bluuebunny/crossref_metadata_2025_split huggingface dataset:
Abstract 522: TGF-beta Neutralization Augments Development of Angiotensin II-induced Aneurysms in Both Ascending and Abdominal Aortic Regions
Authors: Chen Xiaofeng , Rateri Debra L , Howatt Deborah A , Balakrishnan Anju , Moorleghen Jessica J , Cassis Lisa A , Daugherty Alan 
Published: July 2021

Introduction and Objectives

Angiotensin II (AngII) infusion induces ascending and abdominal aortic aneurysms (AAs) in mice. In a mouse model of Marfan Syndrome expressing Fbn1
C1039G/+
, ascending AAs were reduced by administration of a transforming growth factor-beta (TGF-beta) neutralizing antibody. In contrast, administration of TGF-beta neutralizing antibodies to AngII-infused mice increased aortic rupture. The purpose of this study was to compare the effects of TGF-beta neutralization on formation and progression of AngII-induced ascending and abdominal AAs.

Methods and Results
Male C57BL/6 mice were fed a normal diet and infused subcutaneously with AngII (1,000 ng/kg/min). Five days prior to initiating infusion, mice were injected i.p. with either a mouse monoclonal TGF-beta antibody (1D11) or an isotype matched IgG at a dose of either 0.3 or 5 mg/kg x 3/per week. 1D11 administration significantly decreased serum TGF-beta concentrations. TGF-beta neutralization at 5 mg/kg greatly increased the incidence of aortic rupture, which was attributed to rupture in both the ascending and abdominal regions. For mice that remained viable after 28 days of infusion, there were equivalent increases in aortic dilation in both the ascending and abdominal regions. Prior to rupture, aortic diameters determined by ultrasound demonstrated no significant effect on AngII-induced dilation of the ascending or abdominal aorta. We also studied the effects of TGF-beta neutralization in mice with established AngII-induced AAs following AngII-infusion for 28 days. C57BL/6 mice were injected with the mouse TGF-beta neutralizing antibody or IgG control (5 mg/kg x 3/per week, n=10 per group), while AngII infusion was continued for a further 28 days. Although TGF-beta antibody administration significantly decreased serum TGF-beta concentrations in mice with established AAs, there was no effect on aortic rupture or dilation of either the ascending or abdominal aortic region.

Conclusion
TGF-beta inhibition augmented AngII-induced aortic rupture in both the ascending and abdominal regions but had no effect on dilation. Furthermore, TGF-beta neutralization had no effect on either aortic rupture or expansion in established AAs.
Abstract 527: A Novel Stem Cell Delivery Model for Evaluation of Delivery Using Targeted Bifunctional Liposomes with Ultrasound Enhancement
Authors: Naji Ali K , Wassler Michael , Britton George , Shelat Harnath S , Geng Yong-Jian , McPherson David D , Klegerman Melvin E 
Published: July 2021

Introduction.
We have developed bifunctional echogenic liposomes (BF-ELIP) targeted to both CD34 and ICAM-1 to facilitate delivery of CD34+ stem cells to inflammatory endothelium. We previously confirmed that BF-ELIP enhanced CD34+ stem cell adherence to ICAM-1-expressing endothelium in vitro and in vivo and showed that ultrasound (US) facilitated penetration of stem cells into the endothelial cell layer.

Hypothesis.
The transwell culture system can serve as an in vitro model for study of US-enhanced targeted delivery of stem cells to atheroma.

Methods.
BF-ELIP were prepared by evaporation-rehydration-sonication-lyophilization, followed by conjugating antibodies specific for CD34 and ICAM-1 through a thioether linkage. TNFα-pretreated HUVEC monolayers on transwell (6 wells/plate) insert membranes were incubated with nonspecific IgG-ELIP or BF-ELIP (1mg/well) for 15 minutes, followed by human monocytes labeled with Oregon Green. Half the inserts were subjected to 6 MHz color Doppler ultrasound (MI = 0.4) for 5 minutes. Fluorescence of resuspended cells was measured after treatment of both inserts and wells with 0.25% trypsin/0.1% EDTA 24 hours later.

Results.
BF-ELIP enhanced adherence of monocytes to the ICAM-1-expressing HUVEC monolayer relative to untreated controls and IgG-ELIP, but did not increase the number (Fig. 1) or proportion (Fig. 2) of monocytes traversing the monolayer. US greatly increased the number of monocytes both adhering to and passing through the monolayer in all groups.

Conclusions.
We have succeeded in developing a transwell cultured HUVEC system as a model for US-enhanced, BF-ELIP-mediated stem cell delivery to inflammatory endothelium.
Abstract 566: The Transcriptome of the Human Carotid Atherosclerotic Plaque
Authors: Poroyko Valeriy , McCormick Susan , He Qi , Skelly Christopher L 
Published: July 2021

Ischemic stroke is a complex disease process with carotid atherosclerosis as the major etiologic source. To better understand the biology of a vulnerable plaque we carried out whole transcriptome shotgun sequencing of human carotid plaques.

Methods
5 carotid atherosclerotic plaques were obtained at the time of operation (2 symptomatic, 2 asymptomatic, and 1 control). RNA was isolated and 5 cDNA libraries were constructed and sequenced with single-reads100nt in length using one line of flow cell of HySeq 2000 (Illumina Inc). Standard bioinformatic techniques were used to ensure quality screening of raw reads. Ingenuity Systems IPA software was used to determine canonical biological pathways overrepresented in plaques. We compared our data to the data of Illumina Human Body Map processed by a similar analytical pipeline. Linkage analysis was performed.

Results
Among detected pathways were “Atherosclerosis Signaling” and pathways involved in inflammation and cell differentiation. We analyzed gene expression data by creating a distance matrix based on Jaccard similarity measure (Fig 1). When gene expression values were used to create a matrix of Euclidian distances between known transcriptomes, four specimens including the control clustered with heart and muscle. The transcriptome from the stroke patient was more similar to lung, liver and heart.

Conclusion
Whole transcriptome analysis of carotid plaques is feasible and ontologically accurate. We demonstrate linkage differences between symptomatic and asymptomatic plaques and biological pathways involved in vascular lesion formation. Further analysis may provide insight into the vulnerable plaque pathobiology.
Abstract 57: Natural Killer Cells Drive Angiotensin II-Induced Vascular Dysfunction Dependent on the T-bet/IFN-gamma Axis
Authors: Kossmann Sabine , Schwenk Melanie , Hausding Michael , Hu Hanhan , Schmidgen Maria I , Schönfelder Tanja , Oelze Matthias , Daiber Andreas , Muenzel Thomas , Becker Christian , Wenzel Philip 
Published: July 2021

Rationale
Immune cells contribute to angiotensin II (ATII) induced vascular dysfunction and inflammation. However, the mechanisms of recruitment and immune effector pathways remain incompletely understood.

Objective
We tested the hypothesis that interferon-gamma (IFN-γ) and natural killer (NK) cells play a pivotal role in ATII-driven vascular inflammation.

Methods and results

IFN-γ
-/-
and Tbx21
-/-
mice were partially protected from ATII-induced vascular endothelial and smooth muscle dysfunction, whereas mice overexpressing IFN-γ showed constitutive vascular dysfunction. Absence of T-bet, the IFN-γ transcription factor encoded by Tbx21, reduced vascular superoxide and peroxynitrite formation and attenuated expression of NADPH oxidase subunits as well as inducible NO synthase, monocyte chemoattractant protein 1 and interleukin 12 in aortas of ATII-infused mice. Compared to controls, IFN-γ
-/-
and Tbx21
-/-
mice were characterized by reduced ATII-mediated vascular recruitment of both NK1.1
+
NK-cells as the major producers of IFN-γ and CD11b
+
Gr-1
low
IL-12 secreting monocytes. Selective depletion and adoptive transfer experiments identified NK-cells as essential contributors to vascular dysfunction and showed that T-bet
+
LysM
+
myelomonocytic cells were required for NK-cell recruitment into vascular tissue and local IFN-γ production.

Conclusion
We provide first evidence that NK-cells play an essential role in ATII-induced vascular dysfunction. In addition, we disclose the T-bet-IFN-γ pathway and mutual monocyte-NK-cell activation as potential therapeutic targets in cardiovascular disease.
Abstract 297: Segmental Aortic Stiffening is a Mechanism Driving Early Abdominal Aortic Aneurysm Pathogenesis
Authors: Raaz Uwe , Zöllner Alexander M , Toh Ryuji , Nakagami Futoshi , Schellinger Isabel N , Emrich Fabian C , Adam Matti , Maegdefessel Lars , Hertel Thomas K , Deng Alicia , Jagger Ann , Buerke Michael , Dalman Ronald L , Spin Joshua M , Kuhl Ellen , Tsao Philip S 
Published: July 2021

Stiffening of the aortic wall is a phenomenon consistently observed in abdominal aortic aneurysm (AAA). However, its role in AAA pathophysiology is largely undefined.
Using an established murine elastase-induced AAA model, we demonstrate that segmental aortic stiffening (SAS) precedes aneurysm growth. Finite elements analysis (FEA)-based wall stress calculations reveal that early stiffening of the aneurysm-prone aortic segment leads to axial (longitudinal) stress generated by cyclic (systolic) tethering of adjacent, more compliant wall segments. Interventional stiffening of AAA-adjacent segments (via external application of surgical adhesive) significantly reduces aneurysm growth. These changes correlate with reduced segmental stiffness of the AAA-prone aorta (due to equalized stiffness in adjacent aortic segments), reduced axial wall stress, decreased production of reactive oxygen species (ROS), attenuated elastin breakdown, and decreased expression of inflammatory cytokines and macrophage infiltration, as well as attenuated apoptosis within the aortic wall. Cyclic pressurization of stiffened aortic segments ex vivo increases the expression of genes related to inflammation and extracellular matrix (ECM) remodeling. Finally, human ultrasound studies reveal that aging, a significant AAA risk factor, is accompanied by segmental infrarenal aortic stiffening.
The present study introduces the novel concept of segmental aortic stiffening (SAS) as an early pathomechanism generating aortic wall stress and thereby triggering AAA growth. Therefore monitoring SAS by ultrasound might help to better identify patients at risk for AAA disease and better predict the susceptibility of small AAA to further growth. Moreover our results suggest that interventional mechanical stiffening of the AAA-adjacent aorta may be further tested as a novel treatment option to limit early AAA growth.
Abstract 644: CD36 is Essential to Increased Atherosclerosis Mediated by Oral Infection With Porphromonas gingivalis
Authors: Febbraio Maria , Brown Paul M 
Published: July 2021

Epidemiological evidence strongly support a link between periodontal disease & cardiovascular disease, but the mechanism(s) remains poorly understood. Using the human periodontal disease associated bacteria, Porphyromonas gingivalis (Pg) as a model, we carried out studies in macrophages & low density lipoprotein receptor (LDLR) KO mice. Pg associated similarly with macrophages from wild type & CD36 KO mice, but there were differences in responses dependent on Toll-like receptor (TLR) 2. We observed decreased NFkB activation & IL1beta generation following Pg treatment in CD36 KO macrophages, despite similar levels of TLR2 expression. OxLDL strongly inhibited Pg mediated IL-1beta generation in a CD36 dependent manner. Macrophage foam cell formation as a result of incubation with oxLDL & PgLPS was increased in a CD36 dependent manner.
LDLR KO & CD36/LDLR double KO mice were orally infected with Pg & fed a Western diet (12 weeks). There was a significant increase in the cemento-enamel junction of molars of infected compared with uninfected mice, demonstrating the validity of the model. Histological analysis showed inflammatory cell infiltrates in gums of infected mice after 12 weeks, supporting a chronic inflammatory process. Differences in plasma parameters & weight gain did not necessarily track with atherosclerosis burden, however blood neutrophils & cytokines were increased in infected LDLR KO mice compared with all other groups. Infected LDLR KO mice had significantly increased atherosclerotic lesion burden compared with uninfected LDLR KO mice, and all of the increased lesion was CD36-dependent.
Our data suggest that atherosclerosis associated with periodontal disease is mediated by cellular inflammatory responses involving both CD36 & TLR2. Pg enhances oxLDL mediated foam cell formation in a CD36 dependent manner, and this may explain increased lesion burden. Generation of IL1beta, a key pro-atherogenic cytokine, is altered as a result of CD36 expression. Periodontal disease affects more than 20% of the population of the US/Canada, & is associated with increasing age, which is also a risk factor for atherosclerosis. Targeting CD36 may provide important supplemental therapy to current lipid lowering strategies to reduce atherosclerosis.
Abstract 172: Product of Lipid Oxidation During Inflammation is a New Potent Ligand for β2 Integrin-mediated Macrophage Migration
Authors: Yakubenko Valentin , Brown Kathleen , Tischenko Miraslava , West Xiaoxia , Podrez Evgeny , Byzova Tatiana 
Published: July 2021

2-(ω-carboxyethyl)pyrrole (CEP) and ethylpyrrole (EP) derivatives of proteins are formed through adduction of end products of docosahexaenoate (DHA) oxidation[[Unable to Display Character: &#61472;]] with the ε-amino groups of protein lysyl residues, followed by cyclization and dehydration. Our previous studies demonstrated that EP-modified proteins are present in healthy tissue, while CEP-modified proteins are detected in the tissue during inflammation including inflamed peritoneal wall and atherosclerotic lesions.
Potential role of CEP in inflammatory response was evaluated by testing leukocyte recruitment during thioglycollate-induced peritoneal inflammation. We found that injection of anti-CEP antibody dramatically reduced the accumulation of macrophages in the peritoneal cavity at 72 hours, while the earlier accumulation of neutrophils was not affected. Migration of neutrophils was a prerequisite for CEP production in the peritoneum. In vitro adhesion assay revealed strong binding of peritoneal macrophages to CEP, while adhesion to EP was not detected. Moreover, CEP presence in 3D fibrin gel significantly increased migration of macrophages toward MCP-1 gradient. We have previously demonstrated that CEP is ligand for CD36 and TLR2. However, the adhesion of TLR2/CD36-double deficient macrophages to CEP was similar to wild type cells; suggesting that other receptors are involved in CEP-mediated macrophage adhesion/migration. Since αDβ2 and αMβ2 integrins are major macrophage adhesion receptors we tested their ability to bind CEP. Isolated αM and αD I-domains, which are critical for integrin ligand recognition, showed a concentration-dependent binding to immobilized CEP, but not to EP in the biacore assay. Moreover, αDβ2 and αMβ2-transfected but not control HEK293 cells strongly adhered to CEP. This adhesion was significantly inhibited by anti-CEP, anti-β2 and anti-αM (or αD) antibodies, but not by anti-β1 antibody.
In summary, we found that CEP-modified proteins drive the migration of macrophages during inflammation by engaging macrophage integrins αMβ2 and αDβ2. Hence, regulation of CEP-mediated adhesion/migration can control the accumulation of macrophages within the site of inflammation and may regulate the development of chronic inflammatory deseases.
Abstract 386: Thrombin Inhibition by Dabigatran Reduces the Amount of Pro-Inflammatory CD11c-Positive Macrophages in Visceral Adipose Tissue of Low-Density Lipoprotein Receptor-Deficient Mice
Authors: Feldmann Kathrin , Grandoch Maria , Lehr Stefan , Fischer Jens W 
Published: July 2021

Thrombin is involved in inflammatory processes. In obesity, chronic low-grade inflammation promotes the development of insulin resistance and diabetes mellitus type 2 (T2DM) which strongly drive atherosclerosis. Patients with T2DM exhibit increased circulating levels of tissue factor as well as thrombin-antithrombin complexes.
Aim of the present study was to analyse potential effects of thrombin inhibition by dabigatran on adipose tissue inflammation in a mouse-model of diet-induced obesity and atherosclerosis.
Female, 10 weeks old low-density lipoprotein receptor-deficient (LDLR-/-) mice received a Western-type diet containing 5 mg/g dabigatran or matching placebo for 20 weeks.
Flow cytometric analysis of the stromal vascular fraction revealed a reduction in pro-inflammatory macrophages (CD11b+F4/80+CD11c+) in visceral adipose tissue of dabigatran-treated animals (control 8.490 ± 1.996, dabigatran 3.606 ± 0.8884 CD11c+ macrophages/mg adipose tissue, n = 21), while the number of anti-inflammatory CD11c- macrophages was not changed. mRNA expression in the visceral adipose tissue as well as plasma concentrations of the pro-inflammatory cytokines interleukin (IL) 1 beta and IL 6 (quantitative real-time PCR (qPCR), multiplex immunoassay) were reduced in dabigatran-treated animals. No differences in body-weight gain and the amount of body fat (NMR measurement) were observed. Glucose tolerance and insulin resistance did not differ between groups, either. In vitro no direct influence of thrombin on macrophage polarization was detectable in bone marrow-derived macrophages (flow cytometry, qPCR). However, first results indicate that stimulation of the cells with conditioned medium from thrombin-treated, differentiated preadipocytes (3T3L1 cells) enhances M1 polarization of macrophages (flow cytometry).
In conclusion, thrombin inhibition by dabigatran promotes an anti-inflammatory phenotype in adipose tissue macrophages in a mouse model of diet-induced obesity and atherosclerosis, probably resulting from a changed expression profile of adipocyte cytokines. These effects might contribute to the known anti-atherosclerotic effects of thrombin inhibition in this model.
Abstract 395: Elevated Von Willebran Factor in an in vitro Model of Fabry Disease
Authors: Kang Justin J , Kaissarian Nayiri M , Shu Liming , Desch Karl C , Bodary Peter F , Shayman James A 
Published: July 2021

Fabry disease is an X-linked disorder caused by a defect in the gene encoding the lysosomal hydrolase, alpha-galactosidase A (GLA). The loss in GLA function causes an age-dependent accumulation of globotriaosylceramide in the endothelial cells, which results in decreased nitric oxide bioavailability and premature mortality in the affected patients. Based on this, we hypothesized that GLA deficiency promotes endothelial inflammation. We observed a significant age-dependent increase in the plasma von Willebrand Factor (vWF) in Fabry mice compared to age-matched wild type mice (p=0.046 at 2 months and p=0.0004 at 17 months). To evaluate the molecular mechanisms of increased vWF in Fabry disease, small interfering RNA (siRNA) technique was used to knock-down GLA in EA.hy926 cells. Our initial studies with this cell line demonstrated that thrombin (2U/mL) induced a time-dependent vWF secretion into the cell culture medium with an 80-fold increase in the level of vWF after 90 min stimulation (p=0.001 compared to basal level; n=3 replicates per group), measured by a human antibody against vWF and AlphaLISA. Next, the siRNA studies revealed a greater than 95% knockdown of GLA protein (via western blotting) in the GLA-siRNA group compared to the control group with no effect in the scrambled (SCR) sequence-siRNA group (CON vs. GLA vs. SCR arbitrary units: 1.0±0.03 vs. 0.01±0.01 vs. 1.1±0.08, p<0.001; GLA vs. SCR). Importantly, no difference in GAPDH protein was observed between groups. To determine the direct effect of GLA deficiency on vWF, the confluent cells in each group were incubated in serum-free medium overnight. Consistent with the hypothesis, we observed approximately 60% increase in vWF in the cell culture medium in the GLA siRNA group compared with the untreated and scrambled-sequence control groups. (CON vs. GLA vs. SCR arbitrary units: 38.5±4.7 vs. 61.3±1.1 vs. 31.4±3.8, p=0.02; GLA vs. SCR; n=3 replicates per group).
In conclusion, the data provide evidence of endothelial activation in the setting of GLA deficiency, demonstrated by the robust elevation of vWF mediated by GLA gene disruption. Future study is required to further understand the mechanism of the elevated vWF and decreased nitric oxide bioavailability in Fabry disease.
Abstract 650: Characterization of Microparticles Formed by ABCA1
Authors: Hafiane Anouar , Bielicki John K , Johansson Jan O , Genest Jacques 
Published: July 2021

Microparticles (MPS) are lipoprotein-sized structures created by the ABCA1 transporter. Their biological roles in health and in disease remain unknown. Here, we study MPS released from baby hamster kidney (BHK) cells stably expressing ABCA1 and human THP-1 cells also expressing ABCA1.
Media cell culture was first collected from BHK-ABCA1 expressing cells after (45min, 2, 4, 8, and 24h) incubation. After centrifugation (4000xg for 15min and 10000xg for 30min) to remove cell debris, the supernatant was passed through a 10kDa cutoff filter and subsequently subjected to analytical FPLC. FPLC analysis shows creation of a single peak in the presence of ABCA1 but not in mock-transfected BHK cells. In a time-course study, the estimated hydrodynamic diameter remained stable (≥20nm).
After 8h incubation of BHK cell with apoA-I or an apo-E mimetic peptide, CS-6253 (1μM), ABCA1 mediated formation of MPS of similar size with a significant increase in 3[H]-FC content than those generated by ABCA1 alone, (373±23 % cpm, P<0.05) and (277±60 % cpm, P<0.05) respectively. This was associated with highly lipidated nHDL-CS-6253 compararely to nHDL-apoA-I (4535±72 % cpm, P<0.001 versus 1059±14 % cpm) respectively. This data suggests that MPS formations are an integral component of cellular cholesterol efflux. Also, MPS do not contain CS-6253 when ABCA1 cells were incubated with the peptide as confirmed by western blotting similar to MPS generated from ABCA1 cells generated by apoA-I incubation. Cholesterol is effluxed more to nHDL-CS-6253 (14±7.68, % cpm, P<0.01 than to MPS 1±0.10, % cpm, 24h; P<0.01) similar to apoA-I. Depletion of membrane cholesterol by methyl-β-cyclodextrin treatment impaired HDL genesis and decrease MPS release. Detection of flotellin-2 protein enriched in MPS in total cell lysate indicated that these MPS may be related to exosomes. MPS generation was also characterized in THP-1 cells for further molecular characterization. We conclude that MPS are formed by ABCA1 in diverse cell types and the cholesterol content is dependent on activation by apo A-I or mimetic peptides. The physiological role of MPS remains to be understood. These particles may be transporters of lipids and nucleic acids.