bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract P2127: A Novel Arnt Dependent Hif2a Singling In Protecting Against Cardiac Microvascular Endothelial Barrier Dysfunction Following Myocardial Infarction
Authors: Karim Ullah Karim , Ai Lizhuo , Li Yan , Liu Lifeng , Pan Kaichao , Humayun Zainab , Sitikov Albert , Su Qiaozhu , Zhao Qiong , Sharp Willard W , Fang Yun , Wu David , Liao James K , Wu Rongxue R
Published: October 2023

Rationale:
Cardiac microvascular leakage and inflammation are triggered during myocardial infarction (MI) and contribute to heart failure. Hypoxia-inducible factor 2α (
Hif2α
) is highly expressed in endothelial cells (ECs) and rapidly activated by myocardial ischemia, but its impact in microvascular endothelial barrier function during MI is unclear.

Objective:
To test our hypothesis that the expression of Hif2α in ECs regulates cardiac microvascular permeability in infarcted hearts, which is through its binding partner aryl hydrocarbon nuclear translocator. (ARNT).

Methods and Results:
Experiments were conducted with mice carrying an inducible EC-specific
Hif2α
-knockout (

ecHif2α
-/-

) mutation, with mouse cardiac microvascular endothelial cells (CMVECs) isolated from the hearts of

ecHif2α
-/-

mice after the mutation was induced, and with human CMVECs and umbilical-vein endothelial cells transfected with
ecHif2α
siRNA. After MI induction, echocardiographic assessments of cardiac function were significantly lower, while measures of cardiac microvascular leakage (Evans blue assay), plasma IL6 levels, and cardiac neutrophil accumulation and fibrosis (histology) were significantly greater, in

ecHif2α
-/-

mice than in control mice, and RNA-sequencing analysis of heart tissues from both groups indicated that the expression of genes involved in vascular permeability and collagen synthesis was enriched in

ecHif2α
-/-

hearts. In cultured ECs, ec
Hif2α
deficiency was associated with declines in endothelial barrier function (electrical cell impedance assay) and the reduced abundance of tight-junction proteins, as well as an increase in the expression of inflammatory markers, all of which were largely reversed by the overexpression of ARNT. We also found that ARNT, but not Hif2α, binds directly to the IL6 promoter and suppresses IL6 expression.

Conclusions:
Endothelial HIF-2a protects from hypoxia-induced cardiac microvascular barrier dysfunction, promotes inflammation damage and represents a potential therapeutic target for cardioprotection, and prevention of fibrosis following acute ischemic injury.
Abstract P2139: The Adiponectin Receptor Agonist ALY688 Prevents Pressure Overload-induced Cardiac Dysfunction And Adverse Cardiac Remodeling
Authors: Cho Sungji , Dadson Keith , Crawford Kathryn , Hsu Henry , Sweeney Gary
Published: October 2023

Adiponectin has been shown to mediate multiple cardioprotective effects. Since the use of adiponectin protein as a practical therapeutic is associated with multiple challenges, we examined the functional significance of targeting adiponectin signaling with ALY688, a small peptide-based adiponectin mimetic, in a mouse model of cardiac dysfunction caused by left ventricular pressure overload (PO) induced by minimally invasive transverse aortic banding (MTAB). Following daily administration of ALY688 over 5 weeks after induction of PO, echocardiography showed that significantly attenuated PO-induced reduction in ejection fraction and other indices of cardiac function compared with vehicle. ALY688 also significantly reduced cardiac hypertrophic remodeling (heart weight-to-tibia length), cardiomyocyte cross sectional area, and myocardial and circulating levels of serum natriuretic peptide A. ALY688 reduced PO-induced increases in myofibroblasts, and scar tissue collagen content based upon immunofluorescence staining for α-smooth muscle actin and vimentin, scanning electron microscopy, and picrosirius red staining. ALY688 reduced PO-induced myocardial expression of 14 pro-inflammatory genes and circulating IL-12, IL-15, IL-17, and MIP-3α levels. Targeted metabolomics revealed that ALY688 increased myocardial fatty acid oxidation, with increased mobilization of cardiac triacylglycerols and a decrease in sphingomyelin, fatty acids and lysophospholipids. In conclusion, the adiponectin analogue ALY688 is cardioprotective in a preclinical animal model of heart failure and cardiac remodeling, with beneficial metabolic, anti-fibrotic, anti-hypertrophic and anti-inflammatory properties leading to improved cardiac function.
Abstract P2160: High-throughput Imaging Of Cardiomyocytes Using Biomimetic Tools Help Identify Therapeutic Targets
Authors: Kleppe Hannah M , Chandrakumar Rishikesan , Slater Rebecca E , Figarska Sylwia M , Budan Anastasiia , Levinson Alan R , Priest James R , Hoey Tim
Published: October 2023

Cardiovascular disease is a leading cause of death and continues to require novel forms of therapies. Mutations in sarcomeres, the basic contractile units of cardiac muscle, causing disarray and loss of function are causal in many genetic cardiomyopathies. Here, we model sarcomeric disarray in vitro by culturing human induced pluripotent stem cell-derived cardiomyocytes (human iPSC-CMs) on tissue culture plates with biomimetic patterned grooves designed to simulate extracellular matrix. This culture system models the natural structure and phenotypic development of human physiology. Next, we created an Adeno-associated virus open reading frame (AAV-ORF) library of ~100 heart failure associated genes of interest (GOI) identified using publicly available human genetic data. We developed a high throughput screening (HTS) process to test the effects of overexpression of these GOIs on sarcomere morphology. Images of human iPSC-CMs treated with overexpression libraries were run through a custom MATLAB HTS image processing script named ‘Tamarack’, developed for quantifying human iPSC-CM sarcomere morphology. Tamarack enables quantification of sarcomere count, length, and orientation. Analysis of sarcomeres from the Tamarack code in the biomimetic plates allows for enhanced quantification of sarcomere structures, in particular, alignment, a metric missing from standard 2D culture plates. We found that treating cells with siRBM20 induced a sarcomere misalignment phenotype which we looked to rescue with overexpression of our GOI. Genes that increased alignment in an RBM20 background were considered top hits and moved to further analysis.
Abstract P2162: Controlled Porous Structures Of Nitric Oxide-releasing Inhaler For Pulmonary Arterial Hypertension Vascular Reverse Remodeling
Authors: Oh Yoogyeong , Hong Jinkee , Jung Se Yong
Published: October 2023

Pulmonary arterial hypertension (PAH) is an incurable disease characterized by elevated blood pressure in the pulmonary artery. Nitric oxide (NO) is a potent regulator of hemodynamics and macrophage reprogramming. However, the therapeutic effects of NO depend on its concentration and its half-life is very short. Pulmonary delivery has advantages such as the ability to escape first-pass metabolism and the large absorptive surface area. In this study, we prepared NO-releasing inhalers with controlled porous structures to modulate the deposition in the respiratory system. NO-releasing inhalers with micro-sized pores showed improved nebulization efficiency than nano-sized ones. Furthermore, the cGMP expression and macrophage polarization were evaluated to confirm the vasodilatory and anti-inflammatory effects of NO. The customized nebulizer system was prepared for efficiency test of NO-MPs. The fluorescence intensity of Nile red was analyzed using photoluminescence spectroscopy to confirm the amounts of NO-MPs on the slide glass. As with confocal images, 0.8k BPEI/NO-MPs could arrive the lower region than 25k BPEI/NO-MPs. NO produced by endothelial nitric oxide synthase (eNOS) induce vasodilation by increasing cyclic guanosine monophosphate (cGMP) in vascular smooth muscle cells. Hence, the delivery of NO mimicking the physical production of the endothelium in range of 0.05-0.4 nmol cm
-2
min
-1
is required to restore vascular homeostasis as NO has a concentration-dependent effect on cells. Therefore, BPEI0.8k/NO-MPs release 0.277 nmol cm
-2
min
-1
of NO that mimics the environment in healthy endothelium, while BPEI25k/NO-MPs showed a higher concentration of NO release, 0.548 nmol cm
-2
min
-1
. BPEI0.8k/NO-MPs increased the synthesis of cGMP in cells, whereas BPEI25k/NO-MPs presented adverse effects due to an initial burst release. When cell populations were quantified, BPEI0.8k/NO-MPs increased the number of M2-like cells while decreasing the number of M1-like cells. Furthermore, when we analyzed proteins and gene levels of cells, anti-inflammatory factors increased in BPEI0.8k/NO-MPs treatment groups. In conclusion, NO-releasing inhalers could deliver NO to deep lungs and has the potential for pulmonary vascular recovery.
Abstract P2165: Endothelial Leptin Resistance Exacerbates Vascular Dysfunction Through Attenuating PD-L1 Expression
Authors: Cheung Yiu Ming , Wong Wing Tak
Published: October 2023

Background:
Obesity and its vascular complications are escalating globally that threaten millions of people and incurring tremendous financial costs. Fundamentally, endothelial dysfunction is the culprit among these cardiovascular diseases due to the presence of risk factors such as hyperglycemia and hyperlipidemia, which enhance the inflammatory responses and leucocyte infiltration. Persistent endotheliopathy is closely associated to severe cardiovascular events, ultimately leading to mortality. Leptin is a pleiotropic adipokine that is significantly expressed in obese people, resulting in leptin resistance. Programmed cell death ligand 1 (PD-L1; encoded by gene
Cd274
) is the inhibitory immune checkpoint that limits immunogenic responses of lymphocytes by causing functional exhaustion and apoptosis in lymphocytes. The relationship between endothelial PD-L1 expression and progression of vasculopathy has not yet been elucidated.

Methods:
Providing that circulating leptin level is closely associated with body adiposity and feeding behaviors, both hypoleptinemia and hyperleptinemia were generated on C57BL/6 mice by nutritional modulation or genetic knockout on leptin receptor.

Results:
Findings across multiple models suggested, the expression of PD-L1, an immune-coinhibitory molecule, was regulated by leptin, the impairment in leptin signaling resulted in vascular PD-L1 downregulation. By employing the mice with obesogenic hyperleptinemia, we showed that the vascular tissue failed to respond to exogenous leptin stimuli, thereby hindering PD-L1 expression in the inflammatory microenvironment. In fact, PD-L1 expression is crucial in maintaining tissue homeostasis by suppressing the activated PD-1
+
leukocytes, as such hindered PD-L1 expression may favor the vascular inflammation by failing to inhibit immunogenic attack, especially when other risk factors are present. Beyond acting as a ligand to leukocytes, our findings also revealed PD-L1 reverse signaling in endothelium plays a role in resolving inflammation and rescuing endothelial function.

Summary:
These findings propose a new explanation for why obesity results in vasculopathy in relationship with PD-L1.
Abstract P2177: Autophagy-based Drugs Can Be Used To Treat Anthracycline-induced Cardiotoxicity
Authors: Zhang Song , Jacobs Johanna E , Mondaca-Ruff David , Xu Xiaolei , Ding Yonghe , Herrmann Joerg
Published: October 2023

Background:
Anthracyclines are highly effective chemotherapeutics, but their use is limited by Anthracycline-Induced Cardiotoxicity (AIC) with notable decline in ejection fraction (EF). Recovery of cardiac function is slow and often only partial, mandating novel and more efficacious treatment approaches to AIC.

Hypothesis:
Based on high throughput genetic screening in zebrafish we identified autophagy activation, e.g. via Atg7, a key protein in the induction of autophagy, as a potential novel translational therapeutic approach to AIC.

Methods:
In a mouse model of cardiotoxicity (4 x weekly 5 mg/kg intravenous doxorubicin treatment) two approaches were tested starting one week after the last treatment: 1) AdenoAssociated Viral vector 9-mediated overexpression of Atg7 in the myocardium (AAV9-ATG7, one time injection) and 2) 3 FDA approved autophagy activating (FAA) drugs (pravastatin (40 mg/kg), spironolactone (20 mg/kg) and empagliflozin (10 mg/kg), by daily gavage). Echocardiograms were performed at baseline, before start of therapy and after 28 days before sacrifice.

Results:
All three FAA drugs were able to rescue AIC and Atg7 overexpression by AAV9 alone sufficed to rescue from AIC (Figure 1). All three FAA drugs were able to either restore Atg7 levels to baseline (pravastatin and empagliflozin) or 2.5 times above baseline levels (spironolactone) versus more than 60fold above baseline by an AAV9-based approach.

Conclusion:
Atg7 overexpression and autophagy activation suffices to rescue from AIC and can be recapitulated to different levels by pravastatin, empaglifozin and spironolactone.
Abstract P2178: Trends In Age At Diagnosis, Incidence Proportion Of Benign Cardiac Neoplasms And Associated Outcomes: 2012-2021
Authors: Rawlley Bharat , Cordova Sanchez Andres , Menezes Sherna , Gupta Kartik , Chaudhuri Debanik
Published: October 2023

Introduction:
Incidence of benign cardiac neoplasms (BCNs) and outcomes in large scale studies among adults in US are unknown. We study this using TriNetX database.

Methods:
We queried TriNetX US Collaborative network housing data from insurance claims and electronic health records. We include adult patients with any ICD code from 01/01/12 - 12/31/21. ICD codes were used to identify BCN and ventricular arrhythmias, ischemic stroke and acute MI after BCN diagnosis. Mortality was identified by patients categorized as deceased after BCN diagnosis.

Results:
We identified 65.6 million patients (54% female), BCN prevalence was 0.0097% (n=6420). Mean (SD) age at diagnosis for males, females was 58.7 (18.2), 61.1 (17.5) years, respectively. Mean (SD) age at diagnosis increased from 57.2 (20.1) to 62.8 (16.9) years (P < 0.05). Incidence proportion for BCN in females and males was 0.013% and 0.010% respectively, no trend was seen in annual incidence proportion for either sex (P > 0.05). On comparing those with vs without BCN, BCN cases were at higher risk of mortality, Ventricular arrhythmia, Ischemic stroke, and acute MI with RR (95% CI); 3.56 (3.36-3.77), 11.9 (10.9-12.9), 10.09 (9.58-10.62) and 9.41 (8.84-10.02), respectively. The risk of mortality, ventricular arrhythmia, Acute MI and ischemic stroke in males vs females with BCN was 17.4% vs 15.3%, 4.6% vs 4.1%, 7.6% vs 9.2% and 7.5% vs 6.3%, respectively.

Conclusion:
Annual incidence proportion of BCNs remains stable from 2012-21. An increasing trend in age at BCN diagnosis is seen. Females have higher Incidence proportion and age at diagnosis. BCN patients are at risk of adverse outcomes with males having worse outcomes.
Abstract P2179: Endothelial-derived Extracellular Vesicles Communicate With Cardiomyocytes To Elicit Cancer Therapy-related Cardiac Dysfunction
Authors: Ching Crizza , Clift Cassandra L , Kinnear Caroline , Gustafson Dakota , Aikawa Elena , Singh Sasha A , Mital Seema , Thavendiranathan Paaladinesh , Fish Jason
Published: October 2023

Background:
Treatment with the anthracycline, epirubicin, can result in cancer-therapy related cardiac dysfunction (CTRCD), a leading cause of non-malignant mortality in cancer survivors. Endothelial cells (ECs) lining the blood vessels are the first point of contact of epirubicin and secrete factors that are pivotal in regulating cardiac function. Of these factors, extracellular vesicles (EVs) are known to mediate intercellular communication. It remains unknown whether and how EC-EVs affect cardiomyocyte function in the context of CTRCD.

Hypothesis:
We hypothesize that epirubicin exposure leads to altered EC-EV cargo which mediates cardiomyocyte dysfunction.

Methods:
Human umbilical vein endothelial cells (HUVEC) were exposed to the peak and trough concentrations of epirubicin and endothelial function was assessed. EC-EVs were isolated via ultracentrifugation and EV size and concentration were measured through nanoparticle tracking analysis. The protein content of EC-EVs was determined using liquid chromatography with tandem mass spectrometry. Pluripotent stem cell-derived cardiomyocytes (iPSC-CM) were exposed to physiological concentrations of EVs (1x1010 particles/mL) and contractility was assessed in real-time.

Results:
Epirubicin exposure increased endothelial leak, apoptosis, and adhesion molecule expression. We observed an increase in size and concentration of EC-EVs and proteomic analysis revealed altered EV cargo including reduced cardiomyocyte protective factors, CNP and eNOS in EVs from epirubicin treated HUVEC in comparison to control. Treatment of iPSC-cardiomyocytes with EVs from epirubicin treated HUVECs resulted in reduced contractility compared to EVs from healthy ECs. Cleavage of EC-EV surface proteins via proteinase K abrogated the effects of EC-EVs on iPSC-cardiomyocytes. Cardiomyocyte-specific pathways targeted by EC-EVs from epirubicin- compared to control-treated cells are currently on-going.

Conclusion:
We demonstrate that epirubicin treatment leads to endothelial dysfunction and altered release of EVs capable of mediating cardiomyocyte dysfunction. Our results show the potential role of EC-EVs in CTRCD pathogenesis and highlight their influence on cardiomyocyte function.
Abstract P2184: Isocitrate Dehydrogenase 1 Regulates Cardiac Metabolic Adaptation During Oncometabolic Stress
Authors: Kim kyoungmin , Gao Yaqi , Faubert Brandon , Snyder Nathaniel , DeBerardinis Ralph , Karlstaedt Anja
Published: October 2023

Cardiovascular disease and cancer are the two leading causes of morbidity and mortality in the world. Metabolic dysregulation of cancer cells extends beyond the tumor microenvironment and increases risk for cardiovascular diseases. One common somatic mutation in cancer cells affects isocitrate dehydrogenase (IDH) 1 and 2, which catalyzes the oxidative decarboxylation of isocitrate to alpha-ketoglutarate in the cytosol and mitochondria, respectively. IDH1 and 2 mutations cause the production of the oncometabolite D-2-hydroxyglutarate (D2-HG) which allosterically inhibits alpha-ketoglutarate dehydrogenase (alpha-KGDH) and is associated with reduced cardiac contractile function. We combined stable isotope tracer studies with computational modeling to study the fundamental role of IDH isoforms for cardiac adaptation during oncometabolic stress. We uncovered an unexpected cardiac phenotype that expands the role of IDH1 in the heart beyond oxidative metabolism. Using mass spectrometry-based metabolomics, we quantified the stable isotopomer distributions from glucose and glutamine in perfused working rat hearts and isolated adult ventricular cardiomyocytes. Our analysis revealed that defective mitochondrial metabolism causes redirection of carbon flux oxidative towards reductive pathways. Reductive carboxylation of alpha-KGDH increases glutamine uptake and glutamine-derived citrate formation in both working rat heart perfusions and cultured adult mouse ventricular cardiomyocytes. To identify which IDH isoform is responsible for redirecting carbon flux, we developed knockout models of IDH1, 2, and 3 in adult mouse ventricular cardiomyocytes. Loss of IDH1 expression impaired reductive formation of citrate and caused functional defects in cardiomyocytes. Lastly, epigenetic analyses of histone marks revealed that IDH1 causes widespread alterations in histone acetylation and tri-methylation. Our results highlight a novel role for IDH1 in cardiac metabolism and transcriptional control of metabolic adaptation in response to tumor-mediated stress and provide evidence of how reductive-citrate formation may induce epigenetic modifications in the heart.
Abstract P2187: SARS-CoV2 Nonstructural Protein, But Not The Omicron Ba.4/5 Mutant, Suppresses YAP Transcriptional Activity In The Heart
Authors: Meng Fansen , Tsai Chang-Ru , Kim Jong H , Li Xiao , Steimle Jeff , Liu Shijie , Xie Bing , Li Rich Gang , Martin James F
Published: October 2023

Coronavirus disease 2019 (COVID-19), caused by SARS-CoV2, is a worldwide epidemic resulting in global healthcare crises and strained health resources. Currently, the omicron subvariants of SARS-CoV2, BA.4 and BA.5, have become the predominant variant in circulation around the world. Compared to previous variants, omicron BA.4 and BA.5 escape antibodies created by past COVID-19 vaccines and prior infections due to spike protein mutations. However, why omicron subvariants induce milder symptoms after infection are poorly understood. Here we screened SARS-CoV2 proteins and found that one nonstructural protein suppresses the transcriptional activity of YAP, the Hippo pathway signaling effector, independent of negative regulation by upstream Lats kinases in the heart. Our findings correlate with RNA sequencing data from COVID-19 patient lung samples and iPS cardiomyocytes showing decreased expression levels of YAP target genes, which suggested that SARS-CoV2 infection may affect lung epithelial cell and neonatal cardiomyocyte proliferation. Additionally, we found that the Omicron BA.4/5 variant of this viral protein suppresses YAP activity significantly less compared to the wildtype protein whose mechanism are under exploring now. These findings reveal the previously unknown function and mechanism of SARS-CoV2 nonstructural proteins in host cells. Considering the critical role of Hippo signaling in cell proliferation, apoptosis, and immune response, our data on SARS-CoV2 protein function may partially explain the lesser toxicity of Omicron BA.4/5 to the host cells.
Abstract P3002: Role Of Vascular Fibroblast Growth Factor Signaling In Group 3 Hypoxia-induced Pulmonary Hypertension
Authors: Woo Kel Vin , Weinheimer Carla , Nigro Jess , Kovacs Attila , Ornitz David
Published: October 2023

Pulmonary hypertension (PH) is a debilitating disease characterized by pathologic changes in the pulmonary vasculature that increases pulmonary vascular resistance and leads to right heart failure. Group 3 PH is caused by lung disease and/or hypoxia, including bronchopulmonary dysplasia (BPD) and chronic obstructive lung disease (COPD), and is the second largest cause of mortality in PH patients. PH further increases the morbidity and mortality in both COPD and BPD patients.

Fibroblast Growth Factor 2 (FGF2) and FGF Receptor 1 (FGFR1) expression is elevated in lung tissue of PH patients. To study the role of FGF signaling in the pathogenesis of group 3 PH, we developed a physiologically relevant mouse model in which mice are challenged with 10% hypoxia for 2 weeks. Following hypoxia exposure, cardiac catheterization is used to measure right ventricular (RV) pressure that mimics Group 3 PH. Lung tissue is then analyzed for histological and molecular changes.

Our initial work showed that inactivation of FGFRs in endothelial cells (EC) worsened hypoxia-induced PH while expression of a constitutively active FGFR1 (caFGFR1) in ECs protected against hypoxia-induced PH. Here we hypothesize that FGF signaling may also affect the pathogenic response to hypoxia of vascular smooth muscle (VSMC), potentially by limiting vascular muscularization in response to hypoxia. We show that
in vitro
inhibition of FGF2 signaling in human pulmonary artery VSMCs reduced hypoxia-induced proliferation while promoting a contractile phenotype, and
in vivo
inactivation of FGFRs 1 and 2 worsened hypoxia-induced vascular remodeling. These data suggest the potential for a dual beneficial effect of activation of FGF signaling, both in regulating EC-VSMC interactions and directly preventing VSMC remodeling.
Abstract P3004: Vaccination Targets Senescence-associated Glycoprotein Ameliorates Alzheimer’s Pathology And Cognitive Behavior In Mice
Authors: Hsiao Chieh Lun , Katsuumi Goro , Suda Masayoshi , Shimizu Ippei , Yoshida Yohko , Furihata Takaaki , Joki Yusuke , Furuuchi Ryo , Hsiao Yung-ting , Liang Jia-Qi , Abe Manabu , Minamino Tohru
Published: October 2023

Introduction:
From the patients and the mice with atherosclerosis, we found senescence-associated glycoprotein (SAGP-protein) was upregulated in macrophage and vascular endothelial cells. Analysis of transcriptome data from human vascular endothelial cells (HUVECs) revealed the enrichment of SAGP-protein in the HUVECs with senescent status. Based on molecular targeting strategy, we have developed a senolytic vaccine (SAGP-vaccine), which could eliminate SAGP-high senescent cells, leading to amelioration of age-associated pathologic condition including diabetes and atherosclerosis in mice. Since previous studies have reported the positive correlation between SAGP-protein and another age-associated disorder, Alzheimer’s disease (AD), we hypothesize that SAGP-vaccine would also be a treatment to AD.

Purpose:
This study was aimed to examine the efficacy of SAGP-vaccine in improving AD pathology and degeneration in AD mouse model.

Methods:
Mice with knock-in of human mutant amyloid precursor protein (APP
NL-G-F
) (APP-KI) were treated with control vaccine or SAGP-vaccine at 2 and 4 months old. Behavior tests were performed from 6 months old, and sacrificed afterwards. Brain samples were harvested for further molecular-biological analysis.

Results:
APP-KI mice with SAGP-vaccination showed the significant improvement in behavior tests along with the reduction of SAGP expression and APP deposition in brain tissue. Furthermore, SAGP-vaccination significantly reduced expression of inflammatory molecules. Consist with previous studies, SAGP was mainly colocalized with microglia, which is the possible target of SAGP-vaccine.

Conclusions:
SAGP-vaccine would be a sufficient treatment to alleviate amyloid beta-induced pathogenesis in the brain and mitigate the impaired behaviors of AD.
Abstract P3006: Inhibition Of Tnfα Signaling Delay CAVD Progression In Mice
Authors: Thent Zar Chi , Benkaci Sonia , Wu Bingruo , Butcher Jonathan T , Zhou Bin
Published: October 2023

Background:
Inflammation is a prominent feature of calcific aortic valve disease (CAVD), a leading cause of aortic stenosis. Inflammation can activate surface valvular endothelial cells (VEC) to produce pathogenic signaling such as Tumor Necrosis Factor-alpha (TNFα), to valvular interstitial cells (VIC) in aortic valve leaflets, leading to pathological valve remodeling through increased extracellular matrix synthesis, calcium deposition, and cell death which underlies CAVD. Thus, TNFα inflammatory signaling maybe a major candidate therapeutic target for alleviating CAVD progression.

Hypothesis:
We hypothesized that inhibition of TNFα signaling may delay CAVD progression.

Results:
We compared CAVD progression in a mouse model of CAVD with deletion of
Ldlr,
low-density lipoprotein receptor, and fed with standard or high fat diet. We inhibited the TNFα inflammatory signaling in this CAVD model by crossing the
Ldlr
-/-
mice with the
Tnfr1
and
Tnfr2
double null mice. The aortic valve functions were measured by echocardiogram and the results showed that aortic valve velocity was significantly decreased in the triple null (

Ldlr
-/-
;Tnfr1
-/-
;Tnfr2
-/-

) group compared to the

Ldlr
-/-

group, both fed with high fat diet. Contractile function (systole to diastole ratio) at the aortic root was significantly increased in the triple null group. Aortic valve histopathology was examined by various staining methods. In the triple null mice, Hematoxylin and Eosin (H&E) showed significantly reduced the thickness of aortic valve leaflets; Masson’s Trichrome, Picrosirius Red, and Rusell-Movat staining revealed significantly reduced fibrosis and collagen deposition, and Von Kossa and Alizarin Red staining showed significantly decreased calcium deposition. Furthermore, immunofluorescence staining demonstrated significantly decreased DNA damage and cell death, as well as cell proliferation in aortic valve leaflets of the triple null mice.

Conclusion:
Our findings demonstrate a promising role of inhibiting TNFα signaling in slowing CAVD progression in mice, suggesting pharmacological or immunological inhibition of TNFα signaling as a non-invasive therapeutic option to treat CAVD patients.
Abstract P3012: Routine Monitoring For Intracardiac Thrombosis And Pulmonary Embolism With Intraoperative Transesophageal Echocardiography In Orthotopic Liver Transplantation
Authors: Noe Niki , Girish Akanksha , De Marchi Lorenzo
Published: October 2023

Background:
Intracardiac thrombosis (ICT) and pulmonary embolism (PE) are significant intraoperative complications during orthotopic liver transplantation (OLT). Despite the high mortality associated with these events, current monitoring practices are inadequate at detecting these complications during OLT. Transesophageal echocardiography (TEE) allows continuous monitoring of myocardial function and detection of ICT/PE. In this study, we aim to report the incidence of ICT/PE during OLT at our center and to advocate for the use of intraoperative TEE as a standard monitoring tool. We also propose a medical management algorithm stratified by the severity of RV dysfunction and pulmonary artery pressure (PAP) in cases of ICT with/without PE.

Methods:
We conducted a single-center retrospective chart review of OLT patients with intraoperative TEE monitoring from March 13, 2019 - March 13, 2023. Intraoperative data of interest included hemodynamics, RV dysfunction, PAP, presence of ICT and/or PE as identified on TEE and associated medical management. RV dysfunction was graded as mild, moderate, or severe reduction in cardiac contractility. Secondary outcome was mortality during index hospitalization.

Results:
We identified 99 patients at our center who had intraoperative TEE monitoring during OLT. Prevalence of ICT in this population was 9.09% (n=9), of which 44.4% (n=4) developed hemodynamically significant PE. ICT patients who remained hemodynamically stable were observed without intervention (33%, n=3). Heparin was administered to prevent thrombus enlargement in patients with an increase in PAP but no significant RV dysfunction detected on TEE (22%, n=2). Those showing signs of hemodynamically significant PE with moderate or severe RV failure were given tissue plasminogen activator (tPA) for thrombolysis (44%, n=4). Perioperative mortality among all ICT patients was 44.4% (n=4) due to hemorrhagic shock secondary to coagulopathy and heart failure.

Conclusion:
The results of this study show the utility of routine TEE use in intraoperative monitoring for identifying ICT and PE in OLT patients, given the high mortality associated with these complications. TEE also allows for a stepwise approach to medical management based on RV function and PAP.
Abstract P3014: Human Pluripotent Stem Cell-derived Organoids Yield Microvascular-like Endothelial Cells
Authors: Majid Qasim A , Talman Virpi , Harding Sian , Foldes Gabor
Published: October 2023

Introduction:
Coronary microvascular disease (CMD) can manifest as microvascular angina or HFpEF and is implicated in post-STEMI remodelling and COVID-19. Superior in vitro models would aid the development of therapies for CMD. This study outlines the creation of a 3D differentiation protocol that yields stable human pluripotent stem cell-derived microvascular-like endothelial cells (hPSC-MVECs).

Hypothesis:
We hypothesised that the 3D organoid environment would allow for recapitulation of the microvasculature.

Methods:
Growth factor-mediated endothelial differentiation of hPSCs was conducted for 12 days in stirred tank bioreactors devoid of exogenous matrix substrates such as Matrigel. CD31
pos
3D-derived hPSC-ECs were isolated via fluorescence-activated cell sorting and evaluated via RT-qPCR for their gene expression of endothelial markers relative to 2D-derived hPSC-ECs. 3D hPSC-ECs were subsequently treated with high concentration VEGFA for 7 days (3D+V) and assessed for CD31 expression via high content image analysis. A proteome profiler human angiogenesis array was employed to investigate the angiogenic landscape of the generated hPSC-ECs relative to primary Human Cardiac Microvascular Endothelial Cells (HMVEC-Cs). Transcriptomic analysis of the ECs was conducted via Single-Cell RNA sequencing (scRNA-seq).

Results:
Organoids attained on day 12 of the 3D differentiation protocol displayed a complex vascular network. Isolated 3D hPSC-ECs displayed comparable levels of
PECAM1
and
CDH5
relative to 2D hPSC-ECs however, the expression of
VEGFA
was significantly reduced (P<0.01). The resultant high-concentration treatment with exogenous VEGFA gave rise to phenotypically stable hPSC-ECs that had an angiogenic profile comparable to HMVEC-Cs. scRNA-seq for 3D+V hPSC-ECs revealed expression of several reported microvascular markers including
GJA4
(arteriole),
S100A4
(post-arteriole capillary),
TCF15
(capillary),
SOX17
(pre-venular capillary),
CD34
(venule), and the coronary vessel marker
FABP4
.

Conclusions:
These data suggest that 3D organoid differentiation facilitates the generation of cardiac microvascular-like ECs herein termed hPSC-MVECs.
Abstract P3020: O-GlcNAc Transferase Regulates Vascular Smooth Muscle Cell Cycle Progression In Diabetes
Authors: Bhavnani Neha , Khanal Saugat , Mathias Amy , Lallo Jason , Gupta Shreya , Raman Priya
Published: October 2023

Diabetic patients show increased proclivity for vascular smooth muscle cell (VSMC) migration and proliferation. Hyperglycemia, hallmark of diabetes, increases intracellular glucose signaling via O-linked N-acetylglucosamine transferase (OGT), a key regulator of protein O-GlcNAcylation. We previously showed that smooth muscle OGT (smOGT) deletion prevents hyperglycemia-induced atherosclerosis. Goal of this work was to interrogate the role of smOGT in VSMC cell cycle progression in diabetes. For this, we used conditional SMC-specific OGT knockout (smOGT
KO
) and wild-type (smOGT
WT
) mice generated via tamoxifen (tmx)-inducible VSMC-restricted Cre driver mice (

Itga8-CreER
T2

). Briefly, following tmx injection, 8-wks-old smOGT
KO
and smOGT
WT
mice were placed on Western diet feeding regimen for study duration. At 14-wks-age, mice were injected with 50mg/Kg/day streptozotocin i.p. once daily for 5 consecutive days. Mice were harvested at 16-18-wks-age and plasma and aortic tissues were collected for biochemical and molecular studies. Immunoblotting of aortic lysates revealed a significant decrease (59%) in OGT and O-GlcNAc expression, validating the smOGT
KO
mice. Loss of smOGT reduced (40%) CDK6 expression (regulator of G1-S phase cell cycle progression) concomitant to enhanced (62%) p21
Cip1
expression (cyclin-dependent kinase inhibitor) in aortic vessels from diabetic smOGT
KO
vs. smOGT
WT
mice. This was further accompanied with 69% attenuation in SRF expression (transcriptional regulator of SM proliferation) in diabetic smOGT
KO
vs. smOGT
WT
aortic vessels. Similar results were obtained with OGT loss-of-function studies in human coronary artery SMC (HCASMC) primary cultures under diabetic conditions
in vitro
. Briefly, control (siCtrl) and OGT siRNA (siOGT)-transfected HCASMC were treated with or without 25mM glucose+100μM PUGNAc for 48 hours. Immunoblotting revealed that reduced OGT-mediated O-GlcNAcylation in siOGT cells in a diabetic milieu significantly attenuated SRF expression coupled with augmented p21
Cip1
expression compared to diabetic siCtrl cells. Together, our results suggest a direct regulatory role of smooth muscle OGT in VSMC cell cycle progression in diabetes.
Abstract P3027: Potasium Adaptation Down-regulates PSGL-1 And I-CAM-1 Gene Expressions In Salt -Induced Hypertensive Sprague Dawley Rats
Authors: Ajayi Olutayo I , OBAYUWANA MARTHA O
Published: October 2023

Background:
There is an increasing prevalence of hypertension and dietary salt intake has been linked as its risk factor. Although many studies have elucidated this association, the mechanisms by which the increase in salt intake leads to development hypertension have been postulated but yet inconclusive. P - Selectin glycoprotein ligand-1(PSGL-1) and Intercellular adhesion molecule (I-CAM-1) have been implicated in the development of salt sensitive hypertension through vascular inflammation and injury. The role of potassium in blood pressure regulation has been severally reported. Scarce reports of the association of potassium adaptation with high blood pressure development form the basis of this study. We aimed at defining the role of potassium adaptation in the development of salt -induced hypertension in Sprague Dawley rats.

Methods:
This study was divided into two phases: the induction and confirmation of hypertension and the effect of potassium adaptation. The test group received high salt diet (8% NaCl) for 6 weeks while the control animals were fed with standard rat chow and given water ad libitum. Phase II comprised of pre-treated animals with KCl before induction of hypertension and post induction treatment group with KCl. Blood pressures were monitored as well as blood samples taken for analysis of the gene expressions of I-CAM-1 and PSGL-1 using Standard Laboratory methods

Results:
We observed a spontaneous significant increases in the blood pressures of the test animals as from the 4
th
week compared with the control (P<.05, respectively), the potassium -adapted group and KCl post treated groups exhibited relatively stable blood pressure compared with the control. There were significant increases in the expression of PSGL -1 and I-CAM-1 protein in all the test groups but lower in the potassium adapted and post-treated animal groups when compared with hypertensive group. The potassium adapted group did not develop hypertension throughout the period.

Conclusion:
Raised arterial blood pressure is inducible by high salt diet. This is associated with an increased expressions of PSGL-1 and ICAM-1. Potassium adaptation inhibited the development of hypertension and down-regulate PSGL-1 expression significantly.
Abstract P3055: Identification Of Cell-intrinsic Effects Of Clonal Hematopoiesis In Heart Failure
Authors: Abplanalp Wesley T , Schuhmacher Bianca , Cremer Sebastian , Merten Maximilian , Macinkovic Igor , Mariana Szumlakiwska , Zeiher Andreas M , John David , Dimmeler Stefanie
Published: October 2023

Background:
Clonal hematopoiesis of indeterminate potential (CHIP) is caused by somatic mutations in hematopoietic stem cells and associates with worse prognosis in patients with heart failure (HF). While, circulating immune cells derived from patients with HF harboring CHIP mutations show heightened proinflammatory signatures, it is unclear whether these signatures are derived from the relatively low number of cells harboring CHIP-driver mutations, or are indicators of a systemic pro-inflammatory activation associated with CHIP.

Methods and Results:
We assessed the cell-intrinsic effects of the most prevalent CHIP-driver gene (DNMT3A) mutant-carrying circulating immune cells in a total of 5 HF patients. Using a novel protocol to detect DNMT3A mutant cells on a single cell level (MutDetect-Seq), we show that CHIP mutations can be detected in all major immune cell types (e.g. T cells, monocytes, B cells, NK cells), with detected mutation similar to that predicted by traditional targeted DNA sequencing. DNMT3A mutant monocytes demonstrated the highest number of significantly upregulated genes relative to other cell types. DNMT3A mutant monocytes exhibit altered gene expression signatures associated with inflammation and phagolysosome function. Additionally, DNMT3A mutant carrying CD4+, but not CD8+ T cells, showed significant changes in gene expression patterns towards an activated and cytotoxic profile, which was validated in vitro. Finally, increased paracrine signaling pathways involved in amplifying inflammatory circuits are predicted between mutated and non-mutated monocytes, this was validated in vitro with further implications for cardiac fibroblast activation via aSMA and COL1A1 expression.

Conclusion:
This is the first study to decipher the genetic signature of DNMT3A-CHIP-driver mutation carrying circulating immune cells, thereby providing first evidence for the cell intrinsic effects of DNMT3A mutations on peripheral blood cell subsets in patients with HF. The altered gene expression profiles associated with inflammation and phagolysosome function of mutant monocyte, as well as skewing of mutated T cells towards effector cells provide novel insights into how CHIP may promote worse prognosis in heart failure patients.
Abstract P3062: Tet2 Maintains Genome Stablity In Cardiac Myeloid Cells
Authors: Dou Yaling , Zhang Yan , Hong Tingting , Fang Shaohai , Martin James , Huang Yun
Published: October 2023

Myeloid cells play critical roles in the regulation of myocardial injury and repair. Clonal hematopoiesis (CH)-related mutations in genes, such as Ten-eleven Translocation 2 (TET2), can impair myeloid cells and are associated with increased risk of cardiovascular disease. To identify the subsets of myeloid cells that are impacted by Tet2 loss-of-function (LOF) and illuminate how Tet2 LOF disrupts normal myocardial repair, we analyzed the dynamics of myeloid cells in a myeloid-specific Tet2-deficient mouse model after ischemia-induced myocardial infarction (MI). We observed a substantial increase of neutrophils in the post-MI mice, which contributes to adverse outcomes during heart repair. Mechanistically, Tet2-deficient myeloid cells exhibited increased genome instability, accompanied with augmented activation of the intracellular innate immune response. Furthermore, pharmacological inhibiting intracellular innate immunity resulted in significantly reduced neutrophils in Tet2-deficient mice after MI and reversal of deleterious cardiac outcomes. Findings from this study are consistent with the conclusion that targeting the innate immune pathways is a viable therapeutic option to reduce the risk of cardiovascular diseases in CH individuals bearing TET2 LOF mutations.
Abstract P3073: Interleukin-2 Receptor Gamma Signaling Is Required For The Regenerative Adaptive Immune Response In The Adult Zebrafish Heart
Authors: Wang Qianchen , Cardeira-da-Silva João , Stainier Didier
Published: October 2023

Ischemic heart disease remains a leading cause of death worldwide. Unlike adult mammals, zebrafish have the ability to fully regenerate their heart after injury through processes including epicardium and endocardium activation, coronary revascularization, cardiomyocyte dedifferentiation and proliferation, and scar resolution. Immune cells are critical to remove damaged tissue, sustain repair and initiate regeneration after injury. Our previous data show that blocking antigen presentation impairs cardiac regeneration, pointing at a role for the adaptive immune system during this process. However, the molecular mechanisms underlying a pro-regenerative adaptive immune response remain largely unknown. Based on bulk transcriptomics, single-cell transcriptomics, and qPCR data, we have identified
il2rga
(
interleukin-2 receptor gamma a
) and
jak3
(
janus kinase
3) as potential regulators of the adaptive immune response during cardiac regeneration. We indeed found that
il2rga
mutants exhibit a reduced expression of T-cell markers, as well as impaired cardiac regeneration, as observed by decreased cardiomyocyte dedifferentiation and proliferation, and defective scar resolution. In addition,
il2rga
mutants display decreased expression of pro-regenerative factors (e.g.,
igf1, ngfb, nrg1, pdgfaa, tgfb2, vegfc
). Similarly, loss of Jak3, which functions downstream of Il2rga, also leads to reduced cardiomyocyte dedifferentiation and proliferation, as well as impaired endocardium activation. Bulk transcriptomics further shows the downregulation of key genes involved in the adaptive immune response in
il2rga
mutants. Specifically, we found that
il13
, a recently described cardiomyocyte cell cycle regulator and T helper 2 (Th2) cytokine, is strongly downregulated upon the loss of
il2rga
. Altogether, our findings reveal the crucial role of Il2rga/Jak3 signaling during cardiac regeneration, thereby opening new avenues to identify and understand the adaptive immune mechanisms capable of sustaining regenerative responses post-cardiac injury.
Abstract P3080: Macrophages Synthesize Bioactive Lipids To Modulate Cardiomyocyte Metabolism And Proliferation During Cardiac Regeneration
Authors: Lantz Connor , Becker Amanda C , Filipp Mallory , Thorp Edward B
Published: October 2023

Introduction:
Once thought to be a quiescent process, elimination of dying cells by macrophages actively reprograms macrophage metabolism to support downstream functions. Previously, we discovered that the phagocytic receptor MerTK is necessary for regeneration in the neonatal murine heart. Single-cell RNA sequencing and bulk-tissue lipidomics revealed prioritization of arachidonic acid metabolism towards the synthesis of Thromboxane A
2
(TXA
2
) in regenerating hearts, yet the molecular mechanisms directly connecting macrophage-derived TXA
2
to regeneration remained undefined. Therefore, we sought to determine if TXA
2
and activation of its receptor (
Tbxa2r
) on cardiomyocytes is necessary for cardiac regeneration.

Results:
We created a novel mouse to conditionally knockout
Tbxa2r
in cardiomyocytes using the CRE recombinase under the
Myh6
promoter. Loss of
Tbxa2r
did not affect normal cardiac development but significantly impaired cardiac regeneration as seen by significant fibrotic scarring, reduced cardiac output, and decreased cardiomyocyte proliferation. We next asked how
Tbxa2r
initiates intracellular signaling to support cardiomyocyte proliferation. Knowing that the YAP/TAZ pathway has been previously described to induce cardiomyocyte proliferation during regeneration and is regulated by GPCRs, we found that TXA
2
mimetics activate TBXA2R leading to the stabilization of YAP/TAZ in a time- and dose-dependent manner. Furthermore, YAP was translocated into the nucleus of cardiomyocytes in the border zone 3 days post-MI. However, deletion of
Tbxa2r
in cardiomyocytes completely prevents YAP translocation into the nucleus. Lastly, we have found that a TXA
2
mimetic increases the glycolytic capacity of cardiomyocytes further supporting anabolic processes needed for proliferation. Excitingly, we also observed an increase in glycolytic enzyme transcripts at the border zone of the infarct using publicly available spatial transcriptomic data.

Conclusions:
Our findings advance our understanding of how cell death, removal and recycling regulate macrophage metabolism, resulting in the production of bioactive lipids that further regulate the metabolism of neighboring cardiomyocytes during regeneration.