bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 441: Macrophage CD40 in Atherosclerosis, Obesity and Multiple Sclerosis
Authors: Shami Annelie , Aarts Suzanne , Bürger Christina , Kusters Pascal , van den Berg Susan , Seijkens Tom , Kooij Gijs , Weber Christian , Gerdes Norbert , de Winther Menno , Lutgens Esther
Published: July 2021

Introduction:
The co-stimulatory dyad CD40-CD40L plays a central role in fine-tuning immune reactions in atherosclerosis, obesity-induced inflammation and multiple sclerosis (MS). Inhibition of CD40 in atherosclerosis and experimental autoimmune encephalomyelitis (EAE) ameliorates disease outcome, whereas CD40-deficiency in a diet induced obesity (DIO) model worsens insulin resistance and induces excessive adipose tissue inflammation. Although inhibition of CD40 has powerful effects, we do not know which CD40 expressing cell-type is responsible for the amelioration/aggravation of disease. As myeloid CD40 is known to play a role in leukocyte trafficking, which is important in atherosclerosis, obesity and neuro-inflammation, we hypothesize that myeloid CD40 is important in these disease modalities.

Methods:
To investigate the role of myeloid CD40 in atherosclerosis, obesity and EAE we have generated macrophage specific LysM-CD40flfl
-
(ApoE-/-), and dendritic cell/adipose tissue macrophage specific CD11C-CD40flfl). Atherosclerosis was induced by aging the LysM-CD40flfl
-
ApoE-/- mice until 30 weeks. EAE was induced in LysM-CD40flfl mice by subjecting them to myelin oligodendrocyte glycoprotein peptide (MOG35-55), and LysM-CD40flfl and CD11c-CD40flfl mice were subjected to a 60% high fat diet for 18 wks.

Results:
LysM-CD40flfl
-
ApoE-/- mice showed a significant reduction in atherosclerotic plaque area, with a reduced macrophage accumulation. Loss of macrophage CD40 in EAE results in a significant decrease in the neurological symptoms of EAE, and a majority of the LysM-CD40flfl mice were fully protected against EAE. LysM-CD40flfl mice subjected to DIO showed an increase in macrophage accumulation in the visceral adipose tissue, but did not affect adipose tissue mass, insulin tolerance, or plasma triglyceride concentrations. The CD11CcreCD40flfl mice exhibited a decrease in visceral adipose tissue weight, an increased lipid content in the liver and slightly decreases leukocyte numbers and pro-inflammatory gene expression in the adipose tissue.

Conclusions:
Macrophage CD40 is an important driver of atherosclerosis and EAE. Macrophage CD40 is protective in obesity-induced inflammation, but is probably not the key player.
Abstract 448:
Etv2-Mir130a-Jarid2
Cascade Regulates Vascular Patterning During Embryogenesis
Authors: Singh Bhairab N , Tahara Naoyuki , Kawakami Yasuhiko , Koyano-Nakagawa Naoko , Gong Wuming , Garry Mary , Garry Daniel
Published: July 2021

Remodeling of the pre-existing primitive vasculature is necessary for the formation of a complex branched vascular architecture. However, the factors that modulate these processes are incompletely defined. Previously, we defined the role of microRNAs (miRNAs) in endothelial specification. In the present study, we further examined the
Etv2-Cre
mediated ablation of

Dicer
L/L

and characterized the perturbed vascular patterning in the embryo proper and yolk-sac. We mechanistically defined an important role for
miR-130a
, an Etv2 downstream target, in the mediation of vascular patterning and angiogenesis
in vitro
and
in vivo
. Inducible overexpression of
miR-130a
resulted in robust induction of vascular sprouts and angiogenesis with increased uptake of acetylated-LDL. Mechanistically,
miR-130a
directly regulates
Jarid2
expression by binding to its
3’-UTR
region. CRISPR/Cas9 mediated knockout of
miR-130a
showed increased levels of
Jarid2
in the ES/EB system. Further, the levels of
Jarid2
transcripts were increased in the
Etv2-null
embryos at E8.5. In the
in vivo
settings, injection of
miR-130a
specific morpholinos in zebrafish embryos resulted in perturbed vascular patterning with reduced levels of endothelial transcripts in the
miR-130a
morphants. qPCR and
in situ hybridization
techniques demonstrated increased expression of
jarid2a
in the
miR-130a
morphants
in vivo
. These findings demonstrate a critical role for
Etv2-miR-130a-Jarid2
in vascular patterning both
in vitro
and
in vivo
.
Abstract 455: HDL-Small RNA Gene Regulatory Networks Alter T Cell Signalling in Systemic Lupus Erythematosus
Authors: Michell Danielle L , Moore Jared L , Ormseth Michelle J , Zhao Shilin , Sheng Quanhu , Stein C. Michael , Major Amy S , Vickers Kasey C
Published: July 2021

Systemic Lupus Erythematosus (SLE) is a debilitating disease primarily in women involving complex T and B cell dysregulation. SLE presents with dysfunctional HDL and we have previously found that HDL-microRNAs (miRNA) are significantly altered in SLE; however, miRNAs are just one of many types of small non-coding RNAs (sRNA). As such, we hypothesized that HDL-sRNA cargo and cell-to-cell communication in SLE extend beyond miRNAs. Using high-throughput sRNA sequencing (sRNA-seq), we found that tRNA-derived sRNAs (tDRs) were highly abundant on HDL and were significantly altered in SLE subjects (n=9) compared to controls (n=8,
P
<0.05). In addition, circulating levels of angiogenin, an RNaseIII enzyme responsible for tDR cleavage from parent tRNAs, was also found to be significantly increased in plasma (
P
<0.05) from SLE subjects compared to controls. To determine if tDRs are altered in CD4+ T cells in SLE subjects, real-time PCR was used to quantify candidate tDRs, and we found that tDR-GlyGCC levels were significantly increased 4.2-fold in SLE (
P
<0.01) and readily exported to HDL. Strikingly, total RNAseq,
in silico
analysis, and mRNA sequencing suggested that ROCK2, a critical regulator of CD4+ T cell differentiation, is a direct tDR-GlyGCC target gene which was confirmed with gene reported (luciferase) assays. Moreover, activated human CD4+ T cells transfected with tDR-GlyGCC mimetics, demonstrated reduced ROCK2 protein levels and STAT3 phosphorylation, and consequently reduced inflammatory cytokine secretion (IL-17 and IL-21;
P
<0.05). To determine if T cell exported tDR-GlyGCC is transferred between cells by HDL,
ex vivo
studies were completed using Trans-PhotoActivatable-Ribonucleoside-CrossLinking-ImmunoPrecipitation high-throughput Sequencing (Trans-PAR-CLIPseq). Using this approach, we found a cassette of CD4+ T cell-originating sRNAs, including tDR-GlyGCC, that were transferred by HDL to recipient immune cells. Here, we demonstrate that HDL facilitates intercellular transfer of tDRs between immune cells and a critical role for tDR-GlyGCC in regulating T cell signalling.
Abstract 46: The Effects of Membrane Cholesterol on Vascular Smooth Muscle Cell Stiffness and Adhesion to Fibronectin
Authors: Childs Josh , Hong Zhongkui Hong
Published: July 2021

Atherosclerosis remains a major cause of cardiovascular disease (CVD). Cholesterol has been identified as a major contributor to the cause of atherosclerosis. It is well known that the cholesterol accumulation in macrophage-derived foam cells is the major component of atherosclerotic plaque. However, growing evidences suggests that cholesterol loading into vascular smooth muscle cells (VSMC) in atherosclerosis is much larger than previously known, and about 40% of total foam cells in the atherosclerotic plaque are VSMC-derived. Cholesterol may not only contribute as the fatty deposition in the atherosclerotic lesion, but also play a critical role in the VSMC migration toward the intima of the blood vessel wall. In addition, the arterial wall becomes stiffer during atherosclerosis altering the micromechanical environment experienced by the VSMCs leading to changes in VSMC stiffens, adhesion, and phenotype. Migration of VSMCs is a complex process including proliferation and phenotypic switching of VSMCs, thus contributing too many changes in cell membrane adhesion molecules. We tested the hypothesis that membrane cholesterol in VSMCs may play an important role in α
5
β
1
-integrin mediated adhesion, and alter the sensory function of VSMCs to ECM mechanical properties. In this study cholesterol manipulation was achieved using methyl-β-cyclodextrin, and gel substrates with varying stiffness were used to mimic the changing environment in atherosclerosis. Atomic force microscopy (AFM) was used to determine integrin-fibronectin adhesion force and cell stiffness. A custom-written MATLAB program was used to interpret the elasticity of the VSMC cytoskeleton and adhesion force. Cellular adhesion was measured for 50%-70% confluent cells with a sample size of 50 cells on a fibronectin coated AFM stylus probe. Our results show that there is a significant decrease in α5β1-integrin adhesion of VSMCs on substrates above 9 kPa upon membrane cholesterol depletion. Additionally, mechanotransduction of VSMCs upon cholesterol depletion is less efficient. In conclusion, cell membrane cholesterol and extracellular mechanical signals may synergistically regulate cellular mechanical functions of VSMCs and their migration in the progression of atherosclerosis.
Abstract 465: Nicotine Differentially Influences Segmental Aortic Stiffening
Authors: Wagenhaeuser Markus U , Schellinger Isabel N , Guenther Sabina P , Yoshino Takuya , Toyama Kensuke , Kayama Yosuke , Deng Alicia , Zoellner Alexander M , Raaz Uwe , Spin Joshua M , Tsao Philip S
Published: July 2021

Background:
Arterial stiffness is a major risk factor for various cardiovascular diseases and contributes to the development of abdominal aortic aneurysms (AAA). In this context, differential aortic stiffening of adjacent aortic segments increases aortic wall stress and accelerates the disease. Smoking is a major risk factor for AAA, in part due to nicotine. In this study, we investigated aortic stiffening of the thoracic and abdominal aorta and analyzed stiffness-related gene expression.

Methods:
36 WT mice (C57BL/6) mice were infused with nicotine or PBS using osmotic mini pumps for 42 days. Thoracic segment (TS) and abdominal segment (AS) aortic stiffness were analyzed using ultrasound (M-Mode and PW). TS and AS were further investigated by
ex vivo
myograph measurements. Gene expression for TMIP2, MT1-MMP, MMP2, collagen type I and type III was performed for both segments separately.

Results:
Myograph measurements revealed increased strain within the AS after 2 weeks (p<.05) in response to nicotine (vs. PBS), but no stiffening of the TS. After 6 weeks, the AS showed additional increases in strain with nicotine (p<.05); however, only minor increases in stiffness could be observed for the TS. Ultrasound M-Mode results confirmed the myograph results. Nicotine treatment also led to increased aortic pulse wave velocity (PWV) after 2 weeks (p<.05) and 6 weeks (p<.05). Gene expression analysis revealed up-regulation in the TS and AS of MT1-MMP and MMP2 after 2 weeks of nicotine, while TIMP2 was downregulated, and collagen type I and type III were up-regulated in both TS and AS (p<.05). After 6 weeks, there were no longer significant differences in either segment for any of these genes.

Discussion:
Aortic stiffening in response to nicotine varies between the TS and AS segment over a time course of 6 weeks. This leads to an increased stiffness gradient between the TS and AS. Gene expression changes in stiffness-related genes occurred in response to nicotine, although no difference appeared between the segments. We conclude that the difference in stiffness development for TS and AS could be based on a different basic morphological structure involving elastin and collagen load, and that these responses may in part explain nicotine’s role in promoting AAA.
Abstract 467: Fibronectin Extradomain B (FN-EDB) Expression is Specific to the Atherosclerotic Lesion Types III, IV, and V, and the FN-EDB targeting Nanomedicine Enhances Atherosclerotic Plaque Detection and Local Delivery of Model Drug Cargo
Authors: Yu Mikyung , Leitao Renata , Xu Xiaoding , Saw Phei Er , Ortega Carleena Angelica , Si Kevin , Ahn Suyeon , Liu Jerry , Lotfi Azita , Lee In-Hyun , Choi Won-Il , Brayner Mirna , Shi Jinjun , Mahmoudi Morteza , Jon Sanyong , Schoen Frederick , Farokhzad Omid
Published: July 2021

Background:
The Fibronection extradomain B (FN-EDB) is upregulated during tissue remodeling and has been postulated as a potential biomarker for atherosclerosis, yet a systematic investigation for the presence of FN-EDB in plaque has not been reported. We hypothesized that the FN-EDB expression would intensify in advanced plaques; furthermore the engineering of FN-EDB-targeted nanoparticles (FN-EDB-NPs) could enable the imaging/diagnosis and local delivery of therapeutic payloads to plaques.

Methods:
The amount of FN-EDB in atherosclerotic (
n
=143) and normal arteries (
n
=43) of human tissue specimen (ages: 40 to 85 years) was histologically stained and quantified using FN-EDB specific bi-podal peptide (APT
FN-EDB
). Next, FN-EDB-NPs were developed by immobilization of APT
FN-EDB
on nanoparticles surface containing Gd-DTPA for MRI imaging. These NPs were administered to apolipoprotein E-deficient mice fed a western diet and the brachiocephalic arteries were visualized with MRI. The mice were sacrificed and the ascending to the descending thoracic aortas and the aortic roots of the mice were studied for Gd, FN-EDB, and FN-EDB-NPs quantification. The utility of FN-EDB-NPs for drug delivery was evaluated using Cyanine, as a model small molecule drug, and NPs biodistribution and pharmacokinetics were studied.

Results:
The FN-EDB positive area was significantly greater in the atherosclerotic tissues than in the normal arteries (
P
<0.001). It was particularly specific to the Type III (
P
<0.01), IV (
P
<0.01) and V lesions (
P
<0.001) compared to the Type I and II lesions. The FN-EDB expression indicated a positive correlation with macrophage infiltration and neoangiogenesis. T1-weighted images of the atherosclerosis mouse models (
n
=86) revealed substantial FN-EDB-NPs accumulation in plaques compared to the control NPs, Magnevist® or wild type C57BL/6J mice (
n
=21). Additionally, FN-EDB-NPs significantly enhanced the blood circulation time (t
1/2
: ~ 6 h) and plaque accumulation (up to 72 h) of a model drug.

Conclusions:
Our findings offer the first evidence that FN-EDB expression is specific to the Type III, IV and V lesions and suggest that the FN-EDB-NPs could be a promising platform for targeted imaging/diagnosis and therapy of atherosclerosis.
Abstract 478: Altered Blood Flow and Anatomic Variations Increase the Translational Value of the PPE Murine Aneurysm Model
Authors: Busch Albert , Maegdefessel Lars
Published: July 2021

Introduction:
Abdominal aortic aneurysm (AAA) is an individual and socioeconomic burden in the aging society. Treatment relies on surgical exclusion of the dilated aorta by open or endovascular repair only. For research purposes, animal models are necessary and the elastase induced aneurysm model (PPE) has gained much attention for it closely mimics endstage human aneurysm disease. However, to improve the translational value of this model we conducted a couple of modifications in relation to human aneurysm morphology.

Materials and Methods:
In 10-week-old male C57BL/6J wild-type mice the classic PPE procedure with local perfusion of porcine pancreatic elastase in order to induce aneurysm was modified using flow alteration by partial ligation of the iliac artery or the aorta. Additionally, careful exploration of the abdominal aortic branches allowed PPE induction at the suprarenal and iliac level. Molecular biology, ultrasound and immunistochemistry were used to evaluate the results.

Results:
Aortic blood flow restriction (PPE 2.0) significantly increases murine AAA diameter and affects the localization of vascular wall remodeling. Suprarenal aortic dissection allows inducing a more proximal aneurysm (PPE 3.0) similar to the angiotensin II induced aneurysm model. Separate investigation for canonical activation of transforming growth factor ß in these two embryologically distinct segments shows no difference. Creating an aorto-iliac bifurcated aneurysm (PPE 4.0) completes the mimicry of human aneurysm morphologies.

Conclusion:
PPE is the most important model for AAA research and the ß-versions PPE 2.0, 3.0 and 4.0 modifying its morphology further increase the translational value.
Abstract 489: A Data Driven Approach to Identify Subtypes of Abdominal Aortic Aneurysm With Distinct Clinical and Genetic Characteristics
Authors: Ye Zi , Austin Erin , Kullo Iftikhar
Published: July 2021

Background:
Abdominal aortic aneurysm (AAA) is a complex and heterogeneous disorder. We hypothesized that dense phenomapping using electronic health records (EHR) can identify subtypes of AAA with distinct clinical and genetic characteristics.

Methods & Results:
AAA cases (n=1108) and controls (n=4694) were identified from the Mayo vascular disease biorepository and had available high-density genotyping data. 132 Candidate genetic variants (p < 10E-5) associated with atherosclerotic cardiovascular disease (ASCVD) or AAA were selected from GWAS catalog. Using model-based Gaussian-mixture cluster analysis of 46 clinical features from 6 data domains, 24 data elements, 204 variables (including age, sex, vital signs, laboratory data, medication use, family history), spanning over 20 years of observational time in the EHR, we identified two subtypes of AAA: subtype 1(S1, n=421) with faster AAA growth rate (baseline size adjusted mean difference, 95% CI: 0.013, 0.001 to 0.020 cm / year faster, p = 0.03), higher all-cause mortality (age & sex adjusted hazard ratio, 95% CI: 1.36, 1.09 - 1.70, p < 0.01) than subtype 2 (S2, n=687). As compared with the controls, thyroid disorder / rheumatoid or osteoarthritis / infectious conditions were uniquely associated with increased odds ratio for S1; ASCVD / hyperlipidemia / hypertension were uniquely associated with increased odds ratio for S2. After adjustment for all group-specific diseases, genetic variants in
CDKN2B-AS1, NOA1-REST, ERG, ZNF335-MMP9
and
SMAD3
were associated with S1 and variants in
MMP12, DAB2IP, LHFPL2, LPA, FSTL5
and
SORT1
were associated with S2 (all FDR p < 0.05). After adjustment for disease comorbidities and genetic variants differently associated with S1 and S2, the increased risk for all-cause death of S1 than S2 and the difference in AAA expansion rate in subgroups attenuated (Both p-values > 0.09)

Conclusions:
We identified two subtypes of AAA with different rates of aneurysm expansion and all-cause mortality, which were associated with subtype-specific disease comorbidities and genetic markers, suggesting the potential of leveraging EHR to facilitate individualized medicine in patients with AAA.
Abstract 490: Quantitative and Dynamic Measurements of Aortic Wall of Acute Type-A Aortic Dissection With X-Ray Phase-Contrast Tomography
Authors: Yokawa Koki , Tsukube Takuro , Yagi Naoto , Hoshino Masato , Nakashima Yutaka , Nakagawa Kazunori , Okita Yutaka
Published: July 2021

Objectives:
We previously reported excellent findings of X-ray phase-contrast tomography (PCX) for visualization of the formalin-fixed human aortic wall samples, and PCX enabled to demonstrate changes of tunica media in acute type A aortic dissection (AADA) . This study evaluates quantitative and dynamic measurements of fresh aortic wall samples of AADA with this modality.

Methods:
Fresh human aortic samples of the ascending aorta (n=7) were obtained during emergent aortic repair for AADA. Formalin-fixed human aortic walls of AADA (n=15) and normal aorta (n=15) were also investigated. PCX is approximately 1000 times more sensitive than absorption-contrast X-ray imaging and effective resolution of PCX is 11.7 μm. Quantitative and dynamic measurement has been developed to visualize changes in imaging of fresh aortic wall under various tensile force to simulate physiological condition, in which aortic wall is stretched according to blood pressure.

Results:
In normal aorta, quantitative measurement of density of the media was 1.095±0.003(g/cm3), and no different between intimal side (1.083±0.002) and adventitial side (1.085±0.003). On contrast, in formalin-fixed aorta of AADA, the medial density was 1.063±0.027, significantly lower than normal aorta (Figure-1), and different between intimal side and adventitial side (1.061±0.008 vs 1.081±0.011, respectively; p<0.005). In fresh sample of AADA, distribution of the medial density was equal to that of formalin-fixed aorta and differences of the medial density were clearly observed with elevation of tensile force of the aortic wall (Figure-1). These differences in density within tunica media were well correlated with distribution of elastic fibers and existence of cystic medial necrosis in pathological analysis.

Conclusions:
X-ray phase-contrast tomography was a strong modality to understand aortic structures and pathogenesis of acute type A aortic dissection.
Abstract 499: Lipoprotein(A) and Carotid Intimal Medial Thickness
Authors: Ikezaki Hiroaki , Yokota Yuya , Schaefer Ernst J , Horvath Katalin V , Asztalos Bela F , Furusyo Norihiro
Published: July 2021

Introduction:
Atherosclerotic cardiovascular disease (ASCVD) is a leading cause of death and disability in both Japan and the United States. Carotid intimal medial thickness (CIMT) has been used as a marker of ASCVD risk. We examined various specialized lipoprotein parameters to determine their utilities for predictor of CIMT progression.

Methods:
Using plasma samples obtained from fasting men and women (n=2245) living in the Fukuoka area of Japan, (median age 59 years), at baseline we measured direct high density lipoprotein cholesterol (HDL-C), HDL2-C, HDL3-C, low density lipoprotein cholesterol (LDL-C), small dense LDL-C (sdLDL-C), triglycerides (TG), LDL-triglycerides (LDL-TG), lipoprotein(a) or Lp(a), and adiponectin. All assays were measured on an automated high throughput platform (Olympus AU400) using assay kits from the Denka Seiken Corporation (Niigata, Japan) and had within and between run coefficients of variations of < 5%. Blood pressure, body mass index, use of medications, and history of hypertension, dyslipidemia and diabetes were also assessed. CIMT of the common carotid artery was assessed bilaterally at baseline and after 5 years of follow up used trained technicians with analysis coefficients of variation of < 10%. Participants using statin therapy at baseline and during follow up were excluded.

Results:
After controlling for age and gender, only Lp(a) was significantly associated with change in CIMT over a 5 year period in this population. The data indicated that for every 1 log unit higher Lp(a) level at baseline, there was a 0.06 mm increase in CIMT over 5 years (p=0.02). While relationships were seen with other parameters and CIMT change, none of these other relationships reached statistical significance at p<0.05. We also observed significant relationships (p<0.05) between CIMT progression and age, gender, current smoking, history of hypertension, treatment of hypertension.

Conclusions:
Our data indicate that lipoprotein (a) as measured by immunoassay in an important predictor of the progression of atherosclerosis in the common carotid arteries of middle aged and elderly Japanese men and women, and serves as a valuable marker of ASCVD risk.
Abstract 505: Incidence of Major Bleeding in a Real-World Population of 57,070 Nonvalvular Atrial Fibrillation Patients Treated With Rivaroxaban
Authors: Peacock Frank , Patel Manesh , Yuan Zhong , Sicignano Nicholas , Hopf Kathleen , Tamayo Sally
Published: July 2021

Introduction:
Rivaroxaban, a direct factor Xa inhibitor approved in 2011, reduces the risk of stroke and systemic embolism in patients with nonvalvular atrial fibrillation (NVAF). Phase 3 results from the registration trial showed a major bleeding (MB) rate of 3.6 per 100 person-years.

Objective:
To evaluate MB incidence among NVAF patients taking rivaroxaban in a post-approval clinical setting.

Methods:
From January 1, 2013 to June 30, 2016, over 10 million electronic medical records from the Department of Defense Military Health System were queried. A validated MB case-finding algorithm (Cunningham) was used to identify MB-related hospitalizations. Major bleeding incidence, patient characteristics, comorbid conditions, concomitant medications, bleeding management, and fatal bleeds were assessed.

Results:
Of 57,070 rivaroxaban users with NVAF, 1,914 experienced a MB event, with incidence of 2.60 (95% CI 2.49-2.72) per 100 person-years. Patient characteristics and subgroup bleeding rates are displayed in Table 1. There were 62 patients who experienced fatal bleeding, and the mean (SD) age at time of death was 79.3 (7.6) years. Among those who died, 74.2% had intracranial hemorrhage, 22.6% had gastrointestinal hemorrhage, and 3.2% had bleeding in other sites.

Conclusion:
The incidence, pattern, and management of MB in rivaroxaban users with NVAF in a large and diverse population were generally consistent with the findings from the registration trial. Fatal bleeding was rare.
Abstract 509: Comparison of Drug Elluting Stent and Per Cutenaeous Transluminal Angioplasty in Patients With Infra-Popliteal Peripheral Arterial Disease
Authors: Vasudev Rahul , Sheth MeetKumar , Shah Priyank , Rampal Upamanyu , Patel Hiten , Shamoon Fayez , Bikkina Mahesh , Virk Hartaj
Published: July 2021

Introduction:
Drug-eluting Stent(DESs) have demonstrated improved patency and freedom from target lesion revascularization compared with Bare-Metal stents or Percutaneous Transluminal Angioplasty(PTA); however, the effect on clinical outcome parameters, such as limb salvage and wound healing, remains unidentified. We present a direct comparison of clinical outcomes in patients who underwent DES vs PTA.

Methods:
We collected data of patients who underwent infra-popliteal arterial interventions at our institution. Clinical end points analyzed were all cause mortality, target vessel revascularization, primary vessel patency, and target limb major and minor amputations. Differences between two groups were analyzed by chi square for categorical variables and t test for continuous variables. Statistical significance was considered for P values less than .05 in a 2-sided test.

Results:
Total of 83 cases, n=42 in DES group and n=41 in PTA group were analyzed. Mean age was 71.6 years (range 49-95). Out of the total 83 patients in the study 45 had a history of diabetes (54%) and 51 (61%) were current /past smokers. Average follow up period of 14 months for both the groups. Primary vessel patency was defined as absence of ≥50% restenosis on follow up. During the follow up period vessel patency in DES group [69% (n=29/42)] was significantly higher as compared to 36% (15/41) in PTA group (P=0.04, odds ratio 3.867, 95% Confidence interval: 1.5 - 9.6). Target vessel revascularization in DES group was 24% (10/42) as compared to 32% (13/28) in PTA group (P=0.47, odds ratio 0.67, 95% confidence interval: 0.26 - 1.77). Target limb amputation was 10% (4/42) in DES group as compared to 24% (10/41) in PTA group (P = 0.085), odds ratio 0.33, 95% confidence interval: 0.09 - 1.14). All cause mortality in both the groups were at 10%, 4/42 in DES group and 4/41 in PTA group (P=1, odds ratio 0.97, 95% confidence interval: 0.23 - 4.19).

Conclusion:
To conclude primary vessel patency was superior in DES group as compared to PTA group. Target limb amputation rates were higher in PTA group but not statistically significant. Target vessel revascularization and all cause mortality were similar in both the groups. Thus primary treatment with DES should be considered in patients with infra-popliteal PAD.
Abstract 523: Ultrasound Assisted Thrombolysis Versus Catheter Directed Thrombolysis: A Meta-Analysis
Authors: Zaidi Syed Imran M , Khan Abdul Ahad , Panchal Hemang B , Qutrio Baloch Zulfiqar , Josan Enambir , K Paul Timir
Published: July 2021

Introduction:
Deep venous thrombosis (DVT) and pulmonary embolism (PE) have many methods of treatment including anti-coagulation, thrombectomy and thrombolysis. Thrombolysis can be achieved via systemic or local thrombolytic agents, with standard local thrombolysis achieved via catheter insertion in proximity to the thrombus and delivery of thrombolytic agents. Ultrasound-assisted catheter thrombolysis (UAT) is a relatively newer form of thrombolysis which utilizes ultrasonic energy, along with local thrombolytics to help in thrombus breakdown. The objective of our meta-analysis is to compare UAT and catheter directed thrombolysis (CDT) for treatment of DVT and PE.

Methods:
PubMed database was searched through January 2017. Three studies (n=156) comparing UAT (n=99) and CDT (n=57) for thrombolysis were included. End points were > 50% thrombus lysis, bleeding (moderate and severe), and mortality on short term follow up (<1 year). The relative risk (RR) or mean difference (MD) with 95% confidence interval (CI) was computed and p<0.05 was considered as a level of significance.

Results:
Thrombolysis success rate was similar with UAT and CDT (RR 1.06, CI 0.89-1.27, p=0.49). Moderate and severe bleeding events were similar with both groups (RR 0.71, CI 0.27-1.87, p=0.49). Mortality on short term follow up was significantly lower in UAT as compared to CDT (RR 0.47, CI 0.23-0.95, p=0.04).

Conclusions:
The results of our meta-analysis demonstrated no difference in thrombolysis success rate or bleeding events when using UAT Vs CDT, however short term mortality was significantly lower with UAT. Further controlled trials with larger sample sizes are required to assess the possible benefit of using ultrasonic energy for venous thrombolysis.
Abstract 529: Statin Therapy Decreases Vascular Inflammation and Prolongs Patency in Murine Arteriovenous Fistula
Authors: Cui Jie , Jhajj Harkamal S , Kessinger Chase W , McCarthy Jason , Jaffer Farouc A
Published: July 2021

Background:
AVFs are the lifeline of dialysis patients, but can often occlude. The most common cause of AVF failure is inflammation-driven neointimal hyperplasia and thrombosis. Here, we investigated whether statin therapy with well-recognized anti-inflammatory properties can improve murine AVF patency, and further assessed the anti-inflammatory effects.

Methods:
One week prior to AVF creation, mice were randomized to oral atorvastatin 1.14mg/kg/day or control 100ul PBS (both n=10), administered by daily gavage. AVF were created using an end-to-side internal jugular vein and carotid artery anastomosis. AVF blood flow was measured (Transonic blood flow probe) at day 0 and weekly thereafter until occlusion (blood flow < 0.1ml/min). On day 6, CLIO-VT680, an inflammatory cell-targeted fluorescent nanoparticle, was injected intravenously (10mg/kg). 24 hours later, in vivo survival epifluorescence molecular imaging was performed to visualize inflammatory cell in the AVF venous outflow limb. Histopathological assessment of AVF was performed in on day 7 and 14 mice (6 statin each, 6 control each) to assess venous outflow area, AVF scarring via collagen, and adventitial macrophage content.

Results:
At day 7, the in vivo venous outflow cellular inflammation signal (CLIO-VT680 TBR) was significantly lower in statin-treated mice (3.5±0.16 vs 4.5±0.5 PBS, p=0.02). Adventitial macrophage content was significantly lower in statin group at day 14 (p=0.01). Positive AVF remodeling, a desirable feature for clinical AVF patency, was higher in the statin group as compared to the PBS group (p=0.02 at week 1, p=0.002 at week 2). In vivo, statin-treated animals exhibited greater AVF blood flow (BF) preservation from day 7 to day 14 (ΔAVF BF +0.14±0.2 ml/min vs. -0.79±0.32 ml/min PBS, p=0.02). The ΔAVF BF from day 7 to day 14 correlated inversely and significantly with the day 7 CLIO-VT680 cellular inflammation (r=-0.56, p=0.02), indicating that baseline inflammation may predict AVF failure. Kaplan Meier survival analysis showed median AVF patency in mice treated with statin was 28 days compared to 14 days in PBS treated group (p<0.05).

Conclusion:
Statin therapy prolongs AVF patency and preserves AVF blood flow in association with diminished AVF cellular inflammation.
Abstract 530: In vivo DVT Inflammation Presages Vein Wall Injury, and is Ameliorated by Statin Therapy
Authors: Kessinger Chase W , Qi Guanming , Tawakol Ahmed , Henke Peter K , Jaffer Farouc A
Published: July 2021

Objective:
Inflammation mediates early venous thrombosis (VT) resolution and can induce vein wall scarring (VWS), a key driver of the morbid post-thrombotic syndrome (PTS). Statins exhibit anti-inflammatory properties, and may positively impact VWS after VT. However, whether early inflammation contributes to this process and can be detected is not known. In this study, we hypothesized that early VT inflammation detected by 18F-fludeoxyglucose (FDG) positron emission tomography/computed tomography (PET/CT) could predict subsequent VWS and that both VT inflammation and VWS would be attenuated by statin therapy.

Methods:
Stasis VT was induced by complete ligation in male C57BL/6J mice (n=55) in either the infrarenal inferior vena cava (IVC, n=42) or right jugular vein (n=13). IVC VT mice were randomized to statin or control groups. Statin (rosuvastatin 5mg/kg) was given by oral gavage starting one day prior to VT induction; control mice received PBS. All mice underwent survival FDG-PET/CT venography imaging on day 2. FDG-PET inflammation signals (standard uptake value (SUV), SUVmax, target-to-background ratios (TBR)) were measured. Picrosirius red staining of day 14 VT sections measured vein wall collagen/thickness. Ex vivo VT tissue gamma counting of a subgroup was performed at day 2. Whole-thrombus protein/mRNA and VT tissue sections assessed neutrophil content.

Results:
FDG-PET/CT at day 2 revealed increased FDG uptake in jugular VT over the contralateral sham surgery vein (p<0.001). Statin-treated mice showed a decrease in FDG-PET SUV, SUVmax and TBR (p<0.05 for all). Whole-thrombus analyses and tissue section immunostaining showed reduced thrombus neutrophil content at day 2, without reducing GLUT1 or MPO expression (p>0.05). At day 14, statin therapy significantly reduced VWS (p=0.02). In mice undergoing survival imaging, the day 2 FDG-PET VT inflammation signal correlated significantly with the magnitude of day 14 VWS (IVC VT r=0.74, p<0.001) and jugular models of VT (r=0.62, p=0.02).

Conclusions:
Quantitative FDG inflammation imaging demonstrates that early VT inflammation presages subsequent VWS, and is ameliorated by prophylactic statin therapy. The overall findings support the concept that statins and could reduce VWS and PTS.
Abstract 531: Association of D-Dimer With Oxidized Phospholipids, Antibodies to Oxidation-Specific Epitopes and Plaque Morphology in Coronary Artery Disease
Authors: KOTHARI HEMA , Nguyen Anh T , Hisada Yohei , Yang Xiaohong , Tsimikas Sotirios , Mackman Nigel , Taylor Angela , McNamara Coleen A
Published: July 2021

Objective:
Lp(a) is the prominent lipoprotein carrier of oxidized phospholipids (OxPLs) in plasma and OxPLs on Lp(a) are thought to contribute to its atherothrombotic effects. IgM antibodies to oxidation-specific epitopes (OSE) block proinflammatory properties of OxPLs and higher plasma levels in humans associate with reduced CAD and lower risk of cardiovascular (CV) events. Coronary thrombosis upon rupture of an unstable plaque leads to CV events. We hypothesize that plasma D-dimer, a marker of thrombosis, associates inversely with IgM antibodies to OSE and positively with OxPLs on apoB-100 (OxPL-apoB), Lp(a) and indices of unstable plaques as determined by VH-IVUS.

Methods and Results:
OxPL-apoB, Lp(a) and IgM to OSE (phosphocholine-BSA (PC-BSA) and a malondialdehyde (MDA) mimotope) were measured in 106 subjects with stable CAD undergoing angiography. The plasma D-dimer level was 367.8 ng/mL (range 78.3- 2098.8 ng/mL) and 37 subjects had D-dimer levels (mean ± SD; 918.52 ± 482.81 ng/mL) above the threshold for healthy subjects (>500 ng/mL). D-dimer positively associated with OxPL-apoB (r=0.45, p=0.001) and Lp(a) (r=0.44, p=0.001; n=50) and inversely associated with IgM antibodies to PC-BSA and MDA mimotope (r=0.25, p=0.01 and r=0.20, p=0.04 respectively). Moreover, D-dimer associated inversely with plaque fibrosis (r=0.21, p=0.03), positively with coronary artery calcium (r=0.30, p=0.002) and trended to associate positively with necrosis (r=0.15, p=0.13) in univariate analysis. In multivariable analysis, significant association between D-dimer and calcium persisted while association with fibrosis was nearly significant (p=0.07). Notably, in subjects with D-dimer levels above 500 ng/mL, these associations were stronger (fibrosis; r=-0.45, p=0.006, calcium; r= 0.39, p=0.016, necrosis; r=0.45, p= 0.005).

Conclusions:
Our results suggest associations between Lp(a), OxPLs and potentially atheroprotective IgM antibodies to OSE and markers of thrombosis. While associations are modest and a larger study is needed, results raise the interesting possibility that plasma D-dimer may identify subjects prone to Lp(a)/OxPL-induced subclinical thrombosis, which may increase risk for future adverse events.
Abstract 537: Dock2 Deficiency Mitigates High-Fat Diet-Induced Obesity by Reducing Adipose Tissue Inflammation While Increasing Energy Expenditure
Authors: Guo Xia , Li Feifei , Xu Zaiyan , Chen Shi-You
Published: July 2021

Obesity is a public health problem as its association with type 2 diabetes, cardiovascular disorders and many other diseases. Adipose tissue inflammation is frequently observed and plays a vital role in obesity and insulin resistance. Dedicator of cytokinesis 2 (DOCK2) has shown proinflammatory effect in several inflammatory diseases, but its role in obesity remain unknown. To explore the function of DOCK2 in obesity and insulin resistance, wild-type (WT) and DOCK2 knockout (DOCK2-/-) mice were fed with chow or high-fat diet (HFD) for 12 weeks. Metabolic, biochemical and histologic analyses were performed. DOCK2 expression was robustly up-regulated in adipose tissue in WT mice given HFD. DOCK2-/- mice were protected against HFD-enhanced body weight gain with an improved metabolic homeostasis and insulin resistance. In addition, DOCK2 deficiency attenuated adipose tissue and systemic inflammation accompanied by a reduced macrophage infiltration. Moreover, DOCK2 deficiency induced the adipose tissue browning and increased energy expenditure as shown by the up-regulation of metabolic genes in DOCK2-/- mice. Our data indicated that DOCK2 deficiency can protect mice from HFD-induced obesity, metabolic disorders, and insulin resistance. Therefore, targeting DOCK2 may be a potential therapeutic strategy for treating obesity-associated diseases.
Abstract 540: Activation of Srebp1c Processing is Required in Flx-Induced Hepatic Lipid Accumulation
Authors: Xiong Jing
Published: July 2021

Fluoxetine (FLX), a typical drug belonging to the category of selective serotonin reuptake inhibitors (SSRI), is the most widely prescribed psychoactive drug in the treatment of depression and other mood disorders. It has been demonstrated that the administration of FLX increases the possibility of weight gain and dyslipidemia. We find previously that dysregulation of lipogenic and lipolytic genes is critical in FLX-promoted hepatic lipid accumulation. Therefore, a chronic mild stress depression model and cultured primary mouse hepatocytes were used to investigate the effects and underlying mechanisms of FLX on the promoted hepatic lipid accumulation. The evidence have shown that FLX increases the concentrations of triglyceride (TG) and total cholesterol (TC) in the liver tissues of depressive mice, while only increases TG in the liver tissues of normal mice. FLX induces lipid accumulation in both normal and depressive mice by upregulating the lipogeneic genes Acetyl-CoA carboxylase 1 (ACC1) and Fatty acid synthase (FAS) expression and downregulating the lipolytic genes carboxylesterase 1 (CES1) and CES2. Using primary cultured mouse hepatocytes, it is shown that FLX promotes the expression of transcription factor SREBP1c as well as its proteolytic cleavage and nuclear translocation. FLX significantly suppresses SREBP1c proteolytic cleavage in hepatocytes after the incubation lasting as short as 3 hours, which is a more prompt way than the elevated expression of SREBP1c. Further experiments show that the specific inhibitors of proteases S1P and S2P attenuate FLX-induced SREBP1c activation and hepatic lipid accumulation. As SCAP (SREBP cleavage-activating protein) escorts SREBPs from the endoplasmic reticulum to the Golgi complex where proteases cleave SREBPs and therefore is required for SREBP activation, we find that FLX promotes the protein level of SCAP in a concentration- and time-dependent manner. In conclusion, FLX directly acts on the hepatocytes by facilitating the expression and proteolytic activation of SREBP1c to promote hepatic lipid accumulation. The findings not only provide new insight into the understanding of the mechanisms for SSRI-mediated dyslipidemia effects, but also suggest a novel therapeutic target to interfere.
Abstract 544: Diabetes Mellitus Impacts on Vascular Calcification in Peripheral Arterial Disease Patients but Does Not Further Activates Osteogenic Markers in Beta-Glycerophosphate Stimulated Vascular Smooth Muscle Cells
Authors: Farias-Silva Elisangela , Mendes Cynthia A , Pescatore Luciana A , Andrade Maria Claudina C , Carmo Luciana S , Almeida Youri E , Kaminagakura Marcela A , Koga Mônica , Wolosker Nelson , Liberman Marcel
Published: July 2021

Introduction:
Diabetes mellitus (DM) accelerates vascular calcification (VC) in peripheral arterial disease (PAD), increasing limb ischemia and amputation risk. Although DM and VC implicates in PAD, the mechanisms underlying vascular smooth muscle cells (SMC) osteochondrogenic differentiation in this setting are scarce.

Objectives:
To assess VC and osteogenic mRNA/protein expression (Msx2, Runx2 and ALPL) in arteries and in primary SMC isolated from PAD patients with DM (n=3), without DM (n=2) and individuals without PAD (control [CTL], n=2) that underwent amputation.

Methods:
Ethical committee approved study and patients signed informed consent. 5-7
th
passages SMC were incubated without (untreated) or with β-glycerophosphate 10mM (β-GP) for 48h (mRNA expression), 72h (protein expression) and 14 days (calcification). In addition, we performed tissue immunofluorescence and Alizarin Red S.

Results:
Untreated PAD+DM SMC increased calcification (4.6±1.2) in comparison to PAD without DM patients (2.5±0.2) and CTL individuals (1.0±0.07). β-GP further increased calcification in respective groups (51.3±4.7; 13.5±1.1 and 9.8±1.2). Msx2 mRNA expression decreased in CTL SMC after β-GP (48h) and did not change in PAD without DM and in PAD+DM SMC. Msx2 protein expression decreased in all groups after calcifying medium (72h). Moreover, Runx2 mRNA expression increased in β-GP-treated PAD+DM SMC and ALPL mRNA expression augmented in β-GP-treated SMC from PAD without DM patients. However, Runx2 and ALPL protein expression was not modulated by calcifying medium (72h) in all groups. Interestingly, immunofluorescence of arterial samples demonstrated increased osteochondrogenic protein expression (Msx2, Runx2 and ALPL) around calcifying foci in PAD+DM group (+++)
versus
PAD without DM (++) and
versus
CTL (+) patients. Coincidently, PAD+DM group showed augmented vascular calcification (19x10
4
±10x10
4
; 7x10
4
±2x10
4
; 3x10
3
±10
-2
μm
2
).

Conclusion:
We demonstrated increased calcification in arteries from amputated PAD+DM patients and in respective SMC without and with calcifying medium. Although we found augmented osteochondrogenic protein expression in the vessel wall from PAD+DM individuals this was not robustly shown in isolated SMC
in vitro
.
Abstract 559: Exploiting Macrophage Autophagy-Lysosomal Biogenesis With the Natural Sugar Trehalose as a Therapy for Atherosclerosis
Authors: Sergin Ismail , Zhang Xiangyu , Evans Trent D , Razani Babak
Published: July 2021

The autophagy-lysosome system is a catabolic cellular mechanism that degrades dysfunctional proteins and organelles. In atherosclerosis, there is mounting evidence that this process is rendered dysfunctional particularly in plaque macrophages and is an important trigger for plaque progression. In an effort to characterize practical inducers of macrophage degradative capacity, we now describe the unique vascular benefits of a natural sugar called trehalose, a recognized autophagy inducer with a currently unknown mechanism of action. Trehalose-treated macrophages display enhanced autophagy via a process that involves lysosomal stress and resultant activation of TFEB, the master transcriptional regulator of autophagy-lysosomal biogenesis. We find an important downstream effect of trehalose to be the induction of p62-dependent selective autophagy of cytotoxic polyubiquitinated protein aggregates and dampening of IL-1β/inflammasome function. We confirm the relevance of these in vitro observations in several pro-atherogenic (ApoE-/-) mouse models. Administration of trehalose during eight weeks of Western diet feeding potently induces autophagy and TFEB in plaque macrophages with concomitant reductions in polyubiquitinated protein aggregate burden along with significantly reduced plaque size and complexity. Importantly, these findings are completely abrogated in mice deficient in macrophage autophagy (ATG5-/-) or the selective autophagy chaperone (p62-/-). Further detailed pharmacokinetic evaluation of trehalose shows that physiologically relevant concentrations are indeed achievable in mice. Taken together, our data support the serious consideration of this safe and natural sugar as a potent inducer of macrophage degradative capacity in the treatment of atherosclerotic vascular disease.
Abstract 56: Coronary Artery Disease Risk Alleles at the
LIPA
Locus are Associated With Higher Levels of Its mRNA and Enzymatic Activity in Human Macrophages
Authors: Zhang Hanrui , Xue Chenyi , Reilly Muredach P
Published: July 2021

GWASs have identified
LIPA,
encoding lysosomal acid lipase (LAL), as a novel locus for coronary artery diseases (CAD) but not lipid traits. Meta-analyses revealed that CAD risk alleles rs1412444T and rs2246833T (clustered in intron 2/3 in high LD, r
2
=0.985) were associated with higher
LIPA
mRNA in monocytes. It is, however, unclear whether the risk alleles are associated with increased
LIPA
mRNA or LAL enzymatic activity in monocyte-derived macrophages (HMDM), the cell type playing critical roles in atherogenesis. Fine mapping of the
LIPA
region through our CARDIoGRAM+C4D consortium confirmed that the original GWAS SNPs showed the strongest signals. Both SNPs were enriched for H3K27Ac marks and binding sites for PU.1 and CEBPβ in HMDM, suggesting potential regulatory roles.
LIPA
mRNA and LAL activity were higher in HMDM of risk allele carriers (by Kruskal-Wallis test).
LIPA
mRNA was reduced by M1(LPS/IFNγ)-activation and increased by M2(IL-4)-activation. The eQTL relationship of
LIPA
was present in M1- and M2-activated HMDM, suggesting the activity of genetic determinants of
LIPA
expression was retained during macrophage activation. We performed allele-specific expression analysis with RNA-seq of HMDM from 30 subjects of European ancestry (EA) (~70M uniquely-mapped reads/sample) genotyped by Illumina Infinium Multi-Ethnic Global BeadChip imputed with 1000G Phase 3 v5. rs1051338 is within an annotated exon, in high LD with the GWAS lead SNPs (r
2
=0.859 in EA), and had consistent allelic expression favoring G allele, which is on the same haplotype as the risk alleles of the GWAS SNPs. In addition,
LIPA
mRNA was higher in human coronary arteries with atherosclerotic plaque. Immunostaining showed that LAL was abundant within the extracellular space of human atherosclerotic intima and also co-localized with CD68+ macrophages. In summary, the results support a novel, potentially pro-atherogenic role for monocyte-macrophage
LIPA
gain-of-function in CAD.