bluuebunny/crossref_metadata_2025_split huggingface dataset:
Abstract 42: A Talin Mutant that Disrupts Talin-integrin Binding in Platelets Decelerates aIIbß3 Activation Without Pathological Bleeding
Authors: Stefanini Lucia , Snider Adam , Piatt Raymond , Petrich Brian , Bergmeier Wolfgang 
Published: July 2021

Tight regulation of integrin affinity is a critical component of hemostasis. The final step of integrin inside-out activation is talin binding to two distinct sites within the integrin β cytoplasmic tail, an NPLY sequence and a membrane-proximal region (MPR). Selective deletion of talin1 from platelets (Tln1fl/flPf4-Cre+) completely prevents integrin activation and thrombus formation, and results in profound defects in hemostasis. Interestingly, platelet-specific expression of a talin1 mutant (L325R) that lacks the ability to bind the MPR, but can still bind the NPLY sequence, phenocopies talin1-knockout platelets. In this study we sought to analyze the effects on thrombosis and hemostasis of a platelet-specific talin1 mutation (W359A) that dramatically impairs binding to the NPLY sequence and does not rescue the talin1 deficiency in transfected cells.
Homozygous knock-in of talin(W359A) is embryonic lethal in mice. Thus, Tln1W359A/wt mice were crossed with Tln1fl/flPf4-Cre+ mice to generate Tln1W359A/flPf4-Cre+ (TlnWA) and Tln1wt/flPf4-Cre+ (Tlnwt) control mice. Expression of talin(W359A) in platelets partially rescued talin1 deficiency. Compared to Tlnwt controls, agonist-induced αIIbβ3 activation was reduced by ~50% and spreading on fibrinogen was only moderately impaired in TlnWA platelets. However, kinetic studies demonstrated decelerated αIIbβ3 activation in TlnWA platelets, which resulted in delayed aggregation under static conditions and reduced thrombus size at low shear rates. Interestingly, adhesion of TlnWA platelets to collagen at high shear rates was not significantly better than that of talin-deficient cells and TlnWA mice were completely protected from FeCl3-induced carotid artery occlusion. Lastly, in contrast to platelet-specific talin-knockout or TlnLR mice, TlnWA mice showed no detectable gastrointestinal bleeding and only modestly increased tail-bleeding times compared to littermate controls.
In conclusion, we demonstrate that the talin(W359A) mutation decelerates, but does not abolish, talin-dependent integrin activation in platelets. Our studies further suggest that this delay in integrin activation is protective from pathological vessel occlusion while it does not affect hemostasis in mice.
Abstract 422: Atherosclerosis and Rage Expression by Histology and Imaging in Hyperlipidemic Pigs
Authors: Johnson Lynne , Tekabe Yared , Kollaros Maria , Eng George M , Bhatia Ketan , Groner Rachel B , Krueger Christian G 
Published: July 2021

Purpose and methods

To characterize atherosclerotic lesions and RAGE expression in a large animal model and investigate targeting RAGE for in-vivo imaging. Nine hyperlipidemic pigs at 12 mo of age were studied. LDL averaged 513 ± 74 mg/dL and final wt was 63.3 ± 12.4 kg. Pigs were injected with 16.3 ± 3.5 mCi of
99m
Tc anti-RAGE F(ab’)
2
and after 8 hr (blood pool clearance) underwent SPECT/CT imaging of the neck and thorax and then euthanized, the aorta including innominate, proximal carotids, and proximal ileofemoral branches and coronaries were dissected. Vascular tissue was divided into 36 anatomical segments: coronaries: 4 each x 3 = 12, aortic root: 3, aortic arch: 4, thoracic aorta: 3 and abdominal aorta: 3, each carotid and femoral: 2. Sections from each segment were counted and stained for H&E and RAGE. Scans were analyzed for tracer uptake in organs and vascular structures and counts converted in %ID. All sections were classified according to AHA criteria, % stenosis, and % RAGE staining was quantified. One HL pig received non-immune control antibody and 2 farm pigs were injected for blood pool clearance and SPECT imaging.

Results

On pathological examination 75/324 segments or 23% showed class 2 lesion or greater. All class III or IV lesions were in the coronaries, carotids, and femoral arteries and % stenosis ranged from 16 to 65%. In vivo biodistribution showed intense uptake of the antibody in the lungs (15 %ID) indicating high constitutive expression of RAGE and precluded visualizing the thoracic aorta. Heart uptake was low (1.3 x 10
-4
%ID) providing visualization of focal coronary uptake and the carotids were well seen. Focal tracer uptake was visualized on SPECT scans in these 2 vascular territories for all vessels with class III or IV lesions and uptake ranged from 4 -22 %ID x 10
-4
. Correlations were R = 0.85, P = 0.002 for %ID vs. %ID/g and R = 0.83, P = 0.002 for %ID/g vs. % RAGE staining.

Conclusions
There is variability in extent and severity of atherosclerotic disease in HL pigs at one year. RAGE staining is found in all class II-IV lesions and correlated with uptake of radiolabeled antibody targeting RAGE. High RAGE expression in lungs limited imaging the aorta but carotid and coronary artery uptake was seen on SPECT scans.
Abstract 43: The Role of Epsins in Atherosclerosis
Authors: Chen Hong 
Published: July 2021

Background

Epsins are a family of ubiquitin-binding, endocytic clathrin adaptors. Mice lacking both epsins 1 and 2 (Epn1/2) die at embryonic day 10 and exhibit an abnormal vascular phenotype. We have recently published that epsins function as regulators of tumor angiogenesis by controlling the endocytosis of VEGFR2 (
J Clin Invest.
2012). The goal of this project is to define the novel role of epsins in endothelial cells (EC) in regulating inflammation-mediated atherogenesis.

Methods and results
To examine the role of endothelial epsins in atherogenesis, we engineered mice with constitutive deletion of epsins in EC (EC-DKO). Strikingly, these mice are viable but display markedly attenuated atherogenesis on an ApoE null background in response to hypercholesterolemia. Oil red O staining revealed that ApoE-/- mice with endothelial deletion of epsins had a significant reduction in atherosclerotic lesion area in aortic root and macrophage infiltration compared to ApoE-/- mice after being fed a western diet for 10 weeks. FACS analysis revealed that deletion of epsins greatly reduced TNFα and LPS-induced expression of adhesion molecules (ICAM & VCAM), selectins (P- & E-), CCR2 and MCP-1 in primary mouse aortic EC (MAEC) isolated from EC-DKO mice. Mechanistically, epsins promote TNFR/TLRs signaling and NF-κB and MAPK (JNK and p38) activation by stabilizing the receptor complex formation and enhancing recruitment of NEMO, a crucial player in NF-kB activation. Conversely, epsin deficiency does not affect the surface expression of TNFR1/TLR2/4 by FACS analysis, suggesting that epsins may be dispensable for endocytosis of TNFR1/TLR2/4. We also observed that a synthetic peptide comprising the epsin ubiquitin-interacting motif (UIM) potently disrupts the association of epsins with TNFR/TLR signaling complex in vitro, suggesting a potential pharmacological inhibitory role for this peptide in controlling the atherosclerosis in vivo. Our data demonstrate that epsins control atherogenesis by regulating TNFR/TLR signaling in endothelial cells and provide a potential novel strategy to perturb atherogenesis.

Conclusion
Our results demonstrated an essential role of epsins in atherogenesis by regulating TNFR/TLR signaling.
Abstract 500: Basal Peripheral Arterial Blood Flow Increases Concomitantly with Decreased Cardiopulmonary Reserve and Biomarkers Activation in Patients with Heart Failure and Preserved Ejection Fraction
Authors: Vitiello Damien , Harel François , Touyz Rhian M , Sirois Martin G , Lavoie Joel , Myers Jonathan , Gayda Mathieu , Chabot-Blanchet Malorie , Rouleau Jean-Lucien , De Denus Simon , White Michel 
Published: July 2021

Background
The underlying pathophysiology of heart failure with preserved ejection fraction (HFpEF) remains poorly understood. Changes in peripheral arterial function concomitantly with the evaluation of cardiopulmonary reserve and biomarkers related to wall stress, extra-cellular matrix turnover (ECM), subclinical inflammation and oxidative stress have not been investigated in patients with HFpEF nor compared with age-matched healthy volunteers (HV). We hypothesized that patients with HFpEF would have impairments in cardiac reserve with reduced peripheral arterial function associated with broad-spectrum biomarkers activation.

Methods
Eighteen male and female HFpEF patients (aged 70 ± 9 yr) NYHA class II and III were recruited. Data from patients were compared with those from 14 age and sex matched HV. A maximal exercise testing with gas exchange analysis was completed on a treadmill using a RAMP protocol and heart rate recovery (HRR) was measured at 1 and 2 minutes following exercise. Peripheral arterial function was assessed using near infrared radionuclide plethysmography. Biomarkers included BNP, NT-proBNP, hsCRP, TBARS, 8-epi-prostaglandin F2α, MMP 1, 2, 9 and TIMP 1, 2, 3, 4) were analyzed.

Results
Selected data are presented in the Table 1 and 2.

Conclusions
Compared to healthy volunteers, patients with HFpEF demonstrated a significant decrease in aerobic capacity but an increase in basal peripheral arterial blood flow, subclinical inflammation and oxidative stress. The increase in resting arterial blood flow may be a compensatory mechanism for the decrease in cardiac reserve and the pro-inflammatory/oxidant milieu in these patients.
Abstract 517: Effect of Hepatocyte-specific Depletion of AT1a Receptors on Atherosclerosis and Aneurysms
Authors: Rateri Debra L , Balakrishnan Anju , Howatt Deborah A , Moorleghen Jessica J , Cassis Lisa A , Daugherty Alan 
Published: July 2021

Background and Objective
Whole body deficiency of AT1a receptors (AT1aRs) ablates angiotensin II (AngII) induced vascular pathologies that include aortic aneurysms and atherosclerosis. We created AT1aR floxed mice to determine the role of specific cell types expressing this receptor on these vascular pathologies. Unexpectedly, these mice demonstrated a lack of effect of smooth muscle specific AT1aR deficiency on the development of atherosclerosis or aneurysms. Also, endothelial-specific AT1aR deficiency had no effect in atherosclerosis and minimal effect on aneurysms. The purpose of this study was to determined if AngII infusion was releasing substances from the liver that influence vascular pathology.

Methods and Results
Hepatocyte depletion of AT1aRs was accomplished using two methods of introducing Cre into AT1aR floxed mice. One approach was injection of adenovirus Cre (AdCre) and the other by breeding to mice expressing Cre under the control of the albumin promoter. To induce atherosclerosis, groups of wild type and hepatocyte-specific AT1aR depleted male (N=15-21/group) and female (N=21/group) mice were fed a saturated fat enriched diet for 12 weeks. Hepatocyte-specific depletion of AT1aRs was confirmed using detection of Cre and absence of AT1aRs by PCR and RT-PCR, respectively. Body weights, serum cholesterol concentrations, and systemic blood pressures were not different between wild type and hepatocyte-specific AT1aR depleted mice grouped by gender and genotype. Atherosclerotic burden was similar in all groups. To examine AngII-induced atherosclerosis, male wild type and hepatocyte-specific AT1aR depleted mice (N=11-16/group) were fed a fat enriched diet for 5 weeks, and AngII (1,000 mg/kg/min) was infused using osmotic mini-pumps for 28 days. AngII infusion increased systolic blood pressure, promoted atherosclerosis and induced aneurysms to a similar degree in both genotypes.

Conclusion
Deletion of hepatocyte-specific AT1aRs did not affect dietary or AngII-induced vascular pathologies.
Abstract 54: Peptidylarginine Deiminase 4-dependent Generation of Neutrophil Extracellular Traps is Crucial for Deep Vein Thrombosis in Mice
Authors: Martinod Kimberly , Demers Melanie , Fuchs Tobias A , Wong Siu Ling , Brill Alexander , Gallant Maureen , Wang Yanming , Wagner Denisa D 
Published: July 2021

Introduction
Histone hypercitrullination by the enzyme peptidylarginine deiminase 4 (PAD4) leads to nuclear chromatin decondensation that is needed for neutrophil extracellular trap (NET) formation. NETs consist of chromatin and granule proteins that are released into the extracellular environment. NETs were shown to be involved in thrombosis by promoting coagulation and platelet adhesion and were identified in the thrombus scaffold in animal models of deep vein thrombosis (DVT).

Objective
Whether NETs are involved in the pathogenesis of DVT or whether they are merely a consequence of neutrophil recruitment to the thrombus is unknown. We hypothesized that NET formation would be impaired in PAD4-deficient mice during deep vein thrombosis and that this may affect thrombus formation and/or stability.

Methods
PAD4-deficient mice are incapable of citrullinating histones and therefore fail to decondense chromatin during NETosis. We performed the inferior vena cava stenosis model of DVT in wild-type or PAD4-/- mice. Intravital microscopy was done to assess leukocyte vessel wall interaction in PAD4 deficiency.

Results
We induced NET formation in isolated peripheral blood mouse neutrophils with ionomycin and found that PAD4-/- neutrophils had a complete inability to produce NETs (WT, 20.65±2.61% NETs; PAD4-/-, not detected. n=4). Leukocyte-endothelial interactions in PAD4-/- mice were not impaired upon induction of systemic Weibel-Palade body release (WT, 55.2±11.8; PAD4-/-, 62.0±17.5 cells/min, n=5-6). In the DVT model, while a majority (9/10) of wild-type mice formed a thrombus 48 hours after stenosis, only 1 of 11 PAD4-/- mice formed a thrombus. Thrombus formation could be rescued by infusions of isolated WT bone marrow neutrophils into PAD4-/- mice, and extracellular H3Cit+ areas were seen within these thrombi. This data suggests that neutrophil PAD4 was essential for thrombus formation in deep veins.

Conclusion
NETs comprise a crucial part of the pathologic thrombus scaffold, and here we report that the lack of NETs inhibits pathological thrombosis. Chromatin decondensation initiated by PAD4 in neutrophils is a key player in the formation of deep vein thrombi and targeting neutrophil histone modification could be a new way to prevent DVT.
Abstract 540: Loss of Function Studies Highlight the Vasculoprotective Effect of A20 in Transplant Arteriosclerosis
Authors: Lee Andy , Peterson Clayton R , Moll Herwig , Csizmadia Eva , Daniels Kathleen , Notartomaso Ashley , Neidich Alon , Chien Victor , DaSilva Cleide G , Ferran Christiane 
Published: July 2021

Transplant arteriosclerosis (TA) is an accelerated form of atherosclerosis largely driven by the immune response to the allograft. As for all atherosclerotic disease, inflammation is central to vascular remodeling associated with TA. We have shown that overexpression of the NF-κB inhibitory protein in totally mismatched (C57BL/6 to BALB/c) aortic to carotid vascular allografts protects from TA. A20 inhibits TA thanks to its anti-inflammatory and anti-apoptotic functions in endothelial cells (EC), anti-inflammatory and anti-proliferative functions in smooth muscles cells (SMC), pro-apoptotic function in neointimal SMC, and to favoring regulatory FOXP3+ over pathogenic Th1/Th17 T cells.
In this work, we analyzed the impact of A20 knockdown on TA by using aortic allografts from A20 heterozygous (A20-+/- HET) mice. Partial loss of A20 in aortic allografts caused significantly greater TA lesions, measured by intima over media ratios (I/M 1.79±0.18 in HET vs. 1.04±0.17 in wild type (WT) allografts; p<0.01). This was associated with higher inflammation as evaluated by VCAM-1 immunostaining and granulocyte infiltration, lower expression of the anti-inflammatory and tolerogenic cytokine IL-10, and greater incidence of thrombosis (3/11 vs. 0/10) in HET vs. WT allografts. These results are particularly relevant to newly discovered A20 single nucleotide polymorphisms (SNPs) that are less effective in inhibiting inflammation, and have been associated with increased risk of coronary artery disease in diabetic patients.
Based on these results, we conclude that A20 plays an important physiologic role in the vascular remodeling of TA. We propose that tailored A20-based vascular therapies could decrease incidence and severity of TA, and synergize with or minimize use of immunosuppression in solid organ transplantation.
Abstract 565: Nuclear Receptor NR4A1 Represses SDF1a expression in macrophages and Reduces Atherosclerosis.
Authors: Hamers Anouk A 
Published: July 2021

Rationale

NR4A1 is a nuclear receptor, also known as Nur77 and is expressed in human atherosclerotic lesions in macrophages, endothelial cells, T cells and smooth muscle cells. NR4A1-knockout (NR4A1
-/-
) mice lack Ly6C
-
monocytes, but have normal LyC6
+
numbers. Macrophages play a critical role in atherosclerosis and we have demonstrated that NR4A1 has an anti-inflammatory function in THP-1 macrophages. The aim of the current study is to assess the function of this receptor in myeloid cells in atherosclerosis.

Objective
This study aims to delineate the function of Nuclear Receptor NR4A1 in macrophages in atherosclerosis.

Methods and results

Bone marrow-derived macrophages (BMM) from wild-type and NR4A1
-/-
mice were cultured and classically activated with LPS or alternatively activated with IL4. NR4A1
-/-
BMM exhibit changed expression of M2-specific markers and a pro-inflammatory polarization in response to LPS with enhanced expression of IL12, IFNγ, and SDF1α. Nitric oxide synthesis is also strongly induced in (non)-stimulated NR4A1
-/-
BMM. SDF1α is a potent chemotactic factor for B cells and monocytes. The chemoattractive activity of NR4A1
-/-
BMM is abolished by SDF1α inhibiting antibodies as well as by overexpression of NR4A1 in the NR4A1-deficient cells. Luciferase experiments show that NR4A1 binds directly to several response elements present in the mouse and human SDF1α promoter. A ChIP assay confirms these results. The effect of bone marrow-specific NR4A1-deficiency on atherosclerosis was studied in low density lipoprotein receptor-deficient (Ldlr
-/-
) mice. Ldlr
-/-
mice with a NR4A1
-/-
bone marrow develop 2.1-fold larger lesions than wild type bone marrow transplanted mice; containing more macrophages, T cells, smooth muscle cells and larger necrotic cores. SDF1α expression is also higher in these lesions, which may explain the observed aggravation of lesion formation, since SDF1α is not only a chemoattractant for monocytes but also enhances smooth muscle cell proliferation.

Conclusion

In macrophages the nuclear receptor NR4A1 has an anti-inflammatory function, represses SDF1α expression and, most importantly, bone-marrow transplantation studies in Ldlr
-/-
mice revealed that NR4A1 inhibits atherosclerosis.
Abstract 66: SR-BI Mediates Cholesteryl Ester (CE) Sorting in Tetraspanin Microdomains with CD81 Facilitating CE Nano-endocytosis and CE Transport to Lipid Droplets
Authors: Bocharov Alexander V , Vishnyakova Tatyana G , Baranova Irina N , Lizunov Vladimir A , Zimmerberg Joshua , Eggerman Thomas L , Patterson Amy P 
Published: July 2021

SR-BI functions as a neutral lipid transporter facilitating cholesteryl ester (CE) sorting, internalization and accumulation in LDs. The mechanism of CE transport remains obscure. We hypothesized that HDL lipid transport requires CD81 as a co-receptor and tetraspanin enriched domains (TED) for CE sorting, similar to hepatitis C uptake.
Utilizing BODIPY-CE labeled recombinant HDL (rHDL/BP-CE) based on either human apoA-I or an apoA-I mimetic peptide, L-37pA, and confocal and TIRF microscopy, we found that after initially binding to SR-BI at the cellular filopodia and lamellipodia, HDL CE was laterally transported to the bases of filopodia, accumulating in 1-2 μm plasma membrane microdomains. While CE was rapidly internalized, neither apoA-I, L37pA nor SR-BI was, suggesting that these microdomains are an important part of the CE sorting mechanism. Internalized CE was transported in 50-100 nm CE-containing intracellular nano-vesicles. In a process of multiple collisions potentially involving transient nano-vesicle/LD fusion, BODIPY-CE was transferred to LDs. To identify CE-sorting microdomains, SR-BI expressing HeLa cells were transfected with GFP-CD81 or GFP-CD82. After a 1 h incubation with rHDL/BP-CE, CE was co-localized with both tetraspanin proteins demonstrating that these are TED (the observation was confirmed by anti-CD81 and -CD-82 antibody immunostaining). Following a 2 h chase, only GFP-CD81, not CD82, was found internalized and co-localized with CE transporting nano-vesicles. CE transport was not affected by inhibitors of de novo neutral lipid synthesis such as the ACSL-3 inhibitor, triacsin c, ACAT inhibitor, Sandoz 58-035 or ATGL inhibitor, A926500, indicating that CE was directly transported through a novel nano-endocytosis mechanism involving CD81 and not requiring CE hydrolysis and de novo synthesis.
We conclude that TED function as CE sorting sites where CD81 helps to form nano-vesicles delivering CE to LDs. Further understanding of their function will help develop novel anti-atherogenic compounds accelerating reverse cholesterol transport.
Abstract 108: Diagnostic and Prognostic Biomarker Potential of miR-24 in Abdominal Aortic Aneurysm Disease and Rupture
Authors: Chernogubova Ekaterina , Eken Suzanne , Spin Joshua , Jin Hong , Raaz Uwe , Roy Joy , Eriksson Per , Tsao Philip , Maegdefessel Lars 
Published: July 2021

MicroRNAs (miRNAs) have lately received much attention regarding their suitability and feasibility as biomarkers for cardiovascular diseases. Aim of this current study was to explore the diagnostic and prognostic value of detecting circulating levels of miRNAs in abdominal aortic aneurysms (AAAs).
Using a PCR-based array, we profiled the 168 most abundant blood miRNAs in 20 patient plasma samples with AAA disease, undergoing surgical repair of their enlarged aorta vs. 20 samples from a matched screening cohort without aneurysm. We were able to identify a total number of 12 miRNAs that were significantly altered in diseased patient samples as compared to controls. Next we investigated these 12 miRNAs in plasma from ApoE-/- mice with angiotensinII (AngII)-infusion induced AAAs, in order to determine the potential prognostic value of miRNAs being released into circulation. Indeed we were able to detect that the expression of 4 out of the 12 miRNAs (miRs-126 and -668 both increased; miRs-24 and -210 both decreased), was substantially altered in plasma samples drawn from AAA mice immediately before rupture occurred between days 10 and 14 after AngII pump implantation compared to mice with AAA that did not rupture for the remainder of study (28 days), as well as saline-infused controls. The expression of miR-24 was furthermore significantly different in plasma samples from human patients with acutely ruptured AAAs (n=7) compared to patients with non-ruptured AAAs (AA diameter between 55-78 mm; n=7) undergoing surgical repair, as well as un-diseased controls (n=7). Using the AngII model, as well as the elastase infusion model, we were able to show that overexpression of miR-24 protects from aneurysm progression as well as aortic rupture (in the AngII model) by repressing the expression of its target gene chitinase3-like1 (Chi3l1), which regulates macrophage survival and cytokine release in expanding murine AAAs.
The present study explores the biomarker potential of miRNAs being released into circulation during initiation, propagation, and ultimately rupture of AAA disease in mice and humans. The identification of miR-24 potentially offers prognostic value to determine which patients present with increased risk of rapid AAA expansion and subsequent rupture.