Subcutaneous Adipose Tissue Samples Research Articles

Effect of Soybean Oil and Fish Oil on Lipid-Related Transcripts in Subcutaneous Adipose Tissue of Dairy Cows.

Simple SummaryThe objective of this study was to determine the effects of degree of unsaturation of dietary lipids on lipid-related metabolites and transcription of genes involved in lipid metabolism in subcutaneous adipose tissue (SAT) of mid-lactating dairy cows. The objective was achieved by supplementing dairy cows with soybean oil (high in linoleic acid) or fish oil (high in polyunsaturated fatty acids) for 63 days (nine weeks). Results revealed effects of oil supplement on lipid metabolism but a mild effect on the transcriptome of the adipose tissue. Compared to soybean oil, fish oil had a higher lipogenic effect in SAT.The objective of this study was to determine the effect of long-term supplementation of unsaturated oil on lipid metabolism and transcription of genes involved in lipid metabolism in subcutaneous adipose tissue (SAT) of mid-lactating dairy cows. The objective was achieved by supplementing dairy cows with soybean oil (SO; high in linoleic acid) or fish oil (FO; high in EPA and DHA) for 63 days (nine weeks). Cows were fed a control diet with no added lipid, or diets containing SO or FO (n = 5 cows/group). At the onset of the experiment (day 0) and on days 21, 42, and 63 of supplementation, blood and SAT samples were collected from each animal. Oil supplementation increased cholesterol and NEFA in plasma, with a greater effect of SO compared to FO. Concentration of BUN was lower in SO compared to control and FO at the end of the trial. Transcription of few genes was affected by dietary lipids: FABP4 had lowest expression in FO followed by SO and control. ACACA and FASN had higher expression in FO. Transcription of SCAP was higher but expression of INSIG1 was lower in SO. Overall, results revealed that compared to control, SO and FO had lipogenic effect in SAT.

PSIII-39 Gene expression in adipose tissue at 8 months could explain adult overweight status in Beagle dogs

Abstract Individuals are more or less prone to excessive fat deposits and body weight. This could result from differences in the expression of gene involved in adipose tissue metabolism in early life. This study aimed to identify such possible differences in puppy dogs. Twenty-four 4-wk old females Beagle puppies were raised in the same environment, and fed with the same diet, ad libitum but for 3.5 h. Dogs were allotted in 3 groups according to their fat-free mass, fat mass, and pelvic circumference at 24 months of age: IW (n = 9; ideal body condition, OW1 (n = 6; moderately overfat) and OW2 (n = 9; overfat/obese). Subcutaneous and visceral adipose tissue (VAT) samples were collected at the time when animals were neutered (8 months). The mRNA level of genes involved in the regulation of lipolysis (perilipin, HSL), lipogenesis (ACC, ADRP, CPT1, FAS, FABP), adipogenesis (PPARg, SREBP-1c), thermogenesis (UCP1, UCP2), glucose uptake (GLUT4), insulin signaling (IRS2), and adiponectin production was assessed by real time RT-PCR. Gene expression levels in the three groups were compared by a Wilcoxon test. Ethics approval was granted by the French Ministry of Research and Royal Canin’s Ethical Committee. Compared to IW, OW2 dogs had a lower expression of UCP1 in the VAT (P = 0.049). This could result in reduced heat production by non-shivering thermogenesis. Moreover, a low UCP1 expression has been linked to obesity. Compared to IW, OW1 dogs had a higher transcription level of ACC and ADRP genes (P < 0.05). ACC is a point of regulation of the FA synthesis whereas ADRP is involved in the differentiation of the pre-adipocytes, and fat storage. These results suggest that the overweight/obesity phenotype could be associated with a lower heat loss, and the moderately overweight phenotype with a higher ability to fat synthesis compared to the lean phenotype.

Effects of exercise training on markers of adipose tissue remodeling in patients with coronary artery disease and type 2 diabetes mellitus: sub study of the randomized controlled EXCADI trial

BackgroundInvestigate effects of long-term exercise on the remodeling markers MMP-9, TIMP-1, EMMPRIN and Galectin-3 in combined type 2 diabetes mellitus (T2DM) and coronary artery disease (CAD) patients. Any associations between these biomarkers and glucometabolic variables were further assessed at baseline.Methods137 patients (age 41–81 years, 17.2% females) were included and randomized to a 12-months exercise program or to a control group. Fasting blood samples and subcutaneous adipose tissue (AT) samples were taken at inclusion and after 12-months. The intervention was a combination of aerobic and strength training for a minimum of 150 min per week. Circulating protein levels were measured by ELISA methods and RNA was extracted from AT and circulating leukocytes. Expression levels were relatively quantified by PCR.ResultsAfter 12 months of intervention, both AT-expression and circulating levels of EMMPRIN were increased in the exercise group (p < 0.05, both) with significant difference in change between the two groups (p < 0.05 both). No significant effect was observed on MMP-9, TIMP-1 and Galectin-3. Levels of TIMP-1 (AT-expression and circulating) were significantly correlated to insulin, and HOMA2- after Bonferroni correction (p = 0.001, by 48 performed correlations).ConclusionThe increase in levels of EMMPRIN after long-term exercise training, might indicate some degree of AT remodeling in these patients after 12-months of exercise, whether beneficial or not. The remodeling markers were to some extent associated with glucometabolic variables in our population with the combined disease.Trial registration clinicaltrials.gov, NCT01232608. Registered 2 November 2010

Circulating Wnt1-inducible signaling pathway protein-1 (WISP-1/CCN4) is a novel biomarker of adiposity in subjects with type 2 diabetes.

Wnt1-inducible signaling pathway protein 1, or cellular communication network factor 4 (CCN4), a member of CCN family of secreted, extracellular matrix associated signaling proteins, recently was validated as a novel adipose tissue derived cytokine. To assess the relationships between circulating CCN4, adipose tissue distribution and function, and chronic low-grade inflammation in subjects with type 2 diabetes. We observed 156 patients with type 2 diabetes and 24 healthy controls. Serum levels of CCN4, hsCRP and alpha1-acid glycoprotein (alpha1-AGP) were measured by ELISA. Serum concentrations of leptin, resistin, visfatin, adipsin, adiponectin, IL-6, IL-8, IL-18 and TNF-alpha were determined by multiplex analysis. Fat mass and distribution was assessed by DEXA. Mean diameter of adipocytes was estimated in samples of subcutaneous adipose tissue. Patients with diabetes had higher levels of circulating CCN4, leptin, resistin, adipsin, visfatin, hsCRP, alpha1-AGP, and IL-6 (all p<0.02). The CCN4 concentration correlated positively with percentage of fat mass in central abdominal area, as well as with leptin, resistin and visfatin levels; negative correlation was found between CCN4 and mean adipocyte diameter. In multiple regression analysis fat mass in central abdominal area was independent predictor for CCN4 concentration. In subjects with type 2 diabetes serum levels of CCN4 are associated with central abdominal fat mass and adipose tissue dysfunction.

Epicardial adipose tissue volume and annexin A2/fetuin-A signalling are linked to coronary calcification in advanced coronary artery disease: Computed tomography and proteomic biomarkers from the EPICHEART study

Background & aimsThe role of epicardial adipose tissue (EAT) in the pathophysiology of late stage-coronary artery disease (CAD) has not been investigated. We explored the association of EAT volume and its proteome with advanced coronary atherosclerosis. MethodsThe EPICHEART Study prospectively enrolled 574 severe aortic stenosis patients referred to cardiac surgery. Before surgery, EAT volume was quantified by computed tomography (CT). During surgery, epicardial, mediastinal (MAT) and subcutaneous (SAT) adipose tissue samples were collected to explore fat phenotype by analyzing the proteomic profile using SWATH-mass spectrometry; pericardial fluid and peripheral venous blood were also collected. CAD presence was defined as coronary artery stenosis ≥50% in invasive angiography and by CT-derived Agatston coronary calcium score (CCS). ResultsEAT volume adjusted for body fat was associated with higher CCS, but not with the presence of coronary stenosis. In comparison with mediastinal and subcutaneous fat depots, EAT exhibited a pro-calcifying proteomic profile in patients with CAD characterized by upregulation of annexin-A2 and downregulation of fetuin-A; annexin-A2 protein levels in EAT samples were also positively correlated with CCS. We confirmed that the annexin-A2 gene was overexpressed in EAT samples of CAD patients and positively correlated with CCS. Fetuin-A gene was not detected in EAT samples, but systemic fetuin-A was higher in CAD than in non-CAD patients, suggesting that fetuin-A was locally downregulated. ConclusionsIn an elderly cohort of stable patients, CCS was associated with EAT volume and annexin-A2/fetuin-A signaling, suggesting that EAT might orchestrate pro-calcifying conditions in the late phases of CAD.

Increased Adipose Tissue Expression of Interferon Regulatory Factor (IRF)-5 in Obesity: Association with Metabolic Inflammation.

Interferon regulatory factor (IRF)-5 is known to be involved in M1 macrophage polarization, however, changes in the adipose expression of IRF5 in obesity and their relationship with the local expression of proinflammatory cytokines/chemokines are unknown. Therefore, IRF5 gene expression was determined in the subcutaneous adipose tissue samples from 53 non-diabetic individuals (6 lean, 18 overweight, and 29 obese), using real-time RT-PCR. IRF5 protein expression was also assessed using immunohistochemistry and/or confocal microscopy. Adipose gene expression of signature immune metabolic markers was also determined and compared with adipose IRF5 gene expression. Systemic levels of C-reactive protein and adiponectin were measured by ELISA. The data show that adipose IRF5 gene (P = 0.008) and protein (P = 0.004) expression was upregulated in obese compared with lean individuals. IRF5 expression changes correlated positively with body mass index (BMI; r = 0.37/P = 0.008) and body fat percentage (r = 0.51/P = 0.0004). In obese, IRF5 changes associated positively with HbA1c (r = 0.41/P = 0.02). A good agreement was found between gene and protein expression of IRF5 in obese subjects (r = 0.65/P = 0.001). IRF5 gene expression associated positively with adipose inflammatory signatures including local expression of TNF-α, IL-6, CXCL8, CCL-2/5, IL-1β, IL-18, CXCL-9/10, CCL7, CCR-1/2/5, TLR-2/7/8/9, IRF3, MyD88, IRAK-1, and inflammatory macrophage markers (P < 0.05). Interestingly, IRF5 gene expression correlated positively with CRP (r = 0.37, P = 0.03) and negatively with adiponectin levels (r = −0.43, P = 0.009). In conclusion, elevated adipose IRF5 expression in obesity concurs with the typical inflammatory signatures, locally and systemically. Hence, the IRF5 upregulation may represent a novel adipose tissue marker for metabolic inflammation.

Association between ANGPTL-4 and the proinflammatory process in cancer cachexia patients.

BackgroundContradictory results are reported for the role of angiopoietin-like 4 (ANGPTL-4) in the development of cancer-cachexia and inflammation, given its importance in angiogenesis and inflammatory signaling. Our aim was to analyze the levels of ANGPTL-4 in colorectal cancer patients with a stable weight and those with cachexia in order to establish a relationship between ANGPTL-4 and the inflammatory process.ResultsPlasma and tumor levels of ANGPTL-4 were higher in CC in comparison to other groups. A positive association was verified between plasmatic ANGPTL-4 and NFκB levels in tumor from CC. In WSC, we identified an association between the plasmatic ANGPTL-4, IL-15, and IL-10 in tumor and IL-15 in MES. Increased levels of NFκB and TNF-R1 in MES were detected in CC in comparison to WSC. Specifically in CC-group, a positive correlation was found between ANGPTL-4 levels and those of IL-1β, TNF-α, and NFκB in tumor, along with an association between ANGPTL-4 levels with IL-1β and MCP-1 levels in tumor; and ANGPTL-4 and IL-1β levels in MES.MethodsWe studied 102 patients, who were divided into three groups: control patients (C, n=37), cancer patients with a stable weight (WSC, n=23), and cancer-cachexia patients (CC, n=42). Samples of plasma, tumor, mesenteric (MES) and subcutaneous adipose tissue were removed for the determination of ANGPTL-4 levels and other proinflammatory factors.ConclusionsANGPTL-4 levels were higher in plasma and tumor of CC-group, and positively associated with pro-inflammatory and pro-tumorigenic factors. Our results suggest an opposite effect of ANGPTL-4 depending on the concentration and presence of cachexia.

Age- and BMI-Associated Expression of Angiogenic Factors in White Adipose Tissue of Children.

The growth of adipose tissue and its vasculature are tightly associated. Angiogenic factors have been linked to obesity, yet little is known about their expression during early childhood. To identify associations of angiogenic factors with characteristics on individual and tissue level, subcutaneous white adipose tissue samples were taken from 45 children aged 0–9 years undergoing elective surgery. We measured the expression of vascular endothelial growth factor A (VEFGA), fibroblast growth factor 1 and 2 (FGF1, FGF2), angiopoietin 1 and 2 (ANGPT1, ANGPT2), TEK receptor tyrosine kinase (TEK), and von Willebrand factor (VWF). In addition, we determined the mean adipocyte size in histologic tissue sections. We found positive correlations of age with FGF1 and FGF2 and a negative correlation with ANGPT2, with pronounced differences in the first two years of life. FGF1, FGF2, and ANGPT1 correlated positively with adipocyte size. Furthermore, we identified a correlation of ANGPT1 and TEK with body mass index-standard deviation score (BMI-SDS), a measure to define childhood obesity. Except for ANGPT2, all angiogenic factors correlated positively with the endothelial marker VWF. In sum, our findings suggest that differences related to BMI-SDS begin early in childhood, and the analyzed angiogenic factors possess distinct roles in adipose tissue biology.

Coronary disease is not associated with robust alterations in inflammatory gene expression in human epicardial fat.

Epicardial adipose tissue (EAT) is the visceral fat depot of the heart. Inflammation of EAT is thought to contribute to coronary artery disease (CAD). Therefore, we hypothesized that the EAT of patients with CAD would have increased inflammatory gene expression compared with controls without CAD. Cardiac surgery patients with (n = 13) or without CAD (n = 13) were consented, and samples of EAT and subcutaneous adipose tissue (SAT) were obtained. Transcriptomic analysis was performed using Affymetrix Human Gene 1.0 ST arrays. Differential expression was defined as a 1.5-fold change (ANOVA P < 0.05). Six hundred ninety-three genes were differentially expressed between SAT and EAT in controls and 805 in cases. Expression of 326 genes was different between EAT of cases and controls; expression of 14 genes was increased in cases, while 312 were increased in controls. Quantitative reverse transcription PCR confirmed that there was no difference in expression of CCL2, CCR2, TNF-α, IL-6, IL-8, and PAI1 between groups. Immunohistochemistry showed more macrophages in EAT than SAT, but there was no difference in their number or activation state between groups. In contrast to prior studies, we did not find increased inflammatory gene expression in the EAT of patients with CAD. We conclude that the specific adipose tissue depot, rather than CAD status, is responsible for the majority of differential gene expression.

P1628Angiopoietin-like protein (Angptl) 2 secreted from epicardial adipose tissue induces atrial myocardial fibrosis

Abstract Background Using excised human left atrial appendage samples, we previously demonstrated that epicardial adipose tissue (EAT) are highly associated with atrial myocardial fibrosis as a substrate of atrial fibrillation (AF). We also reported the relationship between Angptl2 in EAT and atrial fibrosis. However, the mechanism is not clear. The purpose is to clarify the mechanisms underlying the effect of EAT on the atrial myocardium. Methods Human peri-left atrial EAT and abdominal subcutaneous adipose tissue (SAT) samples were obtained from 6 cases (2 females, 70.2±13.2 years). 50 mg of EAT and SAT were quickly washed with PBS and centrifuged 1min at 1200rpm. After 3 times this procedures, adipose tissues were cultured in DMEM F12 medium with Fetal bovine Serum (FBS) overnight. After pre-incubation, EAT and SAT tissues were washed and centrifuge d three times and cultured in medium without FBS for 24hours. Finally, we collected oozed medium (conditioned medium) and used for experiments. Concentrations of Angptl2 in conditioned medium were measured by ELISA. To study the effects of conditioned medium, we used “organo-culture” system. Isolated atrium from 8week old male Sprague-Dawley rats were placed on the porous membrane with the endothelial face toward the membrane. After that, loading medium (conditioned medium:culture medium = 1:4), culture medium (control), or recombinant Angptl2 were dropped onto the epicardial face of the atrium once a day and incubated for 7 days (37°C, 5% CO2). Then, histological and immunohistochemical analysis were performed. We also performed quantitative reverse transcription–polymerase chain reaction (RT–PCR) analysis. Next, we isolated and cultured neonatal rat fibroblast and loaded Angptl2 for 24 hours.After collected these cells, we performed western blotting analysis. Results Atria organo-culture incubated for 7 days with conditioned medium showed global fibrosis. At epicardial side, fibrotic area of EAT group was significantly greater compared to that of SAT and control group (P&lt;0.05). mRNA of Col1a1, col3a1 and TGFβ1 were significantly increased in EAT group compared with the SAT and control group. And, the concentration of conditioned medium created from EAT was significant higher than that from SAT (P&lt;0.05). Then, we dropped 500 ng/ml of recombinant Angptl2 onto the rat atria. Fibrotic area of Angptl22 group significantly greater than that of control with increasing number of α-SMA positive cells, and mRNA of col3a1 and TGFβ1 were significantly increased in Angptl2 group compared with control group. In cultured fibroblasts, α-SMA and p-ERK expression were increased in Angptl2 group measured by western blotting analysis. Conclusions Our results demonstrated that EAT rather than SAT induces atrial myocardial fibrosis. There is a possibility that Angptl2 effused from EAT plays a part in atrial fibrosis thought EAT paracrine effect. Acknowledgement/Funding ONO PHARMACEUTICAL CO

Adipose tissue mitochondrial dysfunction in human obesity is linked to a specific DNA methylation signature in adipose-derived stem cells

This epigenome-wide association study in stromal/stem cells (ASCs), derived from subcutaneous adipose tissue samples of lean and obese subjects, revealed a specific DNA methylation signature in adipocyte precursors associated with obesity, which has a significant impact on the metabolic phenotype and the mitochondrial function of mature adipocytes.

Adipokines secretion in feline primary adipose tissue culture in response to dietary fatty acids

BackgroundObesity in cats has been associated with alterations in adipokines including: adiponectin, interleukin-6 (IL6), and tumor necrosis factor-α (TNFα). Omega-3 polyunsaturated fatty acids have multiple beneficial effects on obesity-associated disorders, and therefore may alleviate these alterations. This study aimed to determine the effects of body condition, fat depot, troglitazone, and different fatty acids on secretion of adiponectin, IL6 and TNFα from adipose tissue of healthy cats. Subcutaneous and visceral adipose tissue samples were collected from 18 healthy intact female cats, and body condition score (Range 3–7/9) was determined. Concentrations of adiponectin were measured in mature adipocytes cultures and concentrations of IL6 and TNFα were measured in stromovascular cells cultures following treatment with control medium, troglitazone at 10 μM, eicosapentaenoic acid, arachidonic acid, or palmitic acid, at 25, 50, or 100 μM.ResultsStromovascular cells of visceral origin secreted higher concentrations of IL6 than corresponding cells of subcutaneous origin (P = 0.003). Arachidonic acid treatment at 25, 50, and 100 μM increased IL6 secretion in subcutaneous (P = 0.045, P = 0.002, and P < 0.001, respectively) and visceral (P = 0.034, P = 0.001, and P < 0.001, respectively) stromovascular cells. Eicosapentaenoic acid treatment increased TNFα secretion in subcutaneous stromovascular cells at 25, 50, and 100 μM (P = 0.002, P = 0.001, and P = 0.015, respectively) and in visceral stromovascular cells at 50 μM (P < 0.001). No significant effect on medium adiponectin concentration was observed following troglitazone treatment (P = 0.4) or fatty acids treatments at 25 (P = 0.2), 50 (P = 0.8), or 100 (P = 0.7) μM. Body condition score did not have significant effects on medium concentrations of adiponectin (P = 0.4), IL6 (P = 0.1), or TNFα (P = 0.8).ConclusionsThis study demonstrated higher basal secretion of IL6 from visceral compared to subcutaneous adipose tissue, a stimulatory effect of arachidonic acid on secretion of IL6 and a stimulatory effect of eicosapentaenoic acid on TNFα from feline adipose tissue.

Isolation of adipose tissue derived regenerative cells from human subcutaneous tissue with or without the use of an enzymatic reagent.

Freshly isolated, uncultured, autologous adipose derived regenerative cells (ADRCs) have emerged as a promising tool for regenerative cell therapy. The Transpose RT system (InGeneron, Inc., Houston, TX, USA) is a system for isolating ADRCs from adipose tissue, commercially available in Europe as a CE-marked medical device and under clinical evaluation in the United States. This system makes use of the proprietary, enzymatic Matrase Reagent for isolating cells. The present study addressed the question whether the use of Matrase Reagent influences cell yield, cell viability, live cell yield, biological characteristics, physiological functions or structural properties of the ADRCs in final cell suspension. Identical samples of subcutaneous adipose tissue from 12 subjects undergoing elective lipoplasty were processed either with or without the use of Matrase Reagent. Then, characteristics of the ADRCs in the respective final cell suspensions were evaluated. Compared to non-enzymatic isolation, enzymatic isolation resulted in approximately twelve times higher mean cell yield (i.e., numbers of viable cells/ml lipoaspirate) and approximately 16 times more colony forming units. Despite these differences, cells isolated from lipoaspirate both with and without the use of Matrase Reagent were independently able to differentiate into cells of all three germ layers. This indicates that biological characteristics, physiological functions or structural properties relevant for the intended use were not altered or induced using Matrase Reagent. A comprehensive literature review demonstrated that isolation of ADRCs from lipoaspirate using the Transpose RT system and the Matrase Reagent results in the highest viable cell yield among published data regarding isolation of ADRCs from lipoaspirate.

Tissue-Specific Oxidative Stress Modulation by Exercise: A Comparison between MICT and HIIT in an Obese Rat Model.

Background and Aim Exercise is an effective strategy to reduce obesity-induced oxidative stress. The purpose of this study was to compare the effects of two training modalities (moderate-intensity continuous training (MICT) and high-intensity interval training (HIIT)) on the pro/antioxidant status of different tissues in obese Zucker rats. Methods Eight-week-old male Zucker rats (fa/fa, n = 36) were subdivided in three groups: MICT, HIIT, and control (no exercise) groups. Trained animals ran on a treadmill (0° slope), 5 days/week for 10 weeks (MICT: 51 min at 12 m·min−1; HIIT: 6 sets of 3 min at 10 m·min−1 followed by 4 min at 18 m·min−1). Epididymal (visceral) and subcutaneous adipose tissue, gastrocnemius muscle, and plasma samples were collected to measure oxidative stress markers (advanced oxidation protein products (AOPP), oxidized low-density lipoprotein (oxLDL)), antioxidant system markers (ferric-reducing ability of plasma (FRAP), superoxide dismutase (SOD), catalase, and glutathione peroxidase (GPx) activities), and prooxidant enzymes (NADPH oxidase and xanthine oxidase (XO) activities, myeloperoxidase content). Results Compared with the control, MICT increased GPx and catalase activities and the FRAP level in epididymal adipose tissue. HIIT increased the AOPP level in subcutaneous adipose tissue. In the muscle, HIIT increased both SOD and GPx activities and reduced the AOPP level, whereas MICT increased only SOD activity. Finally, plasma myeloperoxidase content was similarly decreased by both training modalities, whereas oxLDL was reduced only in the MICT group. Conclusion Both HIIT and MICT improved the pro/antioxidant status. However, HIIT was more efficient than MICT in the skeletal muscle, whereas MICT was more efficient in epididymal adipose tissue. This suggests that oxidative stress responses to HIIT and MICT are tissue-specific. This could result in ROS generation via different pathways in these tissues. From a practical point of view, the two training modalities should be combined to obtain a global response in people with obesity.

Offspring Exercise Modulates Beige Adipocyte Numbers by Differential Regulation of H3K9 Methyltransferase G9a and Demethylase KDM4C in Mice Fed a Maternal High Fat Diet (P21-067-19)

ObjectivesWe have shown that a maternal low protein and postnatal high fat (HF) diet increases offspring obesity and type 2 diabetes mellitus (T2DM) risk by epigenetically reducing beige adipocyte (BA) numbers via increased protein expression of G9a (Histone3 Lysine9 dimethyl transferase), an inhibitor of the BA marker FGF21. It is not yet known whether offspring exercise reverses the offspring obesity and T2DM risk caused by the maternal HF diet. MethodsTwo month old female C57Bl/6 J mice (F0) were fed a normal fat (NF) 16% diet or a 45% HF diet for 3 months prior to breeding, and subsequent gestation and lactation. Male offspring (F1) were fed the same NF and HF diets and further divided into either cage active (CA) or voluntary wheel running (Ex) groups for an additional 3 months yielding eight groups: NF (maternal treatment condition)-Ex-NF (post weaning treatment conditions), NF-Ex-HF, NF-CA-NF, NF-CA-HF, HF-Ex-NF, HF-Ex-HF, HF-CA-NF, and HF-CA-HF. Subcutaneous adipose tissue samples were collected for protein and mRNA analysis of FGF21, G9a, E4BP4 (G9a coactivator), and H3K9 methylation regulators that are induced by exercise including Activating Transcription Factor 4 (ATF4) and H3K9 demethylases (KDM4C). ResultsPostnatal HF diet decreased FGF21 positive BA numbers regardless of the maternal diet. A postnatal HF diet plus limited postnatal activity (CA) increased G9a and E4BP4 expression to suppress FGF21, while a postnatal HF diet plus exercise condition increased KDM4C expression. Although BA were detectable, the morphology of BA under CA conditions were compromised or scarred. ConclusionsThese findings suggest that a postnatal HF diet has a greater impact on offspring adiposity and BA number than a maternal HF diet. These data also suggest that offspring exercise induces KDM4C to counter the increase in G9a triggered by a postnatal HF diet. Whether KDM4C induces reduction in H3K9-me2 to regulate adipose tissue FGF21 needs investigation. Futures studies should determine whether H3K9 epigenetic modifications in BA have any functional differences that contribute to the observed phenotypic abnormalities. Funding SourcesUSDA Agricultural Research Service Project #3062-51000-052-00D.

797-P: Composition of Morning and Afternoon Meals Affects Metabolism and Inflammation in Human Adipose Tissue

Meal timing is an important factor for metabolic regulation. We recently showed that a diet where fat is mainly eaten in the morning and carbohydrates mainly in the evening worsens glycaemic control in subjects with prediabetes. Here, we investigated the effects of the day time of carbohydrate and fat intake on the subcutaneous adipose tissue (SAT) transcriptome in humans. In a cross-over study 34 non-obese men without diabetes (age 47.4 ± 2.4 years, BMI 27.4 ± 0.7 kg/m2) consumed two 4-week isocaloric diets - (1) HC/HF diet consisted of a high-carb breakfast and lunch (65 EN% carbohydrates, 20 EN% fat, 15 EN% protein) and a high-fat snack and dinner (35 EN% carbohydrates, 50 EN% fat, 15 EN% protein) or (2) inverse sequence of meals (HF/HC). After both dietary interventions, SAT samples were collected at 8.40, 12.20 and 19.00 hr. Gene expression was analyzed by microarrays and validated by PCR. Sinus model was fitted to expression levels of each transcript to predict the 24h rhythm. Out of 1345 SAT transcripts with a significant cosine rhythm, 37.8% transcripts were rhythmic on both diets, whereas 27.3% and 34.9% transcripts were rhythmic only on the HC/HF and HF/HC diet, respectively. Diets neither affected the oscillations of clock genes in SAT nor the salivary cortisol rhythm used as a central clock marker. However, dietary effects on a large set of oscillating and non-oscillating genes involved in glucose (e.g., IRS1, IRS2, PCK1, PDK4) and lipid metabolism (e.g., ACACA, ACAT1, ACOX2, PPARA) were found. Consumption of a HC/HF diet increased the expression of numerous inflammatory genes (e.g., CCL5, IL1B, ITGAX, CD3E) in SAT. Mechanisms underlying this remodeling included the change of activity of the insulin signaling, I-kappa B kinase/NF-kappa B and MAPK pathways. Collectively, our study revealed that the composition of the morning and afternoon meals strongly affect adipose tissue transcriptome in humans including the alteration of glucose and lipid metabolism and inflammatory pathways. Disclosure K. Kessler: None. K. Jürchott: None. N.N. Rudovich: None. A.F. Pfeiffer: Advisory Panel; Self; Abbott, Berlin-Chemie AG, Novo Nordisk A/S. Speaker's Bureau; Self; Lilly Diabetes, Novartis AG, Sanofi-Aventis Deutschland GmbH. O. Ramich: None. Funding German Science Foundation (KFO218 PF164/16-1 to O.R., A.F.P.); German Diabetic Association (to O.R.); German Center for Diabetes Research (82DZD0019G to O.R.)

723-P: A Single Session of Exercise Increases Resident Macrophages in Subcutaneous Adipose Tissue from Healthy Human Subjects

Infiltration of pro-inflammatory adipose tissue macrophages (ATMs) is implicated in systemic low-grade inflammation that underlies metabolic health complications in obesity. Exercise training has been linked to reduced adipose tissue and systemic inflammation, but these observations are often confounded by concomitant weight loss. Many metabolic benefits of exercise are a cumulative effect of each bout of exercise, thus the aim of this study was to determine changes in ATMs (using flow cytometry) in response to a single exercise session. We collected subcutaneous abdominal adipose tissue samples from 10 obese (BMI: 33±3kg/m2; fat mass: 41±20kg) and 14 lean (BMI: 23±13kg/m2; fat mass: 16±5kg) adults, 12 hours after exercising for 60 minutes at 70% VO2max (EX); and again 3 days after their most recent exercise session (No-EX). To characterize the cellular response to exercise without the confounding influence of exercise novelty, all subjects were habitual exercisers. The exercise session did not alter total immune cell number (OBESE: 30±7 vs. 31±7; LEAN: 27±9 vs. 28±8% live cells) or total ATM number (OBESE: 3.9±1.7 vs. 4.0±1.6; LEAN: 3.5±1.5 vs. 4.3±1.9% live cells; for No-EX and EX, respectively). However, there was a main effect for exercise to increase the number of CD11c- ATMs (OBESE: 1.3±1.4 vs. 1.5±1.5; LEAN: 1.0±0.6 vs. 2.0±1.6% live cells; P=0.03), indicating the abundance of resident ATMs increased after exercise. Surprisingly, the response to exercise was not different in lean vs. obese subjects. Although the change in resident ATM content after exercise was rather subtle in this cohort of healthy regular exercisers, these are the first single cell data to suggest each session of exercise may induce a phenotypic shift in subcutaneous ATMs. This could be a mechanism contributing to the anti-inflammatory health benefits of exercise. Disclosure A. Ludzki: None. E.M. Krueger: None. T.C. Baldwin: None. N.M. Taylor: None. L.A. Muir: None. C. Lumeng: None. J.F. Horowitz: Research Support; Self; American Diabetes Association. Funding National Institutes of Health (R01DK077966) Canadian Institutes of Health Research (DFS146190)

731-P: Exercise Training Does Not Alter Resting Fatty Acid Mobilization from Adipose Tissue

Excessive fatty acid (FA) mobilization from subcutaneous adipose tissue into systemic circulation underlies many metabolic health complications associated with obesity, such as insulin resistance. Exercise is often used to help treat and/or prevent insulin resistance, but the direct effects of exercise training (without weight loss) on systemic FA mobilization are unclear. The aim of this study was to determine the effect of both high-intensity interval training (HIIT) and moderate-intensity continuous training (MICT) on FA rate of appearance (FA Ra) into systemic circulation and factors regulating FA mobilization from subcutaneous adipose tissue. 18 obese adults (33±3 kg•m-2) were randomized to 12 weeks (4 d/week) of either HIIT (10 x 1 min at 90% HRmax with 1 min recovery; n=8) or MICT (45 min at 70% HRmax; n=10), and were required to maintain bodyweight throughout. Resting FA Ra (13C palmitate dilution) and abdominal subcutaneous adipose tissue samples were collected in the overnight fasted state before and after training (72h following their final exercise session). The abundance of key lipolytic and lipid storage proteins in adipose tissue were measured via immunoblot. Aerobic fitness (VO2peak) increased ∼10% after training (P = 0.002), with no difference between HIIT and MICT. Body weight remained unchanged after training (HIIT: 98±12 vs. 98±13 kg, MICT: 101±12 vs. 100±13), as did fat mass (HIIT: 40±5 vs. 39±5 kg, MICT: 42±8 vs. 41±9). Training did not affect resting FA Ra in either HIIT (16.5±3.2 vs. 15.3±1.8 μmol•kg FM-1•min-1) or MICT (16.5±2.6 vs. 15.8±1.4 μmol•kg FM-1•min-1). In line with this finding, neither HIIT nor MICT altered adipose tissue abundance or phosphorylation-state of the lipolytic enzymes ATGL and HSL, or other factors involved in FA mobilization and storage in adipose tissue (e.g., GPAT, DGAT, CGI-58, G0S2, CD36). In summary, in the absence of weight loss, 12 weeks HIIT or MICT did not alter the regulation of resting FA mobilization from subcutaneous adipose tissue. Disclosure M.W. Schleh: None. B.J. Ryan: None. J.B. Gillen: None. P. Varshney: None. K. Foug: None. A. Ludzki: None. J.F. Horowitz: Research Support; Self; American Diabetes Association. Funding National Institutes of Health (R01DK077966, P30DK089503, T32DK007245); Canadian Institutes of Health Research (DFS146190)

Impact of HIV/simian immunodeficiency virus infection and viral proteins on adipose tissue fibrosis and adipogenesis

HIV-infected patients receiving antiretroviral treatment (ART) often present adipose tissue accumulation and/or redistribution. adipose tissue has been shown to be an HIV/SIV reservoir and viral proteins as Tat or Nef can be released by infected immune cells and exert a bystander effect on adipocytes or precursors. Our aim was to demonstrate that SIV/HIV infection per se could alter adipose tissue structure and/or function. Morphological and functional alterations of subcutaneous (SCAT) and visceral adipose tissue (VAT) were studied in SIV-infected macaques and HIV-infected ART-controlled patients. To analyze the effect of Tat or Nef, we used human adipose stem cells (ASCs) issued from healthy donors, and analyzed adipogenesis and extracellular matrix component production using two dimensional (2D) and three-dimensional (3D) culture models. Adipocyte size and index of fibrosis were determined on Sirius red-stained adipose tissue samples. Proliferating and adipocyte 2D-differentiating or 3D-differentiating ASCs were treated chronically with Tat or Nef. mRNA, protein expression and secretion were examined by RT-PCR, western-blot and ELISA. SCAT and VAT from SIV-infected macaques displayed small adipocytes, decreased adipogenesis and severe fibrosis with collagen deposition. SCAT and VAT from HIV-infected ART-controlled patients presented similar alterations. In vitro, Tat and/or Nef induced a profibrotic phenotype in undifferentiated ASCs and altered adipogenesis and collagen production in adipocyte-differentiating ASCs. We demonstrate here a specific role for HIV/SIV infection per se on adipose tissue fibrosis and adipogenesis, probably through the release of viral proteins, which could be involved in adipose tissue dysfunction contributing to cardiometabolic alterations of HIV-infected individuals.

Vitamin D receptor in subcutaneous adipose tissue and serum vitamin D levels relate to gestational diabetes mellitus in overweight/obese pregnant women

Objective To investigate the differences in the expression of vitamin D receptor (VDR) and serum vitamin D levels in subcutaneous adipose tissue between overweight/obese and normal-weight gravidas, and the relationship between these two indicators and gestational diabetes mellitus (GDM). Methods Women with full-term singleton pregnancies who underwent elective cesarean section in Changzhou Maternal and Child Health Care Hospital Affiliated to Nanjing Medical University from January 2015 to April 2017 were enrolled. Among them, there were 70 cases GDM women, including 35 normal-weight (NW-GDM group) and 35 overweight/obese women (OW-GDM group). During the same period, another 70 pregnant women with normal glucose tolerance who underwent scheduled cesarean delivery were selected as the control group, including 35 normal weight women (NW-control group) and 35 obese/overweight women (OW-control group). Fasting blood samples were collected before operation to determine the levels of different biomarkers, including vitamin D, lipid, fasting blood glucose, fasting insulin and adiponectin, and to calculate the homeostasis model assessment-insulin resistance (HOMA-IR). Two subcutaneous adipose tissue samples of the abdominal wall were taken during the operation to detect the expression and distribution of VDR protein with immunohistochemistry. Meanwhile, VDR mRNA transcription level was quantitatively analyzed using real-time fluorescence quantitative polymerase chain reaction. One-way analysis of variance, LSD, Kruskal-Wallis test, Mann-Whitney U test, Chi-square test and logistic regression analysis were used for statistical analysis. Results (1) The body mass index (BMI) of the OW-control group and the OW-GDM group before pregnancy and delivery were all higher than that of the NW-control group and the NW-GDM group [BMI before pregnancy: (29.2±2.9), (29.4±3.8) vs (21.1±2.3) and (21.9±2.0) kg/m2, F=87.766; BMI before delivery: (35.2±3.4), (35.1±4.3) vs (27.9±2.8) and (28.8±3.3) kg/m2, F=44.827; all P<0.001]. Newborn birth weight and the proportion of diabetic family history in the OW-GDM group were higher comparing to the NW- and OW- control group [(3 893±498) vs (3 501±402) and (3 625±332) g, F=4.751; 22.9%(8/35) vs 5.7%(2/35) and 5.7%(2/35), χ2=7.869; all P<0.05]. (2) In the OW-control group, the fasting insulin level and HOMA-IR were higher and the adiponectin and vitamin D concentration were lower than those in the NW-control group [13.3(12.3-14.5) vs 12.0(10.4-13.3) mmol/L, 2.7(2.4-3.0) vs 2.2(2.0-2.7), (61.8±20.4) vs (74.9±29.3) ng/ml, (21.6±7.2) vs (25.9±7.3) ng/ml; all P<0.05], and similar results were found between the OW-GDM group and the NW-GDM group [15.3(12.3-19.5) vs 12.0(10.1-15.8) mmol/L, 3.4(2.6-4.1) vs 2.6(2.1-3.2), (50.3±22.3) vs (62.1±23.2) ng/ml, (17.1±6.7) vs (20.6±7.9) ng/ml, all P<0.05]. Compared with the NW-control group, the NW-GDM group had higher fasting glucose and lower high density lipoprotein-cholesterol (HDL-C), adiponectin and vitamin D levels [4.6(4.3-5.1) vs 4.3(4.0-4.5) mmol/L, 1.7(1.6-1.9) vs 2.1(1.6~2.4) mmol/L, (62.1±23.2) vs (74.9±29.3) ng/ml, (20.6±7.9) vs (25.9±7.3) ng/ml; all P<0.05]. Compared with the OW-control group, fasting glucose, fasting insulin and HOMA-IR were higher and HDL-C, adiponectin and vitamin D levels were lower in the OW-GDM group [4.7(4.4-5.4) vs 4.5(4.2-4.7) mmol/L, 15.3(12.3-19.5) vs 13.3(12.3-14.5) mmol/L, 3.4(2.6-4.1) vs 2.7(2.4-3.0), 1.6(1.4-1.8) vs 1.9(1.7-2.2) mmol/L, (50.3±22.3) vs (61.8±20.4) ng/ml, (17.1±6.7) vs (21.6±7.2) ng/ml; all P<0.05]. (3)The overall vitamin D deficiency rate during the third trimester of the four groups was 78.6% (110/140), and the figure was 62.8% (22/35), 82.8% (29/35), 77.1% (27/35) and 91.4% (32/35) in the NW-control group, OW-control group, NW-GDM group and OW-GDM group (χ2=8.994, P=0.029), indicating a higher rate in the OW-GDM group than that in the NW-control group (χ2=8.102, P=0.004). (4) VDR was expressed in the nucleus of adipose tissue in all samples and statistic difference in protein expression was found among the four groups. VDR mRNA expression was higher in both GDM subgroups than that in the two control subgroups, and also higher in the two overweight/obese subgroups than in the corresponding normal-weight subgroups. (5)Serum vitamin D level was negatively correlated with fasting blood glucose and pre-pregnancy BMI, and positively correlated with adiponectin (P<0.05). The incidence of GDM was related to family history of diabetes, VDR mRNA, total cholesterol, HDL-C and HOMA-IR. Conclusions GDM and overweight/obese patients had decreased serum vitamin D level and increased VDR in subcutaneous adipose tissue. These two factors are closely related to GDM. Key words: Diabetes, gestational; Obesity; Overweight; Adipose tissue; Receptors, calcitriol; Calcitriol