bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1053: The Role Of Bile Acid-initiated Signaling In Vascular Smooth Muscle Cell Phenotypic Switching And Atherosclerosis
Authors: Holden Claire M , Paredes Felipe , Bogan Bethany J , Williams Holly C , San Alejandra , Lin Zhiyong
Published: July 2024

Atherosclerosis is a chronic inflammatory disease in which foam cells (lipid-laden macrophage-like cells) accumulate in the arterial wall. Vascular smooth muscle cells (VSMCs) are not terminally differentiated; they switch to a foam cell-like phenotype in response to various stimuli, such as cholesterol (CHOL), increasing susceptibility to vascular diseases. Modulated SMCs are the main contributors to atherosclerotic plaque formation. The intracellular metabolism of CHOL via peroxisomes in hepatocytes produces bile acids like cholic or deoxycholic acid, which agonize liver X receptor (LXR). LXR facilitates the first step of reverse CHOL transport. It is unknown if VSMCs synthesize bile acids and whether peroxisomes and the bile acids they produce play a role in VSMC-derived foam cell formation. Using human aortic SMCs (HASMCs), we found that CHOL treatment decreased expression of several peroxisomal proteins. Peroxisomal loss-of-function experiments demonstrated increased intracellular lipid accumulation, as visualized with Oil Red O (ORO) staining. We have shown that HASMCs synthesize bile acids after exposure to CHOL, which led us to hypothesize that bile acid-initiated signaling participates in the phenotypic modulation initiated by CHOL in VSMCs. Treatment of HASMCs with LXR agonists or bile acids increased expression of the efflux CHOL transporter ABCA1, bile acid-related peroxisomal enzymes, and differentiation markers; they decreased expression of macrophage and foam cell markers and ORO staining. These data show that VSMCs have the metabolic capacity to synthesize bile acids, which is impaired if peroxisomes are non-functional or absent. Our data indicates that activation of bile acid receptors upregulates ABCA1, favoring a differentiated VSMC phenotype. Together, these data suggest that activation of LXR plays a role in CHOL-induced VSMC to foam cell transition and that LXR is a potential therapeutic target which promotes VSMC differentiation and decreases atherosclerotic plaque formation.
Abstract 1055: Atxn2 Inhibition Enhances The Contractile Phenotype Of Vascular Smooth Muscle Cells
Authors: Ashraf Jasni V , Fayed Hend S , Al Haj Zen Ayman
Published: July 2024

Introduction:
Phenotype switching of vascular smooth muscle cells (VSMC) between their synthetic and differentiated contractile state is a critical player in controlling the stability of atherosclerotic plaque. Hundreds of genes associated with coronary artery disease (CAD) have been identified through Genome-Wide Association Studies (GWAS). However, the involvement of most identified genes in the pathogenesis of atherosclerosis remains unknown. ATXN2 was identified as a CAD GWAS candidate gene with unclear biological role in atherosclerosis. ATXN2 regulates the endocytic internalization of the EGF receptor, which is essential in the VSMC phenotype switching. Therefore, we aimed to investigate whether ATXN2 is involved in the VSMC phenotype switching.

Methods and Results:
Using immunofluorescence, we demonstrated that ATXN2 siRNA knockdown resulted in a 5-fold increase in the ratio of the contractile phenotype of human aortic VSMC that expressed a high alpha-smooth muscle cell actin content compared to controls. In addition, we found that ATXN2 inhibition reduced the VSMC proliferation and migration as assessed by the EdU cell proliferation assay and the wound healing migration assay, respectively. Next, we confirmed that ATXN2 siRNA knockdown increased the mRNA expression of contractile markers: calponin, alpha-smooth muscle cell actin, and smooth muscle protein 22 alpha using quantitative PCR. We found that the enhancing effect of VSMC contractile phenotype by ATXN2 siRNA was less pronounced in the absence of TGF-beta. Indeed, the enhancing effect of ATXN2 inhibition on the VSMC contractile phenotype was abolished by SMAD4 gene silencing, suggesting the involvement of TGF-beta signaling. Furthermore, we validated the expression signature of the ATXN2 gene in different phenotypic subsets of VSMC from human atherosclerotic plaque samples using single-cell RNA sequencing analysis. We detected that ATXN2 expression was negatively correlated with the contractile VSMC subset. In conclusion, our findings suggests ATXN2, CAD GWAS candidate gene, as a novel potential therapeutic target to enhance the VSMC contractile phenotype for stabilizing atherosclerotic plaque in coronary artery disease patients.
Abstract 1061: The Phospholipase cγ2 And Protein Kinase cα Signaling Axis Is A Key Regulator Of The Extracellular Digestion Of Lipoprotein Aggregates By Macrophages
Authors: Steinfeld Noah , Ma Cheng-I J , Maxfield Frederick R
Published: July 2024

Atherosclerosis is characterized by cholesterol ester accumulation in macrophages, leading to foam cell formation. In atherosclerotic lesions, most lipoproteins are aggregates (agLDL) that are tightly crosslinked to the extracellular matrix, making digesting them by endocytosis or phagocytosis difficult. Thus, it has been unclear how the cholesteryl esters in the core of agLDL are hydrolyzed. Our laboratory discovered a novel mechanism macrophages use to digest agLDL and generate free cholesterol in an extracellular, acidic, hydrolytic compartment known as the lysosomal synapse. Macrophages form a tight seal around agLDL through actin polymerization and deliver lysosomal contents into this space in a process termed digestive exophagy. Vacuolar ATPase on the plasma membrane lowers the pH of the lysosomal synapse, enabling lysosomal acid lipase (LAL) activity. LAL generates free cholesterol, which accumulates in macrophages and leads to foam cell formation. Our laboratory has begun to characterize the signaling pathways that regulate digestive exophagy, having previously identified TLR4 activation of MyD88/Syk as critical. Here, our goal was to identify digestive exophagy regulators downstream of Syk. Syk activates Bruton’s tyrosine kinase (BTK) and phospholipase Cγ2 (PLCγ2). We tested the hypothesis that Syk phosphorylates and activates PLCγ2 during digestive exophagy.

Using pharmacological treatment and genetic manipulation, we found that PLCγ2 and to a lesser extent BTK regulate digestive exophagy. We confirmed that inhibition of Syk or BTK leads to a loss of PLCγ2 activation. PLCγ2 cleaves PI(4,5)P
2
into diacylglycerol (DAG) and inositol 1,4,5-trisphosphate (IP
3
). Soluble IP
3
then activates the release of Ca
2+
from the endoplasmic reticulum (ER) into the cytoplasm. We demonstrated that Ca
2+
release from the ER is upregulated by agLDL and plays a role in digestive exophagy. Both DAG and Ca
2+
activate protein kinase Cα (PKCα). We also found that PKCα regulates digestive exophagy. In conclusion, we identified new signaling mechanisms regulating digestive exophagy downstream of Syk. We hope that expanding our understanding of the mechanisms of digestive exophagy will be helpful in identifying therapeutics to slow atherosclerosis.
Abstract 1070: Predicting Short Term Mortality In Patients With Acute Pulmonary Embolism With Deep Learning
Authors: cicek Vedat , Orhan Ahmet Lutfullah , Saylik Faysal , tur yalcin , Erdem Almina , Babaoglu Mert , Ayten Omer , Taslicukur Solen , Oz Ahmet , Keser Nurgul , Hayiroglu Mert Ilker , Bagci Ulas , Cinar Tufan
Published: July 2024

Background:
Pulmonary embolism (PE) is one of the leading causes of short-term mortality among cardiovascular diseases. Rapid and accurate stratification of PE-related risk is vital for effective patient management. A strong potential research in this direction is to utilize deep learning (DL) approaches. DL approaches have enhanced risk prediction performance by leveraging novel risk predictors and analyzing complex interrelationships among them. In this study, we developed a DL based new scoring system superior to the current standard, the Pulmonary Embolism Severity Index (PESI) , when predicting the short-term (0-30 days) mortality of PE.

Materials and Methods:
Our study included a total of 207 patients. The diagnosis of acute PE was confirmed using contrast-enhanced computed tomography (CT) scans. DL model was created using CT scans and corresponding ground truth labels (short term mortality). We have benchmarked different DL architectures for this purpose including CNN and Transformers.

Results:
Out of 207 patients, 53 of them died during short term . The research group was categorized into two groups based on their short term survival status. Multivariable logistic regression analysis was used to determine the independent relationships between variables and short term mortality. Short-term mortality was predicted statistically significantly (p<0.001) in the DL model developed from the patient's CT images (Figure 1). Then, we developed a new scoring model by uniquely combining the short-term mortality independent predictors, baseline data and the DL model (Figure 1). The ROC analysis demonstrated an AUC of 0.976 (p<0.001) for the Baseline-DL model and an AUC of 0.865 (p<0.001) for the PESI score The Baseline-DL model was statistically significantly superior to the PESI (p<0.001).

Conclusion:
We developed a DL based new scoring system to predict short term mortality of patients with PE, significantly superior to the PESI.
Abstract 108: Inflammasome Activation Via BRCC3-mediated NLRP3 Deubiquitylation Promotes Atherosclerosis In
Tet2
Clonal Hematopoiesis
Authors: Yalcinkaya Mustafa , Liu Wenli , XIAO TONG , Abramowicz Sandra , Wang Nan , Westerterp Marit , Tall Alan
Published: July 2024

Clonal hematopoiesis (CH) has emerged as an independent risk factor for atherosclerotic cardiovascular disease (CVD) with activation of macrophage inflammasomes as a potential underlying mechanism. The NLRP3 inflammasome has a key role in promoting atherosclerosis in mouse models of
Tet2
CH, while inhibition of the inflammasome product IL-1β appeared to particularly benefit patients with TET2 CH in CANTOS. TET2 is an epigenetic modifier that decreases promoter methylation. However, the mechanisms underlying macrophage NLRP3 inflammasome activation in TET2 deficiency and potential links with epigenetic modifications are poorly understood. We used cholesterol-loaded TET2 deficient murine and embryonic stem cell derived isogenic human macrophages to evaluate mechanisms of NLRP3 inflammasome activation in vitro and hypercholesterolemic

Ldlr
-/-

mice modeling TET2 CH to assess the role of NLRP3 inflammasome activation in atherosclerosis.
Tet2
deficiency acted synergistically with cholesterol loading in cell culture and with hypercholesterolemia in vivo to increase JNK1 phosphorylation and NLRP3 inflammasome activation. The mechanism of JNK activation in TET2 deficiency was increased promoter methylation and decreased expression of the JNK-inactivating dual specificity phosphatase,
Dusp10
. Active Tet1-deadCas9 targeted editing of
Dusp10
promoter methylation abolished cholesterol-induced inflammasome activation in
Tet2
-deficient macrophages. Increased JNK1 signaling led to NLRP3 deubiquitylation and activation by the deubiquitinase BRCC3. Accelerated atherosclerosis and NETosis in
Tet2
CH mice were reversed by holomycin, a BRCC3 deubiquitinase inhibitor, and also by hematopoietic deficiency of
Abro1
, an essential scaffolding protein in the BRCC3-containing cytosolic complex. Hypercholesterolemia and TET2 deficiency converge on a common pathway of NLRP3 inflammasome activation mediated by JNK1 activation and BRCC3-mediated NLRP3 deubiquitylation with potential therapeutic implications for the prevention of CVD in TET2 CH.
Abstract 1085: Vagal Nerve Schwannoma Presenting As A Syncope In A Young Patient
Authors: Aurshina Afsha , Imtiaz Zainab , Rajagopalan Varun , Singh Priyanka , Khan Muhammad S , Vitellas Michail , Nadeem Muhammad
Published: July 2024

Introduction:
Vagal schwannomas are rare, benign masses that usually develop in the cervical region but can arise anywhere along the vagus nerve. Patients may be asymptomatic; however, in most cases, patients present in their third to sixth decades, with neck swelling, hoarseness or cough. Herein, we discuss a case report of a 26-year-old male who presented after an abrupt syncopal episode with sinus bradycardia and was diagnosed with vagal schwannoma.

Case Presentation:
A 26-year-old male with a past medical history of bipolar disorder was brought to the hospital after an abrupt syncopal episode while driving. He denied any palpitations or prodromal symptoms prior to loss of consciousness. Inpatient cardiac monitoring revealed persistent sinus bradycardia, without high degree AV block. Treadmill EKG stress test confirmed chronotropic competence. CT head did not reveal other etiology of syncope. CT chest revealed a 2.4 x 2.0 cm left superior mediastinal mass. Cardiothoracic surgery was consulted for concern of thoracic outlet mass/mediastinal mass. Robotic-assisted left thoracoscopy was performed for mediastinal mass excision, which revealed a left vagus nerve schwannoma on frozen section.

Discussion:
Most vagus nerve schwannomas have been found to arise within the cervical course of the vagus nerve. Intrathoracic vagal schwannomas are rare. These tumors are usually asymptomatic, however varied symptoms such as pain, dysphagia and shortness of breath have been reported. In this case presented, the tumor possibly contributed to the heightened vagal tone and neurogenic syncope (with vasovagal syncope). It is also notable that our patient's resting heart rate normalized following excision of the mass. Of note, with a high reported incidence of peri-perioperative vocal cord paralysis, surgical excision remains a challenging mainstay of treatment in symptomatic patients.

Conclusion:
Vagal neoplasms should be hence considered as a differential diagnosis in patients with otherwise unexplained bradycardia with history of syncope.

Reference:
Neurocardiogenic syncope; Carol Chen-Scarabelli, cardiovascular nurse practitioner and Tiziano M Scarabelli, associate professor of internal medicine. BMJ 2004 Aug 7; 329 (7461):336-341.
Abstract 1095: Association Between Active Cancer And The Risk Of Thrombotic And Cardiovascular Outcomes In Patients With Covid-19
Authors: McGonagle Bridget , Greason Christie M , Krishnathasan Darsiya , Bejjani Antoine , Khairani Candrika , Hamade Nada , Leyva Hannah , Scimeca Giovanni , Campia Umberto , Davies Julia , Porio Nicole , Assi Ali , Armero Andre , Tristani Anthony , Ortiz-Rios Marcos D , Nauffal Victor , Almarzooq Zaid , Wei Eric , Zuluaga-Sanchez Valeria , Zarghami Mehrdad , Achanta Aditya , Jesudasen Sirus , Tiu Bruce , Merli Geno J , Leiva Orly , Fanikos John , Sharma Aditya , Rizzo Samantha , Pfeferman Mariana , Morrison Ruth B , Vishnevsky Alec , Hsia Judith A , Nehler Mark , Welker James , Bonaca Marc P , Carroll Brett , Goldhaber Samuel , Lan Zhou , Bikdeli Behnood , Piazza Gregory
Published: July 2024

Background:
Active cancer and COVID-19 are each independently associated with excess risk of thrombotic and cardiovascular outcomes. Data are limited regarding the risk of these outcomes in patients with both active cancer and COVID-19.

Aim:
To examine the association between active cancer and thrombotic and cardiovascular outcomes among patients with COVID-19.

Methods:
Data from patients with confirmed COVID-19 from the multicenter CORONA-VTE Network registry were used. Active cancer was defined as having a malignancy diagnosis within the past year or receiving related treatment. Outcomes were adjudicated and included (1) a composite of venous and arterial thromboembolism and (2) a composite of major adverse cardiovascular events, including thromboembolism, heart failure, myocarditis, new atrial fibrillation, and cardiovascular death within 90 days of COVID-19 diagnosis.

Results:
The registry included 3,844 inpatients and 6,576 outpatients, of whom 247 (6.4%) and 199 (3.0%) had active cancer, respectively. For inpatients with and without cancer, the cumulative incidences of thromboembolism were 8.6% and 8.3%, respectively (hazard ratio [HR:1.05; 95% Confidence Interval [CI]: 0.68-1.64, P = 0.82) (Figure). The corresponding cumulative incidences of cardiovascular events were 16.1% and 15.3% (HR: 1.04; 95% CI: 0.75-1.45, P = 0.79). For outpatients with and without cancer, the cumulative incidences of thromboembolism were 3.6% and 1.2%, respectively (HR: 3.07; 95% CI: 1.41-6.67, P = 0.005). Corresponding cumulative incidences of cardiovascular events were 4.6% and 1.9% (HR: 2.47; 95% CI: 1.25-4.86, P = 0.009).

Conclusions:
There was no significant difference in the incidence of thrombotic or cardiovascular events among inpatients with and without active cancer, yet outpatients with COVID-19 and active cancer demonstrated a significantly increased hazard of thrombotic and cardiovascular events compared with non-cancer outpatients.
Abstract 1102: Post Gwas Functional Genomics Analysis To Identify Non-coding Single Nucleotide Polymorphisms Controlling The Risk Of Spontaneous Coronary Artery Dissection
Authors: Chan Irene N , Yue Yunshan , Chan Stephen Y , Sun Wei
Published: July 2024

Background:
Spontaneous Coronary Artery Dissection (SCAD) is a non-atherosclerotic coronary artery disease and the leading cause of acute myocardial infarction in young females. SCAD results from the combination of genetic susceptibility and acquired triggering factors including inflammation and sex hormones. Three loci have been revealed by two large-scale genome-wide association studies (GWAS) in SCAD. However, GWAS only reveals the statistical associations between the genetic loci and diseases, and the biological mechanisms remain unknown.

Methods:
GWAS-reported tag single nucleotide polymorphisms (SNPs) and the linked haplotype SNPs were collected. Synthesized oligonucleotides representing the SNP alleles were subjected to electrophoretic mobility shift assays (EMSA) with nuclear proteins extracted from vascular endothelial cells (ECs), smooth muscle cells (SMCs) and fibroblasts. Hi-C analysis was performed to predict the potential target gene(s) based on chromatin interactions. The functionality of the SNP-associated target genes was examined in cultured vascular cells by RNA silencing.

Results:
The risk allele of tag SNPs in three genetic loci (chr1q21.2, 6p24.11 and 12q13.1) were associated with 1.5 to 1.8-fold higher risk of SCAD. Intronic, non-tag SNP rs7549396 (G/A) in chr1q21.2 locus was identified as the functional SNP in EMSA, given its consistent allele-imbalanced gel shifting pattern in ECs, SMCs and fibroblasts. Hi-C analysis in human aortic tissue identified a 490kb long-range chromatin interaction between SNP rs7549396 and the promoter of Annexin A9 (ANXA9), an extracellular matrix (ECM) and an inflammation-related protein potentially associated with the pathogenesis of SCAD. The knockdown of ANXA9 in vascular cells showed significant changes in inflammatory makers (IL-1β, vCAM-1 and CCL-2) and ECM degradation enzyme (MMP-1).

Conclusions:
Leveraging our post-GWAS functional genomics approach, we identified functional non-coding SNPs in disease-associated genetic locus and predicted a novel potential pathogenic gene ANXA9 for SCAD. Further studies on the functions of this SNP and its associated target gene ANXA9 will provide genetics and mechanistic explanation for this mysterious disease.
Abstract 1104: Investigation Into The Potential Role Of Perceived Loneliness And Endothelial Dysfunction In A Community-based African American Cohort At-risk For Cardiovascular Disease: A Translational Approach
Authors: Baumer Yvonne , Stanton Michael V , Tirado Brian A , Ortiz-Whittingham Lola R , Baez Andrew S , Gutierrez-Huerta Cristhian , Jacobs Joshua A , Aquino Peterson Elizabeth M , Andrews Marcus , Mitchell Valerie M , Collins Billy S , Muallem Schwartz Daniella M , Dobrian Anca , Powell-Wiley Tiffany M
Published: July 2024

Background:
Chronic stress is a significant contributor for cardiovascular disease (CVD) and existing health disparities. Loss of endothelial barrier integrity is a hallmark of CVD, but little is known how chronic stress impacts endothelial barrier integrity. This study aimed to investigate the signaling pathways of how loneliness-related chronic stress could impact endothelial barrier integrity.

Methods:
We examined the relationship among validated questionnaires of loneliness-related chronic stress and various hormonal and cytokines in a community-based cohort of African American adults at risk for CVD from resource-limited neighborhoods . Then, we conducted ex vivo as well as in vitro experiments to probe these effects and investigate the mechanism of action.

Results:
We determined that a 1-SD increase in loneliness was associated with a 0.32-increase (p=0.04) in the product of epinephrine (E, a catecholamine increased in individuals experiencing chronic stress) and TNFa (T, a cytokine increased in individuals experiencing loneliness), suggesting a synergism between epinephrine and TNFa. In ex vivo experiments, the product of E*T was associated with a loss in endothelial VE-cadherin expression (p=.03). In subsequent in vitro experiments to investigate this effect we treated human aortic endothelial cells with E, T, or their combination; together, these biomarkers decreased endothelial VE-cadherin expression (E: 0.94 ± 0.11, T: 0.79 ± 0.05 vs E+T: 0.58 ± 0.06-fold change of control, p<0.01) and dampened endothelial barrier integrity by increasing inter-endothelial gap formation (E: 1.59 ± 0.16 and T: 1.65 ± 0.17 vs E+T: 2.25 ± 0.13-fold change of control, p<0.001). Furthermore, we determined that the impact of E+T on endothelial cells is JAK/Stat signaling pathway dependent, establishing a putative mechanism.

Conclusions:
We found that a combination of epinephrine and TNFa decreased endothelial barrier integrity, suggesting an additive effect potentially accelerating CVD development and progression in individuals experiencing loneliness-related chronic stress. We provide a potential mechanism by which chronic stress could impact endothelial barrier integrity and function - ultimately worsening CVD health disparities.
Abstract 1107: Unveiling Sex-based Differences In Plaque Histology In Patients With End-stage Peripheral Arterial Disease
Authors: Drake Madeline , Kansul Fouzul , Csore Judit , Vela Deborah , Roy Trisha
Published: July 2024

Introduction:
Recent studies on peripheral arterial disease (PAD) indicate prevalence of PAD in women to be equally if not greater than men, also suggesting that the latter have poorer outcomes with endovascular treatments.

Hypothesis:
Sex-related differences in treatment outcomes in PAD may be attributed to variations within the vessel wall.

Aims:
To evaluate sex-based differences in calcification patterns and lesion characteristics in PAD patients.

Methods:
35 legs from 34 PAD patients (15 women) undergoing major amputation were collected. Vessels were harvested and cross-sectioned at 3-4 mm intervals, followed by histochemical staining, resulting in 1,278 arterial rings, each analyzed by an expert cardiovascular histopathologist.

Results:
Calcification was more prevalent in men (78.1%) than women (67.2%), though not significantly (p=0.118). The majority of calcified segments were concentric (>75% of circumference) for both men and women (66.7% and 64.4%, respectively). Women had a greater occurrence of predominantly medial calcification than men (73.3% vs. 65.3%, p=0.145). Of atherosclerotic features, men presented more fibrocalcific lesions (46.9% vs. 31.3%, p=0.023), while fibroatheroma (7.5%, vs men 4.2%, p=0.280) and fibrous plaque (10.4%, vs men 4.2% p=0.072) were more prevalent in women). In non-atherosclerotic segments, adaptive intimal thickening was more prevalent in men (37.5%) than women (16.4%, p<.001). Luminal thrombus was typically seen in women (38.7% vs. 25.0%, p=0.016), mostly presenting in an organized, chronic form in women (26.9% vs. men: 10.4%, p=0.016) and an acute form in men (6.3% vs. women: 0.0%, p=0.016).

Conclusions:
The localization of calcium, differences between atherosclerotic and non-atherosclerotic features, as well as the presence and form of luminal thrombus may guide treatment choices. Further studies are warranted to evaluate the impact of these differences on outcomes of endovascular procedures.
Abstract 1122: Endothelial
Cd36
Mediates Lipid-induced Stiffening Of Aortic Endothelium And Western Diet-induced Barrier Disruption
Authors: Aguilar Victor , Le Master Elizabeth , Paul Amit , Ahn Sang J , Lazarko Dana , Lee James , Levitan Irena
Published: July 2024

Introduction:
Western diet (WD) associates with elevations in blood lipids. Endothelial cells (EC) uptake lipids through scavenger receptors such as CD36. In our studies we demonstrated that exposure to lipids induces a stiffening response in ECs, with
in vivo
studies showing that this stiffening is abrogated in mice with a global CD36 knockout.

Hypothesis:
WD associates with aortic barrier disruption.
in vitro
studies have demonstrated that EC stiffening correlates with barrier disruption. It is our hypothesis that EC stiffening mediates WD-induced aortic barrier disruption in an endothelial CD36-dependent manner.

Methods:
To generate a model of endothelial-specific CD36 deficiency, we crossed tamoxifen-inducible Cre mice with an EC-promoter with CD36
fl/fl
mice. Cre only mice are employed as controls. At the end of the study window, aortas are excised for assessment of EC stiffening via atomic force microscopy. Separate cohorts are assessed for barrier disruption via quantification of Evans blue dye (EBD) leakage or VE-cadherin junctional thickness. Inhibition of CD36
in vitro
was accomplished with SSO. For cell experiments we employed long-chain fatty acids (LCFAs), lipid ligands for CD36.

Results:
WD induced significant stiffening of the aortic endothelium [p<0.05]. This phenomenon was only observed in male (n=5-6) and not female (n=3-10) mice. EC stiffening associated with aortic barrier disruption as indicated by increased EBD permeation (n=3-8) and increased VE-cadherin thickness (n=3-4) [p<0.05]. Use of an endothelial-specific CD36 deficiency significantly abrogated stiffening and barrier disruption [p<0.05]. Controls demonstrated no effects by tamoxifen (n=2-3). Exposure to 5 μM of palmitic and stearic acids (saturated LCFAs) results in stiffening while exposure to oleic acid (unsaturated LCFA) does not (n=5-12) [p<0.05]. Palmitic acid-induced EC stiffening was shown to be dependent on CD36 (n=5-6) and activation of RhoA protein kinase (n=3-8) [p<0.05].

Conclusions:
Our results demonstrate that endothelial CD36 is required for WD-induced EC stiffening and subsequent disruption of the aortic barrier. Furthermore, our results suggest that CD36 mediates EC stiffening through its role as scavenger receptor to saturated LCFAs.
Abstract 1133: Docosahexaenoic Acid’s Impact On The HMG CoA Reductase Pathway: Endothelial Cell Growth State-dependency And Implications For Cardiovascular Health
Authors: Huang Shiqi , Zahradka Peter , TAYLOR Carla G
Published: July 2024

Docosahexaenoic acid (DHA), widely assumed to be atheroprotective, has recently faced scrutiny due to divergent findings in clinical trials studying omega-3 fatty acids and cardiovascular disease (CVD). Our prior work unveiled nuanced actions of DHA on endothelial cells, especially demonstrating its differential regulation of endothelial nitric oxide synthase in growing versus quiescent cells, which approximate the dysfunctional and healthy states
in vivo
, respectively. These findings led to the hypothesis that DHA benefits healthy endothelial cells but not dysfunctional ones. To elucidate novel pathways, RNA-seq was done on DHA-treated (20 or 125 μM for 8 h) and control human EA.hy926 cells in both states. The data were processed by the RSEM-STAR-DESeq2 pipeline. Differentially expressed genes (DEGs) were subjected to enrichment analysis with clusterProfiler. Genes selected from candidate enriched pathways were validated by Western blotting. Principal component analysis showed distinct groupings based on cell growth state and DHA concentration. DESeq2 identified 104 and 173 DEGs unique to growing and quiescent cells, respectively, at 20 μM DHA. Pathway analysis revealed significant enrichment for cholesterol biosynthesis-related terms of downregulated DEGs, including
HMGCR
,
SREBF2
, and
INSIG1
, in quiescent cells treated with 20 μM DHA, while
SREBF1
was downregulated in both states. DHA also reduced
HMGCR
expression in human monocytes, concomitant with less cellular cholesterol content. Moreover, many genes associated with the Rho GTPase pathway, downstream of HMG CoA reductase (HMGCR), were downregulated by 20 μM DHA only in quiescent cells. These results attest to similarities between the effects of DHA and statins, inhibitors of HMGCR that address CVD via both cholesterol reduction and pleiotropic actions on Rho GTPase. Our study reveals that only quiescent endothelial cells respond positively to DHA. It pioneers a novel perspective that DHA should be targeted for CVD prevention in healthy people versus as a therapeutic for those with endothelial dysfunction. Further studies are required to validate the
in vivo
responses to DHA in healthy versus dysfunctional endothelium, and the optimal DHA dose to achieve maximal benefits.
Abstract 1138: Impaired Flow-induced Vasodilation In Arterioles By Cholesterol-induced Suppression Of Endothelial Kir2.1
Authors: Ahn Sang Joon , Fancher Ibra , Phillips Shane A , Levitan Irena
Published: July 2024

Dyslipidemia is a critical factor in endothelial dysfunction; however, the mechanisms are still not well understood. Our studies showed that cholesterol-induced suppression of endothelial flow-sensitive Kir2.1 channels contributes significantly to the impairment of flow-induced vasodilation (FIV) in hypercholesterolemic ApoE
-/-
mice. Notably, a CRISPR mouse generated to substitute Kir2.1 with its cholesterol-insensitive mutant via a single-point mutation results in a full rescue of FIV under hypercholesterolemia. To extend our observations to humans, 16 healthy subjects were recruited with LDL-cholesterol ranging from 51-153 mg/dl, and FIV was assessed in resistance arterioles isolated from gluteal adipose. Kir2.1-dependent FIV was assessed by transducing intact resistance arteries with the adenoviral construct expressing dominant-negative Kir2.1 (dn-Kir2.1) driven by an endothelial-specific (
Cdh5
) promoter. Our results show that Kir2.1-dependent FIV of resistance arteries contributed ~50% of the vasodilatory response in participants with normal/low LDL levels (<100 mg/dL) and was sharply reduced in participants with LDL levels >100 mg/dL. A significant negative correlation was observed between FIV and LDL, which was accounted for by the loss of Kir2.1-dependent FIV component. Furthermore, impaired FIV in in arteries from participants with high-LDL was fully rescued by overexpressing WT-Kir2.1 in the endothelium. These data suggest that cholesterol-induced suppression of Kir2.1 is a major mechanism underlying endothelial dysfunction in hypercholesterolemia. Mechanistically, we showed previously that Kir2.1 is required for the flow-activation of Akt1/eNOS. Our new data show that Kir2.1 is also required for the upstream event of flow-induced activation of Piezo1, PECAM1, PI3K and that transducing the arteries with endothelial specific construct of Akt1 stably located at the membrane (myrAkt1) eliminates the dependence of FIV on Kir2.1. Based on our observation, we conclude that Kir2.1 is required for the translocation of Akt1 to the membrane, and the cholesterol suppression of Kir2.1 interferes Akt1-dependent flow-induced NO production.
Abstract 1140: Contrast-induced Nephropathy Among Hospitalized Patients With Chronic Limb Threatening Ischemia Undergoing Percutaneous Transluminal Angioplasty - A Nationwide Analysis
Authors: Nriagu Bede N , Ozbay Mustafa
Published: July 2024

Introduction:
Among patients with chronic limb-threatening ischemia (CTLI) who are not responsive to lifestyle changes and pharmacotherapy, surgical or endovascular treatment are indicated. Contrast-induced nephropathy (CIN) is a known complication of endovascular interventions. In this study, we examine the prevalence of CIN among hospitalized patients with CTLI undergoing percutaneous transluminal angioplasty (PTA).

Methods:
Using the International Classification of Diseases, 10th edition, Clinical Modification (ICD-10-CM) diagnosis and procedure codes, we queried the National Inpatient Sample 2016 to 2020 for hospitalizations among patients ≥18 years old with a diagnosis of contrast-induced nephropathy and chronic limb-threatening ischemia and a procedure code for lower extremity PTA. We identified the prevalence of CIN among patients with CTLI undergoing endovascular treatment. Sociodemographic differences were examined among patients with CTLI undergoing PTA with and without CIN.

Results:
Our study included 500,690 weighted hospitalizations with CTLI, of which 57,020 (11.4%) underwent PTA. CIN was observed among 690 (1.2%) of patients with CTLI and was more common among males compared to females (53.6% vs. 46.4%) Patients with CIN were older compared to those without CIN (mean [SD] age: 70.7 [11.9] vs 68.8 [11.8] years,
p=0.06
). Among hospitalizations for CTLI who underwent PTA, the proportion of patients with CIN was higher than those without CIN for Blacks (28.4% vs. 21.1%) and Hispanics (14.2% vs. 13.2%) but not for Whites (53.0% vs. 60.1%) and Asian/Pacific Islanders (0.8% vs. 1.4%) (
p=0.4
). CIN was more common among those with Medicaid (65.2%) compared to those with private insurance (17.4%), Medicare (10.1%), and self-pay (2.9%) (
p=0.4
).

Conclusion:
Disparities exist in the prevalence of contrast-induced nephropathy among hospitalized patients with chronic limb-threatening ischemia undergoing percutaneous transluminal angioplasty. Results could be used to support future research to examine factors driving these disparities.
Abstract 1145: A Computational Fluid Dynamics Analysis Of Hemodynamic Differences In Patients With High Grade Asymptomatic Carotid Artery Stenosis
Authors: Braet Drew , Assi Ismael , Dandu Vivek , Ranjane Sid , Carrera Joseph , Srinivasan Ashok , Osborne Nicholas H , Burris Nicholas S , Figueroa C A
Published: July 2024

Introduction:
Current assessment/management of internal carotid artery (ICA) stenosis is based on estimation of percentage (%) stenosis which does not account for trans-lesional flow, pressure gradient (PG), wall shear stress (WSS), or oscillatory shear index (OSI). Computation fluid dynamics (CFD) can be used to characterize such indices.

Hypothesis:
We hypothesize that patients asymptomatic >70% ICA stenosis will have hemodynamic differences that are independent of % stenosis.

Aims:
Evaluate hemodynamic indices in patients with asymptomatic >70% ICA stenosis via CFD.

Methods:
Patients with asymptomatic >70% ICA stenosis were included. 3D time-of-flight (TOF) and phase-contrast (PC) magnetic resonance imaging (MRI) sequences were obtained. CRIMSON was used to generate CFD models containing the carotid artery (CCA), ICA, and external carotid artery (ECA). PC MRI-derived flow waveforms were prescribed to the CCA and ECA. The ICA outlet was coupled to a three-element Windkessel model. Blood was modeled as an incompressible Non-Newtonian fluid with a dynamic viscosity of 0.004 kg m
–1
s
–1
and a density of 1,060 kg m
-3
. Incompressible Navier-Stokes equations were used to solve for velocity and pressures.

Results:
Eight patients (average age 73.8 years, 62.5% male) with asymptomatic high-grade ICA stenosis (62.5% left) were included. Hypertension (87.5%), tobacco use (87.5%), and hyperlipidemia (75%) were common. Most patients were medically optimized with anti-hypertensive (100%), statin (87.5%), and aspirin (87.5%). Three patients (37.5%) underwent ICA intervention.. Mean peak systolic velocity was 406 cm/s and mean end-diastolic velocity was 127 cm/s. There was large variability in trans-lesional ICA flow, PG, WSS, and OSI (Table 1).

Conclusions:
Asymptomatic patients with the same % stenosis have differences in trans-lesional ICA flow, PG, WSS, and OSI. Future work is needed to determine the prognostic value of such differences.
Abstract 1147: Crosstalk Between Alk5 And Mtorc1 Signaling Promotes VSMC Differentiation And The Therapeutic Effect Of Rapamycin
Authors: Chakraborty Raja , Chatterjee Payel , Dave Jui , Obrien Brendan , Joshi Divyesh , Schulz Vincent P , Greif Daniel , Hwa John , Gallagher Patrick , Martin Kathleen A
Published: July 2024

Background:
Vascular smooth muscle cells (VSMC) phenotypic switching contributes to vascular repair and remodeling but also to pathologies including intimal hyperplasia. The mTORC1 inhibitor rapamycin is an effective drug-eluting stent agent that promotes VSMC differentiation. TGFβ also promotes VSMC differentiation through SMAD transcription factors. We investigated unexpected convergence between these pathways in VSMC plasticity.

Methods:
We assessed the interactions between rapamycin and the TGFβ1 (ALK5) signaling in human coronary artery SMCs and in vascular remodeling using inducible SMC-specific knockout mice (ALK5iKO).

Results:
Genome-wide histone H3K27 acetylation analysis unexpectedly identified SMAD binding elements as the motif most enriched after rapamycin treatment of hCASMCs. Treatment with rapamycin promoted rapid phosphorylation of SMAD2/3. Extensive signaling studies revealed that this phosphorylation required ALK5 activity but not TGF-β ligand. Surprisingly, ALK5 and SMAD2/3 were required for rapamycin-induced differentiation. We determined that rapamycin relieves FKBP12 inhibition of ALK5 to promote ligand independent Smad signaling, and FKBP12 knockdown was sufficient to induce contractile genes. Notably, rapamycin treatment induced an interaction between TET2 and SMAD2/3, and these SMADs were required for differentiation-associated chromatin remodeling, including DNA hydroxymethylation and H3K27 acetylation at contractile genes, suggesting that SMADs and TET2 function in concert at SBE- and CArG-containing promoter regions. Furthermore, ALK5iKO mice exhibited severe intimal hyperplasia after carotid artery injury compared to controls and were entirely resistant to the therapeutic effect of rapamycin, despite persistent inhibition of mTORC1. Consistent with in vitro findings, treatment with rapamycin elevated pSmad3 staining post-injury in the medial layer of control but not ALK5iKO mice.

Conclusions:
We report the surprising observation that rapamycin requires FKBP12/ALK5/SMAD2/3 signaling in order to promote TET2-dependent chromatin remodeling and SMC differentiation. Understanding these mechanisms may reveal new therapeutic strategies for treating vascular diseases.
Abstract 1151: Concurrent Obesity And Chronic Alcohol Consumption Exacerbates Skeletal Muscle Mitochondrial Dysfunction In A Murine Model Of Peripheral Artery Disease (PAD).
Authors: Fletcher Emma , Miserlis Dimitrios , Papoutsi Evlampia , Bradley Cassandra E , Wilkinson Trevor , Ring Andrew , Pacher Pal , Koutakis Panagiotis
Published: July 2024

PAD causes ischemic mitochondrial and consequent muscle damage in the legs, amplifying patient morbidity and mortality. While obesity and alcoholism are common comorbidities, their impact on PAD myopathy is unclear. We sought to determine the effect of concurrent hindlimb ischemia (HLI), obesity and chronic alcohol consumption on muscle oxidative stress, and mitochondrial and limb function.

Methods:
Twenty-four male, 24 female, 8 month old C57BL/6 mice received high-fat-sucrose (HFS) or low-fat-sucrose (LFS) diets for 16 weeks, followed by either 20% ethanol (EtOH) added to the drinking water / continued water access for another 12 weeks (
n
=12 mice/4 groups). The left femoral artery was then ligated to induce HLI, and mice continued their assigned diets for another 4 weeks. The quadriceps from ischemic and non-ischemic limbs were excised at euthanasia. High-resolution respirometry measured muscle mitochondrial respiration and reactive oxygen species (H
2
O
2
) output. Muscle oxidative stress and mitochondrial quality control (mitophagy) were assessed enzymatically and by western blot.

Results:
Compared to LFS animals with/without EtOH access, two-way ANOVA showed post-HLI limb function was lower in HFS+water and HFS+EtOH animals (p<0.05). Mitochondrial respiration was lower in LFS+EtOH, HFS+water, and HFS+EtOH ischemic muscle during electron transport chain Complex I, state 2 (I.2), Complex I, state 3 (I.3) and Complex III versus LFS+water controls (p<0.05), and was further reduced during Complex II and IV in LFS and HFS-ischemic muscle exposed to EtOH (p<0.01). Mitochondrial H
2
O
2
yield was increased in HFS+water and HFS+EtOH ischemic muscle at complexes I.2, I.3, II, III and IV (p<0.05), but not in LFS+EtOH muscle. Oxidative damage (carbonyl accumulation) (p<0.0001) and reduced antioxidant levels (superoxide dismutase (p<0.05) and PGC-1alpha (p<0.001)) was only evident in HFS ischemic muscle with/without EtOH exposure. However, the mitochondrial detoxifying enzyme, aldehyde dehydrogenase-2 (p=0.03), and mitophagy markers LC3II:I ratio (p=0.002) and mitofusins 1 (p=0.02) and 2 (p=0.002), were only reduced in HFS+EtOH muscle.

Conclusion:
Obesity and chronic alcohol consumption exacerbates muscle mitochondriopathy in mice with HLI.
Abstract 1155: Symptomatic Low- And High-degree Carotid Stenoses Exhibit Similar Features Of Plaque Instability
Authors: Shlemoun Paul , Hellberg Sofie , Waden Katarina , Bergman Otto , Lengquist Mariette , Buckler Andrew J , Matic Ljubica , Chemaly Melody , Marlevi David , Hedin Ulf
Published: July 2024

Background:
Carotid endarterectomy (CEA) for symptomatic carotid stenosis is recommended for patients with the >70% (NASCET) stenosis but not for those with <50%. Since low-degree stenoses may still cause strokes, refined risk stratification is necessary, which could be improved by incorporating biological features of plaque instability. In order to challenge risk-stratification by the degree of stenosis, we compared biological features of carotid plaques from symptomatic patients with <50% stenosis
vs.
>70% and explored mechanisms behind plaque instability in low-degree stenoses.

Methods:
CEA specimens were retrieved from symptomatic patients with >70% (n=204) and <50% stenosis (n=34), all part of the Biobank of Karolinska Endarterectomies (BiKE). Patient demographics, plaque morphology from image analyses of preoperative computed tomography angiography (CTA), and RNA-seq analyses of lesions were used for comparisons. Plaque biology was assessed by transcriptomics using principal component analyses (PCA), differential gene expression (DGE)- and gene-set enrichment analyses (GSEA). Immunohistochemistry (IHC) was used to study proteins corresponding to upregulated genes.

Results:
The demographics of the two groups were statistically similar. They also showed similar plaque morphology with respect to intraplaque hemorrhage (IPH), plaque burden and fibrous cap thickness, whereas calcification was higher in the <50% group and lipid-rich necrotic core higher in the >70% group. PCA analysis indicated poor clustering, while DGE and GSEA identified enrichment of genes and pathways related to tissue hypoxia and angiogenesis in <50% lesions. Corresponding proteins were detected by IHC in neo-vascularized plaque regions.

Conclusions:
Plaques from symptomatic patients with <50% carotid stenoses exhibit largely similar morphological and biological features as those from >70% stenoses, emphasizing the need for improved stratification of patients for CEA of symptomatic carotid stenosis. However, pathways associating plaque burden, hypoxia and angiogenesis were particularly enriched in low-grade stenosis lesions suggesting mechanisms behind plaque instability and symptoms in patients with both low- and high-degree stenosis.
Abstract 1158: Antiplatelet Therapy Versus Intravenous Thrombolysis For Mild Nondisabling Acute Ischemic Stroke: A Living Systematic Review And Meta-analysis
Authors: Qin Mingzhen , Liu Tingting , Shi Xinyi , Feng Luda , Li Tingting , Cheng Zixin , Cheng Sisong , Zhou Congren , Zou Mingrun , Jia Qi , Zhang Chi , Gao Ying
Published: July 2024

Background:
Previous studies have shown contradictory results between early application of antiplatelet therapy and intravenous thrombolysis (IVT) for mild nondisabling (defined as NIHSS score of 0 to 5) acute ischemic stroke (AIS).

Objective:
To compare the benefits and risks of antiplatelet therapy and IVT in patients with mild nondisabling AIS.

Methods:
A systematic search of MEDLINE, Embase, and Cochrane Library was conducted from database inception until July 26, 2023, without language restriction. Randomized clinical trials (RCTs) or observational studies were selected. The primary outcomes were 90-day functional outcomes, measured by the modified Rankin Scale (mRS) score. The protocol has been registered before data collection (PROSPERO CRD42023447862).

Results:
Two RCTs and four observational studies with relatively low risk of bias that enrolled 3975 patients were analyzed (2454 in antiplatelet therapy and 1521 in IVT therapy). There were no significant differences between antiplatelet therapy and IVT in 90-day functional outcomes (mRS 0-1, odds ratio [OR], 1.08 [95% CI, 0.73-1.58]; mRS 0-2, OR, 1.04 [95% CI, 0.63-1.73]), death (OR, 0.64 [95% CI, 0.19-2.13]), and stroke recurrence (OR, 0.71 [95% CI, 0.28-1.79]). Antiplatelet therapy was associated with a reduced risk of sICH compared with IVT (OR, 0.20 [95% CI, 0.06-0.69]).

Conclusions:
Among patients with mild nondisabling AIS, compared with IVT, early application of antiplatelet therapy was not significantly associated with improved functional outcomes, reduced mortality, or stroke recurrence, but was significantly associated with a reduced risk of sICH. Regular updates will provide the best available evidence as new researches are published.