25-Hydroxylase (CH25H) converts cholesterol into 25-Hydroxycholesterol (25-HC), an oxysterol that modulates immune responses. Using human and murine models, we investigated the role of CH25H in metabolic inflammation. METHODS: We performed transcriptomic (RNASeq) analysis on the human whole AT biopsies and sorted ATMs from obese non-diabetic (NDM) and obese diabetic (DM) subjects to inquire if CH25H was increased in DM. We challenged mice lacking Ch25h with a high-fat diet (HFD) to characterize their metabolic and immunologic profiling. Ch25h KO mice and human adipose tissue biopsies from NDM and DM subjects were analyzed. LC-MS was conducted to measure 25-HC level in AT. In vitro analysis permitted us to investigate the effect of 25-HC on cytokine expression. RESULTS: In our RNASeq analysis of human visceral and subcutaneous biopsies, gene pathways related to inflammation were increased in obese DM vs. non-DM subjects that included CH25H. CH25H was enriched in the stromal vascular fraction of human adipose tissue and highly expressed in CD206+ human ATMs by flow cytometry analysis. We measured the levels of the oxysterols, 25-HC and 7α25diHC, in human visceral adipose tissue samples and showed a correlation between BMI and 25-HC. Using mouse models of diet-induced obesity (DIO), we found that HFD-induced Ch25h expression in eWAT and increased levels of 25-HC in AT. On HFD, Ch25h KO mice became obese but exhibited reduced plasma insulin levels, improved insulin action, and decreased ectopic lipid deposit. Improved insulin sensitivity in Ch25h KO mice was due to attenuation of CD11c+ adipose tissue macrophage infiltration in eWAT. Finally, by testing AT explants, bone marrow-derived macrophages (BMDMs) and SVF cells from Ch25h deficient mice, we observed that 25-HC is required for the expression of pro-inflammatory genes. 25-HC was also able to induce inflammatory genes in preadipocytes. CONCLUSIONS: Our data suggest a critical role for CH25H/25-HC in the progression of meta-inflammation and insulin resistance in obese humans and mouse models of obesity. In response to obesogenic stimuli, CH25H/25-HC could exert a pro-inflammatory role.

Viscoelastic characterization of diabetic and non-diabetic human adipose tissue.

BACKGROUND: Obesity-induced chronic inflammation and fibrosis in adipose tissue contributes to the progression of type 2 diabetes mellitus (DM). While fibrosis is known to induce mechanical stiffening of numerous tissue types, it is unknown whether DM is associated with alterations in adipose tissue mechanical properties. OBJECTIVE: The purpose of this study was to investigate whether DM is associated with differences in bulk viscoelastic properties of adipose tissue from diabetic (DM) and non-diabetic (NDM) obese subjects. METHODS: Bulk shear rheology was performed on visceral (VAT) and subcutaneous (SAT) adipose tissue, collected from obese subjects undergoing elective bariatric surgery. Rheology was also performed on the remaining extracellular matrix (ECM) from decellularized VAT (VAT ECM). Linear mixed models were used to assess whether correlations existed between adipose tissue mechanical properties and DM status, sex, age, and body mass index (BMI). RESULTS: DM was not associated with significant differences in adipose tissue viscoelastic properties for any of the tissue types investigated. Tissue type dependent differences were however detected, with VAT having significantly lower shear storage and loss moduli than SAT and VAT ECM independent of DM status. CONCLUSION: Although DM is typically associated with adipose tissue fibrosis, it is not associated with differences in macroscopic adipose tissue mechanical properties.

Adipose tissue dendritic cell signals are required to maintain T cell homeostasis and obesity-induced expansion.

Adipose tissue derived chronic inflammation is a critical component of obesity induced type II diabetes. Major histocompatibility complex II (MHCII) mediated T cell activation within adipose tissue is one mechanism that contributes to this phenotype. However, the contribution of dendritic cells as professional antigen presenting cells in adipose issue has not previously been explored. Using ItgaxCre x MHCIIfl/fl (M11cKO) mice we observed adipose tissue specific changes in adipose tissue leukocytes. While there was a complete knockout of MHCII in dendritic cells, MHCII was also absent on the majority of macrophages. This resulted in reduction of TCR expression in CD4+ T cells in obese adipose tissue, and an increase in CD8+ and CD4+ CD8+ double positive T cells with decreased CD4+ T cells independent of diet type. Increased CD8+ cells were not observed in the spleen, suggesting adipose tissue T cell regulation is tissue specific. In vitro studies demonstrated more potent antigen presentation function in adipose tissue dendritic cells compared to macrophages. Obese M11cKO mice had decreased CD11c+ adipose tissue macrophages. Despite the changes of immune cellularity in adipose tissue, M11cKO largely did not change inflammatory gene expression in adipose tissue and did not demonstrate differences in glucose and insulin intolerance. Overall MHCII expression on CD11c+ cells is important for maintaining CD4+ and CD8+ adipose tissue T cells, but these cellular changes fail to alter inflammatory output and systemic metabolism.

Landscape of Intercellular Crosstalk in Healthy and NASH Liver Revealed by Single-Cell Secretome Gene Analysis.

Cell-cell communication via ligand-receptor signaling is a fundamental feature of complex organs. Despite this, the global landscape of intercellular signaling in mammalian liver has not been elucidated. Here we perform single-cell RNA sequencing on non-parenchymal cells isolated from healthy and NASH mouse livers. Secretome gene analysis revealed a highly connected network of intrahepatic signaling and disruption of vascular signaling in NASH. We uncovered the emergence of NASH-associated macrophages (NAMs), which are marked by high expression of triggering receptors expressed on myeloid cells 2 (Trem2), as a feature of mouse and human NASH that is linked to disease severity and highly responsive to pharmacological and dietary interventions. Finally, hepatic stellate cells (HSCs) serve as a hub of intrahepatic signaling via HSC-derived stellakines and their responsiveness to vasoactive hormones. These results provide unprecedented insights into the landscape of intercellular crosstalk and reprogramming of liver cells in health and disease.

GM-CSF Administration Improves Defects in Innate Immunity and Sepsis Survival in Obese Diabetic Mice.

Sepsis is the leading cause of death in the intensive care unit with an overall mortality rate of 20%. Individuals who are obese and have type 2 diabetes have increased recurrent, chronic, nosocomial infections that worsen the long-term morbidity and mortality from sepsis. Additionally, animal models of sepsis have shown that obese, diabetic mice have lower survival rates compared with nondiabetic mice. Neutrophils are essential for eradication of bacteria, prevention of infectious complications, and sepsis survival. In diabetic states, there is a reduction in neutrophil chemotaxis, phagocytosis, and reactive oxygen species (ROS) generation; however, few studies have investigated the extent to which these deficits compromise infection eradication and mortality. Using a cecal ligation and puncture model of sepsis in lean and in diet-induced obese mice, we demonstrate that obese diabetic mice have decreased "emergency hematopoiesis" after an acute infection. Additionally, both neutrophils and monocytes in obese, diabetic mice have functional defects, with decreased phagocytic ability and a decreased capacity to generate ROS. Neutrophils isolated from obese diabetic mice have decreased transcripts of Axl and Mertk, which partially explains the phagocytic dysfunction. Furthermore, we found that exogenous GM-CSF administration improves sepsis survival through enhanced neutrophil and monocytes phagocytosis and ROS generation abilities in obese, diabetic mice with sepsis.

Properties and functions of adipose tissue macrophages in obesity.

The expansion of adipose tissue (AT) in obesity is accompanied by the accumulation of immune cells that contribute to a state of low-grade, chronic inflammation and dysregulated metabolism. Adipose tissue macrophages (ATMs) represent the most abundant class of leukocytes in AT and are involved in the regulation of several regulatory physiological processes, such as tissue remodeling and insulin sensitivity. With progressive obesity, ATMs are key mediators of meta-inflammation, insulin resistance and impairment of adipocyte function. While macrophage recruitment from blood monocytes is a critical component of the generation of AT inflammation, new studies have revealed a role for ATM proliferation in the early stages of obesity and in sustaining AT inflammation. In addition, studies have revealed a more complex range of macrophage activation states than the previous M1/M2 model, and the existence of different macrophage profiles between human and animal models. This review will summarize the current understanding of the regulatory mechanisms of ATM function in relation to obesity, type 2 diabetes, depot of origin, and to other leukocytes such as AT dendritic cells, with hopes of emphasizing the regulatory nodes that can potentially be targeted to prevent and treat obesity-related metabolic disorders.

The long noncoding RNA Blnc1 orchestrates homeostatic adipose tissue remodeling to preserve metabolic health.

OBJECTIVE: Long noncoding RNAs (lncRNAs) are emerging as powerful regulators of adipocyte differentiation and gene expression. However, their significance in adipose tissue metabolism and physiology has not been demonstrated in vivo. We previously identified Blnc1 as a conserved lncRNA regulator of brown and beige adipocyte differentiation. In this study, we investigated the physiological role of Blnc1 in thermogenesis, adipose remodeling and systemic metabolism. METHODS: We generated fat-specific Blnc1 transgenic and conditional knockout mouse strains and investigated how adipocyte Blnc1 levels are causally linked to key aspects of metabolic health following diet-induced obesity. We performed studies using cultured adipocytes to establish cell-autonomous role of Blnc1 in regulating adipocyte gene programs. RESULTS: Blnc1 is highly induced in both brown and white fats from obese mice. Fat-specific inactivation of Blnc1 impairs cold-induced thermogenesis and browning and exacerbates obesity-associated brown fat whitening, adipose tissue inflammation and fibrosis, leading to more severe insulin resistance and hepatic steatosis. On the contrary, transgenic expression of Blnc1 in adipose tissue elicits the opposite and beneficial metabolic effects, supporting a critical role of Blnc1 in driving adipose adaptation and homeostatic remodeling during obesity. Mechanistically, Blnc1 cell-autonomously attenuates proinflammatory cytokine signaling and promotes fuel storage in adipocytes through its protein partner Zbtb7b. CONCLUSIONS: This study illustrates a surprisingly pleiotropic and dominant role of lncRNA in driving adaptive adipose tissue remodeling and preserving metabolic health.

TLR4, TRIF, and MyD88 are essential for myelopoiesis and CD11c+ adipose tissue macrophage production in obese mice.

Obesity-induced chronic inflammation is associated with metabolic disease. Results from mouse models utilizing a high-fat diet (HFD) have indicated that an increase in activated macrophages, including CD11c+ adipose tissue macrophages (ATMs), contributes to insulin resistance. Obesity primes myeloid cell production from hematopoietic stem cells (HSCs) and Toll-like receptor 4 (TLR4), and the downstream TIR domain-containing adapter protein-inducing interferon-ß (TRIF)- and MyD88-mediated pathways regulate production of similar myeloid cells after lipopolysaccharide stimulation. However, the role of these pathways in HFD-induced myelopoiesis is unknown. We hypothesized that saturated fatty acids and HFD alter myelopoiesis by activating TLR4 pathways in HSCs, differentially producing pro-inflammatory CD11c+ myeloid cells that contribute to obesity-induced metabolic disease. Results from reciprocal bone marrow transplants (BMTs) with Tlr4-/- and WT mice indicated that TLR4 is required for HFD-induced myelopoiesis and production of CD11c+ ATMs. Experiments with homozygous knockouts of Irakm (encoding a suppressor of MyD88 inactivation) and Trif in competitive BMTs revealed that MyD88 is required for HFD expansion of granulocyte macrophage progenitors and that Trif is required for pregranulocyte macrophage progenitor expansion. A comparison of WT, Tlr4-/-, Myd88-/-, and Trif-/- mice on HFD demonstrated that TLR4 plays a role in the production of CD11c+ ATMs, and both Myd88-/- and Trif-/- mice produced fewer ATMs than WT mice. Moreover, HFD-induced TLR4 activation inhibited macrophage proliferation, leading to greater accumulation of recruited CD11c+ ATMs. Our results indicate that HFD potentiates TLR4 and both its MyD88- and TRIF-mediated downstream pathways within progenitors and adipose tissue and leads to macrophage polarization.

Frontline Science: Rapid adipose tissue expansion triggers unique proliferation and lipid accumulation profiles in adipose tissue macrophages.

Obesity-related changes in adipose tissue leukocytes, in particular adipose tissue macrophages (ATMs) and dendritic cells (ATDCs), are implicated in metabolic inflammation, insulin resistance, and altered regulation of adipocyte function. We evaluated stromal cell and white adipose tissue (WAT) expansion dynamics with high fat diet (HFD) feeding for 3-56 days, quantifying ATMs, ATDCs, endothelial cells (ECs), and preadipocytes (PAs) in visceral epididymal WAT and subcutaneous inguinal WAT. To better understand mechanisms of the early response to obesity, we evaluated ATM proliferation and lipid accumulation. ATMs, ATDCs, and ECs increased with rapid WAT expansion, with ATMs derived primarily from a CCR2-independent resident population. WAT expansion stimulated proliferation in resident ATMs and ECs, but not CD11c+ ATMs or ATDCs. ATM proliferation was unperturbed in Csf2- and Rag1-deficient mice with WAT expansion. Additionally, ATM apoptosis decreased with WAT expansion, and proliferation and apoptosis reverted to baseline with weight loss. Adipocytes reached maximal hypertrophy at 28 days of HFD, coinciding with a plateau in resident ATM accumulation and the appearance of lipid-laden CD11c+ ATMs in visceral epididymal WAT. ATM increases were proportional to tissue expansion and adipocyte hypertrophy, supporting adipocyte-mediated regulation of resident ATMs. The appearance of lipid-laden CD11c+ ATMs at peak adipocyte size supports a role in responding to ectopic lipid accumulation within adipose tissue. In contrast, ATDCs increase independently of proliferation and may be derived from circulating precursors. These changes precede and establish the setting in which large-scale adipose tissue infiltration of CD11c+ ATMs, inflammation, and adipose tissue dysfunction contributes to insulin resistance.

The IKK-related kinase TBK1 activates mTORC1 directly in response to growth factors and innate immune agonists.

The innate immune kinase TBK1 initiates inflammatory responses to combat infectious pathogens by driving production of type I interferons. TBK1 also controls metabolic processes and promotes oncogene-induced cell proliferation and survival. Here, we demonstrate that TBK1 activates mTOR complex 1 (mTORC1) directly. In cultured cells, TBK1 associates with and activates mTORC1 through site-specific mTOR phosphorylation (on S2159) in response to certain growth factor receptors (i.e., EGF-receptor but not insulin receptor) and pathogen recognition receptors (PRRs) (i.e., TLR3; TLR4), revealing a stimulus-selective role for TBK1 in mTORC1 regulation. By studying cultured macrophages and those isolated from genome edited mTOR S2159A knock-in mice, we show that mTOR S2159 phosphorylation promotes mTORC1 signaling, IRF3 nuclear translocation, and IFN-ß production. These data demonstrate a direct mechanistic link between TBK1 and mTORC1 function as well as physiologic significance of the TBK1-mTORC1 axis in control of innate immune function. These data unveil TBK1 as a direct mTORC1 activator and suggest unanticipated roles for mTORC1 downstream of TBK1 in control of innate immunity, tumorigenesis, and disorders linked to chronic inflammation.

Weight loss independent changes in adipose tissue macrophage and T cell populations after sleeve gastrectomy in mice.

OBJECTIVE: In addition to adipocytes, adipose tissue contains large numbers of immune cells. A wide range of evidence links the activity of these cells to regulation of adipocyte and systemic metabolic function. Bariatric surgery improves several aspects of metabolic derangements and at least some of these effects occur in a weight-loss independent manner. We sought to investigate the impact of vertical sleeve gastrectomy (VSG) on adipose immune cell frequencies. METHODS: We analyzed the frequencies of immune cells within distinct adipose tissue depots in obese mice that had VSG or sham surgery with a portion of the latter group pair-fed such that their body mass was matched to the VSG animals. RESULTS: We demonstrate that VSG induced a shift in the epididymal adipose tissue leukocyte profile including increased frequencies of CD11c- macrophages, increased frequencies of T cells (CD4+, CD8+, and CD4-/CD8- T cells all increased), but a significantly decreased frequency of adipose tissue dendritic cells (ATDC) that, despite the continued high fat feeding of the VSG group, dropped below control diet levels. CONCLUSIONS: These results indicate that VSG induces substantial changes in the immune populations residing in the adipose depots independent of weight loss.

Adipocyte hypertrophy-hyperplasia balance contributes to weight loss after bariatric surgery.

Predictors of weight loss responses are not well-defined. We hypothesized that adipose tissue phenotypic features related to remodeling would be associated with bariatric surgery weight loss responses. Visceral and subcutaneous adipose tissues collected from patients during bariatric surgery were studied with flow cytometry, immunohistochemistry, and QRTPCR, and results correlated with weight loss outcomes. Age, male sex, and a diagnosis of type 2 diabetes were associated with less weight loss. Adipocyte size was increased and preadipocyte frequency was decreased in visceral adipose tissue from diabetic subjects. Decreased adipose tissue preadipocyte frequency was associated with less weight loss in women but not men. These data suggest that phenotypic features of adipose tissue remodeling may predict responses to weight loss interventions.