tbuckley/cabot-search huggingface dataset:
Abstract Tu114: Prostaglandin E2 Alters Mitochondrial Energy Metabolism in the Murine Heart
Authors: Timothy Bryson, DruAnne Maxwell, David Taube, Pamela Harding
Journal: Circulation Research (2024)

Prostaglandin E 2 (PGE2) is an autacoid that acts via 4 receptors (EP1-EP4), with EP3 and EP4 being the most prominent in the heart. We previously reported that cardiomyocyte specific knockout of EP4 (EP4 KO) results in dilated cardiomyopathy. Additionally, transgenic mice that overexpress EP3 in the cardiomyocyte (EP3 TG) develop heart failure. Recently published data from our laboratory suggests EP4 KO results in mitochondrial dysfunction, likely via EP3 receptor signaling. We therefore hypothesized that PGE2 alters energy metabolism in the heart via EP3. To test our hypothesis, we performed gene array analysis on left ventricles from adult EP4 KO and wild type (WT) mice. Significant reductions (2.1-fold) in carnitine palmitoyltransferase 2 ( Cpt2 ) were observed in EP4 KO vs. EP4 WT. CPT2 is a key enzyme in fatty acid oxidation and deficiencies in humans have been shown to cause cardiomyopathy. Substantial reductions in CPT2 were also observed in C57 mice after 2 weeks of myocardial infarction (0.22 ± 0.04 a.u. in sham controls vs. 0.039 ± 0.06 a.u. after MI. p&lt;0.01) by western blot analysis. Moreover, treatment with the EP3 receptor agonist, sulprostone, reduced CPT2 protein expression significantly in cardiomyocytes isolated from C57 mice (1.0 ± 0.0 a.u. in vehicle control vs. 0.67 ± 0.07 a.u. in sulprostone treated. p&lt;0.005). These data suggest that PGE2 via its EP3 receptor reduces CPT2 expression. In fact, RNAseq analysis on left ventricles from EP3 TG and EP3 WT mice showed significant reductions in Cpt 2 expression as early as 5 wks of age (log 2 FC = -1.17. p&lt;0.01). RNAseq also showed reductions in other key fatty acid oxidation/transport genes in EP3 TG mice ( Acsl1 ; log 2 FC= -0.73, Acads , log 2 FC= -0.59, Cd36 , log 2 FC= -0.64. p&lt;0.005). Lastly, to further investigate the role of the EP receptors on energy metabolism in the heart, we isolated cardiomyocytes from EP4 KO and EP4 WT mice and performed metabolomics. As anticipated, we observed significant reductions in L-carnitine (7.6 ± 1.13 fmol/cell in EP4 WT vs. 2.1 ± 0.32 fmol/cell in EP4 KO. p&lt;0.005) and several acyl carnitine products. Altogether our data supports our hypothesis that PGE2 signaling via its EP3 receptor impairs mitochondrial energy metabolism in the heart by reducing fatty acid oxidation.
Abstract Tu018: Integrin alpha 1 plays a critical role in angiotensin II-induced abdominal aortic aneurysm rupture
Authors: Naofumi Amioka, Toru Miyoshi, Toshihiro Iida, Yukihiro Saito, Megumi Kondo, Kaoru Akazawa, Satoshi Akagi, Kazufumi Nakamura, Shinsuke Yuasa
Journal: Circulation Research (2024)

Background: Abdominal aortic aneurysm (AAA) is a life-threatening condition without an established specific therapy. This study aims to identify a new therapeutic target for preventing AAA rupture. Methods and Results: Angiotensin II (AngII)-infused ApoE-/- mice develop AAA, an established model of AAA. To identify molecular pathways contributing to AAA development, we conducted single-cell RNA sequence analysis using a dataset of aortic tissues harvested from mice infused with AngII for 4 weeks (GSE186865). Gene set enrichment analysis revealed upregulation of integrin- and TGFB-related pathways in AngII-stimulated smooth muscle cells (SMCs). Integrin α1 (Itga1) constitutes half of the α1β1 integrin duplex and serves as a major collagen receptor in SMCs. Male wild-type (n=37) and Itga1 knockout (n=36) mice with an ApoE-/- background received a continuous infusion of AngII for 4 weeks to develop AAA. Kaplan-Meier analysis indicated that ablating Itga1 significantly suppressed AAA rupture (p=0.009), without affecting the maximum diameter of the abdominal aorta. Considering extracellular matrix degradation as a critical hallmark of AAA rupture, we examined the abundance of major matrix metalloproteinases (MMPs) in the aortic wall, specifically MMP2 and MMP9, two major MMPs in the aorta. Gelatin zymography revealed that ablation of Itga1 suppressed AngII-induced MMP9 increase, without affecting MMP2 abundance. The expression of MMP9 is controlled by inflammation and proliferation signals; hence, we focused on these signals. qPCR analysis (n=8 per group) demonstrated that AngII-induced upregulation of Tnfa (p&lt;0.001) and Tgfb1 (p=0.026) was mitigated by Itga1 ablation in aortic tissues. Conclusion: The integrin pathway is activated in the tissue of AAA tissue and deletion of Itga1 suppresses AAA rupture. Here we showed that the Itga1-Tgfb1-MMP9 axis would be a novel molecular target to inhibit AAA rupture.
Abstract Tu005: Small Extracellular Vesicles Derived from Bovine Milk Contain an Endogenous Carboxyl Terminal Polypeptide of The Gap Junction Protein Connexin-43
Authors: Md Ruhul Amin, Spencer Marsh, Claire Beard, Jane Jourdan, Robert Gourdie
Journal: Circulation Research (2024)

Background: Connexin 43 (Cx43) is the body's most widely expressed gap junctional connexin. Researchers have shown evidence of non-canonical functions of Cx43 and the therapeutic promise of Cx43 mimetic peptides in cardiac pathologies. However, some critical barriers, including in vivo stability and potential immunotoxicity of Cx43 mimetic peptides, hinder clinical translation of these pro-drugs. To address these challenges, we are pioneering bovine milk-derived small extracellular vesicles (mEVs) as delivery vehicles for peptidic drugs. While exploring mEVs as drug delivery tools for αCT1 – a peptide mimicking Cx43 carboxyl terminus (CT) undergoing Phase III testing and known for promoting wound healing and cardioprotection - mEVs without loaded drug cargo exhibited similar bioactivity. This study aimed to test the hypothesis that Cx43, particularly CT fragments of Cx43, are naturally present in mEVs. Methods: mEVs were isolated from bovine milk using a novel protocol developed by our lab based on Tangential Flow Filtration and characterized by Western Blotting (WB), Nanoparticle Tracking Analysis, and Transmission Electron Microscopy. To determine the presence of Cx43 in mEVs, we used WB, immunoprecipitation and Single-Molecule Localization Microscopy (SMLM). Results: Characterization of mEVs confirmed relatively pure and ultrastructurally definitive mEVs isolated from milk at high density. WB for Cx43 in mEVs lysates based on two antibodies - one against the cytoplasmic loop domain and the other against CT-most 20 amino acids of Cx43, showed that full-length Cx43 was present in mEVs. Interestingly, the Cx43 CT antibody also detected a band at around 16 KDa, and this was confirmed by WB by using two more Cx43 CT antibodies. We also immunoprecipitated this Cx43 CT polypeptide from mEVs lysates using the CT antibodies. SMLM suggested that a subpopulation of mEVs contain Cx43 and demonstrated vesicle-to-vesicle heterogeneity to Cx43 immunolabeling. Conclusions: We show for the first time that an endogenous Cx43 CT polypeptide is present in mEVs. Our results provide new insight into the role of Cx43 in mEV bioactivity and potentially offer a path to safe, effective, and orally available therapeutic options based on mEVs.
Abnormal Spirometry and Risk of Dementia: A Focus on Preserved Ratio Impaired Spirometry
Authors: Jinyuan Wang, Jiongxue Chen, Yanting Chen, Dong Pan, Zhenhong Deng, Huanquan Liao, Zecong Lin, Daji Guo, Yongteng Xu, Yamei Tang
Journal: American Journal of Respiratory and Critical Care Medicine (2024)
Abstract Tu071: Computational Modeling to Predict Mechanisms of DYRK1A-mediated Inhibition of Cardiomyocyte Proliferation
Authors: Bryce Murillo, Catherine Zhao, Michelle Wu, Bryana Harris, Kaitlyn Wintruba, Jeffrey Saucerman, Matthew Wolf
Journal: Circulation Research (2024)

Introduction and Background: Adult cardiomyocytes lack strong proliferative capability with only a small subset being able to undergo mitosis. This can lead to poor functional recovery after heart injuries such as myocardial infarctions (MI). We have previously demonstrated that inhibition of DYRK1A, a regulator of the DREAM complex and cell quiescence, increases cardiomyocyte cycling. However, the mechanism of DYRK1A inhibition that induces this increased cycling has yet to be elucidated. Therefore, we built a network model that encapsulates potential components that can lead to cardiomyocyte proliferation. Methods: We developed a logic-based differential equations model to simulate DYRK1A-mediated contributors of cardiomyocyte proliferation and G1/S phase transition. Various knock-down/knock-out and overexpression simulations were performed to predict targets of cell cycle regulation. Literature validation was performed to compare experiments across cancer, cardiomyocytes, and broader cell types to independent model simulations. A sensitivity analysis was used to identify the influence of particular proteins/genes on cardiomyocyte proliferation. Results: Our model simulations validated against (n = # of primary literature articles) 84.6% of literature experiments in cancer cell lines (n = 13), 88.8% in broader cell types (n = 9), and 100% in cardiomyocytes (n = 5). Simulated knockouts of DYRK1A and overexpression of Cyclin-D, Cyclin-E, and B-MyB increased DNA replication, indicating exit from G 0 phase. Sensitivity analysis showed that a DYRK1A knockout was the most influential in terms of increasing DNA replication. Conclusion: Our model’s predictions were successfully validated against literature in several different types of cell lines. Furthermore, it gives valuable insight on possible targets that can induce cardiomyocyte proliferation in addition to DYRK1A. Further in vitro and in vivo experiments are necessary to fully evaluate our simulations in cardiomyocytes.
Abstract Tu008: Efficient Intramyocardial Delivery of EV-Encapsulated AAVs to Target Cardiomyocytes in a Pre-Clinical Swine Model
Authors: Alex Gallinat, Sabrina La Salvia, Xisheng Li, Nikhil Raisinghani, shihong zhang, Alan Shao, Anh Phan, Spyros Mavropoulos, Kiyotake Ishikawa, Susmita Sahoo
Journal: Circulation Research (2024)

Background: Adeno-associated viruses (AAV) have been widely used to deliver therapeutic genes to the heart due to their safety and efficacy. However, a major obstacle to their successful gene delivery is the presence of neutralizing antibody (NAb) which form during natural exposure to AAV or following AAV administration. Our recent publication has shown that EV-encapsulated AAVs (EV-AAVs) shields AAVs from NAb neutralization, and overcoming this key roadblock associated with AAV-mediated gene therapy. Hypothesis: In this study, we addressed whether EV-AAVs can target cardiomyocytes in vivo . Aims: To test whether cardiomyocytes effectively uptake EV-AAVs in vivo , in a pre-clinical swine model. Methods and Results: Adult pigs prescreened for a lower NAb titer were randomly assigned to receive intramyocardial administration of either vehicle, AAV6-Luciferase or EV-AAV6-Luciferase. A total of six injections across three injection sites within the left ventricle (LV) were administered per animal. All animals were sacrificed at four weeks and liver and the LV tissues were collected. Luciferase activity assay, flow cytometry analysis of isolated cardiomyocytes (CM) and non-myocytes (NM), and immunofluorescence microscopy were performed to evaluate transgene expression and cellular tropism. Echocardiography was performed at baseline and at 4W to monitor cardiac function. At 4W, EV-AAV6s resulted in a higher luciferase expression in LV compared to nude AAV6 particles. Expression in the LV was higher compared to the liver. Flow cytometry analysis revealed a higher luciferase expression in CM compared to NM, which was corroborated by Immunofluorescence. No differences were detected in the global heart function across groups. Biodistribution of EV-AAV uptake and transgene expression are being studied with intramyocardial and intracoronary delivery mechanisms. Conclusion: EV-AAVs deliver higher amounts of genes than nude AAV6 via intramyocardial administration in a pre-clinical swine model. CMs preferentially uptake EV-AAVs compared to NMs. Our studies suggest that EV-AAVs are a superior cardiac gene delivery vector for future clinical studies.
Abstract Tu003: The angiotensinogen/vasorin ratio in peripheral blood, a biomarker for preeclampsia
Authors: Saravanakumar Murugesan, Michelle Tubinis, Mark Powell, Hanna Hussey, Lakshmi Saravanakumar, Ryan Melvin, Rachel Sinkey, James Mobley, Alan Tita, Tamas Jilling, Dan Berkowitz
Journal: Circulation Research (2024)

Background: Preeclampsia (PE) significantly impacts maternal and perinatal health worldwide. Despite limited treatment options, predictive tests could help in early diagnosis and personalized treatment/prevention strategies. Current biomarker, such as soluble fms-like tyrosine kinase-1 (sFlt-1) and placenta growth factor (PIGF) offer reasonable negative predictive values, but their use as positive predictive markers is limited. Using unbiased discovery proteomics on urine extracellular vesicles (EV), followed by verification by ELISA and western blotting we identified a novel biomarker candidate for PE, vasorin (VASN), which is significantly decreased in PE, along with a known mediator of PE, angiotensinogen (AGT), that was significantly increased in PE as biomarker candidates. Hypothesis: Levels of AGT, VASN and their ratios in peripheral blood can be utilized as biomarkers for PE. Goals/Aims: 1) To determine gestational age (GA)-dependence of AGT, VASN and AGT/VASN plasma levels in human and murine pregnancy. 2) To assess disease association of AGT, VASN and AGT/VASN in a cohort of pregnant women with PE with severe features (sPE) and gestational age-matched normotensive pregnant women (NTP) and in a murine model of PE. Methods: Plasma was collected from GA-matched NTP and sPE. Timed pregnant mice were injected with adenoviral vector (AD) encoding sFLT-1 on embryonic day 10.5 (mouse PE), or with AD encoding enhanced green fluorescent protein injection (mouse control). AGT and VASN were measured by ELISA in human samples and were quantified by western blotting in murine samples. Results: AGT and AGT/VASN showed negative correlations with GA, while VASN levels showed positive correlation with GA in NTP and in mouse controls. PlGF and sFLT-1 levels had weak/no correlations with GA. AGT and AGT/VASN were significantly higher in sPE and mouse PE, while VASN was significantly decreased in sPE and mouse PE as compared to NTP and mouse controls, respectively. To differentiate between sPE vs NTP with receiver operating characteristic curve (ROC) for AGT/VASN: AUC (0.90), Sensitivity =92%, Specificity = 85% at AGT/VASN&gt;81.6. Conclusion: AGT and AGT/VASN ratio increase, and VASN decrease can serve as biomarkers for PE.
Abstract Tu113: Metabolomic Profile of Human End-Stage Ischemic Cardiomyopathy Reveals Few Differences from Non-Ischemic Cardiomyopathy
Authors: Sho Tanosaki, Kenneth Bedi, Kenneth Margulies, Zoltan Arany
Journal: Circulation Research (2024)

Background: The heart is highly metabolic, relying on a range of fuels to sustain its high generation of ATP. The primary source of energy in the human heart is fatty acids, but fatty acid oxidation is reduced in failing hearts, contributing to a decrease in ATP production. Previously, we provided a comprehensive metabolic profile of human non-ischemic cardiomyopathy (NICM) hearts, including maintained ATP levels despite decreased energy charge (EC), lower acylcarnitines, acyl CoAs, and free fatty acids, suggesting a defect in fatty acid import. Research Questions: The pathogenesis of ischemic cardiomyopathy (ICM) is dramatically different from that of NICM, raising the question of whether metabolic defects differ between these two conditions. Aims: Our current study was therefore aimed at elucidating the metabolomic profile of end-stage ICM compared to that of NICM. Methods: Whole human hearts were obtained either from patients with end-stage heart failure (NICM or ICM) undergoing cardiac transplantation or from brain-dead organ donors. Hearts were arrested in situ and kept in ice-cold cardioplegia solution at all times. Transmural tissue samples were obtained from left ventricular free wall, snap frozen, and extracted metabolites were quantified by liquid chromatography coupled with mass spectrometer. Results: The comparison of metabolomics data from non-failing (NF) control heart samples and NICM samples were consistent with our previous study, with marked decrease in acylcarnitines and increase in lysophosphatidylcholine. On the other hand, the metabolomics profiles of ICM and NICM samples were remarkably similar. Phenylacetylglutamine and trimethylamine N-oxide, both related to atherosclerotic disease risk, were statistically significantly higher in ICM samples compared to NICM samples. Conclusion(s): The human cardiac metabolome of end-stage ICM is nearly identical to that of NICM, despite dramatically different pathogeneses, suggesting that end-stage heart failure constitutes a final common pathway. It would be of great interest to evaluate metabolomic profiles of the diseases differ at earlier timepoints.
Abstract Tu091: Mechanism of beta1-adrenergic receptor signaling by IgG3 subclass of autoantibodies
Authors: Maradumane Mohan, Chia-Feng Liu, Anushruti Ashok, Dhanachandra Singh Khuraijam, Sadashiva Karnik, Wai Hong Tang, Sathyamangla Naga Prasad
Journal: Circulation Research (2024)

Autoimmune response to self-antigens results in autoantibodies (AAbs), wherein AAbs against extra cellular loop (ECL) 2 of β1-adrenergic receptor (β1AR) is known to underlie dilated cardiomyopathy (DCM). Contrarily, recent studies show that patients with β1AR AAbs belonging to the IgG3 subclass have beneficial outcomes. However, the signaling mechanisms that underlie the beneficial outcomes are not well understood. We have previously shown that IgG3(+) β1AR AAbs facilitates uniquely biased β1AR signaling in response to β-blocker compared to non-IgG3(+) β1AR AAbs. Since IgG3(+) β1AR AAbs are associated with favorable patient outcomes, it is critical to determine the mechanistic basis of the unique beneficial signaling pathway that may have therapeutic potential. HEK 293 cells stably expressing human β1AR (HEK-β1AR) was generated and treated with β1AR AAbs. Minimal changes in phosphorylation of β1AR was observed following IgG3(+) β1AR AAbs, while significant phosphorylation of β1ARs was observed with non-IgG3(+) β1AR AAbs. To dissect the diverse signaling responses mediated by IgG3(+) and non-IgG(+) β1AR AAbs, HEK-β1AR cells were treated with IgG3(+) or non-IgG3(+) β1AR AAbs. β1ARs were immunoprecipitated and subjected to mass-spectrometry analysis to identify unique co-immunoprecipitating proteins that could complex with receptor in presence of IgG3(+) or non-IgG3(+) β1AR AAbs. One of the proteins that was differentially recruited to the β1AR complex was Insulin Receptor Substrate 4 (IRS4). Traditionally, IRS4 is a signaling hub downstream of insulin/insulin growth factor receptors mediating glucose uptake. The differential interaction of IRS4 with β1AR in the presence of IgG3(+) vs. non-IgG3(+) β1AR AAbs reflects that IRS4 could be a key signaling regulator of uniquely biased signal transduction pathway in the presence of IgG3(+) vs. non-IgG3(+) β1AR AAbs. Mechanisms underlying this unique IRS4 mediated signaling modulation will be discussed.
Abstract Tu096: The Aryl Hydrocarbon Receptor Agonist, L-Kynurenine, Modulates Cardiac Fibroblast Activation and Antigen Presentation
Authors: Brandon Theall, Sasha Smolgovsky, Abraham Bayer, Erin Sanders, Maria Zambrano, Zachary Robbe, mark aronovitz, Kuljeet kaur, Pilar Alcaide
Journal: Circulation Research (2024)

Background: The Aryl Hydrocarbon Receptor (AHR) is a ubiquitously expressed transcription factor that regulates the cellular response to external stimuli through the binding of ligands including tryptophan metabolites such as L-Kynurenine (L-Kyn). Alterations in tryptophan metabolic pathways and downregulation of myocardial AHR are associated with cardiac inflammation and fibrosis in experimental heart failure (HF) induced by transverse aortic constriction (TAC). We hypothesized cardiac fibroblast (CFB) AHR and its agonism by L-Kyn prevent the pro-fibrotic and pro-inflammatory activity of CFBs in experimental HF. Methods and Results: We performed TAC or sham surgery on Ahr -/- mice and WT littermates for 4 weeks and found that Ahr-/- mice showed significantly decreased contractile function measured by echocardiography, and increased collagen deposition in left ventricular histological sections after TAC compared to WT mice. We performed bulk RNA sequencing in sorted CFB (CD31-CD45-MEFSK4+) from Ahr -/- and WT mice. Gene set enrichment analysis showed Ahr -/- CFB had increased expression in pathways for fibroblast activation, chemokine production, and immune cell recruitment. To investigate the role of L-Kyn agonism of CFB AHR in transformation and contractile function, murine CFB were treated with Transforming Growth Factor β (TGFβ) (100ng/ml) or seeded in collagen discs, in the presence or absence of L-Kyn(100ug/ml). L-Kyn attenuated TGFβ mediated induction of αSMA and collagen 1 protein expression by immunofluorescence and western blot, as well as decreased disc contraction 72h after mechanical activation and release. The role of AHR agonism by L-Kyn on the pro-inflammatory function of CFB was determined in WT CFB treated with IFNγ (100U/ml) in the presence or absence of L-Kyn. IFNγ induction of chemokines Cxcl9 and Cxcl10, as well as MHC-II surface expression were attenuated by L-Kyn, so was T cell adhesion to CFB in co-culture assays. In contrast, L-Kyn had no effect on the ability of CFB to take up and process antigens, as determined by DQ-Ovalbumin internalization quantified by flow cytometry. Conclusions: Our results demonstrate that AHR regulates activation and pro-inflammatory signatures in CFB at baseline, and that stimulation of CFB transformation and pro-inflammatory activity is dampened by agonism with L-Kyn. Current studies are aimed at investigating AHR agonism in the context of pressure overload induced HF in CFB reporter mice.