Reduced Adipose Tissue Inflammation Research Articles

Therapeutic targets to reduce the contribution of pulmonary neutrophilic inflammation towards obesity-associated co-morbidities: a mini-review.

Epidemiology and experimental models have shown a close link between adipose tissue inflammation, systemic inflammation and pulmonary neutrophilic inflammation, which predispose obese patients to pulmonary diseases, obesity-associated co-morbidities and cancer. Increased content and activation of neutrophils in the lung microvasculature, resulting from peripheral activation of neutrophils, and increased adhesion of neutrophils to the lung microvasculature are important factors explaining the increased susceptibility of obese patients towards respiratory diseases and loss of insulin sensitivity. Mechanism-based therapies to break this link are urgently needed to reduce pulmonary damage in obesity, due to the growing prevalence of obesity world-wide. Current research suggests that these approaches should be focused on, one or more of the following: reduction of macrophage activation at the adipose tissue, healthy growing of adipose tissue by induction of Nrf-2, inhibition of NF-κB activation, reduction of circulating neutrophil activation, blocking adhesins/selectins, inhibition of neutrophil activation by targeting NADPH oxidase-2 activation, inhibition of myeloperoxidase activity and scavenging of hypochlorous acid. These strategies are expected to reduce adipose tissue inflammation, peripheral inflammation, pulmonary neutrophilic inflammation and obesity-associated co-morbidities.

Impaired insulin signaling in the B10.D2-Hc0 H2d H2-T18c/oSnJ mouse model of complement factor 5 deficiency.

Given the chemoattractant potential of complement factor 5 (C5) and its increased expression in adipose tissue (AT) of obese mice, we determined whether this protein of the innate immune system impacts insulin action. C5 control (C5cont) and spontaneously C5-deficient (C5def, B10.D2-Hc0 H2d H2-T18c/oSnJ) mice were placed on low- and high-fat diets to investigate their inflammatory and metabolic phenotypes. Adenoviral delivery was used to evaluate the effects of exogenous C5 on systemic metabolism. C5def mice gained less weight than controls while fed a high-fat diet, accompanied by reduced AT inflammation, liver mass, and liver triglyceride content. Despite these beneficial metabolic effects, C5def mice demonstrated severe glucose intolerance and systemic insulin resistance, as well as impaired insulin signaling in liver and AT. C5def mice also exhibited decreased expression of insulin receptor (INSR) gene and protein, as well as improper processing of pro-INSR. These changes were not due to the C5 deficiency alone as other C5-deficient models did not recapitulate the INSR processing defect; rather, in addition to the mutation in the C5 gene, whole genome sequencing revealed an intronic 31-bp deletion in the Insr gene in the B10.D2-Hc0 H2d H2-T18c/oSnJ model. Irrespective of the genetic defect, adenoviral delivery of C5 improved insulin sensitivity in both C5cont and C5def mice, indicating an insulin-sensitizing function of C5.

Siah2 modulates sex-dependent metabolic and inflammatory responses in adipose tissue to a high-fat diet challenge

BackgroundThe obesity-related risk of developing metabolic syndrome is higher in males than in females of reproductive age, likely due to estrogen-mediated reduced adipose tissue inflammation and fibrosis with hypertrophied adipocytes. Depletion of the ubiquitin ligase Siah2 reduced white adipose tissue inflammation and improved glucose metabolism in obese male mice. Siah2 is a transcriptional target of estrogen, but data is lacking about the effect of Siah2 on adipose tissue of females. We therefore evaluated the impact of Siah2 deficiency on white and brown adipose tissue in females of reproductive age.MethodsBody composition, adipose tissue morphology, brown adipose tissue gene, and protein expression and adipocyte sizing were evaluated in wild-type and Siah2KO female and male mice fed a low-fat or high-fat diet. Glucose and insulin tolerance, fasting glucose, insulin, fatty acids and triglycerides, and gene expression of inflammation markers in perigonadal fat were evaluated in wild-type and Siah2KO female mice. Microarray analysis of brown fat gene expression was carried out in both sexes. Statistical analysis was assessed by unpaired two-tailed t test and repeated measures ANOVA.ResultsSiah2 deficiency improves glucose and insulin tolerance in the presence of hypertrophied white adipocytes in high-fat-fed female mice with percent fat comparable to male mice. While previous studies showed Siah2KO reduces the white adipose tissue inflammatory response in male mice, the response in females is biased toward the upregulation of M2-like markers in white adipose tissue. In contrast, loss of Siah2 leads to increased whitening of brown fat in males, but not in females. This corresponded to increased expression of markers of inflammation (F4/80, Ccl2) and thermogenic genes (Pgc1alpha, Dio2, Ucp-1) and proteins (PGC-1α, UCP-1) in females. Contrary to expectations, increased expression of thermogenic markers in females was coupled with a downregulation of ERalpha and ERRgamma protein levels.ConclusionsThe most striking sex-related effect of Siah2 deficiency is reduced whitening of brown fat in high-fat-fed females. Protection from accumulating unilocular adipocytes in the brown fat corresponds to increased expression of thermogenic genes and proteins in female, but not in male mice. These results raise the possibility that Siah2 contributes to the estrogen-related effects on brown fat function in males and females.

Inhibitory effects of lingonberry (Vaccinium vitis-idaea L.) fruit extract on obesity-induced inflammation in 3T3-L1 adipocytes and RAW 264.7 macrophages

Abstract In this study, the lingonberry fruit extract (LGBE) has been investigated for the ability to suppress the inflammatory response and mitigate oxidative stress in activated 3T3-L1 adipocytes and RAW 264.7 macrophages. The obtained results showed that LGBE significantly reduced IL-6, MCP-1 and IL-1β production, and counteracted the decrease in adiponectin and IL-10 expression in TNF-α-induced adipocytes. Moreover, LGBE exhibited a high anti-inflammatory potential in the macrophage cell culture by down-regulation the expression of pro-inflammatory mediators (IL-6, TNF-α, IL-1β, MCP-1, COX-2, iNOS). Furthermore, LGBE decreased intracellular ROS generation in inflamed adipocytes by enhancing expression of antioxidant defense enzymes (SOD, catalase, GPx) and inhibiting an oxidant enzyme (NADPH oxidase 4), which is one of the primary sources of ROS. In conclusion, we have demonstrated that lingonberry fruit may reduce adipose tissue inflammation and support immune cell homeostasis, and thus can be considered a natural tool for inflammation control.

Contribution of Adipose Tissue Inflammation to the Development of Type 2 Diabetes Mellitus.

The objective of this comprehensive review is to summarize and discuss the available evidence of how adipose tissue inflammation affects insulin sensitivity and glucose tolerance. Low-grade, chronic adipose tissue inflammation is characterized by infiltration of macrophages and other immune cell populations into adipose tissue, and a shift toward more proinflammatory subtypes of leukocytes. The infiltration of proinflammatory cells in adipose tissue is associated with an increased production of key chemokines such as C-C motif chemokine ligand 2, proinflammatory cytokines including tumor necrosis factor α and interleukins 1β and 6 as well as reduced expression of the key insulin-sensitizing adipokine, adiponectin. In both rodent models and humans, adipose tissue inflammation is consistently associated with excess fat mass and insulin resistance. In humans, associations with insulin resistance are stronger and more consistent for inflammation in visceral as opposed to subcutaneous fat. Further, genetic alterations in mouse models of obesity that reduce adipose tissue inflammation are-almost without exception-associated with improved insulin sensitivity. However, a dissociation between adipose tissue inflammation and insulin resistance can be observed in very few rodent models of obesity as well as in humans following bariatric surgery- or low-calorie-diet-induced weight loss, illustrating that the etiology of insulin resistance is multifactorial. Taken together, adipose tissue inflammation is a key factor in the development of insulin resistance and type 2 diabetes in obesity, along with other factors that likely include inflammation and fat accumulation in other metabolically active tissues. © 2019 American Physiological Society. Compr Physiol 9:1-58, 2019.

Tart Cherry Reduces Inflammation in Adipose Tissue of Zucker Fatty Rats and Cultured 3T3-L1 Adipocytes.

Obesity increases adipose tissue inflammation and secretion of pro-inflammatory adipokines, which have systemic effects on the organism’s health status. Our objective was to dissect mechanisms of anti-inflammatory effects of tart cherry (TC) in adipose tissue of Zucker fatty rats, and cultured 3T3-L1 adipocytes. Rats were fed either a control diet, or 4% TC powder diets for eight weeks. Body and epididymal fat pad weights were not significantly different between control and TC groups. However, rats fed the TC diet had significantly reduced adipose tissue inflammation (p < 0.05), as determined by reduced mRNA levels of pro-inflammatory markers including interleukin-6 (IL-6), tumor necrosis factor alpha (TNFα), interleukin-1beta (IL-1β), monocyte chemoattractant protein 1 (MCP-1), inducible nitric oxide synthase (iNOS), and CD-11b, and increased mRNA levels of type-1 arginase (Arg-1) anti-inflammatory marker. Consistent with these in vivo results, TC significantly decreased expression of IL-6 mRNA and protein levels in lipopolysaccharide (LPS) stimulated adipocytes compared to those stimulated with LPS, but no TC. Moreover, both in vivo (rat adipose tissue) and in vitro (3T3-L1 adipocytes), phosphorylation of p65-NF-κB subunit was significantly reduced by TC. Additionally, TC decreased mRNA expression of fatty acid synthase (FASN), and increased expression of peroxisome proliferator-activated receptor alpha (PPARα), master regulator of lipid oxidation, and anti-oxidant markers nuclear factor erythroid-derived 2-related factor (NRFs) in both models. In conclusion, our findings indicate that TC downregulates inflammation in part via the nuclear factor kappa B (NF-κB) pathway in adipose tissue. Thus, TC may serve as a potential intervention to reduce obesity-associated inflammation.

PS02.124: NOVEL APPROACHES AND CONSIDERATIONS FOR IMMUNOTHERAPY IN OBESITY-ASSOCIATED OESOPHAGEAL ADENOCARCINOMA

Abstract Background Immunotherapies are transforming cancer treatment for inoperable and advanced disease. However, the incidence of obesity-associated cancers, including oesophageal adenocarcinoma (OAC) continue to increase. Treatment with immune-based therapies present a unique challenge for immunologists, as they need to enhance anti-tumour immunity without exacerbating pre-existing carcinogenic inflammation. Therefore, we examined the effect of PD-1 blockade in omentum and liver of OAC patients, sites of activated inflammatory T cells, which play a key role in obesity-associated inflammation. In addition, we examined novel ways to reduce the infiltration of inflammatory T cells to these sites in a bid to reduce adipose tissue inflammation. Methods Blood, omentum and liver samples were obtained from consenting OAC patients and treated with anti-PD-1 antibody. CD4+ and CD8+ T cell activation, cytokine expression and cytotoxic potential were assessed by flow cytometry. To identify potential chemokine pathways to alter T cell trafficking to the omentum and liver, chemokine receptor expression was examined, along with levels of secreted chemokines using flow cytometry and ELISAs. Pre-treatment of T cells with chemokine receptor antagonists was performed prior to chemotaxis assays using an in vitro transwell system. Results In addition to OAC tumour, omentum and liver were found to be enriched with substantial populations of PD-1 expressing T cells. Treatment of omental and hepatic T cells with anti-PD-1 did not enhance inflammatory cytokine expression or proliferation, but transiently increased CD107a expression by CD8+ T cells. MIP-1α, MIP-1β and IP-10 mediate T cell trafficking to the omentum and liver in OAC. OAC-derived T cells preferentially migrate to the adipose and liver tissue conditioned media of obese OAC patients and this can be significantly reduced using a MIP-1α receptor antagonist. Conclusion This study provides evidence that anti-PD-1 treatment is unlikely to exacerbate pre-existing T cell-mediated inflammation outside the tumour in obesity-associated cancers. Furthermore, novel CCR1 antagonists may be used to attenuate pathological inflammation in obesity. This dual targeting approach aims to enhance anti-tumour immunity through the use of immune checkpoint inhibitors, and has identified a novel immunotherapeutic approach to reduce obesity-associated inflammation by targeting key chemokine pathways. In vivo testing is required to determine the efficacy of this approach. Disclosure All authors have declared no conflicts of interest.

Effects of Bariatric Surgery on FGF-19 and FGF-21 Levels on Obese Diabetic Women

Background: Fibroblast growth factor 19 (FGF-19) and FGF-21 regulate glucose tolerance and insulin sensitivity, which in type 2 diabetes mellitus (T2DM) conditions becomes dysregulated. Whilst weight loss can restore this functionality, bariatric surgery can have a specific impact due to mechanical changes on gut/liver physiology. We aimed to analyse the effects of different bariatric surgeries: sleeve gastrectomy (SG), laparoscopic adjusted gastric banding (LAGB) and biliopancreatic diversion (BPD) on changes in FGF-19 and FGF-21 levels. Methods: Obese and T2DM women (n=41 BMI&amp;gt;35 Kg/m2) undergoing SG (n=14), LABG (n=13), and BPD (n=14) participated in an ethics approved study. Anthropometry, biochemical data, and adipose tissue (AT) biopsies were collected before and 6 months after surgery. FGF-19, FGF-21 levels and AT inflammation (TLR2, MYD88, TRAF6, IKK and NFkb) was assessed. Results: Post surgery SG led to 63% reduction in FGF-21 whilst LABG and BPD led to 41% and 4% decrease respectively. There was a significant differences in FGF-21 levels between SG and BPD (p=0.021). Moreover, FGF-21 change in SG positivly correlated with changes in glucose (p=0.019), insulin (p=0.047), HOMA-IR (p=0.022) and triglyceride (p=0.027); absent in BPD and LABG. FGF-21 levels in SG appeared as a significant predictor of HOMA-IR change post surgery (p&amp;lt;0.05). FGF-19 levels post surgery increased by 31% in SG compared to 10% rise in BPD and 39% decrease in LAGB with significant differences between SG and LAGB (p=0.012). FGF-19 change in BPD inversly correlated with changes in weight (p=0.012), BMI (p=0.037), waist circumfrance (p=0.044); absent in SG and LAGB. Inflamatory markers post surgery were significantly reduced in all surgeries (TLR2: p&amp;lt;0.01, MYD88: p&amp;lt;0.01, TRAF6: p&amp;lt;0.01, IKK: p&amp;lt;0.01 and NFkb: p&amp;lt;0.01). Conclusion: SG appears to have a beneficial influence on FGF-21 and FGF-19 levels irrespective of reduced AT inflammation. Our FGF findings therfore suggests that SG provides better glucose homeostatis outcomes than LAGB and BPD. Disclosure S. Azharian: None. A. Murphy: None. P.G. McTernan: None. M.K. Piya: None. G. Tripathi: None.

The Effect of Saxagliptin on Adipose Tissue Inflammation in Overweight and Obese Nondiabetic Subjects

Inflammation plays a crucial role in early stages of both insulin resistance and atherosclerosis development. Preclinical evidence suggests an important role for DPP-4 in the regulation of adipose tissue (AT) inflammation. The present study tests the hypothesis that DPP-4 inhibition by saxagliptin reduces adipose tissue inflammation independent of changes in glucose metabolism. Forty-four overweight or obese individuals without diabetes were randomized to receive once daily saxagliptin (5 mg) or matching placebo (2:1 randomization) for 6 weeks. Adipose tissue biopsies, 4 hour fat-enriched meal challenge tests, and endothelial function measurement were performed at baseline and at the end of each treatment. Forty-two participants completed the study (27 saxagliptin/15 placebo). There were no group differences in the AT expression of inflammation-related genes (MCP-1, CD36, CD68, IL-6, IL-8, TxNIP and adiponectin) and protein pathway activation (ERK, JNK and NFκB). Secretion of IL-8 from adipose tissue explants was reduced after saxagliptin compared to placebo (median fold-baseline 0.8 saxagliptin vs. 2.3 placebo, p=0.01) and there was also a trend for reduced IL-6 release (0.7 vs. 1.5, p=0.06). After the high-fat meal, plasma glucose was slightly lower after saxagliptin (4 hour AUC, 0.96 vs. 1.0, p=0.04) while insulin and triglycerides were similar between the two groups. There were no treatment differences in endothelial function. In conclusion, reduced AT secretion of IL-8 and IL-6 (trend) in obese nondiabetic subjects suggests saxagliptin may directly reduce AT inflammation. However, this effect was relatively modest, with many other indicators of tissue inflammation remaining unchanged. The reduction in AT inflammation did not appear sufficiently potent to influence postprandial metabolism or endothelial function. Disclosure J. Koska: None. T. Osredkar: None. K. D'Souza: None. S. Sinha: None. C. Meyer: Employee; Self; Merck &amp; Co., Inc. P. Reaven: Research Support; Self; AstraZeneca, Amgen Inc., Bristol-Myers Squibb Company.

Insulin Sensitizing Effects of Vitamin D Mediated through Reduced Adipose Tissue Inflammation and Fibrosis

Adipose tissue inflammation and fibrosis appear to be mediated by adipocytes and contribute to insulin resistance. Since vitamin D (25(OH)D) has anti-inflammatory and anti-fibrotic effects, expression of its receptor in adipocytes and macrophages suggests that 25(OH)D signaling could mediate paracrine effects within adipose tissue and improve insulin resistance. We performed a randomized, double-blinded placebo-controlled trial to examine the effects of repleting vitamin D in 25(OH)D-deficient (&amp;lt;20 ng/ml), insulin resistant, overweight-to-obese humans (n=19). 25(OH)D repletion to &amp;gt;30 ng/ml was associated with reductions in adipose tissue expression of inflammatory (0.6-0.7-fold decreased expression of TNF-α, IL-6, iNOS and PAI-1) and pro-fibrotic (0.4-0.8-fold decreased expression of TGF-β1, HiF1α, Collagen I, V, VI and MMP7) genes, decreased collagen VI immunofluorescence (19% reduction, p=0.02) and improved hepatic insulin sensitivity, assessed as suppression of endogenous glucose production (EGP) during hyperinsulinemic clamp studies (1.28 ± 0.20 vs. 0.88 ± 0.18 mg/kg/min, p=0.03). To determine whether vitamin D’s effects are mediated through adipocytes, we studied an adipocyte-specific vitamin D receptor knockout mouse model (Adiponectin-Cre+VDR+/ fl) after 12 weeks on high fat diet. Despite no differences in body weight or adiposity, VDR KO mice exhibited increased adipose tissue expression of several pro-inflammatory (Tnf-α, iNos, Pai-1, Mcp-1 and F4/80; 4-10 fold) and pro-fibrotic genes (Tgf-β1, Collagen VI, and Tsp1; 2-4 fold), in concert with hepatic insulin resistance during 2h hyperinsulinemic (4 mU/kg/min)-euglycemic clamps (EGP 10 ± 3 vs. 3 ± 2 mg/kg/min in WT, p = 0.021). These human and rodent studies establish a beneficial role of vitamin D in restraining adipose tissue inflammation and fibrosis as well as hepatic insulin resistance, and suggest that normalizing 25(OH)D levels could have metabolic benefits in targeted individuals. Disclosure E. Lontchi-Yimagou: None. S. Kang: None. K. Zhang: None. A. Goyal: None. J. You: None. P. Kishore: None. E. Rosen: None. M. Hawkins: None.

Siah2 Expression in Adipocyte Progenitor Cells

Obesity-related insulin resistance is associated with adipose tissue inflammation and visceral adipose tissue expansion via adipocyte hypertrophy and impaired recruitment of new progenitor cells to undergo adipogenesis (hyperplasia). Deletion of the ubiquitin ligase Seven-in-absentia homolog 2 (Siah2) in obese mice significantly reduced adipose tissue inflammation and insulin resistance compared to obese wild type mice. However, larger adipocyte size accompanied this phenotype and subsequent experiments with adipose tissue stromal vascular fraction (SVF) cells showed adipogenesis required Siah2. In an earlier study, we found Siah2 affects adipogenesis by enhancing gene expression of the PPARγ coactivator, Zfp423 via proteasome-dependent degradation of ZFP521, a transcriptional repressor of Zfp423. ZFP423 is an early marker of preadipocyte determination that is expressed in adipocyte progenitor cells. Although Siah2 mediates adipogenesis and is highly expressed in the adipose tissue stromal vascular cell fraction, cell-specific expression of Siah2 in adipose tissue is not well characterized. We found that Siah2 is highly expressed in macrophage. RNA-seq and microarray data from the Immunological Genome Project (Immgen.org) also reported Siah2 expression in lymphocytes. To identify Siah2 expression profile in adipose tissue, unsorted cells and magnetically sorted macrophage, T-cell, B-cell, and SCA-1 positive progenitor cells from mouse adipose tissues SVF were analyzed for Siah2 expression. In parallel, in situ hybridization RNAscope of fat pads was employed to inspect Siah2 mRNA localization in adipose tissue. Since our evidence indicates Siah2 acts upstream of ZFP423, we predict Siah2 is expressed in adipocyte progenitor cells as well as immune cells in adipose tissue. This work will provide the groundwork to better understand how Siah2 controls commitment of progenitor cells to the adipocyte lineage and links obesity to adipose tissue inflammation and insulin resistance. Disclosure T. Dang: None. G.E. Kilroy: None. J.L. Taylor: None. Y. Yu: None. E. Floyd: None.

P-174 - Genetic deletion of RAGE in Dbdb mice interferes with AGEs receptors and detoxifying systems in liver

Advanced glycation end products (AGEs) are toxic compounds involved in the development of diabetes complications and the onset of insulin resistance in obesity. In particular, AGEs are preferentially trapped by adipose tissue through the binding with the AGE-receptor RAGE, leading to the activation of proinflammatory signaling and oxidative stress in adipocytes that can interfere with peripheral insulin sensitivity. The genetically-induced deletion of RAGE in leptin receptor deleted (DbDb) mice, a model of diet-induced diabetes/obesity, is reported to increase circulating AGEs levels and reduce adipose tissue inflammation and insulin resistance. However, we here report that the deletion of RAGE in DbDb mice, in contrast to other tissues, impairs hepatic AGE-receptor-1 and glyoxalase-1, two major AGEs detoxifying systems, and increases galectin-3 expression, another AGEs-receptor, as a compensatory response to remove plasma AGEs that might sustain hepatosteatosis and inflammation. These results thus indicate a complex tissue specific control mechanism among AGEs receptors and detoxifying systems that need to be better elucidated to identify molecular targets and therapeutic tools to modulate the pathogenic contribution of AGEs to hepatic metabolic disturbances.

Inducible heterozygous knockout of Pik3r1 in adipocytes reverses glucose intolerance and enhances adipocyte insulin signaling in obese mice

Insulin resistance is a central and defining feature of the metabolic diseases associated with obesity. Phosphatidylinositol 3‐kinase (PI3K), a heterodimeric enzyme composed of a regulatory subunit (p85α/β, p55α/γ, p50α) and a catalytic subunit (p110α/β), is an essential component of the conical insulin signaling cascade in adipose tissue. PI3K activity is decreased in obese adipose tissue (AT); however, a global heterozygous (HZ) deletion of Pik3r1 (encodes p85/p55/050α) prevents the decrease in insulin‐stimulated PI3K activity, and reduces AT inflammation and systemic insulin resistance relative to obese control mice. Therefore, we hypothesized that adipocyte‐specific deletion of Pik3r1 in already obese mice would reverse obesity‐induced systemic insulin resistance by increasing insulin signaling downstream of PI3K in adipocytes. Inducible adipocyte‐specific Pik3r1 heterozygous (HZ) or homozygous (KO) knockout mice were generated by crossing Pik3r1 flox mice (LoxP sites flanking exon7 of the Pik3r1 gene) with Adipoq‐CreERT2 mice, which utilize tamoxifen‐dependent recombination. Pik3r1 flox/flox (FL/FL) mice were used as controls. At 8 weeks of age, mice were fed a high‐fat diet (HFD; 45% fat) or sucrose‐matched control diet (10% fat) for 10 weeks. At week 11, tamoxifen was delivered by oral gavage (100μL of 10 mg/mL) for 5 days and allowed to recover for 1 week) glucose tolerance was measured before and 1–2 weeks post tamoxifen. Total activity, energy expenditure and respiratory exchange ratio (RER) were collected for 3 days following one‐week acclimation and recovery. Isolated adipocytes from lean and obese mice were stimulated +/− 1.2nM insulin and signaling proteins were measured by immunoassay. There were differences in body weight, body composition before or after gene knockout with tamoxifen in HFD vs. CON, for WT, HZ and KO. Total activity (WT: 90±6; HZ: 115±12; KO: 106±10), energy expenditure (WT: 143±11; HZ: 122±12; KO: 134±12), and RER (WT: 0.73±0.001; HZ: 0.775±0.03; KO: 0.81±0.024) were not different for WT, HZ, and KO during HFD. Glucose AUC calculated from oral glucose tolerance test was significantly higher in all groups with HFD vs. CON. We find that after tamoxifen dosing, glucose (g) AUC was reduced by 59% in obese HZ mice as compared to pre‐tamoxifen levels. No changes in gAUC were found in obese WT or KO. Insulin‐stimulated AKT(S473) phosphorylation was decreased by 41% and 63% in adipocytes from obese WT and KO, respectively, compared to lean; however, only 17% in obese HZ. Taken together, these data show that adipocyte Pik3r1 plays a critical role in mediating systemic glucose metabolism and that novel treatments that target Pik3r1expression may be successful in improving insulin sensitivity in individuals with obesity.Support or Funding InformationSupported by Eugene &amp; Clarissa Evonuk Memorial Fellowship, University of Oregon Graduate Student Food Studies Grant (Z.S.C) and National Institute of Diabetes Digestive and Kidney Diseases under Award Number RO1DK095926 (C.E.M)This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Regular exercise reduces adipose tissue inflammation and improves glycemic control in Western diet‐fed mice despite hyperendothelinemia

The upregulation of endothelin‐1 (ET‐1), a peptide primarily expressed and secreted into the circulation from endothelial cells, is a key molecular signature in obesity and type 2 diabetes. Given its potent pro‐oxidant and inflammatory effects, ET‐1 is thought to be implicated in the etiology of metabolic dysfunction. Based on previous evidence that regular exercise decreases ET‐1 expression and signaling, it is conceivable that the well‐established exercise‐induced anti‐inflammatory effects and associated improvement in glycemic control is, in part, mediated by a reduction in ET‐1 actions; however, this remains unknown. Accordingly, we tested the hypothesis that overproduction of ET‐1 from endothelial cells and resulting hyperendothelinemia limits the beneficial metabolic adaptations of exercise. Six‐week‐old male transgenic mice overexpressing human preproET‐1 selectively in the endothelium (eET‐1) and non‐transgenic (i.e., wild‐type) littermates were fed a Western diet (46% Kcal fat, 36% Kcal carbohydrate) to induce obesity, while they either remained sedentary or had free access to a running wheel for 12 weeks, creating four groups (n=7–12/group): wild‐type sedentary, wild‐type exercise, eET‐1 sedentary, eET‐1 exercise. Plasma ET‐1 was significantly elevated in eET‐1 mice (sedentary: 15.12±3.77 pg/ml; exercise 14.90±2.30 pg/ml) versus wild‐type (sedentary: 0.02±0.02 pg/ml; exercise: 0.29±0.14 pg/ml) in both sedentary and exercise‐trained mice. Exercise reduced final body weight, adiposity, fasting plasma insulin, HOMA‐IR, cholesterol and triglycerides compared to sedentary mice, and these effects were independent of genotype. Also independent of genotype, exercise reduced visceral adipocyte hypertrophy, immune cell infiltration, inflammatory gene expression (e.g., IL‐1β and CD11c), as well as enhanced visceral adipose tissue insulin signaling (i.e., increased ex vivo insulin‐stimulated pAkt/Akt). Furthermore, exercise improved glucose tolerance and this improvement was mildly enhanced in eET‐1 mice (i.e., significant genotype × exercise interaction). In summary, contrary to our hypothesis, we find that experimentally‐induced hyperendothelinemia does not impede exercise‐induced improvements in metabolic function, supporting the notion that reduced ET‐1 signaling is not required for the beneficial metabolic adaptations of exercise to manifest.Support or Funding InformationNational Institute of HealthThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Toll-like receptor 4 (TLR4) deficient mice are protected from adipose tissue inflammation in aging.

Adipose tissue (AT) inflammation is a central mechanism for metabolic dysfunction in both diet-induced obesity and age-associated obesity. Studies in diet-induced obesity have characterized the role of Fetuin A (Fet A) in Free Fatty Acids (FFA)-mediated TLR4 activation and adipose tissue inflammation. However, the role of Fet A & TLR4 in aging-related adipose tissue inflammation is unknown. In the current study, analysis of epidymymal fat pads of C57/Bl6 male mice, we found that, in contrast to data from diet-induced obesity models, adipose tissue from aged mice have normal Fet A and TLR4 expression. Interestingly, aged TLR4-deficient mice have diminished adipose tissue inflammation compared to normal controls. We further demonstrated that reduced AT inflammation in old TLR4-deficient mice is linked to impaired ER stress, augmented autophagy activity, and diminished senescence phenomenon. Importantly, old TLR4-deficient mice have improved glucose tolerance compared to age-matched wild type mice, suggesting that the observed reduced AT inflammation in aged TLR4-deficient mice has important physiological consequences. Taken together, our present study establishes novel aspect of aging-associated AT inflammation that is distinct from diet-induced AT inflammation. Our results also provide strong evidence that TLR4 plays a significant role in promoting aging adipose tissue inflammation.

DOCK2 deficiency mitigates HFD-induced obesity by reducing adipose tissue inflammation and increasing energy expenditure

Obesity is the major risk factor for type 2 diabetes, cardiovascular disorders, and many other diseases. Adipose tissue inflammation is frequently associated with obesity and contributes to the morbidity and mortality. Dedicator of cytokinesis 2 (DOCK2) is involved in several inflammatory diseases, but its role in obesity remains unknown. To explore the function of DOCK2 in obesity and insulin resistance, WT and DOCK2-deficient (DOCK2-/-) mice were given chow or high-fat diet (HFD) for 12 weeks followed by metabolic, biochemical, and histologic analyses. DOCK2 was robustly induced in adipose tissues of WT mice given HFD. DOCK2-/- mice with HFD showed decreased body weight gain and improved metabolic homeostasis and insulin resistance compared with WT mice. DOCK2 deficiency also attenuated adipose tissue and systemic inflammation accompanied by reduced macrophage infiltration. Moreover, DOCK2-/- mice exhibited increased expression of metabolic genes in adipose tissues with greater energy expenditure. Mechanistically, DOCK2 appeared to regulate brown adipocyte differentiation because increased preadipocyte differentiation to brown adipocytes in interscapular and inguinal fat was observed in DOCK2-/- mice, as compared with WT. These data indicated that DOCK2 deficiency protects mice from HFD-induced obesity, at least in part, by stimulating brown adipocyte differentiation. Therefore, targeting DOCK2 may be a potential therapeutic strategy for treating obesity-associated diseases.

IgG is elevated in obese white adipose tissue but does not induce glucose intolerance via Fcγ-receptor or complement.

In obesity, B cells accumulate in white adipose tissue (WAT) and produce IgG, which may contribute to the development of glucose intolerance. IgG signals by binding to Fcγ receptors (FcγR) and by activating the complement system. The aim of our study was to investigate whether activation of FcγR and/or complement C3 mediates the development of high-fat diet-induced glucose intolerance. We studied mice lacking all four FcγRs (FcγRI/II/III/IV-/-), only the inhibitory FcγRIIb (FcγRIIb-/-), only the central component of the complement system C3 (C3-/-), and mice lacking both FcγRs and C3 (FcγRI/II/III/IV/C3-/-). All mouse models and wild-type controls were fed a high-fat diet (HFD) for 15 weeks to induce obesity. Glucose metabolism was assessed and adipose tissue was characterized for inflammation and adipocyte functionality. In obese WAT of wild-type mice, B cells (+142%, P<0.01) and IgG (+128% P<0.01) were increased compared to lean WAT. Macrophages of FcγRI/II/III/IV-/-mice released lower levels of cytokines compared to wild-type mice upon IgG stimulation. Only C3-/- mice showed reduced HFD-induced weight gain as compared to controls (-18%, P<0.01). Surprisingly, FcγRI/II/III/IV-/- mice had deteriorated glucose tolerance (AUC +125%, P<0.001) despite reduced leukocyte number (-30%, P<0.05) in gonadal WAT (gWAT), whereas glucose tolerance and leukocytes within gWAT in the other models were unaffected compared to controls. Although IgG in gWAT was increased (+44 to +174%, P<0.05) in all mouse models lacking FcγRIIb, only FcγRI/II/III/IV/C3-/- mice exhibited appreciable alterations in immune cells in gWAT, for example, increased macrophages (+36%, P<0.001). Lack of FcγRs reduces the activity of macrophages upon IgG stimulation, but neither FcγR nor C3 deficiency protects against HFD-induced glucose intolerance or reduces adipose tissue inflammation. This indicates that if obesity-induced IgG contributes to the development of glucose intolerance, this is not mediated by FcγR or complement activation.

BMP4 Gene Therapy in Mature Mice Reduces BAT Activation but Protects from Obesity by Browning Subcutaneous Adipose Tissue

We examined the effect of Bone Morphogenetic Protein 4 (BMP4) on energy expenditure in adult mature mice by targeting the liver with adeno-associated viral (AAV) BMP4 vectors to increase circulating levels. We verified the direct effect of BMP4 in inducing a brown oxidative phenotype in differentiating preadipocytes invitro. AAV-BMP4-treated mice display marked browning of subcutaneous adipocytes, with increased mitochondria and Uncoupling Protein 1 (UCP1). These mice are protected from obesity on a high-fat diet and have increased whole-body energy expenditure, improved insulin sensitivity, reduced liver fat, and reduced adipose tissue inflammation. On a control diet, they show unchanged body weight but improved insulin sensitivity. In contrast, AAV-BMP4-treated mice showed beiging of BAT with reduced UCP1, increased lipids, and reduced hormone-sensitive lipase (HSL). Thus, BMP4 exerts different effects on WAT and BAT, but the overall effect is to enhance insulin sensitivity and whole-body energy expenditure by browning subcutaneous adipose tissue.

Anti-inflammatory effects of exercise training in adipose tissue do not require FGF21.

Exercise enhances insulin sensitivity; it also improves adipocyte metabolism and reduces adipose tissue inflammation through poorly defined mechanisms. Fibroblast growth factor 21 (FGF21) is a pleiotropic hormone-like protein whose insulin-sensitizing properties are predominantly mediated via receptor signaling in adipose tissue (AT). Recently, FGF21 has also been demonstrated to have anti-inflammatory properties. Meanwhile, an association between exercise and increased circulating FGF21 levels has been reported in some, but not all studies. Thus, the role that FGF21 plays in mediating the positive metabolic effects of exercise in AT are unclear. In this study, FGF21-knockout (KO) mice were used to directly assess the role of FGF21 in mediating the metabolic and anti-inflammatory effects of exercise on white AT (WAT) and brown AT (BAT). Male FGF21KO and wild-type mice were provided running wheels or remained sedentary for 8 weeks (n = 9-15/group) and compared for adiposity, insulin sensitivity (i.e., HOMA-IR, Adipo-IR) and AT inflammation and metabolic function (e.g., mitochondrial enzyme activity, subunit content). Adiposity and Adipo-IR were increased in FGF21KO mice and decreased by EX. The BAT of FGF21KO animals had reduced mitochondrial content and decreased relative mass, both normalized by EX. WAT and BAT inflammation was elevated in FGF21KO mice, reduced in both genotypes by EX. EX increased WAT Pgc1alpha gene expression, citrate synthase activity, COX I content and total AMPK content in WT but not FGF21KO mice. Collectively, these findings reveal a previously unappreciated anti-inflammatory role for FGF21 in WAT and BAT, but do not support that FGF21 is necessary for EX-mediated anti-inflammatory effects.

Stromal cell cadherin-11 regulates adipose tissue inflammation and diabetes.

M2 macrophages, innate lymphoid type 2 cells (ILC2s), eosinophils, Tregs, and invariant NK T cells (iNKT cells) all help to control adipose tissue inflammation, while M1 macrophages, TNF, and other inflammatory cytokines drive inflammation and insulin resistance in obesity. Stromal cells regulate leukocyte responses in lymph nodes, but the role of stromal cells in adipose tissue inflammation is unknown. PDGFRα+ stromal cells are major producers of IL-33 in adipose tissue. Here, we show that mesenchymal cadherin-11 modulates stromal fibroblast function. Cadherin-11-deficient mice displayed increased stromal production of IL-33, with concomitant enhancements in ILC2s and M2 macrophages that helped control adipose tissue inflammation. Higher expression levels of IL-33 in cadherin-11-deficient mice mediated ILC2 activation, resulting in higher IL-13 expression levels and M2 macrophage expansion in adipose tissue. Consistent with reduced adipose tissue inflammation, cadherin-11-deficient mice were protected from obesity-induced glucose intolerance and adipose tissue fibrosis. Importantly, anti-cadherin-11 mAb blockade similarly improved inflammation and glycemic control in obese WT mice. These results suggest that stromal fibroblasts expressing cadherin-11 regulate adipose tissue inflammation and thus highlight cadherin-11 as a potential therapeutic target for the management of obesity.