Development Of Adipose Tissue Inflammation Research Articles

Notch signaling in adipose tissue macrophages prevents diet-induced inflammation and metabolic dysregulation.

The importance of macrophages in adipose tissue (AT) homeostasis and inflammation is well established. However, the potential cues that regulate their function remain incompletely understood. To bridge this important gap, we sought to characterize novel pathways involved using a mouse model of diet-induced obesity. By performing transcriptomics analysis of AT macrophages (ATMs), we found that late-stage ATMs from high-fat diet mice presented with perturbed Notch signaling accompanied by robust proinflammatory and metabolic changes. To explore the hypothesis that the deregulated Notch pathway contributes to the development of AT inflammation and diet-induced obesity, we employed a genetic approach to abrogate myeloid Notch1 and Notch2 receptors. Our results revealed that the combined loss of Notch1 and Notch2 worsened obesity-related metabolic dysregulation. Body and AT weight gain was higher, blood glucose levels increased and metabolic parameters were substantially worsened in deficient mice fed high-fat diet. Moreover, serum insulin and leptin were elevated as were triglycerides. Molecular analysis of ATMs showed that deletion of Notch receptors escalated inflammation through the induction of an M1-like pro-inflammatory phenotype. Our findings thus support a protective role of myeloid Notch signaling in adipose tissue inflammation and metabolic dysregulation.

Estrogen contributes to the sex difference in the occurrence of senescence-related T cells during the development of visceral adipose tissue inflammation.

It remains unclear whether sex differences exist during the development of visceral adipose tissue (VAT) inflammation associated with obesity. The purpose of this study was to clarify sex differences in the occurrence of senescence-related T cells (CD44high PD-1+ CD4+), which play a key role in the progression of VAT inflammation associated with high-fat diet (HFD)-induced obesity. Phase 1: C57BL/6N mice were fed either a control diet (HFC) or HFD for 5 wk. The area under the curve of the oral glucose-tolerance test (oGTT) was maximal at 15 wk in HFD-fed males and at 21 wk in females. At 17 wk, VAT weights were similar, but an increase in the number of macrophages in the VAT was observed only in HFD-fed males. In addition, the numbers of regulatory and senescence-related T cells were consistently higher in males than in females. Phases 2 and 3: 6-wk-old female mice were randomly divided into sham operation and bilateral ovariectomy (OVX) groups and fed either an HFC or HFD from 7 wk. OVX mice were subjected to 17β-estradiol releasing or placebo pellet implantation and fed an HFC. Body and VAT weights were higher in the OVX group than in the sham. The number of macrophages did not change in the OVX group with either diet. HFC-fed OVX mice exhibited high senescence-related T cells in the VAT, resembling HFC-fed male mice. This change was abolished by 17β-estradiol replacement. Thus, we demonstrated different accumulation patterns of VAT immune cells between the sexes, revealing a role for estrogen in the appearance of senescence-related T cells.NEW & NOTEWORTHY The accumulation pattern of adipose tissue differs between the sexes; however, it is unclear whether sex differences exist during the development of adipose tissue inflammation and whether estrogen plays a role. We demonstrated sex differences in immune cells' subpopulation of visceral adipose tissue. The proinflammatory environment appeared earlier in males than in females. In addition, our results suggest that estrogen plays a role in visceral adipose tissue inflammation, particularly by regulating the appearance of senescence-related T cells.

Grape pomace reduces the severity of non-alcoholic hepatic steatosis and the development of steatohepatitis by improving insulin sensitivity and reducing ectopic fat deposition in mice

While non-alcoholic fatty liver disease (NAFLD) represents the common cause of chronic liver disease, specific therapies are currently unavailable. The wine industry produces millions of tons of residue (pomace), which contains high levels of bioactive phytochemicals. The aim of this study was to clarify the potential benefits of grape pomace for the treatment of NAFLD at different levels of severity, and to clarify the mechanism of action. C57Bl/6 mice were given high fat diet (HFD) or western diet (WD) as models of obesity and hepatic steatosis or steatohepatitis, respectively, with or without pomace supplementation (50–250 mg/day). Pomace inhibited food intake, and reduced serum leptin and body weight gain. Ectopic fat deposition was reduced, while white adipose tissue mass was preserved. In addition, pomace improved glucose tolerance and insulin sensitivity, prevented the development of adipose tissue inflammation, and reduced hepatic steatosis. Higher expression of genes involved in fatty acids transport and oxidation was observed in adipose tissue, while lipogenic genes were attenuated in the liver of pomace-treated mice. In WD-fed mice, pomace reduced the severity of hepatic steatosis and inflammation and improved blood lipid profile, but was ineffective in reversing hepatic damage of advanced NASH. In conclusion, pomace improved insulin sensitivity and reduced ectopic fat deposition, leading to a healthier metabolic profile. Pomace may hold the potential as a supplement with beneficial health outcomes for the prevention and treatment of hepatic steatosis and other obesity-related pathologies.

Toll-like receptors in the pathophysiology of obesity

Obesity is a complex and relevant global medical and social problem. The adipose tissue is not only a place of deposition of energy substrates but also a source of secretion of pro-inflammatory and anti-inflammatory mediators involved in the development of the chronic latent systemic inflammatory process in the organism with obesity. The metabolic signal in obesity contributes to the polarization of macrophages in the M1 direction and triggers the Th1 immune response, causing the development of adipose tissue inflammation. A chronic inflammatory condition plays a key role in the pathophysiology of obesity-induced insulin resistance. Toll-like receptors (TLRs) may be a possible pathophysiological link in the development of insulin resistance in inflammation. At the same time, inflammation-induced lipolysis is necessary for the release of energy resources during the development of the infectious process. Thus, low-grade inflammation is important to protect against adipocyte dysfunction. These results suggest that pro-inflammatory signaling is not exclusively pathogenic in obesity. In this regard, the study of inflammatory signaling pathways involved in the modulation of chronic inflammation of adipose tissue is particularly relevant. This review summarizes current views on the structure, function of TLRs and their involvement in the pathogenesis of chronic inflammation in obesity. The possibility of using TLRs as a therapeutic target in this pathology is discussed. Obviously, further study of inflammatory signaling pathways involving TLRs initiating the development of chronic inflammation of adipose tissue will allow the development of new and effective therapeutic strategies for obesity and its metabolic complications.

Obesity induces preadipocyte CD36 expression promoting inflammation via the disruption of lysosomal calcium homeostasis and lysosome function.

BackgroundPreadipocyte is closely related to obesity-induced inflammation. The impairment of autophagic flux by defective lysosomal function has been observed in adipose tissue from obese mice. While the fatty acid translocase CD36 is an important immuno-metabolic receptor, it remains unclear whether preadipocyte CD36 is involved in adipose tissue inflammation and whether CD36 regulates lysosomal function.MethodsUsing visceral adipose tissue from obese patients, a high-fat diet (HFD)-induced obese mice model, primary mouse preadipocytes and 3T3L1 cells we analyzed whether and how preadipocyte CD36 modulates lysosomal function and adipose tissue inflammation.FindingsCD36 expression in preadipocytes is induced in obese patients and HFD-fed mice, accompanied with the disruption of lysosome function. CD36 knockout protects primary preadipocytes of HFD-fed mice from lysosomal impairment. In vitro, CD36 interacts with Fyn to phosphorylate and activate Inositol (1,4,5)-trisphosphate receptor 1 (IP3R1), causing excess calcium transport from endoplasmic reticulum (ER) to lysosome, which results in lysosomal impairment and inflammation. Moreover, IP3R inhibitor 2-aminoethoxydiphenyl borate (2APB) attenuates lysosomal impairment, inflammation and lipid accumulation in CD36-overexpressing preadipocytes.InterpretationOur data support that the abnormal upregulation of CD36 in preadipocytes may contribute to the development of adipose tissue inflammation. CD36/Fyn/IP3R1-mediated lysosomal calcium overload leads to lysosomal impairment and inflammation in preadipocyte. Thus targeting improving lysosomal calcium homeostasis may represent a novel strategy for treating obesity-induced inflammation.

Bidirectional crosstalk between neutrophils and adipocytes promotes adipose tissue inflammation

Abstract Obesity-associated adipose tissue inflammation contributes to the development of type 2 diabetes. Chronic activation of IL-1β system in adipose tissue on metabolic disorders is well demonstrated. However, a mechanism for its expression and activation in the tissue has remained unexplored. Here we demonstrate that IL-1β transcript was enriched in neutrophils of white adipose tissue (WAT), and the expression of IL-1β was up-regulated by a high-fat diet in the WAT but not in other tissues, including bone marrow, peripheral blood, and spleen. The co-culture of neutrophils with 3T3-L1 adipocytes indicate that contact interaction with adipocytes is critical for the expression of IL-1β via NF-κB activation in adipose tissue neutrophils. Lipolysis of adipocytes accumulated neutrophils prior to macrophages in the WAT, and produced high levels of IL-1β via an ASC-caspase-1-mediated inflammasome pathway. Leukotriene B4 production in the WAT also contributed to neutrophil accumulation. Furthermore, neutrophils and IL-1β contributed to the expression of chemotactic molecules involved in high-fat diet-induced macrophage infiltration into the WAT. We have identified previously unappreciated roles for neutrophils in the development of adipose tissue inflammation: robust IL-1β production and infiltration of macrophages to initiate chronic inflammation.

Chronic Adipose Tissue Inflammation Linking Obesity to Insulin Resistance and Type 2 Diabetes.

Obesity is one of the major health burdens of the 21st century as it contributes to the growing prevalence of its related comorbidities, including insulin resistance and type 2 diabetes. Growing evidence suggests a critical role for overnutrition in the development of low-grade inflammation. Specifically, chronic inflammation in adipose tissue is considered a crucial risk factor for the development of insulin resistance and type 2 diabetes in obese individuals. The triggers for adipose tissue inflammation are still poorly defined. However, obesity-induced adipose tissue expansion provides a plethora of intrinsic signals (e.g., adipocyte death, hypoxia, and mechanical stress) capable of initiating the inflammatory response. Immune dysregulation in adipose tissue of obese subjects results in a chronic low-grade inflammation characterized by increased infiltration and activation of innate and adaptive immune cells. Macrophages are the most abundant innate immune cells infiltrating and accumulating into adipose tissue of obese individuals; they constitute up to 40% of all adipose tissue cells in obesity. In obesity, adipose tissue macrophages are polarized into pro-inflammatory M1 macrophages and secrete many pro-inflammatory cytokines capable of impairing insulin signaling, therefore promoting the progression of insulin resistance. Besides macrophages, many other immune cells (e.g., dendritic cells, mast cells, neutrophils, B cells, and T cells) reside in adipose tissue during obesity, playing a key role in the development of adipose tissue inflammation and insulin resistance. The association of obesity, adipose tissue inflammation, and metabolic diseases makes inflammatory pathways an appealing target for the treatment of obesity-related metabolic complications. In this review, we summarize the molecular mechanisms responsible for the obesity-induced adipose tissue inflammation and progression toward obesity-associated comorbidities and highlight the current therapeutic strategies.

Obesity Induces Preadipocyte CD36 Expression Promoting Inflammation <i>via</i> the Disruption of Lysosomal Calcium Homeostasis and Lysosome Function

Background: Preadipocyte is one of the major contributors to obesity-induced inflammation. The impairment of autophagic flux by defective lysosomal function has been observed in adipose tissue from obese mice. While the fatty acid translocase CD36 is an important immuno-metabolic receptor, it remains unclear whether preadipocyte CD36 is involved in adipose tissue inflammation and whether CD36 regulates lysosomal function. Methods: Using visceral adipose tissue from obese patients, a high-fat diet (HFD)-induced obese mice model, primary mouse preadipocytes and 3T3L1 cells we analyzed whether and how preadipocyte CD36 modulates lysosomal function and adipose tissue inflammation. Findings: CD36 expression in preadipocytes is induced in obese patients and HFD-fed mice, accompanied with decreased lysosomal numbers and impaired lysosomal acidification. CD36 knockout protects lysosomal impairment in primary preadipocytes from mice. In vitro, CD36 interacts with Fyn to phosphorylate and activate Inositol (1,4,5)-trisphosphate receptor 1 (IP3R1), causing excess calcium transport from endoplasmic reticulum (ER) to lysosome, which results in lysosomal impairment and inflammation. Moreover, IP3R inhibitor 2-aminoethoxydiphenyl borate (2APB) attenuated lysosomal impairment, inflammation and lipid accumulation in CD36-overexpressing preadipocytes. Interpretation: Our data support that increased CD36 expression in preadipocytes is essential for the development of adipose tissue inflammation. CD36/Fyn/IP3R1-mediated lysosomal calcium overload leads to lysosomal impairment and inflammation in preadipocyte. Thus targeting improving lysosomal calcium homeostasis may represent a novel strategy for treating obesity-induced inflammation. Funding Statement: This work was supported by National Key R&D Program of China (2018YFC1312700), the National Natural Science Foundation of China (81873569, 81970510, 31971084) and the Science and Technology Research Program of Chongqing Municipal Education Commission (Grant No.KJZD-K201800401, KJQN201900438). Declaration of Interests: The authors declare that they have no conflict of interest. Ethics Approval Statement: The study was approved by the Ethics Committees of CQMU, and all subjects provided written informed consent. All mouse care and experimental procedures were approved by the Institutional Animal Care and Use Committee of CQMU.

CD11c-MHC2low Macrophages Are a New Inflammatory and Dynamic Subset in Murine Adipose Tissue.

BackgroundThe prevalence of obesity is rising and leads to increased morbidity and mortality. Adipose tissue inflammation, due to accumulation and activation of adipose tissue macrophages (ATMs), is a key driver of this phenomenon. Macrophages are heterogeneous cells, adapting quickly to the microenvironment, resulting in so-called M1 or M2 macrophages. In this study, we describe the dynamics and inflammatory properties of a newly identified ATM subset in obese mice.MethodsLDLR-/- mice received a high fat diet (HFD) for 5 weeks or 16 weeks to induce obesity. Adipose tissues were isolated and immune cell subsets were analyzed with flow cytometry or microarray analysis. Bone marrow transplantation (BMT) using CD45.1 and CD45.2 LDLR-/- mice was performed to determine ATM origin.ResultsUpon HFD, there is a massive increase of ATM subsets in the adipose tissue. CD11c−M2 ATMs could be subdivided based on their MHC2 expression into CD11c−MHC2high ATMs and previously unidentified CD11c−MHC2low ATMs. CD11c−MHC2low ATMs accumulated very rapidly after 10 days of HFD, after which they increased even further with prolonged HFD. Microarray data showed that CD11c−MHC2low ATMs resembled CD11c−MHC2high ATMs in the steady state, but became more inflammatory during development of obesity. In vitro stimulation of bone marrow-derived macrophages with palmitate, abundantly present in HFD, resulted in the induction of the CD11c−MHC2low phenotype.ConclusionsAmong M2 macrophages, a novel pro-inflammatory subset of macrophages was found based on their low level of MHC2 expression. This subset may play a role in the development of adipose tissue inflammation.

Anti-inflammatory effects of Sarcopoterium spinosum extract.

Sarcopoterium spinosum is a Mediterranean plant, widely used by the Beduin traditional medicine. While its major use is for the treatment of diabetes, there are also documentations for its use as anti-inflammatory agent. This therapeutic potential of Sarcopoterium spinosum was not validated before. To investigate the anti-inflammatory properties of Sarcopoterium spinosum extract (SSE). Experiments were performed on RAW264.7 macrophages and bone marrow-derived macrophages (BMDM) and the effect of SSE on markers of inflammation was investigated. In addition, the effect of SSE on the development of inflammation in adipose-tissue of obese, insulin resistant mice was measured in KK-Ay mice and high fat diet (HFD)-fed mice. Lipopolysaccharide (LPS) and SSE increased the viability of RAW264.7. In addition, the cells acquired distinct dendritic-like morphology, however, while LPS induced NO production and the mRNA expression of pro-inflammatory cytokines, SSE increased the mRNA expression of anti-inflammatory genes and blocked LPS effects. All three pathways of MAPK were activated by LPS and SSE, as demonstrated by the phosphorylation of ERK, p38 and JNK. NFκB was activated and Akt was phosphorylated by LPS, while SSE blocked this effects. STAT proteins were differently phosphorylated by SSE and LPS. Immunomodulatory effects of SSE were also found in BMDM. In adipose tissue of SSE-treated mice, less crown-like structures were found, and lower expression of pro-inflammatory adipocytokines was observed, although adipocytes hypertrophy was not affected. SSE has an immunomodulatory effects that affect macrophage function, and reduces adipose tissue inflammation. Identifying active component and clarifying its mechanism of action might support the development of new anti-inflammatory agent.

Bidirectional crosstalk between neutrophils and adipocytes promotes adipose tissue inflammation.

Chronic activation of the IL-1β system in adipose tissue on metabolic disorders is well demonstrated. However, a mechanism for its expression and activation in the tissue has remained unexplored. Here, we demonstrate that IL-1β transcript was enriched in neutrophils of white adipose tissue (WAT) from lean mice. Mechanistically, the interaction of neutrophils with adipocytes induced IL-1β expression via NF-κB pathway. Lipolysis of adipocytes accumulated neutrophils prior to macrophages in WAT and produced high levels of IL-1β via an inflammasome pathway. Leukotriene B4 (LTB4) production in WAT also contributed to neutrophil accumulation. Furthermore, an LTB4-inflammasome axis contributed to the expression of chemotactic molecules involved in high-fat diet-induced macrophage infiltration into WAT. We have identified previously unappreciated roles for neutrophils in the development of adipose tissue inflammation: robust IL-1β production and infiltration of macrophages to initiate chronic inflammation.-Watanabe, Y., Nagai, Y., Honda, H., Okamoto, N., Yanagibashi, T., Ogasawara, M., Yamamoto, S., Imamura, R., Takasaki, I., Hara, H., Sasahara, M., Arita, M., Hida, S., Taniguchi, S., Suda, T., Takatsu, K. Bidirectional crosstalk between neutrophils and adipocytes promotes adipose tissue inflammation.

The Dualistic Effect of COX-2-Mediated Signaling in Obesity and Insulin Resistance.

Obesity and insulin resistance are two major risk factors for the development of metabolic syndrome, type 2 diabetes and associated cardiovascular diseases (CVDs). Cyclooxygenase (COX), a rate-limiting enzyme responsible for the biosynthesis of prostaglandins (PGs), exists in two isoforms: COX-1, the constitutive form, and COX-2, mainly the inducible form. COX-2 is the key enzyme in eicosanoid metabolism that converts eicosanoids into a number of PGs, including PGD2, PGE2, PGF2α, and prostacyclin (PGI2), all of which exert diverse hormone-like effects via autocrine or paracrine mechanisms. The COX-2 gene and immunoreactive proteins have been documented to be highly expressed and elevated in adipose tissue (AT) under morbid obesity conditions. On the other hand, the environmental stress-induced expression and constitutive over-expression of COX-2 have been reported to play distinctive roles under different pathological and physiological conditions; i.e., over-expression of the COX-2 gene in white AT (WAT) has been shown to induce de novo brown AT (BAT) recruitment in WAT and then facilitate systemic energy expenditure to protect mice against high-fat diet-induced obesity. Hepatic COX-2 expression was found to protect against diet-induced steatosis, obesity, and insulin resistance. However, COX-2 activation in the epidydimal AT is strongly correlated with the development of AT inflammation, insulin resistance, and fatty liver in high-fat-diet-induced obese rats. This review will provide updated information regarding the role of COX-2-derived signals in the regulation of energy metabolism and the pathogenesis of obesity and MS.

248-LB: Deficiency of APPL1 in Macrophages Triggers Adipose Tissue Inflammation and Insulin Resistance by Potentiating NLRP3 Inflammasome Activation

Adipose tissue inflammation participates in the pathogenesis of type 2 diabetes in obesity. While the activation of the multiprotein complex known as NLRP3 inflammasome and subsequent release of IL-1β contribute to the development of adipose tissue inflammation and insulin resistance, the mechanisms of its regulation remain unclear. Here we revealed that APPL1, an adaptor protein essential for adiponectin and insulin signaling, is upregulated in the stromal vascular fraction of visceral fat of db/db diabetic mice. Also, lack of APPL1 in bone marrow-derived macrophages specifically enhanced inflammatory response mediated by NLRP3 inflammasome. Mice with hematopoietic cells-specific APPL1 deficiency (APPL1-BMT-KO) were then generated using bone marrow transplantation approach and subjected to high-fat diet (HFD) feeding. After 36 weeks of HFD feeding, APPL1-BMT-KO mice exhibited increased insulin resistance and glucose intolerance, accompanied with elevated IL-1β in serum, liver and epididymal adipose tissue and impaired Akt phosphorylation in the subcutaneous adipose tissue and liver. In vitro experiments suggest that ablation of APPL1 in bone marrow-derived macrophages promotes IL-1β secretion by enhancing ROS generation, which is an important intracellular signal for NLRP3 inflammasome activation. Pharmacological inhibition of intracellular and mitochondrial ROS successfully abrogated the upregulation of IL-1β secretion in APPL1 deficient macrophages. In this study, we demonstrated the novel role of APPL1 in alleviating macrophage-mediated adipose tissue inflammation and insulin resistance. Lack of APPL1 in macrophages promotes ROS generation and NLRP3 inflammasome activation, which causes enhanced IL-1β secretion in adipose tissue and the development of insulin resistance. Disclosure K.K.L. Wu: None. A. Xu: None. K.K. Cheng: None. Funding Research Grants Council of Hong Kong (171018/15M)

Coronary Artery Disease Is Associated with an Increased Amount of T Lymphocytes in Human Epicardial Adipose Tissue.

Immunocompetent cells including lymphocytes play a key role in the development of adipose tissue inflammation and obesity-related cardiovascular complications. The aim of the study was to explore the relationship between epicardial adipose tissue lymphocytes and coronary artery disease (CAD). To this end, we studied the content and phenotype of lymphocytes in peripheral blood, subcutaneous adipose tissue (SAT), and epicardial adipose tissue (EAT) in subjects with and without CAD undergoing elective cardiac surgery. Eleven subjects without CAD (non-CAD group) and 22 age-, BMI-, and HbA1C-matched individuals with CAD were included into the study. Blood, SAT, and EAT samples were obtained at the beginning of surgery. Lymphocyte populations were quantified as % of CD45+ cells using flow cytometry. Subjects with CAD had a higher total lymphocyte amount in EAT compared with SAT (32.24 ± 7.45 vs. 11.22 ± 1.34%, p = 0.025) with a similar trend observed in non-CAD subjects (29.68 ± 7.61 vs. 10.13 ± 2.01%, p = 0.067). T (CD3+) cells were increased (75.33 ± 2.18 vs. 65.24 ± 4.49%, p = 0.032) and CD3- cells decreased (21.17 ± 2.26 vs. 31.64 ± 4.40%, p = 0.028) in EAT of CAD relative to the non-CAD group. In both groups, EAT showed an elevated percentage of B cells (5.22 ± 2.43 vs. 0.96 ± 0.21%, p = 0.039 for CAD and 12.49 ± 5.83 vs. 1.16 ± 0.19%, p = 0.016 for non-CAD) and reduced natural killer (NK) cells (5.96 ± 1.32 vs. 13.22 ± 2.10%, p = 0.012 for CAD and 5.32 ± 1.97 vs. 13.81 ± 2.72%, p = 0.022 for non-CAD) relative to SAT. In conclusion, epicardial adipose tissue in subjects with CAD shows an increased amount of T lymphocytes relative to non-CAD individuals as well as a higher number of total and B lymphocytes and reduced NK cells as compared with corresponding SAT. These changes could contribute to the development of local inflammation and coronary atherosclerosis.

Anaphylatoxin Receptors C3aR and C5aR1 Are Important Factors That Influence the Impact of Ethanol on the Adipose Secretome

Background and aims: Chronic ethanol exposure results in inflammation in adipose tissue; this response is associated with activation of complement as well as the development of alcohol-related liver disease (ALD). Adipose communicates with other organs, including liver, via the release of soluble mediators, such as adipokines and cytokines, characterized as the “adipose secretome.” Here we investigated the role of the anaphaylatoxin receptors C3aR and C5aR1 in the development of adipose tissue inflammation and regulation of the adipose secretome in murine ALD (mALD).Methods: Wild-type C57BL/6 (WT), C3aR−/−, and C5aR1−/− mice were fed Lieber-DeCarli ethanol diet for 25 days (6% v/v, 32% kcal) or isocaloric control diets; indicators of inflammation and injury were assessed in gonadal adipose tissue. The adipose secretome was characterized in isolated adipocytes and stromal vascular cells.Results: Ethanol feeding increased the expression of adipokines, chemokines and leukocyte markers in gonadal adipose tissue from WT mice; C3aR−/− were partially protected while C5aR1−/− mice were completely protected. In contrast, induction of CYP2E1 and accumulation of TUNEL-positive cells in adipose in response to ethanol feeding was independent of genotype. Bone marrow chimeras, generated with WT and C5aR1−/− mice, revealed C5aR1 expression on non-myeloid cells, likely to be adipocytes, contributed to ethanol-induced adipose inflammation. Chronic ethanol feeding regulated both the quantity and distribution of adipokines secreted from adipocytes in a C5aR1-dependent mechanism. In WT mice, chronic ethanol feeding induced a predominant release of pro-inflammatory adipokines from adipocytes, while the adipose secretome from C5aR1−/− mice was characterized by an anti-inflammatory/protective profile. Further, the cargo of adipocyte-derived extracellular vesicles (EVs) was distinct from the soluble secretome; in WT EVs, ethanol increased the abundance of pro-inflammatory mediators while EV cargo from C5aR1−/− adipocytes contained a greater diversity and more robust expression of adipokines.Conclusions: C3aR and C5aR1 are potent regulators of ethanol-induced adipose inflammation in mALD. C5aR1 modulated the impact of chronic ethanol on the content of the adipose secretome, as well as influencing the cargo of an extensive array of adipokines from adipocyte-derived EVs. Taken together, our data demonstrate that C5aR1 contributes to ethanol-mediated changes in the adipose secretome, likely contributing to intra-organ injury in ALD.

The Role of Epicardial Adipose Tissue Lymphocytes in Low-Grade Inflammation and Coronary Artery Disease

Lymphocytes play a role in the development of adipose tissue inflammation. Epicardial adipose tissue was shown to be a potential source of inflammatory mediators contributing to coronary atherosclerosis. The aim of the study was to assess lymphocyte content in peripheral blood, subcutaneous (SAT) and epicardial adipose tissue (EAT) in subjects with coronary artery disease (CAD) along with the effects of elective cardiac surgery. Twelve subjects without CAD (non-CAD group) and 24 age-, BMI- and HbA1C-matched subjects with CAD were included into the study. Blood, SAT and EAT samples were obtained before and at the end of elective cardiac surgery. Lymphocyte populations were quantified as percent of CD45+ cells using flow cytometry. Subjects with CAD had higher total lymphocyte (CD45+) amount in EAT compared to SAT (32.24±7.45 vs. 11.22±1.34%, p=0.025) with a similar trend observed in non-CAD subjects (29.68±7.61 vs. 10.13±2.01%, p=0.067). CD3+ cells were increased and CD3- cells decreased in EAT of CAD relative to non-CAD group with no difference between EAT and SAT. In both groups EAT showed reduced percentage of NK cells (CD15/56+CD3-) and elevated B cells (CD19+) relative to SAT (NK cells: 5.96±1.32 vs. 13.22±2.10%, p=0.012 for CAD and 5.32±1.97 vs. 13.81±2.72%, p=0.022 for non-CAD; B cells: 5.22±2.43 vs. 0.96±0.21%, p=0.039 for CAD and 12.49±5.83 vs. 1.16±0.19%, p=0.016 for non-CAD). CD8+ and NKT (CD16/56+CD3+) cells in EAT correlated positively with fasting glucose and HbA1C, while showing no association in SAT. Cardiac surgery decreased total lymphocytes in peripheral blood, SAT and EAT of both groups, while having only minor effects on lymphocyte subpopulations. To conclude, epicardial adipose tissue in subjects with CAD shows increased amount of total and CD3+ lymphocytes as compared with SAT or non-CAD subjects. These changes could contribute to the development of local inflammation and coronary atherosclerosis. Disclosure M. Mraz: None. A. Cinkajzlova: None. Z. Lacinov : None. J. Klouckova: None. H. Kratochvilova: None. M. Lips: None. P. Kopecky: None. M. Porizka: None. J. Lindner: None. M. Haluzik: None.

Targeted disruption of the iNOS gene improves adipose tissue inflammation and fibrosis in leptin-deficient ob/ob mice: role of tenascin C

Background/Objectives:Obesity is related to a dynamic extracellular matrix (ECM) remodeling, which involves the synthesis and degradation of different proteins, such as tenascin C (TNC) in the adipose tissue (AT). Given the functional relationship between leptin and inducible nitric oxide synthase (iNOS), our aim was to analyze the impact of the absence of the iNOS gene in AT inflammation and ECM remodeling in ob/ob mice.Subjects/Methods:The expression of genes involved in inflammation and ECM remodeling was evaluated in 10-week-old male double knockout (DBKO) mice simultaneously lacking the ob and iNOS genes as well as in ob/ob mice classified into three groups [control, leptin-treated (1 mg kg−1 day−1) and pair-fed].Results:Leptin deficiency increased inflammation and fibrosis in AT. As expected, leptin treatment improved the obesity phenotype. iNOS deficiency in ob/ob mice improved insulin sensitivity, AT inflammation, and ECM remodeling, as evidenced by lower AT macrophage infiltration and collagen deposition, a downregulation of proinflammatory and profibrogenic genes Tnf, Emr1, Hif1a, Col6a1, Col6a3, and Tnc, as well as lower circulating TNC levels. Interestingly, leptin upregulated TNC expression and release in 3T3-L1 adipocytes, and iNOS knockdown in 3T3-L1 fat cells produced a significant decrease in basal and leptin-induced Tnc expression.Conclusions:Ablation of iNOS in leptin-deficient mice improved AT inflammation and ECM remodeling-related genes, attenuating fibrosis, and metabolic dysfunction. The activation of iNOS by leptin is necessary for the synthesis and secretion of TNC in adipocytes, suggesting an important role of this alarmin in the development of AT inflammation and fibrosis.

SUCNR1-mediated chemotaxis of macrophages aggravates obesity-induced inflammation and diabetes

Aims/hypothesisObesity induces macrophages to drive inflammation in adipose tissue, a crucial step towards the development of type 2 diabetes. The tricarboxylic acid (TCA) cycle intermediate succinate is released from cells under metabolic stress and has recently emerged as a metabolic signal induced by proinflammatory stimuli. We therefore investigated whether succinate receptor 1 (SUCNR1) could play a role in the development of adipose tissue inflammation and type 2 diabetes.MethodsSuccinate levels were determined in human plasma samples from individuals with type 2 diabetes and non-diabetic participants. Succinate release from adipose tissue explants was studied. Sucnr1−/− and wild-type (WT) littermate mice were fed a high-fat diet (HFD) or low-fat diet (LFD) for 16 weeks. Serum metabolic variables, adipose tissue inflammation, macrophage migration and glucose tolerance were determined.ResultsWe show that hypoxia and hyperglycaemia independently drive the release of succinate from mouse adipose tissue (17-fold and up to 18-fold, respectively) and that plasma levels of succinate were higher in participants with type 2 diabetes compared with non-diabetic individuals (+53%; p < 0.01). Sucnr1−/− mice had significantly reduced numbers of macrophages (0.56 ± 0.07 vs 0.92 ± 0.15 F4/80 cells/adipocytes, p < 0.05) and crown-like structures (0.06 ± 0.02 vs 0.14 ± 0.02, CLS/adipocytes p < 0.01) in adipose tissue and significantly improved glucose tolerance (p < 0.001) compared with WT mice fed an HFD, despite similarly increased body weights. Consistently, macrophages from Sucnr1−/− mice showed reduced chemotaxis towards medium collected from apoptotic and hypoxic adipocytes (−59%; p < 0.05).Conclusions/interpretationOur results reveal that activation of SUCNR1 in macrophages is important for both infiltration and inflammation of adipose tissue in obesity, and suggest that SUCNR1 is a promising therapeutic target in obesity-induced type 2 diabetes.Data availabilityThe dataset generated and analysed during the current study is available in GEO with the accession number GSE64104, www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE64104.

Circulating classical monocytes are associated with CD11c+ macrophages in human visceral adipose tissue

Immune cell accumulation in adipose tissue (AT) is associated with the development of AT inflammation, resulting in metabolic dysfunction. Circulating immune cell patterns may reflect immune cell accumulation in expanding AT. However, data linking human leukocytes in blood and AT is lacking. We investigated whether blood immune cell populations are associated with their counterparts in subcutaneous (scAT) or visceral AT (vAT). Flow cytometry was performed on blood, scAT and vAT from 16 lean and 29 obese men. Circulating natural killer (NK)-cells, classical monocytes and nonclassical monocytes were higher in obese individuals. vAT, but not scAT, of obese individuals contained more inflammatory CD11c+ “M1” macrophages and NK cells compared to lean individuals. Blood classical monocytes were associated with CD11c+ macrophages in vAT but not scAT. This association was unrelated to expression of the adhesion molecules CD11b and CD11c or of the chemokine receptor CX3CR1 on these monocytes. Other AT immune cells were not associated with their respective counterparts in blood. Finally, CD11c+ macrophages and CD4+ T-cells in vAT were associated with their counterparts in scAT. In conclusion, blood classical monocytes reflect CD11c+ macrophages in vAT.

Adipocyte-Macrophage Cross-Talk in Obesity.

Obesity is characterized by the chronic low-grade activation of the innate immune system. In this respect, macrophage-elicited metabolic inflammation and adipocyte-macrophage interaction has a primary importance in obesity. Large amounts of macrophages are accumulated by different mechanisms in obese adipose tissue. Hypertrophic adipocyte-derived chemotactic monocyte chemoattractant protein-1 (MCP-1)/C-C chemokine receptor 2 (CCR2) pathway also promotes more macrophage accumulation into the obese adipose tissue. However, increased local extracellular lipid concentrations is a final mechanism for adipose tissue macrophage accumulation. A paracrine loop involving free fatty acids and tumor necrosis factor-alpha (TNF-alpha) between adipocytes and macrophages establishes a vicious cycle that aggravates inflammatory changes in the adipose tissue. Adipocyte-specific caspase-1 and production of interleukin-1beta (IL-1beta) by macrophages; both adipocyte and macrophage induction by toll like receptor-4 (TLR4) through nuclear factor-kappaB (NF-kappaB) activation; free fatty acid-induced and TLR-mediated activation of c-Jun N-terminal kinase (JNK)-related pro-inflammatory pathways in CD11c+ immune cells; are effective in macrophage accumulation and in the development of adipose tissue inflammation. Old adipocytes are removed by macrophages through trogocytosis or sending an "eat me" signal. The obesity-induced changes in adipose tissue macrophage numbers are mainly due to increases in the triple-positive CD11b+ F4/80+ CD11c+ adipose tissue macrophage subpopulation. The ratio of M1-to-M2 macrophages is increased in obesity. Furthermore, hypoxia along with higher concentrations of free fatty acids exacerbates macrophage-mediated inflammation in obesity. The metabolic status of adipocytes is a major determinant of macrophage inflammatory output. Macrophage/adipocyte fatty-acid-binding proteins act at the interface of metabolic and inflammatory pathways. Both macrophages and adipocytes are the sites for active lipid metabolism and signaling.