ResearchPad - adipose-tissue-appetite-and-obesity https://www.researchpad.co Default RSS Feed en-us © 2020 Newgen KnowledgeWorks <![CDATA[SUN-596 A Rare Presentation of Early Onset Wernicke Encephalopathy Following Sleeve Gastrectomy]]> https://www.researchpad.co/article/elastic_article_8747 Background: Wernicke encephalopathy (WE) has been reported after malabsorptive bariatric surgeries but is an uncommon complication of sleeve gastrectomy. Since 2011, the number of patients receiving a sleeve gastrectomy has tripled, with almost 60% of patients undergoing bariatric surgery receiving a sleeve gastrectomy in 2017 (1). We present a case of WE in a young woman as a rare and early complication of sleeve gastrectomy.

Clinical Case: A 32-year old female with a past medical history significant for hypertension, pseudotumor cerebri, and morbid obesity status post sleeve gastrectomy two months prior presented to the emergency department with complaints of blurry vision and lower extremity numbness. Physical examination showed sluggish light reflex and decreased extraocular movements. Given history of pseudotumor cerebri, patient underwent a therapeutic lumbar puncture with removal of 13 ml of CSF. Opening pressure was 20 cm of water and patient experienced no relief of her symptoms. Ophthalmology consult did not offer an explanation for the blurry vision. MRI-brain with and without contrast showed findings highly suggestive of WE. It showed faint linear symmetric hyperintensities along the bilateral mesial thalamus, dorsal midbrain and periaqueductal gray matter, which were determined to be acute in nature in comparison with a MRI performed three weeks prior. Upon further investigation, thiamine level was low at 43.6 nmol/L [66.5 – 200] confirming the diagnosis of WE. Thiamine supplementation was started immediately and patient reported improvement of her vision the next day with return to baseline in 3 days.

Conclusion: There have been a handful of cases of WE reported in literature as a complication of sleeve gastrectomy. Zheng, L also reported a case of WE 7 weeks after sleeve gastrectomy (2). Although sleeve gastrectomy does not directly affect the primary absorptive pathway of thiamine in the gastrointestinal tract, it is imperative to consider WE in patients presenting with suspicious neurologic symptoms after a recent sleeve gastrectomy. WE was suspected in our case due to typical MRI findings and neurological presentation after bariatric surgery, which was later confirmed by low serum thiamine level. Early detection and thiamine supplementation resulted in complete reversal of symptoms in our patient. WE is a rare but severe and preventable consequence of bariatric surgery that warrants attention given its rapid onset and detrimental course.

Reference: (1) Estimate of Bariatric Surgery Numbers, 2011-2017. (2018, June 26). Retrieved from https://asmbs.org/resources/estimate-of-bariatric-surgery-numbers. (2) Zheng, L. (2016).

Wernicke Encephalopathy and Sleeve Gastrectomy. American Journal of Therapeutics, 23(6).

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<![CDATA[SUN-586 CXCR2 Repression by Glucocorticoids in Adipose Tissue]]> https://www.researchpad.co/article/elastic_article_8741 Obesity-induced type 2 diabetes (T2D) is a significant risk factor of cardiovascular disease (CVD), which affects 28.1 million adults in the United States. Adipose tissue chronic inflammation is one of the main factors that drive obesity-induced insulin resistance (IR) and T2D. Despite several studies that have shown a link between obesity, adipose tissue inflammation, and IR/T2D, the mechanisms underlying this association are not well understood. Synthetic glucocorticoids are widely used for their potent anti-inflammatory actions; however, their use is hampered due to off-target side effects. Glucocorticoids exert profound effects on adipose tissue, including the regulation of adipocyte metabolism and immune functions. However, whether their effects on adipose tissue are positive or negative it is still a controversial topic. Genome-wide microarray data obtained from adipocyte-specific glucocorticoid receptor (GR) knockout (AdipoGRKO) mice showed that lack of GR leads to a significant increase in the expression of pro-inflammatory genes in white adipose tissue (WAT). Moreover, WAT isolated from adipoGRKO mice demonstrated significant increase in immune cell infiltration, which correlates with our gene expression data. Among the most up-regulated genes, we found the C-X-C Motif Chemokine Receptor 2 (CXCR2), which is a critical mediator of chemotaxis to the sites of inflammation. Although studies have shown the presence of CXCR2 in adipocytes and suggested the contribution of CXCR2 signaling in adipocyte development, its role in obesity-driven adipose tissue inflammation is unknown. This led us to hypothesize that adipocyte specific administration of glucocorticoids can reduce obesity-induced adipocyte inflammation by inhibiting CXCR2 gene transcription and signaling. Our in vitro studies using 3T3-L1 cells derived adipocytes showed that treatment with the synthetic glucocorticoid, Dexamethasone (Dex) led to a significant repression of CXCR2 mRNA and protein levels. Correlating with these results, Dex treatment significantly inhibited macrophage migration to adipocytes in a mechanism dependent on GR activation and repression of CXCR2. Furthermore, these results were recapitulated in vivo. Together our findings suggest that local delivery of glucocorticoids to adipose tissue could ameliorate inflammation and reduce the risk of developing IR and T2D.

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<![CDATA[SAT-605 Characterization of Dual Projection Patterns of Refeeding-Activated Neurons in the Parasubthalamic Nucleus]]> https://www.researchpad.co/article/elastic_article_8625 We have observed that following a fast, animals terminate their food intake within 2h after refeeding accompanied by a pattern of neuronal activation as identified by c-fos immunostaining that involves a number of brain regions associated with the regulation of food intake including the nucleus tractus solitarius (NTS), parabrachial nucleus (PBN), central nucleus of the amygdala (CEA), hypothalamic arcuate and paraventricular nuclei, and bed nucleus of the stria terminalis. We also observed striking c-fos activation in the posterior-lateral hypothalamus called the parasubthalamic nucleus or PSTN, raising the possibility that it may also be an important anorectic center in the brain. To establish how the PSTN is integrated into the CNS, we performed dual-label retrograde tract tracing studies to characterize whether refeeding-activated PSTN neurons project to one, or more than one target area in the CNS. Adult, Sprague-Dawley rats received dual stereotaxic injections of Alexa Fluor 488- and Alexa Fluor 555-conjugated cholera toxin β subunit (CTB; 0.1%, 0.5–1 µl volume) into the 1) PBN and NTS, 2) PBN and CEA and 3) NTS and CEA. After 7–12 days, the animals were fasted for 24 h and then given free access to food for 2 h before euthanasia by transcardial perfusion with 4% paraformaldehyde. Brains with successful dual injections were further processed for c-fos immunohistochemistry. The results showed that 26.5±3.8% of PSTN neurons projecting to the PBN also project to the CEA, and 34.6±7.6% of PSTN neurons that project to the CEA also project to the PBN. In addition, 20.2±2.7% of PSTN neurons that project to the PBN also project to the NTS, and 38.1±9.7% of PSTN neurons that project to the NTS also project to the PBN. Furthermore, 35.0±12.5% of PSTN neurons that project to the CEA project to the NTS and 37.1±4.0% of PSTN neurons that project to the NTS project to the CEA. Finally, up to 15% of the neurons with dual projections to the PBN and CEA contained c-fos after refeeding; up to 18% of the neurons with dual projections to the PBN and NTS contained c-fos; and up to 30% of neurons with dual projections to the NTS and CEA contained c-fos. We conclude that a large number of PSTN neurons have more than one projection site within the brain, thus the PSTN appears to have the capability of simultaneously communicating information about appetite to several, major feeding-related sites within the brain, presumably to terminate feeding.

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<![CDATA[SAT-606 Distribution of Beta Klotho Gene Expression in the Mouse Brain]]> https://www.researchpad.co/article/elastic_article_8605 Fibroblast growth factor 21 (FGF21) has emerged as a critical endocrine factor for understanding the neurobiology of obesity, such as by the regulation thermogenesis, food preference, and metabolism, as well as for neuroprotection in Alzheimer’s disease and traumatic brain injury. FGF21 is synthesized primarily by the liver and pancreas then crosses the blood brain barrier to exert its effects in the brain. However, the sites of FGF21 action in the brain is not well-defined. FGF21 action requires the activation of FGF receptor 1c as well as its obligate co-receptor beta klotho (KLB). In order to determine the sites of FGF21 action, we mapped the distribution of Klb mRNA by in situ hybridization using RNAscope technology. We labeled Klb distribution throughout the rostrocaudal axis of male wildtype mice by amplifying Klb hybridization using tyramine signal amplification and visualizing Klb hybridization using Cyanine 3 fluorescence. The resulting Klb signal appears as punctate red “dots,” and each Klb neuron may express low (1–4 dots), medium (5–9 dots), or high levels (10+ dots) of Klb hybridization. We then mapped individual Klb expressing neuron to the atlas plates provided by the Allen Brain Atlas in order to determine Klb distribution within the substructures of each brain region, which are defined by Nissl-based parcellations of cytoarchitectural boundaries. The distribution of Klb mRNA is widespread throughout the brain, and the brain regions analyzed thus far point to notable expression in the hypothalamus, amygdala, hippocampus, and the cerebral cortex. The highest expression of Klb was localized to the suprachiasmatic nucleus in the hypothalamus, which contained low and medium Klb-expressing neurons in the lateral hypothalamic area and ventromedial hypothalamic nucleus while low expressing Klb neurons were seen in the paraventricular and dorsmedial hypothalamic nucleus. Hippocampal Klb expression was limited to the dorsal region and largely restricted to the pyramidal cell layer of the dentate gyrus, CA3, CA2, and CA1 but at low levels only. In the amygdala, low and medium Klb expressing cells were seen in lateral amygdala nucleus while low levels were observed in the basolateral amygdala nucleus. Cortical Klb expression analyzed thus far included low Klb-expressing neurons in the olfactory areas, including layers 2 and 3 of piriform cortex and nucleus of the lateral olfactory tract. These findings are consistent with the known roles of FGF21 in the central regulation of energy balance, but also implicates potentially wide-ranging effects of FGF21 such as in executive functions.

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<![CDATA[SAT-LB102 Obesity Is Associated With Reduced Expression of the Anorexigenic Neuropeptide Nucleobindin-2/Nesfatin-1 in the Human Nucleus of the Solitary Tract]]> https://www.researchpad.co/article/elastic_article_8594 Introduction: Feeding is a complex behavior coordinated by interrelated forebrain, hypothalamic, and brainstem neuronal networks. Brainstem neurons constitute an important input for the neural circuitry integrating nutrient signals to control ingestive behavior. Orexigenic and anorexigenic neuropeptides act in concert to regulate energy balance. Data from animal models suggest that altered neuropeptidergic expression contributes to obesity. Nucleobindin-2/nesfatin-1, an appetite-suppressing neuropeptide and negative regulator of body weight, is reduced in the hypothalamus of mouse obesity models. In obese and overweight humans, we have recently reported decreased nucleobindin-2/nesfatin-1 immunoexpression in the lateral hypothalamic area, which is critically involved in appetite and metabolic regulation and has extensive connections with brainstem feeding circuits. Objective: The present study explored nucleobindin-2/nesfatin-1 localization pattern as well as the association between nucleobindin-2/nesfatin-1 protein expression and body weight in human brainstem nuclei. Methods: Sections of 20 human brainstems (13 males, 7 females; 8 normal weight, 6 overweight, 6 obese) were examined by means of immunohistochemistry and double immunofluorescence labeling. Results: Nucleobindin-2/nesfatin-1 widespread distribution was observed in various brainstem areas, including nuclei with well-defined roles in energy homeostasis and in autonomic and behavioral processes, such as the nucleus of the solitary tract, dorsal motor nucleus of vagus, area postrema, inferior olive, raphe nuclei, reticular formation, locus coeruleus, parabrachial nuclei, and pontine nuclei, and in Purkinje cells of the cerebellum. Interestingly, nucleobindin-2/nesfatin-1 immunofluorescence signal extensively localized in neuronal subpopulations expressing neuropeptide Y and cocaine- and amphetamine-regulated transcript (peptides known to exert potent actions on food intake and energy balance) in nucleus of the solitary tract, inferior olive, locus coeruleus, and dorsal raphe nucleus. Of note, nucleobindin-2/nesfatin-1 immunoexpression was significantly lower in obese than normal weight subjects in the nucleus of the solitary tract (p<0.05). Conclusions: These data provide for the first time neuroanatomical support for the potential role of nucleobindin-2/nesfatin-1 in human brainstem circuits controlling energy homeostasis. In nucleus of the solitary tract, a key integrator of nutrient state signals and a neural substrate of food reward-related processes, altered neurochemistry such as nucleobindin-2/nesfatin-1 deficiency may contribute to dysregulation of homeostatic and/or hedonic feeding behavior and ultimately to obesity.

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<![CDATA[MON-LB108 Measurement Of Carotid Intima,hepatic Steatosis And Inflammatory Markers In Obese Children]]> https://www.researchpad.co/article/elastic_article_8593 Measurement of carotid intima,hepatic steatosis and inflammatory markers in obese children. Elevated levels of inflammatory factors and increased mean intimal carotid thickness (IMT) would increase the risk of atherothrombotic events and contribute to the progression of cardiovascular disease in obese children. Objectives: Evaluate inflammatory factors, metabolic syndrome and non-alcoholic liver steatosis and carotid IMT as an early cardiovascular risk marker. Patients and methods: Descriptive cross-sectional exploratory study. Consider 41 obese children both sexes between 6- 12 years old. Evaluated: anthropometry and determinations of lipid and liver profile, blood glucose, insulin, HOMA, ultrasensitive CRP, fibrinogen. Hepatic ultrasound and measurement of carotid IMT with ESAOTE Mylab 50 Exdicion equipment. . Results: From 41 studied patients, 57% were female. 51% presented MS and 68% elevated triglycerides. CRP> 1 was found in 71% of cases. Hepatic steatosis was observed in 60%, which only 10% had altered transaminases. 12% presented high fibrinogen. Patients with MS had a significant positive difference in the IMTCC (X = 0.41 ± 0.12; p 0.024), HDL (X 37.89 ± 1.72; p 0.004) triglycerides (X 149.42 ± 10.69; p 0.002) in relation to patients without MS. Conclusion: CRP is an inflammatory risk factor associated with elevated BMI and MS. There was a higher prevalence of MS in our study. The increase in the average intimal thickness is significantly related to the presence of MS and RCP>1. The determination of marker molecules of an inflammatory state and measurement of carotid IMT would contribute to the implementation of strategies to prevent cardiovascular, hepatic and metabolic risk since childhood.

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<![CDATA[MON-590 Presence of Diabetes Diminishes the Ultimate Weight Loss After Bariatric Surgery]]> https://www.researchpad.co/article/elastic_article_8574 Background

Obesity and diabetes as well as their related complications result in both individual and global health burdens. Among patients who present with both obesity and diabetes, bariatric surgery can lead to remission of both these diseases. However, the possible impact of diabetes on the magnitude of weight loss outcomes after bariatric surgery has not been quantified.

Methods

To address this question, we extracted data from Michigan Bariatric Surgery Cohort (MI-BASiC) to see whether diabetes at baseline could be a predictor of weight loss outcomes. Consecutive patients 18 years of age or older undergoing gastric bypass (GB) or sleeve gastrectomy (SG) for obesity at the University of Michigan between January 2008 and November 2013 were included in our retrospective cohort. All patients had either body mass index (BMI) > 40 kg/m2 or BMI 35 – 39.9 kg/m2 with comorbid condition. Firstly, we performed Generalized Linear Mixed Model (GLMM) analysis to compare the probability of achieving BMI under 30kg/m2 or achieving excess body weight loss (EBWL) 50% or more for patients with or without diabetes. We then further tested the effect of presence of diabetes for the BMI outcomes across time using Linear Mixed Model (LMM) analysis. Finally, we conducted a LMM analysis to determine if diabetes is a predictor of the future weight loss, percentage of total weight loss or percentage of excess weight loss over 5 years of follow up.

Results

Based on our criteria, 380 patients were included for GB [female 305 (80.3%), mean age 43.6±0.6 years, mean BMI 47.3±0.4kg/m2, diabetes 149 (39.2%), on insulin 45 (11.8%)] and 334 for SG [female 259 (77.5%), mean age 45.3±0.6 years, mean BMI 49.9±0.5kg/m2, diabetes 108 (32.3%), on insulin 29(8.7%)]. From GLMM analysis, the presence of diabetes at baseline did not impact the probability of achieving BMI under 30kg/m2 (p=0.0848), but substantially impacted the probability of achieving 50% or more EBWL (p=0.0021) with individuals without diabetes having a 1.6 (odds ratio 1.56, 95% CL [1.18-2.08]) times higher chance to achieve this threshold. We also showed that individuals with diabetes at baseline had a significant effect to modify BMI points lost, regardless of the surgery type (p=0.0178). The presence of diabetes at baseline diminished weight loss by 1.2 BMI points (95% CL [0.21- 2.20]) which is roughly 10 to 15% of the total BMI points to be lost. LMM analysis further confirmed that after adjusting the time, surgery type, age, gender and baseline weight, there was still a significant difference of absolute weight loss (p=0.0110), percentage of total weight loss (p=0.0089) and percentage of excess weight loss (p=0.0365) between individuals with diabetes versus individuals without diabetes.

Conclusion

In conclusion, our data demonstrate that diabetes diminishes the ultimate weight loss effect of bariatric surgery. Further research is needed to understand why this is the case.

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<![CDATA[SAT-LB105 A Transcribed Ultraconserved Noncoding RNA, Uc.336-As, Promotes White to Brown Conversion in 3T3-L1 Cells]]> https://www.researchpad.co/article/elastic_article_8553 Brown adipose tissue (BAT) has gained its popularity since it shows great potential in counteracting obesity and metabolic diseases development. Transcribed ultraconserved regions (T-UCRs), a novel class of long non-coding RNA (lncRNAs), have been implicated in regulating diverse biological processes, including the process of white fat browning. However, the functional and mechanistic details of T-UCRs in the browning process are poorly understood. Here, we identified that a T-UCR, uc.336-as, played an important role during the browning process. Uc.336-as was significantly elevated during browning process induced by glucagon-like peptide-1 receptor agonist (exendin-4) or β3-adrenergic agonist (CL316,243). Overexpression of uc.336-as reduces the differentiation of 3T3-L1 preadipocytes into white adipocytes (inhibited lipid accumulation and decreased the expression of several adipogenesis markers) and induces brown characteristics during differentiation of 3T3-L1 preadipocytes (spurred browning adipocytes phenotypes and increased the expression of the browning associated genes). Moreover, we found that uc.336-as inhibited adipogenesis and promoted browning process via influencing the serine/threonine kinase (AKT)-mammalian target of rapamycin (mTOR) axis, an essential signal pathway in adipocyte metabolism. Taken together, our data show that uc.336-as acts as a negative regulator in white adipocyte differentiation and promotes the browning process, suggesting a potential therapeutic role for uc.336-as in controlling obesity.

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<![CDATA[MON-596 Effects of Angiotensin Type 1 Receptor Blockers (ARBs) on the Expression and Secretion of Adiponectin and Leptin in Human White Adipocytes]]> https://www.researchpad.co/article/elastic_article_8534 [Introduction]

Adiponectin and leptin are adipokines that are mainly produced in adipocytes and exert various functions. Adiponectin decreases atherosclerosis, oxidative stress, angiogenesis, inflammation, and apoptosis, whereas leptin works oppositely. Angiotensin type-1 receptor (AT1R) blockers (ARBs) are widely used as antihypertensive drugs. Some ARBs are known to activate peroxisome proliferator-activated receptor (PPAR) γ, which is a key regulator of fatty acid metabolism. It is reported that adiponectin secretion increases by pioglitazone, a full agonist of PPARγ, and some ARBs via PPAR γ activation. However, the effects of ARBs on leptin secretion are controversial. The present study aimed to examine the effects of ARBs on the expression and secretion of adiponectin and leptin in human white adipocytes.

[Materials and Methods]

Human white preadipocytes (Promo Cell) were differentiated into mature adipocytes in the medium containing insulin, dexamethasone, thyroxin and isobutylmethylxanthine. Pioglitazone and ARBs including telmisartan, irbesartan, azilsartan, candesartan, losartan, olmesartan and valsartan (1µM) were administered in the culture medium on day 4 and 8. The medium was collected on day 12 and the concentrations of adiponectin and leptin were measured by enzyme immunoassay. Real time PCR was performed to quantitate the mRNA expression of adiponectin and leptin in adipocytes. The experiments were performed in quadruplicate.

[Results]

Pioglitazone significantly increased adiponectin secretion (386.7 ± 133.7 vs. 7.3 ± 1.9 ng/ml in control) from human adipocytes. Among ARBs, adiponectin secretion significantly increased by telmisartan (136.7 ± 16.3 ng/ml) and irbesartan (69.7 ± 23.1 ng/ml), while the other 5 ARBs did not have any influence on adiponectin secretion. Real-time PCR also showed that mRNA expression increased 5.1-fold, 3.8-fold and 1.5-fold by pioglitazone, telmisartan and irbesartan, respectively. Leptin secretion significantly decreased by pioglitazone (27.7 ± 5.0 vs. 82.5 ± 3.8 ng/ml in control). Among ARBs, only telmisartan (38.7 ± 4.2 ng/ml) decreased leptin secretion. Real-time PCR also showed that mRNA expression decreased to be 0.5-fold and 0.7-fold by pioglitazone and telmisartan, respectively. GW9662, a selective antagonist of PPARγ, potently blocked pioglitazone-induced changes of adiponectin and leptin expression and secretion. On the other hand, GW9662 did not reverse telmisartan and irbesartan induced changes.

[Conclusion]

The changes in adiponectin and leptin secretion by pioglitazone are via PPARγ activation, while those by telmisartan and irbesartan may occur in PPARγ-independent manner.

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<![CDATA[SAT-598 Shared Signaling Profile Between Human MRAPα-Induced Human MC4R Constitutive Activity and Obesity-Associated Human MC4R Constitutive Activity]]> https://www.researchpad.co/article/elastic_article_7161 The human melanocortin 4 receptor (hMC4R) plays a critical role in the regulation of energy balance with more than 150 distinct human obesity-associated mutations. Most exhibit defective MC4R functionality but six have been reported to associate with constitutive activity. This represents a conundrum since a lean phenotype is expected for enhanced MC4R signaling. Human melanocortin 2 receptor accessory protein alpha (hMRAPα) induces hMC4R constitutive activity in transfected HEK293 cells (1,2). We do not know whether the hMRAPα-induced gain-in-function for hMC4R would cause, or prevent, obesity because of this conundrum. Here, we hypothesize that wild-type hMC4R, obesity-associated constitutively active hMC4R and hMRAPα-induced constitutive active hMC4R can exist in distinct conformational states and elicit distinct signaling profiles. To test this, we compared transiently expressed HA-hMC4R in HEK293 cells for basal and agonist activation for adenylyl cyclase, Cre driven β-galactosidase reporter transcription, and receptor protein expression. Six previously reported obesity-associated hMC4R constitutively active variants were compared with two hMC4R constitutively active mutations not associated with obesity, two hMC4R variants associated with protection from development of obesity, five non-constitutively active hMC4R mutations associated with obesity, hMRAPα co-expressed with hMC4R, and wild-type hMC4R. Our data confirm hMC4R constitutive activity coupling to both adenylyl cyclase and Cre β-galactosidase reporter for only two hMC4R variants associated with obesity (H76R & L250Q), one hMC4R mutation (H158R) not associated with obesity, and hMRAPα co-expressed with hMC4R. We show α-MSH stimulated concentration curves for wild-type hMC4R, H76R, L250Q & H158R hMC4R variants and hMRAPα co-expressed with hMC4R coupling to adenylyl cyclase. Surprisingly, out of these, only wild-type hMC4R and H158R hMC4R variant exhibited α-MSH-stimulated Cre β-galactosidase reporter concentration curves. Western blotting and ELISA showed ~70% reduced cell surface and total receptor protein expression for hMC4R co-expressed with hMRAPα and obesity-associated constitutively active hMC4R variants, compared to wild-type hMC4R. To summarize, two constitutively active hMC4R variants (H76R and L250Q) associated with obesity, and hMC4R co-expressed with hMRAPα, share a signaling profile comprising protein expression and α-MSH stimulated functional coupling to adenylyl cyclase and Cre-reporter gene expression. We conclude (1) if hMC4R is co-expressed with hMRAPα in vivo it would likely contribute to human obesity, and (2) obesity-associated constitutively active hMC4R variants exhibit a signaling anomaly that may underpin development of anti-obesity therapeutics.

1. Kay EI, et al. J Mol Endocrinol. 2013;50:203-215.

2. Kay EI, et al. PLoS ONE. 2015;10(10):e0140320.

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<![CDATA[SUN-591 PAPP-A Inhibition - a Novel Anti-Obesity Therapeutic Approach]]> https://www.researchpad.co/article/elastic_article_7101 Background: Adipose tissue is a heterogeneous endocrine organ with tremendous capability for expansion. The antithetical pathogenicity of visceral adipose tissue (VAT), compared to subcutaneous adipose tissue (SAT), has been linked to the metabolic stress of enlarging mature adipocytes and a limited ability to recruit new adipocytes. One of the major distinguishing features of VAT preadipocytes is the high expression of Pregnancy Associated Plasma Protein–A (PAPP-A) when compared to SAT. PAPP-A is a zinc metalloprotease that is secreted, and can associate with the cell surface in an autocrine or paracrine fashion. It is the only known physiological IGFBP-4 (Insulin-like Growth Factor Binding Protein) protease. It cleaves the IGF/IGFBP-4 complex, releasing IGF, making it more bio-available for receptor engagement and downstream signaling. The role of IGFs in adipogenic differentiation is well established. While there is quantitative depot-specific variability in PAPP-A expression among preadipocytes, mature adipocytes do not express any PAPP-A. These findings suggest that there may be a relationship between PAPP-A inhibition and adipogenic differentiation and maturation. Similar to human VAT, PAPP-A expression is highest in visceral fat in murine models. The PAPP-A KO mice, when fed a high fat diet, showed restrained visceral adiposity and decreased visceral adipocyte size, suggesting that PAPP-A could regulate adipogenesis locally in tissues that express high PAPP-A.

Hypothesis: PAPP-A inhibition is a novel anti-obesity treatment strategy. Methods/Results: We fed 20 male and 20 female wild type mice 42% high fat diet (HFD) starting at 10 weeks of age. Concomitantly, we treated 10 mice in each group with either mAb-PA1/41 (a PAPP-A neutralizing monoclonal antibody) or IgG2a (control isotope), intraperitoneally at a dose of 30 mg/kg weekly for the duration of the HFD. At the end of 15 weeks, the mice were sacrificed and the adipose tissue, serum and solid organs were harvested.

Compared to the control (IgG2a) mice, the mAb-PA1/41 treated male and female mice gained 40% less weight (P = 0.03) and had smaller visceral fat depots (mesenteric and pericardial). Also, when we looked at individual adipocyte size, the drug treated mice had 45% smaller mesenteric adipocytes (P = 0.002) and 44% smaller pericardial adipocytes (P= 0.003). Also, the visceral depots in the drug treated mice had 30% more cells (P = 0.006). In both groups, there was decreased liver lipid content (P=0.005). The mAb-PA1/41 treatment had no significant effect on subcutaneous fat depots.

Conclusion: Pharmacologic inhibition of PAPP-A decreased weight gain, visceral fat depot weight, visceral adipocyte size, hepatic lipid deposition and increased visceral adipocyte cell number in both male and female mice that were fed a high fat diet.

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<![CDATA[MON-600 Hydroethanolic Extract of Lampaya Medicinalis Phil. (Verbenaceae) Decreases Intracellular Triglycerides and Proinflammatory Marker Expression in Fatty Acid-Exposed HepG2 Hepatocytes]]> https://www.researchpad.co/article/elastic_article_7037 Background: Non-alcoholic fatty liver disease (NAFLD) is the most common hepatic chronic disease worldwide. NAFLD is characterized by an abnormal triglyceride (TG) accumulation (steatosis) in the liver, that may lead to hepatic inflammation (1). DGAT2 is a key enzyme that catalyzes the final step of TG synthesis and whose expression is elevated in NAFLD (2). FABP4 is a transporter of intracellular lipids and its levels are related with inflammation, characterized by a high expression of proinflammatory cytokines such as TNF-α, IL-6 and IL-1β. Palmitic acid (PA, C16:0) and oleic acid (OA; C18:1) are two of the most abundant fatty acids that participate in the formation of TGs in hepatic cells in vivo and in vitro (3). Lampaya medicinalis Phil. (Verbenaceae) is a small bush that grows in the “Puna atacameña” in the North of Chile. The infusion from leaves and aerial parts of the plant has been used by local ethnic groups to treat and cure inflammatory diseases (4). The aim of this study was to assess in vitro the effect of the hydroalcoholic extract of Lampaya medicinalis (HEL) against OA/PA-induced steatosis and proinflammatory marker expression in HepG2 hepatocytes.

Methods: HEL (0.01, 0.1, 1, 10 μg/mL) cytotoxicity potential (48 h) was evaluated by Trypan blue exclusion. Cells were exposed for 48 h to 1 mM OA/PA (2:1) in the presence or not of 0.01 or 10 μg/mL HEL. Intracellular TGs were assessed with Oil Red O staining and quantified with Nile Red reagent by fluorimetry. mRNA expression of DGAT2, TNF-α, IL-6 and IL-1β was evaluated by qRT-PCR. FABP4 content was assessed by Western blot. The levels of TNF-α and IL-6 in the culture media were analysed by ELISA.

Results: HEL was not cytotoxic at any concentration assessed (n=4; p>0.05). The increased content of TG induced by OA/PA was reduced in the presence of HEL (n=7; p<0.05), showing a strong consistency with Oil Red O staining. The increase in the protein content of FABP4 as well as the increment in mRNA expression of DGAT2, TNF-α, IL-6 and IL-1β induced by OA/PA were lower in cells co-exposed to HEL (n=6-9; p<0.05). The incubation with HEL+OA/PA reduced proinflammatory cytokine levels in culture media compared to cells exposed to OA/PA alone (n=6-7; p<0.05).

Conclusion: HEL reduces the OA/PA-induced increase in intracellular TG, DGAT2 and proinflammatory cytokine expression and FABP4 content, as well as the levels of secreted proinflammatory cytokines in HepG2 cells. These findings suggest a protective role for HEL against OA/PA-induced steatosis and inflammation, and therefore that Lampaya medicinalis may represent a promising therapeutic tool for pathologies such as NAFLD.

References: (1) Gluchowski L, et al. Nat Rev Gastroenterol Hepatol. 2017;14(6):343-355. (2) Perry, et al. Nature. 2014;510(7503):84-91. (3) Cunningham P, et al. J Nutr. 2009;139(4):636–639. (4) Morales G, et al. Biol Res. 2014;47:6.

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<![CDATA[OR33-07 ARNT2: A Potential Novel Candidate Gene for Monogenic Obesity in Humans]]> https://www.researchpad.co/article/elastic_article_7007 Introduction: Aryl hydrocarbon nuclear translocator 2 (ARNT2) is a basic helix-loop-helix (bHLH)-PAS (Per/Arnt/Sim) transcription factor shown to be critical to the development of paraventricular nucleus of the hypothalamus (PVN), key region for energy homeostasis and feeding response. In vivo and in vitro studies have shown that ARNT2 is an obligate heterodimer for SIM1, known cause of monogenic obesity. Null mutations in Arnt2 in animals are not viable, but hypomorphic mutation results in hyperphagic obesity and its associated consequences (1). Due to the critical role of ARNT2 in the development of PVN, we hypothesize that hypomorphic mutations may result in early onset obesity in humans.

Methods: The Genetics of Early Childhood Obesity (GECO) study recruits children with severe obesity (BMI > 120% of 95th percentile) of early onset (< 6 years). Whole exome sequencing (WES) was performed in a subset of proband-parent trios. The functional validation of the mutation(s) in ARNT2 is ongoing with co-transfection of tagged Arnt2 and Sim1 in HEK293 cells, with the induction of a luciferase reporter gene under the control of 6 repeats of bHLH-PAS core binding element by the Arnt2-Sim1 complex.

Results: Two adolescents from unrelated families were found to have genetic variants in ARNT2. Subject 1 has a novel de novo heterozygous coding variant in ARNT2, c.388 C>G (p.P130A, CADD 25), predicted to be deleterious by 8/12 in silico algorithms. She is a 14-year old Caucasian girl with severe early onset obesity, BMI 28.1 kg/m2 (BMIz +4.72) at 2.5 years of age that has increased to 53.54 kg/m2 (BMIz + 3.25) at 14-years, and height > 95th %tile. She is non-dysmorphic, has developmental delay, absence seizures, behavior abnormalities & glucose intolerance/dyslipidemia secondary to obesity. Using genematcher, we identified another proband with the phenotype of obesity: an African American girl (BMIz +1.9) with biallelic inherited heterozygous variants in ARNT2, c.1228T>A (p.W410R, CADD 29) and c.916G>A (p.G306S, CADD 22). An only child conceived by IVF, she is non-dysmorphic and on treatment for bilateral focal epilepsy. All 3 variants are rare, with mean allele frequency < 0.005 in population-based databases such as gNOMAD. Both the patients have early onset obesity and a significant neurological phenotype. ARNT2 is a highly constrained gene of 717 amino acids with a significant depletion of missense variants in the N-terminus (1-244 aa) and overall fewer loss of function variants in ~282,644 alleles sequenced in gNOMAD.

Conclusions: We propose that hypomorphic mutations in ARNT2 could be a potential novel cause of monogenic obesity in humans. Future studies will investigate the molecular mechanisms causing weight dysregulation in patient specific disease relevant hypothalamic neurons.

Reference: (1) Turer et al., Dis Model Mech. 2018; 11(12)

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<![CDATA[MON-588 Anthropometric Parameters, Body Fat Percentage and Metabolic Profile in Sarcopenic Women with Recommendation for Bariatric Surgery]]> https://www.researchpad.co/article/elastic_article_6939 INTRODUCTION: Sarcopenia (SARC) is a musculoskeletal disorder that predisposes several complications, including metabolic ones. Obesity also provides higher risk for metabolic complications, however, there is lack of evidences regarding the association of obesity with SARC on metabolic parameters in non-elderly individuals. OBJECTIVE: To evaluate anthropometric parameters, body fat percentage (BFP) and metabolic parameters in women with and without SARC preceding Bariatric Surgery (BS). METHODS: A cross-sectional study involving 60 obese women in the outpatient care in a public Brazilian University Hospital between March to September 2018. Body composition was given by bio-impedance (inbody-370), multifrequency (5, 50, 250Hz) with 12 hours fasting, dominant Handgrip Strength (HS) was evaluated by Jamar dynamometer (3 measurements; 30 sec interval). Were also evaluated fasting blood glucose, HbA1c, homeostatic model assessment-insulin resistance (HOMA-IR), total cholesterol (TC), low density lipoprotein (LDL), high density lipoprotein (HDL), triglycerides and high-sensitive C-reactive protein (hs-CRP). SARC was defined by the association of a low muscle mass index (weight-adjusted appendicular skeletal muscle mass: ASMM/weight x 100%) and decreased HS, using as cutoff points the smallest quintile for each variable. Data were expressed as mean ± standard deviation and independent t-test was used for comparison between groups. Statistics were made by SPSS software, 20th version (IBM Corp., Armonk, NY). RESULTS: The mean age, weight, body mass index and BFP of sarcopenic and non-sarcopenic women were: 40.75 ± 11 x 39.23 ± 8.92 years old (p=0.665), 102.93 ± 9.58 x 109.19 ± 14.25 Kg (p=0.237), 44.88 ± 2.7 x 42.24 ± 4.79 Kg/m2 and 54.12 ± 1.11 x 51.44 ± 3.43% (p=0.052), respectively. Regarding the laboratory parameters of women with and without SARC: fasting blood glucose 89.25 ± 14.48 x 98.40 ± 27.48 mg/dL (p=0.359), HbA1c 5.83 ± 0.33 x 6.21 ± 1.18% (p=0.185), HOMA-IR 3.61 ± 1.28 x 5.31 ± 4.74 (p=0.160), TC 170.87 ± 39.36 x 180.82 ± 34.51 mg/dL, LDL 94.67 ± 26.63 x 105.60 ± 30.85 mg/dL, HDL 53.37 ± 18.50 x 50.84 ± 10.32 mg/dL, triglycerides 114.12 ± 38.84 x 127.30 ± 75.04 mg/dL and hs-CRP 8.51 ± 6.50 x 7.51 ±6.52 mg/L (p=0.792). CONCLUSION: Women with SARC and recommendation for BS when compared to women without SARC had similar anthropometric, metabolic and BFP parameters.

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<![CDATA[OR04-04 Identification of a Novel Transcriptional Regulator of Metabolic Disease in Circulating and Central Myeloid Cells]]> https://www.researchpad.co/article/elastic_article_6924 Derangement in systemic metabolic homeostasis is tightly associated with widespread activation of resident and circulating immune cells, a phenomenon known as ‘metaflammation’. Numerous studies have explored the role of tissue resident and circulating macrophages in contributing to metaflammation, obesity, and their sequelae; however, there is a dearth of information regarding targetable transcriptional regulators of the genesis and persistence of metabolic disease. Here, we identify myeloid Krüppel-like factor 2 (KLF2) as a novel regulator of metabolic disease. Previous reports demonstrate that KLF2 serves as a critical regulator of myeloid cell quiescence and is downregulated in numerous acute and chronic inflammatory states. Specifically in the context of chronic metaflammation, we note that KLF2 expression is decreased in circulating immune cells of obese patients and in adipose tissue macrophages of high fat diet (HFD) fed mice, which is consistent with the hypothesis that KLF2 regulates metaflammation. To explore this further, we utilized mice with myeloid cell-specific deletion of KLF2 (K2KO) which exhibit accelerated obesity and insulin resistance. K2KO mice have widespread central (i.e. CNS) and peripheral metaflammation both in the basal and HFD-stimulated states. To discern whether the effect of myeloid deletion of KLF2 on metabolism is due to deletion in microglia in the feeding centers of the hypothalamus or in peripheral immune cells, bone marrow chimeras with head shielding were created. 50% reconstitution of circulating immune cells with K2KO cells in wildtype (WT) mice was sufficient to maintain the metabolic disease phenotype, while mice with K2KO microglia + WT circulating cells had only slightly improved outcomes compared to K2KO mice. Conversely, ablation of microglia in K2KO mice using PLX5622 formulated in HFD also successfully attenuated the aberrant feeding behavior, weight gain, and glucose dyshomeostasis seen in K2KO mice. Together, these data demonstrate a role for loss of KLF2 in hematopoietic and CNS resident cells in causing metabolic disease. Given that myeloid KLF2 expression decreases under metabolic stress in WT mice and humans, we sought to explore whether maintenance of KLF2 expression in these cells would be protective against diet-induced metabolic disease. Indeed, mice with myeloid-specific overexpression of KLF2 demonstrated a markedly improved metabolic phenotype when challenged with HFD, providing evidence that targeting KLF2 expression in myeloid cells may prove to be a therapeutic option against metaflammation.

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<![CDATA[SAT-595 Early Weight Gain Impact onto the Arcuate Leptin Receptor Expressing Neurons Activity During Development]]> https://www.researchpad.co/article/elastic_article_6897 The arcuate nucleus (ARH) is considered the main mediator of the effects of leptin on energy homeostasis, as well, part of the hypothalamic circuitry through which the sex steroids mediate the ovulatory cycle and therefore reproduction. While it is known that aging influences synaptic plasticity of ARH neurons, the effects of increased weight gain early in life onto in the activity of ARH neurons are still poorly understood. In order to demonstrate whether prepubertal adiposity gain is able to modulate the synaptic transmission in leptin receptor (LepR)-expressing cells of the ARH, we employed a postnatal over-nutrition model by raising mice in small litters (SL), while control mice were maintained in regular litters (6-8 pups per litter). The spontaneous currents of LepR-expressing neurons in the ARH were measured by whole-cell patch-clamp recordings in hypothalamic slices obtained from prepubertal, pubertal and adult female LepR-reporter mice. As expected, mice raised in SL exhibited increased body weight compared to control mice at prepubertal and pubertal stage (prepubertal, t(59) = 6.9, P < 0.001; pubertal, t(59) = 5.3, P < 0.001), despite no difference at adult age (t(59) = 1.5, P = 0.4). By evaluating ARH neuronal activity we observed an increased average of excitatory and inhibitory currents frequency during development, in both control females (sEPSC: F(2, 46)= 17.76, P< 0.0001, sIPSC: F(2, 21)= 4.064, P= 0.0322) and in mice raised in SL (sEPSC: F(2, 49)= 27.76, P< 0.0001, sIPSC: F(2, 19)= 6.714, P= 0.0062). However, SL and control mice exhibited similar sEPSC amplitudes at all stages of development. A significant interaction between litter size and excitatory transmission frequency onto LepR-expressing cells were noted at the pubertal and adult stages (F(2, 95)= 3.164, P= 0.046), despite no changes in the amplitudes of these signals (interaction: F(2, 95) = 0.1516, P= 0.8596; age: F(2, 95) = 0.9961, P= 0.3731; litter size: F(1, 95) = 0.076, P= 0.7832). By evaluating the inhibitory transmission to ARH LepR-expressing neurons, no significant interaction between litter size and inhibitory transmission frequency were observed (F(2, 40) = 0.09271, P= 0.9117). However, the average sIPSC amplitude were significantly reduced in the ARH cells recorded from mice raised in SL, when compared to the control group (F(2, 42) = 22.86, P< 0.0001), despite no identifiable interaction between litter size and inhibitory transmission (F(2, 42) = 0.6619, P= 0.5212). Taken together our results suggest that early weight gain influences the excitatory transmission pattern in LepR-expressing neurons by increasing presynaptic excitatory inputs and suppressing postsynaptically transmission to ARH neurons.

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<![CDATA[OR33-02 The Treatment of Partial Lipodystrophy in the Setting of Neutralizing Antibody Against Metreleptin with Leptin Receptor Agonist REGN4461]]> https://www.researchpad.co/article/elastic_article_6892 Background: An 18-year-old patient with atypical partial lipodystrophy had a transient initial metabolic response to metreleptin that deteriorated when neutralizing antibodies against metreleptin developed. A therapeutic trial with setmelanotide did not result in any metabolic benefit as desired. Because her status continued to deteriorate, we attempted to treat her with REGN4461, a fully human monoclonal antibody that is an agonist to the human leptin receptor (LEPR). Clinical Case: A compassionate use protocol (IND No. 144013) was initiated to treat the patient with REGN4461. The treatment consisted of 5 mg/kg intravenous infusion followed by 300 mg weekly subcutaneous injections of REGN4461. The primary endpoint was achievement of fasting triglycerides < 500 mg/dL without the need for ongoing plasmapheresis. Treatment-emergent adverse events were also followed. Here, we report her first 21-week response to treatment with REGN4461. The treatment resulted in a reduction of triglycerides from 1288 mg/dL to 344 mg/dl and allowed her to discontinue plasmapheresis. She lost 3.4 kilograms so far, and her liver size reduced per liver span measured by physical examination. Also, the liver MRI at week 12 showed a significant reduction in liver size and fat content (from 29.9% to 16.6%). Although her glucose control is still challenging, a slight reduction in her HbA1c was observed at week 12 along with improvements in her average glucose levels and total daily insulin requirement so far. No untoward signals were detected in her safety measurements. Conclusion: To date, treatment with REGN4461 offered substantial clinical benefit by improving the metabolic abnormalities in this unique patient. This experience represents the longest human exposure with REGN4461. The improvements noted in metabolic parameters and hepatic steatosis are similar to previous observations in lipodystrophic humanized LEPR mice. Our results suggest that REGN4461 showed clinical benefit even in the presence of neutralizing antibodies to metreleptin.

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<![CDATA[SAT-LB103 Glucose-Dependent Insulinotropic Polypeptide Promotes Proliferation, Inhibits Apoptosis and Modifies Adipogenesis in Human Omental Adipose-Derived Stem Cells]]> https://www.researchpad.co/article/elastic_article_6881 Increased visceral fat correlates with a high risk of morbidity and mortality from diabetes and other metabolic diseases. To cope with changes of nutritional status, the adipose tissue undergoes dynamic remodeling, during which adipose derived stem cells (ADSCs) participate through cell proliferation and adipogenic differentiation into mature adipocytes. Besides, beige adipocytes formation from ADSCs, to dissipate energy as heat in mitochondrial via uncoupling protein1 (UCP1) has been proved to improve energy expenditure. Thus, modifying adipose remodeling and promoting beige adipogenesis of ADSCs in visceral fat bring much metabolic benefits. Newly listed LY3298176, an agonist targeted on glucose-dependent insulinotropic polypeptide (GIP) /glucagon-like peptide-1 (GLP-1) receptor, shows outstanding effect of reducing glucose and weight. Due to superior efficacy in dual-target agonist to GLP-1 monotherapy, and the unknown role of GIP in human visceral adipose, we aimed to clarify GIP’s role in undifferentiated ADSCs in vivo. We selected cell model derived from abdominal omental adipose tissue by obtaining ADSCs via primary culture from patients, because of wide-distributed GIP receptors in fat, and the dominant role of abdominal fat in metabolism. Then the cells were allowed to proliferate, or differentiate into adipocytes in the differentiation medium (DM), with or without co-treated with GIP or GIP3-42 (GIP receptor antagonist), followed by subsequently measurement. CCK-8, EdU incorporation, and cell cycle analysis were conducted to assess cellular proliferation. Annexin V FITC/PI stain, TUNEL and cleaved caspase3 detection were performed to evaluate apoptosis. The related signaling pathway was measured by Western blot and the validation was conducted by using pathway inhibitors followed with the above proliferation and apoptosis analysis. Besides, at the early stage of adipogenesis, mitotic clonal expansion (MCE) was reflected by cell cycle detection. Western blot analysis, quantitative real time-PCR (qRT-PCR), and Oil Red O staining were performed to evaluate adipogenesis. We found that GIP facilitated ADSCs viability and DNA synthesis, accelerated cell cycle progress and reduced palmitate-induced apoptosis by promoting phosphorylation of ERK1/2, AKT, PKA and AMPK. We further confirmed that ADSCs after confluence underwent MCE once induced by DM. GIP also modified adipogenesis by accelerating MCE, upregulating core transcription factor (PPARγ and C/EBPα), increasing beige-related markers (UCP1, PGC1α, PRDM16, et al) while suppressing white-related genes (ZFP423 and TLE3). In summary, we illustrated the efficacies of GIP on proliferation, apoptosis and adipogenesis (especially the beige adipocyte formation) of ADSCs, providing evidence of the additional metabolic benefits of GIP/GLP-1 dual-target agonist over GLP-1 agonist monotherapy in vivo.

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<![CDATA[SAT-587 Molecular Markers of Beige Adipose Following ESR1 Knockdown in the Mediobasal Hypothalamus of Adult Female Rhesus Monkeys]]> https://www.researchpad.co/article/elastic_article_6873 Our studies in female marmoset monkeys show that the ablation of ovarian estradiol (E2) production fails to alter energy homeostasis or body fat accumulation. Peripheral E2 may therefore not play a crucial role in metabolic regulation in female primates. shRNA-mediated knockdown of ESR1 expression in the hypothalamic ventromedial nucleus (VMN) in adult female rodents, however, induces obesity and suggests ESR1 is a hypothalamic target for E2 regulation of energy homeostasis, and likely mediates thermogenesis in brown/beige adipose depots. In female primates, including humans, the hypothalamic estrogen receptor mediating metabolic regulation is unknown. To test the hypothesis that ESR1 mediates female primate regulation of energy homeostasis, 11 ovary intact, adult female rhesus macaques, pair housed with female peers, received five 12µl MRI-guided MBH infusions into the rostral-to-caudal extent of both right and left VMN. Each infusion comprised a gadolinium contrast agent and ~3–4 x 1010 adeno-associated virus 8 (AAV8) particles containing either an shRNA specific for ESR1 (n=6, ERaKD) or scrambled shRNA (n=5, control). Mid-surgery MRI scans identified targeting accuracy. ~ 1.5 yrs following AAV8 infusion, pronounced gain in BMI enabled conversion of 83% of ERaKD females to overweight/obese compared to 20% of controls (p=0.08). Percent increase in BMI remained intermittently greater (p<0.05) than controls thereafter. Adipose depots were harvested at necropsy ~2.5–3 yrs following treatment. Total RNA was isolated using the Qiagen AllPrep DNA/RNA/miRNA Universal kit. RNA was reverse transcribed with High-Capacity cDNA Reverse Transcription kit (Applied Biosystems). All quantitative real-time PCR (qRT-PCR) were performed on a StepOnePlus System using Power SYBR Green master mix (Applied Biosystems). Primer sequences were designed using NCBI Primer-Blast. Expression of TATA-box binding protein (TBP) was used as the internal control housekeeping gene. The relative expression of target genes was measured using the comparative cycle threshold (Ct) method with results expressed as target mRNA expression relative to TBP using the formula 2^-delta Ct. Upper body beige adipose represents an organ system in primates, including humans, involved in thermogenesis. Axillary beige adipose depots in ERaKD females, however, did not exhibit significantly diminished gene expression for selected markers of beige adipocytes, including PAT2, CD137 and C/EBPβ, compared to control females. More crucially, thermogenically relevant UCP1 expression also did not differ between ERaKD females and controls. Taken together, these results suggest that knockdown of VMN ESR1 in adult female monkeys, while inducing modest weight gain after 1.5 years, may not markedly alter beige adipose gene expression of initially selected thermogenically relevant genes.

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<![CDATA[SAT-593 Sex-Specific Modifications in MicroRNAs Contained in Exosomes of Astrocytes in Response to Palmitic Acid]]> https://www.researchpad.co/article/elastic_article_6768 Communication between astrocytes and neurons is fundamental for correct functioning of the brain, in both physiological and pathophysiological situations. It is clear that astrocytes play an active role in metabolic control, but much is yet to be learned regarding how these glial cells and the neurons involved in energy intake/expenditure communicate to regulate energy homeostasis. We hypothesized that miRNAs contained in exosomes are an important means of cross-talk between these cells. Our objectives here were to determine whether the miRNA content of exosomes released by hypothalamic astrocytes changes in function of nutrient signals and if these signals are similar between males and females. To this end, primary hypothalamic astrocyte cultures were prepared from 2-day old male and female mice, using a standard protocol, and treated with palmitic acid (PA, 0.5 mM) or vehicle for 24 hours. The exosomes secreted into the culture media were purified and next generation sequencing analysis of the miRNAs contained in these microvesicles performed. Over 200 known miRNAs were identified in the samples. Heat map analysis of the 50 miRNAs most highly expressed across all samples showed significant differences based on sex and PA exposure, as well as differential changes between sexes in response to PA. Of the 25 most highly expressed miRNAs, 24 were significantly different between males and females (Benjamini-Hochberg FDR corrected p-values, between p<0.05 and p<0.0001). In response to PA, 190 miRNAs changed significantly in female astrocytes, but only 92 in male astrocytes; hence, after exposure to PA, 59 miRNAs were identified to be differentially expressed in exosomes of male and female astrocytes. Gene ontology enrichment analysis indicated that modifications in the miRNAs identified here could be related to biological processes such as response to cell injury, as might be expected, but also protein polymerization, receptor trafficking, intracellular signaling, microtubule polymerization, vasodilation, and cytoskeleton organization. Our results suggest that astrocytes communicate changes in nutrient availability to other cell types through miRNAs. Verification and determination of the specific responses to the modification in these miRNAs are now necessary.

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