bluuebunny/crossref_metadata_2025_split huggingface dataset:
S1P receptor-1 signaling reduces arterial thrombosis by upregulation of endothelial thrombomodulin expression
Authors: Mourikis P H I L I P , Benkhoff M A R C E L , Barcik M A I K E , Knoop B E T U E L , Trojovsky K A J E T A , Ahlbrecht S A M A N T , Metry S A R A , Helten C A R O L I , Dannenberg L I S A , Hohlfeld T H O M A S , Zeus T O B I A S , Kelm M A L T E , Levkau B O D O , Polzin A M I N 
Published: November 2023

Abstract

Background
Sphingosine-1-phosphate (S1P) is a bioactive lipid known for its cardioprotective effects. The extent to which S1P also has effects on arterial thrombosis has not yet been fully elucidated. Another important factor in arterial thrombosis is thrombomodulin (TM), an important endothelial antithrombotic factor. To what extent S1P, TM, and, in addition, platelet adhesion as an initial part influences arterial thrombosis is unclear and the aim of this work.

Methods
The influence of S1P on human umbilical cord venous endothelial cells (HUVECs) was investigated in cell culture. To investigate effects of S1P and TM on platelet adhesion, flow chamber experiments were performed. In addition, murine arterial thrombus formation was analyzed by intravital microscopy.

Results
S1P increased TM expression on HUVECs. This effect was abolished by S1P receptor1 (S1PR1) inhibition with W146 (S1P: 1.081 ± 0.091 vs. S1P + W146: 0.951 ± 0.073, normalized to control, p&lt;0.0001). Sphingosine kinase-2 deficient mice (SphK1-/-) lack the kinase primarily producing sphingosine. Accordingly, these animals have less S1P. Here, TM expression was significantly lower in the aorta compared to wild-type mice (WT: 207.2 ± 45.8 pg/g aorta vs. SphK1-/- 124.5 ± 40.2 pg/g aorta, p&lt;0.0001). S1P treatment decreased platelet adhesion from whole blood to endothelial cells in the flow chamber (Con: 100.00 ± 28.09% vs. S1P: 80.91 ± 22.85%, p&lt;0.0001). W146 reversed this effect by S1PR1 inhibition (S1P: 80.91 ± 22.85%, vs. S1P + W146: 118.60 ± 33.65%, p=0.0067). Platelet adhesion to endothelial cells was assessed in the presence of a TM-neutralizing antibody. The TM antibody abolished the anti-adhesive effects of S1P (S1P: 54.79 ± 29.59% vs. S1P+AB: 105.54 ± 35.43%, p=0.0174). TM inhibited intravital murine arterial thrombus formation (time to first occlusion: con: 9.38 ± 1.02 min vs. TM: 60 ± 0 min, p=0.0286). Furthermore, the duration of the first occlusion was significantly shorter and the embolic rate as a marker for thrombus stability was higher.

Conclusion
We demonstrated that S1P inhibits arterial thrombosis via regulation of endothelial TM. In particular, platelet adhesion to endothelial cells is reduced. This effect is mediated via S1PR1. The extent to which S1PR1 represents a potential antithrombotic target must be clarified in future studies.
Sirtuin-1 directly binds and deacetylates hepatic PCSK9 to inhibit low-density lipoprotein receptor degradation
Authors: Velagapudi S , Xavier Miranda M , Mohammed S , Kraler S , Robert J , Rohrer L , Lee H , Jang H D , Camici G , Von Eckardstein A , Kim H S , Akhmedov A , Luscher T 
Published: November 2023

Abstract

Background
Low-density lipoprotein cholesterol (LDL-C) is causally involved in atherosclerotic cardiovascular disease (ASCVD). We have previously shown that pharmacological activation of Sirtuin-1 (SIRT1), a NAD+-dependent deacetylase, reduces plasma LDL-C levels by decreasing PCSK9 and increasing hepatic LDLR expression, thus exerts atheroprotective effects in mice; however, the mechanism of SIRT1 action remains elusive.

Purpose
We found diminished levels of circulating SIRT1 in atherosclerotic mice compared to wild-type healthy controls. We hypothesized that this decrease in SIRT1 levels may promote dysregulated lipid metabolism and ASCVD in humans. Herein, we aimed to restore the reduced systemic Sirtuin-1 plasma levels in ASCVD by injecting recombinant murine Sirtuin-1 (rmSIRT1) in atherosclerotic mice.

Methods
Twelve-week-old apolipoprotein E-deficient (ApoE-/-) male mice fed a high-cholesterol diet (1.25% w/w) were randomized to receive rmSIRT1 (n=6; 0.3mg/kg BW i.p.) or vehicle treatment (n=6) every third day over 4-weeks. Mice were euthanized and organs were harvested to study lipid metabolism and ASCVD progression. Human hepatocytes (Huh7) cells were used to perform radioactive assays to study cellular uptake of radioiodinated LDL, surface plasmon resonance to study binding and site-directed mutagenesis to dissect the underlying molecular mechanism exerted by circulating SIRT1.

Results
Boosting circulating SIRT1 levels increased hepatic LDLR expression, reduced plasma LDL-C levels and progression of plaques in ApoE-/- mice (Figure 1). rmSIRT1 treatment did not change hepatic expression of total PCSK9 but increased its deacetylated status. Mechanistically, rmSIRT1 directly bound to hepatic PCSK9 and deacetylated PCSK9 at 3 sites: Lys243, Lys421 and Lys506 as shown by Surface Plasmon Resonance and mass spectrometry, respectively. In vitro mutagenesis to triple deacetylation mimetic (3KR) in hepatocytes (Huh7 cells) reduced SIRT1-induced PCSK9 activity, as evidenced by increased cellular binding and association of 125I-LDL through LDLR in Huh7cells. Finally, plasma levels of SIRT1 and PCSK9 were assessed at baseline in patients with acute coronary syndromes (ACS) where plasma SIRT1 showed an inverse correlation with PCSK9 and conferred a reduced risk of major adverse cardiovascular events.

Conclusion
SIRT1 directly binds PCSK9 and decreases its activity by deacetylation, thereby enhancing LDL-C clearance by hepatic LDLR upregulation. Increased circulating SIRT1 exerts atheroprotective effects and is associated with improved prognosis in patients with ACS.Figure 1
Canagliflozin enhances endogenous eliminatiton of senescent cells through downregulation of PD-L1
Authors: Katsuumi G , Suda M , Shimizu I , Yoshida Y , Furuuchi R , Hsiao C L , Hsiao Y T , Minamino T 
Published: November 2023

Abstract
Sodium-glucose co-transporter 2 (SGLT2) inhibitors have been developed as a new class of anti-diabetic drug, and shown cardio- and renal-protective effects in patients with diabetes, heart failure, and chronic kidney diseases. Also, there's evidence that canagliflozin, one of the SGLT2 inhibitors has expanded lifespan in aged mice, but their detailed protective machinery is still to be clarified. Here we demonstrate that canagliflozin has a effect to eliminate senescent cells from pathological tissues, so-called "senolysis" in some murine disease models and alleviates their pathology.
We treated canagliflozin mixed in food at 0.03% (w/w) to high-fat-diet(HFD)-fed obese mice and found that canagliflozin reduced senescent cells in visceral adipose tissue within 1 week. Also, we found that senolytic effect of canagliflozin was provided through enhancing endogenous senolytic function by NK or T cells. This machinery was mediated by enhanced AMPK signaling, which leads to the downregulation of PD-L1 expression on senescent cells. Furthermore, we treated canagliflozin to ApoE-knockout atherosclerotic mice with western diet feeding, progeroid mice and chronological aged mice. Canagliflozin reduced their senescent cell burden, resulting in alleviation of atherosclerosis, extended lifespan of progeroid, and altered physical function due to chronological aging.
Our results would be promising evidence that SGLT2 inhibitors can be used as not only cardio- or renal-protective drugs but also anti-aging drugs.
The COVID-19 mRNA vaccine Comirnaty induces anaphylactic shock in an anti-PEG hyperimmune large animal model
Authors: Barta B A , Radovits T , Dobos A B , Kozma G T , Meszaros T , Berenyi P , Facsko R , Merkely B , Szebeni J 
Published: November 2023

Abstract

Background
The polyethylene-glycol (PEG)-containing COVID-19 mRNA vaccines can cause hypersensitivity reactions (HSRs), or rarely, life-threatening anaphylaxis in a small fraction of patients. The role of anti-PEG antibodies (Abs) has been proposed, but causality has not yet been proven in an animal model.

Purpose
The aim of this study was to provide such evidence using an anti-PEG hyperimmune porcine model wherein the Ab levels are increased by PEGylated liposomes called Doxebo. We also aimed to find evidence for a role of complement activation and thromboxane A2 release in blood.

Methods
Experimental pigs (n=6) were immunized with 0.1 mg/kg BW Doxebo i.v., and the rise of anti-PEG Abs were measured with ELISA. 2-3 weeks later the animals were injected with 1/3 human dose of Pfizer’s COVID-19 mRNA vaccine Comirnaty i.v., and the hemodynamic, hematological and blood immune mediator changes were measured to assess the severity of hypersensitivity reaction (HSR). The endpoints were PAP, SAP, HR, EtCO2, WBC, granulocyte, lymphocyte and platelet counts, C3a and plasma TXB2.

Results
The level of anti-PEG IgM rose 5-10-thousand-fold in all pigs immunized with Doxebo by day 10. Each of the 6 animals developed an anaphylactic shock to injection of 1/3 human dose of Comirnaty that required cardiopulmonary resuscitation in 4 cases. The reaction, starting within 1 min involved maximal pulmonary hypertension (PAP 13±2 before vs. 37±6mmHg after Comirnaty, median±MAD, P&lt;0.05) and severe systemic hypotension (SAP 63±8 before vs. 42±11mmHg after Comirnaty, median±MAD, P&lt;0.05), tachycardia, granulo- and thrombocytopenia, and skin flushing or rash. These hemodynamic changes were paralleled by C3a and TXB2 levels in blood.

Conclusion
We have established the first relevant large animal model of anaphylactic reaction induced by an mRNA vaccine. Consistent with previous studies, our current data show a causal role of anti-PEG Abs in the anaphylaxis to Comirnaty, which involves complement activation, and, hence, it represents CARPA (complement activation-related pseudo-allergy).
Inclisiran reduces expression of Toll-like receptor-4 on circulating monocytes
Authors: Bernhard J , Engelberger K , Galli L , Haider P , Ungersbaeck K , Hengstenberg C , Speidl W S , Krychtiuk K A 
Published: November 2023

Abstract

Background
Toll-like receptor 4 (TLR4) is a key regulator of both innate and adaptive immune responses and its functions affects the initiation and progression of atherosclerosis. In addition, it has recently been shown that TLR4 regulates foam cell formation by catabolism of aggregated LDL and that TLR4 is induced by proprotein convertase subtilisin/kexin type 9 (PCSK9) in vitro. The aim of this study was to examine the effects of siRNA-induced knockdown of PCSK9 by inclisiran on the expression of TLR4 in vivo.

Methods
Patients on stable lipid lowering therapy for at least 4 weeks were treated with a single dose of inclisiran (284 mg s. c.) and blood samples were taken directly before treatment and after 1 and 3 months. Monocyte expression of TLR4 was analyzed by flow-cytometry.

Results
We included 22 consecutive patients, 54.5% of patients were male and median age was 63 (IQR 60-75) years. 15 patients were treated with high dose statin and 7 patients did not tolerate statin treatment. At baseline TLR4 expression tended to be lower in statin-treated patients (MFI: 2136 IQR 1427-4011 vs. 3115 IQR 2187 vs. 5851; p=0.06). TLR4 expression after one month was comparable to baseline. However, 3 months after after inclisiran treatment, TLR4 expression on monocytes significantly decreased as compared to baseline expression (MFI baseline: 2396 IQR 1802-4408; MFI 3 months: 1575 IQR 1263-2233; p=0.006). This effect was more pronounced in statin-naïve as compared to statin-treated patients (TLR4 MFI 3 months vs baseline no statin: -58.8 IQR -41.9 to -81.2 %; statin: -27.1 IQR +2.2 to -42.1 %).

Conclusions
siRNA-induced knockdown of PCSK9 by inclisiran was associated with significant decreased expression of TLR4 on circulating monocytes. This effect was more pronounced in patients with statin intolerance.
Semaglutide effects on human epicardial and subcutaneous fat released exosomes, aortic endothelial cells and neutrophils
Authors: Sanchez-Lopez D , Lestegas-Soto A , Garcia-Vega D , Vinuela J E , Reija-Lopez L , Martinez-Cereijo J M , Fernandez-Lopez A L , Gonzalez-Juanatey J R , Eiras S 
Published: November 2023

Abstract

Background
Obesity has increased in recent years with consequences on diabetes and other comorbidities. Thus, 1 out of 3 diabetic patients have cardiovascular disease (CVD) and obese patients have a 95% risk of heart failure (HF) and, consequently, of death. The proinflammatory profile of epicardial fat (EAT) participates in this pathological mechanism. New drugs based on GLP-1 receptor agonists (GLP-1-RA) have demonstrated benefits on CVD. Semaglutide is a new GLP-1-RA that can reduce body weight and atherogenesis process.

Purpose
To study the semaglutide effects on human epicardial fat, aortic endothelial cells and neutrophils as main involved-cardiovascular cells on atherogenesis.

Materials and Methods
Epicardial (EAT) and subcutaneous fat (SAT) biopsies were collected from patients undergoing open-heart surgery. All the patients signed an informed consent. Differential fat-released exosomes, isolated by Exo-Spin Exosome Purification Kit, regarding semaglutide or/and insulin treatment were characterized by proteomics Triple TOF (n=3). Modulation of human neutrophils (dHL60 cells) phenotype, obtained after promyelocytic cell differentiation, and adhesion into human aortic endothelial cells (HAEC) by Semaglutide were analyzed by flow cytometry and functional analysis, respectively. Semaglutide effects on angiogenesis were performed on HAEC according to the manufacturer’s protocol. GLP-1 receptor expression levels were confirmed by real time-PCR.

Results
Fat biopsies, neutrophils and HAEC expressed GLP-1 receptor, measured by qPCR. We did not observe differential exosomes-cargo after semaglutide treatment on EAT. However, the released-exosomes from SAT with ANXA5, FABP4, GSN, SERPINF1 and PRDX1 were modulated by Semaglutide treatment. In addition, the CD11b molecule expression on neutrophils (p=0.0482) and their adhesion (p=0.0148) into HAEC treated with high glucose (22.2 mM) were modulated by Semaglutide treatment. Finally, GLP-1-RA improves angiogenesis, measured by loop formation after 20 hours.

Conclusion
Semaglutide modulates the released exosomes by SAT, the neutrophils adhesion into endothelial cells and enhances the angiogenesis process. These results suggest a new anti-inflammatory and anti-atherogenic role of this molecule in the human cardiovascular cell system.Figure 1Figure 2
Characterization of AT-02, a pan-amyloid-binding peptide fusion immunoglobulin with high binding potency, complement activation, and immune cell stimulation
Authors: Wall J S , Selvarajah S , Foster J S , Williams A , Richey T , Balachandran M , Jackson J , Hancock T , Stuckey A , Martin E B , Kennel S J 
Published: November 2023

Abstract

Background
Cardiac amyloidosis is an ominous manifestation of systemic amyloidosis, notably in AL and ATTR-associated types, resulting in a restrictive hypertrophic cardiomyopathy with electrical conduction and atrioventricular strain dysfunction (1). Amyloid is not naturally cleared by the immune system due to the presence of extracellular matrix components in the deposits (2). However, recruiting and stimulating the innate immune system, notably phagocytic macrophages, is a promising strategy to effect amyloid removal. To facilitate this, we have generated AT-02, a humanized IgG1 incorporating the pan-amyloid-binding peptide, p5R (3), at the light chain C-terminus. This reagent binds diverse types of amyloid via the peptide interactions and retains immunomodulatory capacity.

Purpose
The goal of these studies was to characterize the bioactivity of AT-02 that underlies its potential therapeutic activity.

Methods
AT-02 was produced from a CHO pool using a perfusion cell culture production process, purified by Protein A, anion exchange, and cation exchange chromatographies, and characterized by mass spectrometry and SDS-capillary electrophoresis. AT-02 was biotinylated and used for immunohistochemical detection of both AL and ATTR amyloidosis in formalin fixed tissues (Figure 1A). Binding to heparin (as a surrogate for amyloid-associated hypersulfated heparan sulfate proteoglycans) and synthetic AL fibrils was studied using both ELISA and surface plasmon resonance assays (Figure 1B). Activation of complement following binding to amyloid substrates was determined using a C5b9 liberation assay in the presence of human plasma. The effect of AT-02 on amyloid fibril growth was determined using a fluorescence fibril elongation assay. In vivo clearance of amyloid and cell activation was demonstrated using a murine model of AL amyloidoma.

Results
AT-02 bound human cardiac amyloid in tissue sections with high specificity and intensity. The potency for heparin and synthetic amyloid fibrils, as well as human cardiac amyloid extracts, was sub-nanomolar in the ELISA assay with kinetics consistent with a high affinity antibody. In the presence of plasma, complement activation of amyloid-bound AT-02 was significantly higher as compared to controls, and the reagent inhibited synthetic amyloid fibril growth by ∼40%. Treatment of human amyloid with AT-02 expedited phagocytosis by murine macrophages in vivo and enhanced the intensity of immune response to the implanted amyloid.

Conclusion
Clearance of cardiac amyloid is a significant clinical unmet need for patients with systemic amyloidosis. The humanized IgG1-peptide fusion, AT-02, potently binds cardiac amyloid and may serve as an opsonin and induce macrophage-mediated phagocytosis and clearance of cardiac amyloid deposits.
CAR-T cells inhibit vascular smooth muscle phenotypic switching and reduce intimal hyperplasia in vein grafts
Authors: Xiao B , Jiang Y , Guo Y 
Published: November 2023

Abstract

Background
Intimal hyperplasia (IH) is a prevalent cause of vein graft stenosis after coronary artery bypass grafting (CABG) in the middle and long term. Vascular smooth muscle cell (VSMC) phenotypic switching from a contractile phenotype to a synthetic phenotype is a hallmark of this process, along with the degradation of extracellular matrix (ECM) and smooth muscle migration. Chimeric antigen receptor (CAR)-T cell therapy is a targeted cellular immunotherapy that uses genetically engineered T cells to specifically eliminate the antigen-bearing cells. There is a potential to apply CAR-T therapy to cardiovascular diseases.

Purpose
In this study, we aim to design a novel CAR-T cell therapy to eliminate synthetic vascular smooth muscle cells in intimal hyperplasia, inhibit phenotypic switching, in order to reduce vein graft stenosis.

Methods and results
Using bulk RNA-seq dataset from the GEO database, we have identified human organic cation transporter (OCT1) as a key marker for synthetic vascular smooth muscle cell in vein grafts (Fig.1A, B). We then engineered switchable fourth-generation CAR-T cells incorporating CD28 and CD3 ζ costimulatory domains (Fig.1C, E). In addition, these CAR-T cells secrete tissue inhibitor of metalloproteinase-3 (TIMP3), a protein that had been shown to inhibit matrix metalloproteinases (MMPs) activity and the ERK1/2 pathway (Fig. 1F). Targeting FITC instead of OCT1 itself allows a controllable and switchable working process for the CAR-T cells, as they only start to identify synthetic VSMCs in combination with a FITC-conjugated OCT1 antibody (Fig.1D). Immunostaining of PDGF-induced synthetic VSMCs co-cultured with CAR-T cells and FITC-conjugated OCT1 antibody revealed increased expression of contractile markers, and reduced expression of synthetic markers (Fig1.G), along with decreased mitochondrial membrane potentials, which could be reverted after pre-treatment with a pan caspase inhibitor (Fig.1H),. In vivo studies using mice vein graft model showed reduced intimal thickness, increased intimal/medial ratio in cross-sections, in addition to increased peak velocity and inner wall diameter on ultrasound after CAR-T cell therapy (Fig.2).

Conclusion
Switchable CAR-T cell therapy targeting synthetic vascular smooth muscle cells might be a promising approach to alleviate intimal hyperplasia in vein graft stenosis.
Impact of sodium-glucose cotransporter-2 (SLGT2) inhibitors treatment initiation on the reduction of atrial and ventricular arrhythmias in patients with implanted cardiac devices
Authors: Minguito Carazo C , Sanchez Munoz E , Martinez Sande J L , Rodriguez Manero M , Fidalgo Andres M L , Garcia Seara J , Fernandez Lopez X A , Gonzalez Rebollo J M , Gonzalez Ferrero T , Rodriguez Santamarta M , Gonzalez Melchor L , Martinon Martinez J , Garcia Rodeja F , Fernandez Vazquez F , Gonzalez Juanatey J R 
Published: November 2023

Abstract

Introduction
Sodium-glucose cotransporter-2 (SLGT2) inhibitors have demonstrated benefits on survival and hospital admissions in patients with heart failure and reduced ejection fraction. However, their impact on the burden of ventricular (VA) and atrial arrhythmias (AA) is unknown.

Methods
Retrospective multicentric study of heart failure patients with implantable cardiac defibrillator device (ICD) from 2015 to 2020 with or without cardiac resynchronization therapy (CRT) receiving SGLT2 inhibitors. Device-registered arrhythmic events were analyzed and compared during one year following SLGT2 inhibitor initiation and during one year before initiation. Relevant VA were defined as the occurrence of any sustained VT (SVT) (&gt;30 seconds), ventricular fibrillation (VF), or appropriate therapy (antytachycardia pacing or shock). All VA included all the relevant VA and the occurrence of non-sustained ventricular tachycardia (NSVT). AA included atrial fibrillation (AF) burden, and episodes of more than 24 hour of AF.

Results
147 patients (67 ± 10 years, 82.3% males, LVEF 30.3 ± 11%, CRT 39.3%), were included in the study. 77.6% had complete remote tele-monitoring during the entire follow up and there was no difference in the antiarrhythmic therapy between the two periods. Following SGLT2 inhibitor initiation, there was a significant reduction in the percentage of patients with relevant VA (19.7% pre vs 10.9% post; p=0.019) and all VA (47.6% pre vs 30.6% post; p&lt;0.001) (Figure 1 and 2). There was a reduction in the prevalence of NSVT (40.1% pre vs. 27.2% post; p&lt;0.001), of SVT (17.7% vs 8.8%; p&lt;0.001), of VF (4.1% vs 2%; p=NS) and appropriate therapies (14.3% vs 9.5%; p=NS). Among patients with any VA, there was a significant reduction in the incidence of NSVT (p&lt;0.001), but not in the incidence of SVT or appropriate therapies (p=NS). There was no impact on atrial fibrillation burden or in the number of AF episodes lasting more than 24 hours (p=NS).

Conclusions
In our study, initiation of a SGLT2 inhibitor in patients with heart failure, was associated with a reduction in the prevalence of relevant VA and any VA, with a significant reduction in the number of NSVT per patient. There was no impact on AF burden or the number of AF episodes lasting more than 24 hours. Prospective studies should be performed to sustain these conclusions.Relevant ventricular arrhythmiasAny ventricular arrhythmia