DIO Mice Research Articles

Targeted Retrograde Chemogenetic Approach to Treat Sleep Apnea

IntroductionObstructive Sleep Apnea (OSA) is a prevalent condition and a major cause of morbidity and mortality. The tongue and specifically genioglossus (GG) muscle have been implicated in the pathogenesis of upper airway (UA) obstruction during sleep. Nasal positive airway pressure can treat OSA but poor adherence severely limits its effectiveness. There is an urgent need for pharmacotherapy that reverses neuromuscular defects in upper airway function. We have previously shown that diet induced obese C57BL/6J (DIO) mice have OSA, similar to humans. We have also recently demonstrated that Designer Receptors Exclusively Activated by Designer Drugs (DREADDs) deployed by intracerebral injection can be expressed in hypoglossal motor neurons and that activation of these DREADDs by specific ligand increased GG activity and improved pharyngeal patency. The goal of our novel approach is to avoid intracranial injection and improve specificity of DREADD delivery to hypoglossal motor neurons innervating genioglossus muscle. We hypothesize that tongue injecting retrograde adeno‐associated virus type 9 (AAV9) carrying excitatory DREADDs will (1) deploy DREADDs in hypoglossal motor neurons; (2) improve UA patency and (3) and treat sleep apnea in DIO mice.MethodsDREADDs (AAV9‐hSyn‐hM3(Gq)‐mCherry n=10) or Control AAV9‐hSyn‐GFP (n=5) were delivered bilaterally by tongue injection in DIO mice. Six weeks after gene delivery, mice were injected with specific ligand (J60) and saline intraperitoneally (IP) one week apart in a random order: we performed GG electromyography (EMGGG), UA dynamic MRI, positron emission tomography (PET) scan and sleep studies.ResultsDREADDs or control virus were expressed in the tongue and hypoglossal nucleus in all mice. In DREADD‐transfected mice, administration of J60 resulted in (1) 7.05 fold increase in EMGGG activity (p<0.001); (2) pharyngeal dilation at the rim of the soft palate (0.46 ±0.03mm2 to 1.16±0.05mm2, p<0.05) by MRI; (3) increased tongue metabolic activity (2.21± 0.28 to 4.14 ± 0.31 g/ml, p<0.001) by PET scan; (4) increased minute ventilation (22.53 ± 10.39 to 60.18 ± 33.28 mL/min) and maximal inspiratory flow (1.58 ± 0.58 to 4.11 ± 1.98 mL/sec) during sleep, treating sleep apnea. No changes were seen following j60 injection in mice injected with control virus or saline injection.ConclusionTongue injection of retrograde AAV9 carrying DREADDs successfully deployed DREADDs in hypoglossal motor neurons. Activation of these DREADDs by specific ligand (J60) treated OSA.Support or Funding InformationRO1HL138932, AHA 19CDA34660245, ATS Foundation Unrestricted Grant(A) Whole body PET scan in mouse treated with retrograde excitatory DREADD after Saline (left) or J60 (right). Note the increase in tongue metabolic activity marked by the intense uptake after J60 (right). (B) Plotted Standardized Uptake Values (SUVs) in these animals. *, p < 0.001.Figure 1(A) Representative genioglossal electromyography (EMGGG), recorded at baseline (left) and after J60 administration (right). Note the increase in both phasic and tonic EMG activity. (B) Plotted EMG responses to J60 or saline in the same DREADD treated animals. *, p < 0.001.Figure 2

Alliin, an Allium sativum Nutraceutical, ReducesMetaflammation Markers in DIO Mice.

Obesity generates a chronic low-grade inflammatory state which promotes oxidative stress and triggers comorbidities. Alliin is the main organosulfur compound in garlic and has been shown to induce a decrease in the expression of proinflammatory cytokines; its systemic effect on metabolic parameters and adipose tissue is not yet known, however. After nine weeks of HFD and with obesity established in C57BL/6 mice, we observed that a daily treatment with alliin for 3.5 weeks (15 mg/kg) did not affect body weight, but significantly improved insulin sensitivity and glucose tolerance, both evaluated through a blood glucose monitoring system. Once alliin treatment was completed, serum, adipose tissue, and organs of interest related to metabolism were removed for further analysis. We observed that alliin significantly decreased the size of adipocytes from epididymal adipose tissue, evaluated via microscopy. A decrease in gene expression and serum protein levels of the adipocytokines leptin and resistin, as well as decreased serum IL-6 concentration, were detected by qRT-PCR and ELISA, respectively. It did not, however, affect mRNA expression of antioxidant enzymes in the liver. Taken altogether, these results indicate that treatment with alliin reduces metaflammation markers in DIO mice and improves some metabolic parameters without affecting others.

NADPH Oxidase 2 Mediates Myocardial Oxygen Wasting in Obesity.

Obesity and diabetes are independent risk factors for cardiovascular diseases, and they are associated with the development of a specific cardiomyopathy with elevated myocardial oxygen consumption (MVO2) and impaired cardiac efficiency. Although the pathophysiology of this cardiomyopathy is multifactorial and complex, reactive oxygen species (ROS) may play an important role. One of the major ROS-generating enzymes in the cardiomyocytes is nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 2 (NOX2), and many potential systemic activators of NOX2 are elevated in obesity and diabetes. We hypothesized that NOX2 activity would influence cardiac energetics and/or the progression of ventricular dysfunction following obesity. Myocardial ROS content and mechanoenergetics were measured in the hearts from diet-induced-obese wild type (DIOWT) and global NOK2 knock-out mice (DIOKO) and in diet-induced obese C57BL/6J mice given normal water (DIO) or water supplemented with the NOX2-inhibitor apocynin (DIOAPO). Mitochondrial function and ROS production were also assessed in DIO and DIOAPO mice. This study demonstrated that ablation and pharmacological inhibition of NOX2 both improved mechanical efficiency and reduced MVO2 for non-mechanical cardiac work. Mitochondrial ROS production was also reduced following NOX2 inhibition, while cardiac mitochondrial function was not markedly altered by apocynin-treatment. Therefore, these results indicate a link between obesity-induced myocardial oxygen wasting, NOX2 activation, and mitochondrial ROS.

Hydrocarboxylic acid receptor 1 in BAT regulates glucose uptake in mice fed a high-fat diet.

Interscapular brown adipose tissue (BAT) has the capability to take up glucose from the circulation. Despite the important role of BAT in the control of glucose homeostasis, the metabolic fate and function of glucose in BAT remain elusive as there is clear dissociation between glucose uptake and BAT thermogenesis. Interestingly, intracellular glycolysis and lactate production appear to be required for glucose uptake by BAT. Here, we specifically examine whether activation of lactate receptors in BAT plays a key role in regulating glucose homeostasis in mice fed a high-fat diet (HFD). When C57BL/6J mice are given HFD for 5 weeks at 28°C, male, but not female, mice gain body weight and develop hyperglycemia. Importantly, high-fat feeding upregulates expression of the lactate receptor hydroxycarboxylic acid receptor 1 (HCAR1) in female C57BL/6J mice, whereas male C57BL/6J mice show reduced HCAR1 expression in BAT. Treatment with the HCAR1 agonist lowers systemic glucose levels in male DIO mice. This reduction is associated with increased glucose uptake in BAT. Therefore, our results suggest that HCAR1 in BAT may contribute to the development of hyperglycemia in male C57BL/6J DIO mice.

Ursolic acid induces the production of IL6 and chemokines in both adipocytes and adipose tissue

ABSTRACT Adipose tissue inflammation plays an important role in the regulation of glucose and lipids metabolism. It is unknown whether Ursolic acid (UA) could regulate adipose tissue inflammation, though it can regulate inflammation in many other tissues. In this study, 3T3-L1 adipocytes, DIO mice and lean mice were treated with UA or vehicle. Gene expression of inflammatory factors, chemokines and immune markers in adipocytes and adipose tissue, cytokines in cell culture medium and serum, and inflammation regulatory pathways in adipocytes were detected. Results showed that UA increased the expression of interleukins and chemokines, but not TNFα, in both adipocytes and adipose tissue. IL6 and MCP1 levels in the cell culture medium and mouse serum were induced by UA treatment. Cd14 expression level and number of CD14+ monocytes were higher in UA treated adipose tissue than those in the control group. Glucose tolerance test was impaired by UA treatment in DIO mice. Mechanistically, UA induced the expression of Tlr4 and the phosphorylation levels of ERK and NFκB in adipocytes. In conclusion, our study indicated that short-term UA administration could induce CD14+ monocytes infiltration by increasing the production of interleukins and chemokines in mouse adipose tissue, which might further impair glucose tolerance test.

Hypotriglyceridemic effects of brown seaweed consumption via regulation of bile acid excretion and hepatic lipogenesis in high fat diet-induced obese mice.

BACKGROUND/OBJECTIVESThe present study aimed to further investigate the potential health beneficial effects of long-term seaweed supplementation on lipid metabolism and hepatic functions in DIO mice.MATERIALS/METHODSFour brown seaweeds (Undaria pinnatifida [UP], Laminaria japonica [LJ], Sargassum fulvellum [SF], or Hizikia fusiforme [HF]) were added to a high fat diet (HFD) at a 5% ratio and supplemented to C57BL/6N mice for 16 weeks. Triglycerides (TGs) and total cholesterol (TC) in the liver, feces, and plasma were measured. Fecal bile acid (BA) levels in feces were monitored. Hepatic insulin signaling- and lipogenesis-related proteins were evaluated by Western blot analysis.RESULTSFasting blood glucose levels were significantly reduced in the LJ, SF, and HF groups compared to the HFD group by the end of 16-week feeding period. Plasma TG levels and hepatic lipid accumulation were significantly reduced in all 4 seaweed supplemented groups, whereas plasma TC levels were only suppressed in the UP and HF groups compared to the HFD group. Fecal BA levels were significantly elevated by UP, LJ, and SF supplementation compared to HFD feeding only. Lastly, regarding hepatic insulin signaling-related proteins, phosphorylation of 5′-AMP-activated protein kinase was significantly up-regulated by all 4 types of seaweed, whereas phosphorylation of protein kinase B was up-regulated only in the SF and HF groups. Lipogenesis-related proteins in the liver were effectively down-regulated by HF supplementation in DIO mice.CONCLUSIONSBrown seaweed consumption showed hypotriglyceridemic effects in the prolonged DIO mouse model. Specifically, combinatory regulation of BA excretion and lipogenesis-related proteins in the liver by seaweed supplementation contributed to the reduction of plasma and hepatic TG levels, which inhibited hyperglycemia in DIO mice. Thus, the discrepant and species-specific functions of brown seaweeds provide novel insights for the selection of future targets for therapeutic agents.

The Glucose-Dependent Insulinotropic Polypeptide (GIP) Regulates Body Weight and Food Intake Via CNS-GIPR Signaling

Uncertainty exists as to whether the glucose-dependent insulinotropic polypeptide receptor (GIPR) should be activated or inhibited for the treatment of obesity. Gipr was recently demonstrated in hypothalamic feeding centers, but the physiological relevance of CNS Gipr remains unknown. We show that HFD-fed CNS-Gipr ko mice and humanized (h)GIPR knock-in mice with CNS-hGIPR deletion show improved body weight and glycemia, but these metabolic improvements vanish upon adult-onset Gipr deletion. In DIO mice, acute central administration of acyl-GIP increases cFos neuronal activity in the arcuate, dorsomedial, paraventricular and lateral hypothalamus and leads to improved body weight, food intake, and glycemia. Chronic administration of acyl-GIP improves body weight and food intake in wildtype mice, but shows blunted/absent efficacy in CNS-Gipr ko mice. Also, the superior metabolic effect of GLP-1/GIP co-agonism relative to GLP-1 was extinguished in CNS-Gipr ko mice. Our data establish a key role of CNS Gipr for control of energy metabolism.

Inhibition of XBP1s ubiquitination enhances its protein stability and improves glucose homeostasis

BackgroundHepatic ER stress is a risk factor of insulin resistance and type 2 diabetes. X-box binding protein 1 spliced (XBP1s), a transcription factor, plays a key role in ameliorating insulin resistance and maintaining glucose homeostasis. Unfortunately, the short half-life of the protein dampens its clinical application, and the specific site of lysine residue that could be ubiquitinated and involved in the degradation of XBP1s remains elusive. Methods and resultsHere, we identified K60 and K77 on XBP1s as two pivotal ubiquitin sites required for its proteasome-dependent degradation. We also constructed a double mutant form of XBP1s (K60/77R) and found that it showed higher capacity in resisting against ubiquitin-mediated protein degradation, increasing nuclear translocation, enhancing transcriptional activity, suppressing ER stress and promoting Foxo1 degradation, compared to that of wild type XBP1s (WT). Consistently, overexpression of the K60/77R XBP1s mutant in DIO mice increased the ability to reduce ER stress and decrease Foxo1 levels, thus contributed to maintaining glucose homeostasis. ConclusionOur results suggest that delaying the degradation of XBP1s by preventing ubiquitination might provide a strategic approach for reducing ER stress as an anti-diabetes therapy.

Design and biological evaluation of tetrahydropyridine derivatives as novel human GPR119 agonists

A series of novel tetrahydropyridine derivatives were prepared and evaluated using cell-based measurements. Systematic optimization of general structure G-1 led to the identification of compound 35 (EC50 = 4.9 nM) and 37 (EC50 = 8.8 nM) with high GPR119 agonism activity and moderate clog P. Through single and long-term pharmacodynamic experiments, we found that compound35 showed a hypoglycemic effect and may have an effect on improving basal metabolic rate in DIO mice. Both in vitro and in vivo tests indicated that compound 35 was a potential potent GPR119 agonist in allusion to T2DM treatment.

DR10601, a novel recombinant long-acting dual glucagon-like peptide-1 and glucagon receptor agonist for the treatment of obesity and type 2 diabetes mellitus.

Both glucagon-like peptide-1 (GLP-1) and glucagon (GCG) belong to the incretin family. This study aimed to investigate the pharmacokinetics and pharmacodynamics of DR10601, a fully recombinant hybrid peptide with dual GLP-1/GCG receptor agonistic activity. The agonistic ability of DR10601 was indirectly assessed by inducing cAMP accumulation in Chinese hamster ovary cells transfected with GLP-1R or GCGR in vitro. Following s.c. administration, the plasma pharmacokinetics of DR10601 were analysed in male Sprague-Dawley rats. The antiobesity effects and improved glycaemic control of DR10601 in vivo were evaluated by administering DR10601 to high-fat DIO mice and ICR mice as a single dose or repeated s.c. doses once every 4days for 24days. DR10601 exhibits dual agonistic activity on GLP-1 and glucagon receptors. The plasma half-life of DR10601 in Sprague-Dawley rats following s.c. administration was 51.9 ± 12.2h. In an IPGTT, a single s.c. dose of DR10601 (30nmol/kg) produced similar glycaemic control effects and a longer duration of action compared to dulaglutide (10nmol/kg). Compared with that achieved with liraglutide (40nmol/kg) s.c. administered daily, DR10601 administered s.c. once every 4days at 90nmol/kg exerted a nearly equivalent effect on food intake and significantly reduced the body weights of high-fat DIO mice at 24days. Repeated administration of DR1060 provides potent and sustained glycemic control and body weight loss effect in high-fat DIO mice. DR10601 is a promising long-acting agent deserving further investigation for the treatment of type 2 diabetes and obesity.

Activation of Kappa Opioid Receptor Regulates the Hypothermic Response to Calorie Restriction and Limits Body Weight Loss.

Mammals maintain a nearly constant core body temperature (Tb) by balancing heat production and heat dissipation. This comes at a high metabolic cost that is sustainable if adequate calorie intake is maintained. When nutrients are scarce or experimentally reduced such as during calorie restriction (CR), endotherms can reduce energy expenditure by lowering Tb [1-6]. This adaptive response conserves energy, limiting the loss of body weight due to low calorie intake [7-10]. Here we show that this response is regulated by the kappa opioid receptor (KOR). CR is associated with increased hypothalamic levels of the endogenous opioid Leu-enkephalin, which is derived from the KOR agonist precursor dynorphin [11]. Pharmacological inhibition of KOR, but not of the delta or the mu opioid receptor subtypes, fully blocked CR-induced hypothermia and increased weight loss during CR independent of calorie intake. Similar results were seen with DIO mice subjected to CR. In contrast, inhibiting KOR did not change Tb in animals fed ad libitum (AL). Chemogenetic inhibition of KOR neurons in the hypothalamic preoptic area reduced the CR-induced hypothermia, whereas chemogenetic activation of prodynorphin-expressing neurons in the arcuate or the parabrachial nucleus lowered Tb. These data indicate that KOR signaling is a pivotal regulator of energy homeostasis and can affect body weight during dieting by modulating Tb and energy expenditure.

Delayed Pulmonary Apoptosis of Diet-Induced Obesity Mice following Escherichia coli Infection through the Mitochondrial Apoptotic Pathway.

Escherichia coli (E. coli) is one of pathogens causing nosocomial pneumonia and could induce pulmonary excessive apoptosis. Although much has been learned about metabolic diseases induced by obesity, the information linking bacterial pneumonia to obesity is limited. Accordingly, we investigated the apoptosis of normal (lean) and diet-induced obesity (DIO, fed a high-fat diet) mice after nasal instillation with E. coli. Lung tissues were obtained at 0 (preinfection), 12, 24, and 72 h after infection, and acute pulmonary inflammation was observed at 12 h. Elevated cell apoptosis and percentage of pulmonary cells depolarized with collapse of the mitochondrial transmembrane potential (Δψm) occurred in response to bacterial infection. The relative mRNA and protein expressions of Bax, caspase-3, and caspase-9 increased, but Bcl-2 decreased in the lung. Interestingly, the apoptotic percentage and most of apoptosis-associated factors mentioned above peaked at 12 or 24 h in the lean-E. coli group, while at 24 or 72 h in the DIO-E. coli group. Taken together, these findings indicated that the E. coli pneumonia caused excessive pulmonary apoptosis through the mitochondria-mediated pathway, and the apoptosis was delayed in the DIO mice with E. coli pneumonia.

Bilirubin reduces visceral obesity and insulin resistance by suppression of inflammatory cytokines.

ObjectiveAlthough previous studies have reported a negative relationship between serum bilirubin concentration and the development of diabetes mellitus (DM), the relationship between bilirubin and insulin resistance has not been thoroughly assessed. This study was designed to determine the relationships between bilirubin, body fat distribution, and adipose tissue inflammation in patients with type 2 DM and the effect of bilirubin in an obese animal model.MethodBody fat distribution was measured using an abdominal dual bioelectrical impedance analyzer in patients with type 2 DM. We also measured glycemic control, lipid profile, serum bilirubin concentration and other clinical characteristics, and determined their relationships with body fat distribution. In the animal study, biliverdin (20 mg/kg daily) was orally administered to high-fat diet (HFD)-induced obese (DIO) mice for 2 weeks, after which intraperitoneal insulin tolerance testing was performed. Then, adipocyte area, adipocytokine expression, and macrophage polarization were evaluated in epididymal adipose tissues.ResultsIn the clinical study, univariate analysis showed that a lower bilirubin concentration was significantly correlated with higher body mass index, waist circumference, triglyceride, uric acid, creatinine, visceral fat area and lower HDL-C. In multivariate analyses, bilirubin concentration significantly correlated with diastolic blood pressure, creatinine, and visceral fat area. However, there was no association between bilirubin concentration and subcutaneous fat area. In the animal study, DIO mice treated with biliverdin had smaller adipocytes than untreated DIO mice and biliverdin improved HFD-induced insulin resistance. Biliverdin treatment reversed the higher gene expression of Cd11c, encoding an M1 macrophage marker, and Tnfa, encoding the proinflammatory cytokine tumor necrosis factor-α, in the adipose tissues of DIO mice. These data suggest biliverdin administration alleviates insulin resistance by ameliorating inflammation and the dysregulation of adipocytokine expression in adipose tissues of DIO mice.ConclusionsBilirubin may protect against insulin resistance by ameliorating visceral obesity and adipose tissue inflammation.

Abstract 112: Soluble (pro) Renin Receptor Serves as Novel Insulin Sensitizing Agent in Mice With Diet-Induced Obesity

Thiazolidinediones, the PPARgamma agonists, represent a unique class of insulin sensitizers for management of hyperglycemia in type 2 diabetes but are limited by the side effect of fluid retention. Here we report therapeutic potential of histidine-tagged recombinant soluble (pro)renin receptor (sPRR), termed as sPRR-His, in a mouse obesity model induced by 9-month high fat diet (DIO) for management of glucose dysregulation. In the DIO mice, a 2-wk administration of sPRR-His lowered body weight (47.0 ± 1.4 vs. 52.8 ± 1.2 g) and remarkably improved multiple metabolic parameters including hyperglycemia (serum glucose: 148.8 ± 19.1 vs. 224.8 ± 12.5 mg/dL), hyperinsulinemia (serum insulin: 3.7 ± 0.4 vs. 10.9 ± 0.9 ng/ml), glucose intolerance, albuminuria (urine albumin/ Creatinine: 63.4 ± 3.5 vs. 97.7 ± 10.2 mg/g), and glomerular hyperfiltration (GFR: 264.7 ± 12.2 vs. 521.3 ± 18.2 μl/min) in the absence of fluid retention (plasma volume: 28.7 ± 1.2 vs. 27.8 ± 0.9 μl/g). GFR and plasma volume were measured by using FITC-sinistrin (7.5mg/100g i.v.) and FITC-dextran (2 mg/100 g i.v.) respectively. Conversely, inhibition of endogenous sPRR production by PF429242, an inhibitor of site-1 protease induced diabetes and insulin resistance which were reversed by sPRR-His supplement. At cellular level, sPRR-His enhanced insulin-induced increases in glucose uptake via upregulation of phosphorylated AKT and protein abundance of Glut4. Promoter and gene expression analysis revealed PRR as a direct target gene of PPARγ. Adipocyte-specific PPARγ deletion induced severe diabetes (blood glucose: 243.6 ± 39.8 vs. 108.4 ± 2.1 mg/dL) and insulin resistance (plasma insulin: 48.9 ± 4 vs. 1.5 ± 0.2 ng/ml) associated with reduced adipose PRR expression and circulating sPRR (plasma sPRR: 265.7 ± 14 vs. 893.0 ± 86.8 pg/ml). sPRR-His supplement in the null mice nearly normalized blood glucose (142.4 ± 12.2 mg/dL) and insulin level (14.3 ± 7.9 ng/ml). Additionally, sPRR-His treatment suppressed DIO-induced renal sodium-glucose cotransporter-2 (SGLT2) expression. Overall, sPRR-His exerts potent hypoglycemic action via serving as a mediator of insulin-sensitizing action of thiazolidinedione as well as a negative regulator of renal SGLT2 in the absence of fluid retention side effect.

Novel lipid side chain modified exenatide analogs emerged prolonged glucoregulatory activity and potential body weight management properties

Exenatide is known as the first marketed GLP-1 agonist for antidiabetic treatment, but it need twice injection a day because of its fast clearance. This work aims to prolong the half-life of exenatide by modified with novel lipid chain. Four optimized exenatide analogs named as Cys12-Exenatide (1–39)-NH2, Cys40-Exenatide (1–39)-NH2, Cys12-Tyr22-Gln24-Glu28-Arg35-Exenatide (1–39)-NH2 and Tyr22-Gln24-Glu28-Arg35-Cys40-Exenatide (1–39)-NH2 were selected and applied for conjugation. Then a series of evaluations including GLP-1R activation assay were conducted, conjugation C2 was selected for further investigation. Glucoregulatory and insulin secretion assay and hypoglycemic duration test were accessed and showed that C2 was capable of comparable insulinotropic activities and glucose-lowering abilities with those of liraglutide and exenatide. Cell protective effects in INS-1 cells confirmed that C2 had relatively protection effects. Meanwhile, once daily injection of C2 to STZ-induced diabetic mice achieved long-term beneficial effects on glucose tolerance, body weight and blood chemistry. Acute feeding studies were evaluated in DIO mice. These results suggested that C2 is a promising agent for further investigation of its potential to treat diabetes patients with obese.

Abstract 813: Downregulation of the Cardiac Sodium Channel Nav1.5 Mediates Obesity-Induced Atrial Fibrillation

Background: Obesity increases atrial fibrillation (AF) risk, but the underlying mechanisms are unclear. Emerging evidence supports reduced cardiac Na + channel (Nav1.5) expression and current ( I Na ) as one potential mechanism. Objective: We used an acquired (diet-induced obese, DIO) obese mouse model to test the hypothesis that obesity increases AF risk by downregulating Nav1.5. Methods: Weight, BP, plasma glucose, and F 2 -isoprostanes were measured in DIO mice and compared to controls. Echocardiography, EP studies, immunohistochemistry, Western blotting, cellular patch clamping and optical mapping studies were performed. Results: DIO (37.5±3.8) mice were heavier than controls ( 24.3 ±4.1;P <0.001). AF burden was 355±383 sec in DIO mice vs. 8.3± 21.4 sec in controls ( P <0.0001). Nav1.5 expression, I Na , atrial action potential duration (APD) and conduction velocity (CV) were reduced, and F 2 -IsoPs were increased. AF burden was reduced in the DIO model after chronic treatment with MitoTEMPO antioxidant, with partial restoration of the APD90, I Na and AP upstroke and reduction in F 2 -IsoPs. Conclusions: Mice with either acquired or genetic susceptibility to obesity are more prone to develop AF. And this is partially mediated by increased oxidative stress leading to downregulation of Nav1.5, shortened atrial APD and reduced atrial conduction velocity suggesting a possible reentrant mechanism of AF.

Abstract SY28-04: Obesity-associated leptin signaling promotes chemotherapy resistance in triple-negative breast cancer: The role of tumor-initiating cell enrichment

Abstract SY28-04: Obesity-associated leptin signaling promotes chemotherapy resistance in triple-negative breast cancer: The role of tumor-initiating cell enrichment

Abstract 2843: Advanced age and obesity separately and interactively potentiate triple negative breast cancer progression

Abstract Advanced age and obesity are major risk factors for breast cancer (BC) mortality that are independently associated with aggressive disease and therapeutic resistance. Despite this, preclinical models of BC mainly utilize young lean mice. The mechanisms underlying age- and obesity-related BC progression are currently unknown, highlighting a critical gap in our understanding of BC biology. To address this gap, we generated aged (16 mos) and young (6 mos) cohorts of lean and obese mice by placement on either low-fat control (10% kcal from fat) or diet-induced obesity (DIO; 60% kcal from fat) diet, respectively, beginning at 8-weeks of age. This resulted in four experimental groups: aged lean, aged DIO, young lean, young DIO. Mice were orthotopically injected with one of two triple negative BC cell lines (E0771 or Met-MWNTLung), which represent aggressive BC subtypes associated with poor prognosis in humans. Outcomes included primary tumor weight and lung metastasis quantification. Biochemical and molecular analyses included serum cytokines and gene expression within tumor and adjacent normal mammary tissue. Our TNBC models indicate tumor growth is similarly increased in young DIO and aged lean mice, with both significantly higher than young lean mice. Moreover, the combination of advanced age and obesity in aged DIO mice significantly increased tumor growth relative to all other experimental groups. Preliminary pathology analyses in Met-MWNTLung-bearing mice reveal that advanced age enhances lung metastasis beyond that of young mice regardless of diet. Increased tumor burden in both the aged lean and young DIO mice is accompanied by increased serum proinflammatory cytokines (IL-1b, CXCL13, CCL11, among others). Consistent with these findings, Gene Set Enrichment Analysis (GSEA) of Met-MWNTLungtumor and adjacent normal mammary microarray data demonstrated enrichment of inflammatory pathways, including IFNγ response, IFNα response, IL-2/STAT5 signaling, and IL-6/JAK/STAT3 signaling in aged lean and young DIO mice relative to young lean mice. Interestingly, tumor-adjacent mammary from aged lean and young DIO mice was also enriched for markers of EMT, consistent with age- and DIO-associated phenotypic switching of cells within the tumor-adjacent microenvironment. Taken together, our findings reveal that obesity and advanced age augment BC progression and promote inflammation within the local (mammary & tumor) and systemic (serum) microenvironments. Ongoing analyses are investigating the role of cellular senescence, a stress-induced cell cycle arrest in which cells acquire a proinflammatory secretome. Knowledge gained from this study will help to elucidate the mechanisms underlying advanced age- and obesity-related BC progression, which is urgently needed to develop new strategies for reducing adverse TNBC outcomes associated with advanced age and obesity in human populations. Citation Format: Laura A. Smith, Magdalena A. Rainey, Nishita T. Sheth, Stephanie A. Montgomery, Stephen D. Hursting. Advanced age and obesity separately and interactively potentiate triple negative breast cancer progression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2843.

The mitochondrial-derived peptide MOTS-c is a regulator of plasma metabolites and enhances insulin sensitivity.

MOTS‐c is an exercise mimetic and improves insulin sensitivity in aged and diet‐induced obese mice. Although plasma markers are good markers for the metabolic condition, whether MOTS‐c changes plasma markers in diet‐induced obese mice has not been examined. Here, we used an unbiased metabolomics approach to examine the effect of MOTS‐c on plasma markers of metabolic dysfunction. We found that three pathways – sphingolipid metabolism, monoacylglycerol metabolism, and dicarboxylate metabolism – were reduced in MOTS‐c–injected mice. Interestingly, these pathways are upregulated in obese and T2D models. MOTS‐c improves insulin sensitivity and increases beta‐oxidation to prevent fat accumulation in DIO mice through these pathways. These results provide us a better understanding of the mechanism of how MOTS‐c improves insulin sensitivity and reduces the body weight and fatty liver and opens a new venue for further study.

Differential Anti-Adipogenic Effects of Eicosapentaenoic and Docosahexaenoic Acids in Obesity.

To assess the associations of plasma eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) with body fat in a population-based sample and explore the mechanism of action based on browning of white adipose tissue (WAT) in high-fat-diet-induced obese (DIO) mice and 3T3-L1 adipocytes. Plasma EPA and DHA of 1719 adults in the National Health and Nutrition Examination Survey (2003-2004) are determined by gas chromatography mass spectrometry, while total body fat is measured by dual-energy X-ray absorptiometry. DIO mice are fed a high-fat diet supplemented with EPA or DHA (1% wt/wt) for 15 weeks and 3T3-L1 preadipocytes are treated with EPA or DHA during differentiation. Plasma DHA but not EPA is associated with lower body fat mass (ptrend < 0.0001), which persists in overweight/obese subjects (ptrend = 0.02). DHA supplementation reduces inguinal WAT and exhibits a more pronounced thermogenic effect than EPA in DIO mice. In vitro, the browning process is induced after 2-day and 6-day treatment with DHA and EPA, respectively. Plasma DHA but not EPA is inversely associated with body fat mass. The more potent anti-adipogenic effect of DHA than EPA may involve a better capability of inducing browning of WAT for DHA.