bluuebunny/crossref_metadata_2025_split huggingface dataset:
SUN-254 Angiotensin II Stimulates Microglia Cell Inflammatory Responses
Authors: Velez Jean Paul Moliere , Inostroza-Nieves Dra Yaritza , Romero Jose R , Arenas Claudia , Capo Diego 
Published: May 2020

Abstract
Angiotensin II (AngII) is the principal effector molecule of the renin-angiotensin system (RAS). It’s effects on the cardiovascular and renal system are well-documented. AngII acts mainly via interaction with the AngII type-1 receptor (AT1R). Disordered levels of AngII lead to hypertension and cardiovascular disease. Increasing evidence suggests that AngII may also play a role in the pathophysiology of neurodegenerative diseases through unclear mechanisms. We investigated AngII, AT1R and AT2R levels in a mouse model of neurodegenerative disease, the experimentally induced autoimmune encephalomyelitis (EAE) mouse. In EAE mice, AngII and AT1R gene expression in brain tissue were significantly increased when compared to control mice (3.2 folds ±1.9, p&lt;0.05, n=5; and 2.6 folds ±1.1, p&lt;0.01, n=5 respectively). In addition, iNOS mRNA expression by qRT-PCR was likewise upregulated in EAE mice compared to control (3.4 ± 1.4 folds, p&lt;0.01, n=5). We then studied the effects of AngII in human microglial cells (HMC3) -resident innate immune cells of the central nervous system (CNS). In HMC3 cells, treatment with AngII up-regulated the expression IL-6 (3.9 folds ± 1.2, p&lt;0.01, n=4) and increased IL-6 concentration by 83% (p&lt;0.05, n=4) by ELISA; effects that were blocked by the AT1R antagonist, Losartan. Also, AngII induced TNF-α production, increasing its concentration by 90% (p&lt;0.05, n=4), an increase that was blocked by Losartan. We also quantified Nitric Oxide (NO) production by using Griess Reagent and reactive oxygen species (ROS) production by the MUSE Oxidative Stress assay. In these cells, NO and ROS production were significantly increased by AngII (p&lt;0.05, n=4) and treatment with Losartan reduced their production (p&lt;0.05, n=4). In addition, AngII treatment induced iNOS overexpression (2.5 folds ±0.8, p&lt;0.05, n=4); results that are consistent with increases in the EAE mice. These data suggest that AngII can activate microglia cell inflammatory responses and as such may contribute to the pathophysiology of CNS inflammation and neurodegenerative diseases.
MON-709 Single-Cell RNA-Sequencing Deciphers POMC Neuron Destiny
Authors: Yu Hui , Jones Graham , Thompson Zoe , Rubinstein Marcelo , Low Malcolm James 
Published: May 2020

Abstract
The hypothalamus is one of the critical brain nodes regulating body weight and energy homeostasis. Within this node, Pomc neurons sense nutrient and hormonal signals to release melanocortin peptides that induce satiety, whereas AgRP/NPY neurons exert opposite effects by releasing AGRP that promotes feeding. Immature neurons in the hypothalamic ventricular zone start to express Pomc at E10.5, reach a maximum number at E14.5 and then decrease to stabilize at E18.5. However, it remains elusive how Pomc expressing precursors adopt their final cell fates. Therefore, the goal of this study was to decipher the temporal sequence of transcription factor (TF) expression leading to the terminal differentiation of POMC neurons. Red fluorescent cells collected from dissociated hypothalami of Pomc-tDimer-dsRed mice at six critical developmental time points - E11.5, E13.5, E15.5, E17.5, P5 and P12- were FACS sorted for the 10X genomics scRNA-seq pipeline. Unsupervised cell clustering identified 11 distinct clusters based on their transcriptional profiles. Eight of the clusters were highly-enriched for neuronal signature genes and were further characterized based on their transcript levels for Pomc (high, medium or low) and other distinct feature genes. Cells in the Pomchigh cluster expressed genes identified previously to modulate Pomc expression, including Isl1, Nkx2-1, and Tbx3, together with several novel candidate TFs. Unexpectedly, Nr5a1, the ventromedial hypothalamic nucleus marker gene encoding SF1, was highly expressed in the Pomchigh cluster at early stages. One of the Pomclow clusters highly expressed Otp, Agrp, Npy, Sst and Calcr while a second was highly enriched with Tac2, Kiss1, Pdyn, Prlr, Ar and Esr1 transcripts. All the clusters showed direct correlations of embryonic stage with the expression of progressively more mature markers of differentiation, thereby extending previous reports of these clusters based on single time points. Moreover, our results uncovered five novel Pomc neuron clusters with unique patterns of TF gene expression. For comparison of these data to the adult hypothalamus, we performed a TRAP-Seq study using PomcCreERT,Rosa26eGFP-L10a mice. Prdm12 and Tbx3 were among the most highly differentially expressed TFs in the POMC neuron affinity purified translatome. Similarly, Cited1, Npy2r, and Asb4 were highly expressed in both the Pomchigh cluster and the TRAP-Seq derived POMC translatome. This comprehensive molecular characterization of POMC cells during development sheds new light on the molecular diversification of early POMC neuron precursors and provides a valuable resource for elucidating the regulatory mechanisms defining POMC neuron subgroups in the hypothalamus.
MON-535 Deep-Machine Learning for Objective Quantification of Nerves in Immunohistochemistry Specimens of Thyroid Cancer
Authors: Astono Indriani , Rowe Christopher W , Welsh James , Jobling Phillip 
Published: May 2020

Abstract
Introduction: Nerves in the cancer microenvironment have prognostic significance, and nerve-cancer crosstalk may contribute to tumour progression, but the role of nerves in thyroid cancer is not known (1). Reproducible techniques to quantify innervation are lacking, with reliance on manual counting or basic single-parameter digital quantification.
Aims: To determine if a deep machine learning algorithm could objectively quantify nerves in a digital histological dataset of thyroid cancers immunostained for the specific pan-neuronal marker PGP9.5.
Methods: A training dataset of 30 digitised papillary thyroid cancer immunohistochemistry slides were manually screened for PGP9.5 positive nerves, annotated using QuPath (2). 1500 true positive nerves were identified. This dataset was used to train the deep-learning algorithm. First, a colour filter identified pixels positive for PGP9.5 (Model 1). Then, a manually tuned colour filter and clustering method identified Regions of Interest (ROIs): clusters of PGP9.5 positive pixels that may represent nerves (Model 2). These ROIs were classified by the deep learning model (Model 3), based on a Convolutional Neural Network with approximately 2.7 million trainable parameters. The full model was run on a testing dataset of thyroid cancer slides (n=5), containing 7-35 manually identified nerves per slide. Model predictions were validated by human assessment of a random subset of 100 ROIs. The code was written in Python and the model was developed in Keras.
Results: Model 2 (colour filter + clustering only) identified median 2247 ROIs per slide (range 349-4748), which included 94% of the manually identified nerves. However, most Model 2 ROIs were false positives (FP) (median 85% FP, range 68-95%), indicating that Model 2 was sensitive but poorly specific for nerve identification. Model 3 (deep learning) identified fewer ROIs per slide (median 1068, range 150-3091), but still correctly identified 94% of manually annotated nerves. Of the additionally detected ROIs in Model 3, median FP rate was 35%. However, in slides where higher non-specific immunostaining was present, then the number of FP ROIs was &gt;90%.
Conclusion: Simple image analysis based on colour filtration/cluster analysis does not accurately identify immunohistochemically labelled nerves in thyroid cancers. Addition of deep-learning improves sensitivity with acceptable specificity, and significantly increases the number of true positive nerves detected compared to manual counting. However, the current deep learning model lacks specificity in the setting of non-specific immunostaining, which is a basis for improving further iterations of this model to facilitate study of the significance of innervation of thyroid and other cancers.
References: (1) Faulkner et al. Cancer Discovery (2019) doi: 10.1158/2159-8290.CD-18-1398. (2) Bankhead P et al. Sci Rep 2017;7(1):16878.
SAT-588 Role of Adipose Tissue in Reproduction, Mammary Gland Development, Function and Cancer
Authors: Brenot Audrey , Hutson Irina , Harris Charles Andrew 
Published: May 2020

Abstract
To investigate the role of adipose tissue in the function of the mammary gland (MG) and reproductive system, we have examined lipodystrophic (LD) mice. LD mice of both sexes are sterile, but fertility was restored in both sexes with leptin injections. In addition, leptin was only needed for initial stages of pregnancy and not for parturition. A transplant of mouse embryonic fibroblasts (MEFs) led to the formation of an ectopic fat pad which also rescued the fertility in both sexes. However, pups born to rescued LD mothers died shortly after birth. We therefore examined the mammary glands of these mothers. MGs from LD mice were rudimentary and lacked terminal end buds. Leptin-injected and MEF rescued LD mice were able to become pregnant, showed normal pregnancy-associated glandular proliferation despite a smaller glandular area, were able to produce a small amount of milk that had grossly normal content of milk proteins and neutral lipids, but could not sustain pups to weaning. In order to separate the individual requirements for 1) adipokines such as leptin, 2) estradiol, and 3) epithelial-adipocyte interactions, we performed a series of experiments with both LD and ob (leptin-deficient) mice that received either estradiol or preadipocyte transplant. The resulting fat pad did not rescue the defect in MG development in LD mice. The defect also was not rescued with estradiol pellets. Ob/ob mice, like LD mice, lack leptin and estradiol, but retain adipose tissue. Ob mice have defective MG development. However, in striking contrast to LD mice, reconstitution of a WT fat pad in ob mice rescued the defect in MG development. Estradiol treatment did not rescue MG development in ob mice. Therefore direct interaction between mammary gland epithelia and adipocytes is a requirement for full invasion and expansion of the gland during puberty, but is not required for glandular proliferation during pregnancy and milk production.
Given that excess adipose tissue is a risk factor for breast cancer we wanted to determine if breast cancer was affected by the absence of adipose tissue. LD mice were bred to MMTV-PyMT mice that develop spontaneous breast cancer. Remarkably, LD PyMT+ mice had accelerated growth of primary tumors compared to WT PyMT+ mice. Using our MEF transplant model future studies will be directed to understanding whether the accelerated breast cancer growth is due to loss of adipokines or altered epithelial-stromal interactions.
SAT-463 Thyrotoxicosis from Nivolumab in a Patient with Preexisting Graves’ Disease
Authors: Polavarapu Preethi , Akkireddy Padmaja 
Published: May 2020

Abstract
Introduction
Thyroid dysfunction is one of the common immune-related adverse events associated with immune checkpoint inhibitors like Nivolumab. Thyroiditis or primary hypothyroidism is the most commonly reported presentation. Graves’ disease is less frequently reported. We report a case of preexisting Graves’ disease patient, on antithyroid meds who developed thyrotoxicosis followed by hypothyroidism after receiving Nivolumab therapy.
Case
66 y/o female patient with newly diagnosed metastatic melanoma presented to us for evaluation of abnormal thyroid test after her second cycle of Nivolumab. She has a long-standing history of Graves’ disease and has been on methimazole since her diagnosis. Her baseline thyroid labs before the start of Nivolumab were within normal limits (on methimazole 2.5 mg daily). She presented with weight loss, palpitations, and tremors four weeks after the start of Nivolumab. On exam, she was tachycardic with tremors noted to outstretched hands and had diffusely enlarged thyroid. Repeat lab work done before her second cycle revealed suppressed TSH 0.02 (0.4-4.5 uIU/ml) with elevated free T4 and T3. Her TSI titers were elevated. Methimazole dose was increased to 10 mg daily, and follow up labs done in a month revealed TSH of 89 uIU/ml, Free T4 0.16 (0.76-1.8 ng/dl). Methimazole was completely stopped at this time. She continued to have elevated TSH despite being off of methimazole for more than a month, concerning for the development of hypothyroidism. She was started on levothyroxine, after which labs returned to normal. The patient continued on immunotherapy during this period.
Discussion
Immune checkpoint inhibitors have been increasingly used for cancer therapy. Endocrinopathies are the most common immune-related adverse events associated with the use of these agents, with thyroid dysfunction being more common. Our patient had well-controlled Graves’ disease and was on a stable dose of methimazole for years. She developed autoimmune thyroiditis four weeks after receiving immunotherapy and subsequently developed hypothyroidism. The literature search did not reveal cases of autoimmune thyroiditis in a patient with preexisting Graves’ disease. One study reported that the timeline for developing the thyrotoxic phase is five weeks, which is followed by the rapid development of either euthyroid or hypothyroid phase. Management during the thyrotoxic phase is usually beta-blockers. Current guidelines recommend checking thyroid function test before initiation of therapy and every two weeks after the diagnosis of thyrotoxicosis until they become euthyroid or hypothyroid.
Our case illustrates that patients with preexisting Graves’ disease can develop thyroiditis after receiving immune checkpoint inhibitors, and hence, frequent monitoring with thyroid function tests is needed.
SAT-632 Involvement of NF-κB-p65 in BAG3 Regulation After Stress Stimuli
Authors: Dhanabalan Karthik , Singh Madhu V , Wong Thomas , Dokun Ayotunde O 
Published: May 2020

Abstract
We previously identified the limb salvage QTL1 (LSQ-1) on mouse chromosome 7 as a locus that offered protection against ischemic injury following induction of hind limb ischemia (HLI) a model of experimental peripheral arterial disease (PAD) in mice. To better understand the role of the LSQ-1 locus in post ischemic adaptation we characterized several genes within this locus and identified a number of genes that were important in tissue adaptation to ischemia, including BCL2-associated athanogene 3 (BAG3). BAG3 is an anti-apoptotic protein that plays an important role in cell survival through the regulation of autophagy. BAG3 expression is induced in the gastrocnemius muscles of mice after hind limb ischemia but how ischemia regulates BAG3 expression is poorly understood. Additionally, the activation of NF-κB transcription factor is essential for cell survival after stress stimuli. We hypothesized that BAG3 upregulation following stress stimuli is regulated by NF-κB. We determined whether NF-κB is involved in BAG3-mediated survival of primary human skeletal muscle cells (HSMC) during ischemia and diabetic conditions. Within 6 hours of treatment, ischemia induced BAG3 mRNA (no ischemia vs ischemia: 1.0 ±0.09 vs 1.41±0.02; p&lt;0.01) and protein expression (BAG3/total protein, no ischemia vs ischemia 1.0±0.01 vs 1.38±0.06; p&lt;0.01). Knockdown of BAG3 expression by shRNA induced early cell damage in HMSC under ischemic conditions as measured by HMGB1 expression (HMGB1/total protein; control plasmid vs shRNA, 1.0±0.09 vs 1.71±0.04; p&lt;0.01). Knockdown of p65 subunit of the NF-κB by shRNA significantly diminished BAG3 mRNA expression after ischemia (control plasmid vs shRNA: 2.11±0.18 vs 1.48±0.05; p&lt;0.05). Moreover, treatment of HSMC with 750 uM palmitic acid (PA) and 100nM insulin for 3 days to mimic diabetic conditions significantly increased the expression of BAG3 mRNA (control vs PAL+Ins, 1.0±0.14 vs 2.27±0.08; p&lt;0.01) and protein (BAG3/total protein, control vs PAL+Ins, 1.0±0.03 vs 1.39±0.11, p&lt;0.01). Knocking down p65 attenuated these increase in BAG3 mRNA (PAL+Ins vs shRNA+PAL+Ins, 2.27±0.08 vs 1.56±0.02 p&lt;0.01) and protein (BAG3/total protein; PAL+Ins vs shRNA+PAL+Ins, 1.39±0.11 vs 0.99±0.1; p&lt;0.05) Thus, 1) BAG3 expression is important in cell survival under ischemic conditions, and 2) NF-kB plays a key role in upregulating the expression of BAG3 under diabetic and ischemic conditions.
MON-687 Diabetic Muscle Infarction (DMI) a Rare Under-Recognised Complication of Diabetes Mellitus (DM): A Series of Three Cases
Authors: Dhanasekaran Maheswaran , Kishore Preeti , Schubart Ulrich K 
Published: May 2020

Abstract
Background: Diabetic muscle infarction (DMI) also known as diabetic myonecrosis is an acute, rare, microangiopathic complication of long-standing poorly controlled Diabetes Mellitus (DM). DMI presents as severe pain and swelling of the affected muscle group, usually of the lower extremities. It generally occurs in patients with established vasculopathy from uncontrolled diabetes including retinopathy and nephropathy. However, a clear association of DMI to long term mortality has not yet been defined. Proposed pathophysiologic mechanisms for DMI are microvascular endothelial damage complicated by thromboembolic events triggering an inflammatory cascade, leading to local tissue ischemia and eventual infarction. Alterations in the coagulation-fibrinolysis system and vasculitis have also been invoked. About 50% of the cases have DMI recurrence in the setting of uncontrolled DM.
Clinical Case: We report 3 cases of DMI diagnosed in patients with long-standing poorly controlled DM with at least two established micro-vascular complications. The lower extremity muscle group was affected, especially the thigh muscles and paraspinal muscle in one case. Biochemical parameters were significant for normal white blood cell counts, elevated erythrocyte sedimentation rate (112, 105, 145 mm/hr respectively; n 0–15 mm/hr) and C-reactive protein (CRP) (54.3, 260, 162 mg/L respectively; n 0–5.0 mg/L) and elevated creatine kinase in two cases (539 and 379 U/L; n 5–150 U/L). Infectious work-up including blood cultures were negative. Lower extremity doppler was negative in all three cases. Diagnosis was established with the help of MRI in two cases showing diffuse hyperintensities of the muscle groups involved and muscle biopsy in the third showing neutrophilic and eosinophilic inflammation and areas of necrosis, fibrosis and hemorrhage. Treatment involved adequate pain management and anti-platelet therapy along with tight glycemic control. No recurrence thus far was noticed in all three patients.
Conclusion: DMI should be considered in the differential diagnosis of patients with the aforementioned risk factors who did not show a response to antibiotics, as administered for presumed cellulitis. Though self-limiting, studies have shown a long term temporal correlation with increased all-cause mortality in patients with DMI. Hence DMI is considered as another cardiovascular risk equivalent. Further studies are needed to explore strategies aimed at preventing DMI and its recurrence.
MON-617 Improving Care Transitions: Optimizing Diabetes Medication Reconciliation
Authors: Oraibi Omar , Holmes Kimberly , Pichardo-Lowden Ariana 
Published: May 2020

Abstract
Introduction
Medication reconciliation (MR) is essential in caring for hospitalized patients with diabetes (DM). Inaccuracies prevail, affecting transitions of care. Factors contributing to errors are driven by providers, patients, and systems of practice and increase in transitions out of hospital. A study showed that MR errors occurred in about 38 % of admissions. Common discrepancies among DM subjects were related to DM and cardiovascular drugs. An intervention aiming to reduce MR discrepancies at discharge achieved 70% less errors due to communication between inpatient providers, primary care and patients within 24 hours. Systems-based interventions and multidisciplinary approaches show promise to improve processes. However, a comprehensive assessment of the elements of practice to target and how interventions may be more effective is lacking. Our purpose was to examine aspects of practice among personnel responsible for the MR steps, and within the hospital workflow in order to identify gaps in the process. We intend to recognize factors to be targeted to optimize MR for DM, and to provide a multipronged approach to inform changes in hospital processes.
Methods:
We used quantitative and qualitative methods to investigate errors in the MR process as part of a hospital quality improvement program. We randomly included patients 18 years or older with type 1 or 2 DM evaluated by the endocrine team for 6 months. Chart reviews were conducted to assess type and frequency errors. Interviews of nurses, pharmacists, clinicians, and DM educators were sought to understand unique situations and the roles of health providers in the MR process.
Results
Twenty-two subjects were identified with one or more of the following gaps in their MR pertaining to DM medications: a) missing, b) redundancy, and c) dosing error. Scenarios included ≥2 discrepancies (4 of 22); ≥1 medication inappropriately changed from home regimen (18 of 22); redundantly adding a medication (4 of 22); wrong dose of medication (1 of 22); incomplete prescription for DM supplies (8 of 22).
Conclusion:
We identified deficits and their attribution to professionals and categorized errors in hospital workflows. Observations, providers’ insights and literature review enabled an assessment of MR failures. We developed a conceptual model where types of error, professionals’ roles, and solutions intersect Interface of prioritizing actions, hospital resources, use of checklists, interdisciplinary collaborations, and staff education is essential to advance adequate MR in the system of practice. Our future steps include a Plan-Do-Study-Act cycle to advance care.