bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 433: Coronary Artery Ectasia in Patients with Obstructive Coronary Artery Disease
Authors: Xu Shasha , Jiang Jianjun , Zhu Huanhuan , Wang Bing , Fang Congfeng , Xue Yinsheng , Chen Xiaofeng
Published: July 2021

Background and Objective:
The pathophysiology of isolated coronary artery ectasia (CAE) remains poorly understood although associations between CAE and a broad spectrum of different diseases, especially atherosclerotic coronary artery disease have been reported. This study aimed to investigate the clinical characteristics of CAE and its relationship to obstructive artery disease in the Chinese Population.

Methods and Results:
This study recruited 3793 consecutive patients who had undergone coronary angiography for suspected coronary artery disease (CAD) between January 2009 and December 2014. The median age of the patients was 63 years (range, 27 to 94 years) and the majority (69.71%) was male. There were 3068 patients with obstructive coronary artery disease, including angina pectoris (n=1611), acute myocardial infarction (n=1265), and old myocardial infarction (n=192). A total of 120 cases (Male, n=93, female, n=27) with a median age of 63 years (range, 28-86) were identified as CAE in patients undergoing coronary angiography. Co-existent CAD was present in 94% of CAE patients, including 56 angina pectoris; 53 acute myocardial infarction; and 4 old myocardial infarction cases. The prevalence of CAE in patients with obstructive coronary artery disease was 3.67%. The frequency of arterial involvement was: right coronary artery (RCA), 63%; left anterior descending artery (LAD), 43%; left circumflex artery (LCX), 38%; and left main artery (LM), 14%. CAE affected only 1 major vessel in 83% of cases, 2 vessels in 12%, and all 3 vessels in 5%. Using multivariate analysis, serum levels of uric acid were independently associated with present of CAE (P<0.001), while other cardiovascular risk factors such age, arterial hypertension, dyslipidemia, smoking, and diabetes mellitus did not show statistically significant associations (P>0.05).

Conclusion:
The prevalence of CAE in patients with obstructive coronary artery disease was 3.67%. The RCA was the most commonly affected and most patients had single vessel involvement. Increased serum uric acid level may be a potential risk factor for presence of CAE.
Abstract 439: Biomechanical Stimulation Promotes Translocation of Nuclear Factor of Activated T-cells 5 Into The Nucleus of Vascular Smooth Muscle Cells
Authors: Hödebeck Maren , Hecker Markus , Korff Thomas
Published: July 2021

Chronic alterations in the biomechanical stimulation of vascular smooth muscle cells (VSMC) as experienced during hypertension lead to changes in VSMC phenotype and function and further enable the structural remodeling of the vessel wall. In this context, we recently reported that an increase in wall stress or biomechanical stretch is sufficient to activate nuclear factor of activated T-cells 5 (NFAT5). This transcription factor promotes the expression of gene products such as tenascin-C and κ-actin, both involved in VSMC migration. Based on these findings, we hypothesized that biomechanical stretch elicits NFAT5 mRNA expression and induces biochemical modifications of NFAT5 on the post-translational level, a prerequisite for its entry into the nucleus and transcriptional activity. To scrutinize this hypothesis, human arterial VSMC were exposed to biomechanical stretch (13%, 0.5 Hz) and subjected to detailed mRNA expression analyses. While a ~3-fold reduction in NFAT5 splice variant 1 (isoform A) mRNA expression was observed in stretch-stimulated VSMC as compared to the static controls (n=3, p<0.05), splice variant 3 (isoform C) mRNA levels were induced ~1.8-fold (n=3, p<0.05). Overexpression of corresponding Flag-tagged NFAT5 proteins in VSMC and subsequent immunofluorescence as well as biochemical analyses revealed that isoform A was primarily located in the cytoplasm of static and stretch-stimulated VSMC while isoform C was preferentially localized in the nucleus under baseline conditions and further accumulated in the nucleus upon biomechanical stimulation (n=3, p<0.05). Nuclear translocation of isoform C was amplified for phosphorylation-deficient mutants generated by exchanging serine to alanine at position 1197 even under static culture conditions while a phosphomimetic mutation at this residue (serine to glutamate) inhibited NFAT5c nuclear translocation (n=3, p<0.05).

Collectively, our findings indicate that exposure of VSMC to biomechanical stretch triggers the expression of NFAT5 isoform C and controls its entry into the nucleus via phosphorylation at S-1197. Current investigations are focusing on the impact of NFAT5 on hypertensive remodeling utilizing inducible smooth muscle cell-specific NFAT5-deficient mice.
Abstract 44: Organ-specific Stochastic Phenotype Switching is Required for Endothelial Health
Authors: Yuan Lei , Chan Gary C , Beeler David , Janes Lauren , Spokes Katherine C , Mojiri Anahita , Adams William J , Sciuto Tracey , Garcia-Cardeña Guillermo , Molema Grietje , Jahroudi Nadia , Marsden Philip A , Dvorak Ann , Regan Erzsébet R , Aird William C
Published: July 2021

Among unicellular organisms, stochastic phenotype switching is a documented strategy for survival. These populations "hedge their bets": while the majority of their cells are adapted to their present environment, a minority remains poised to thrive under drastically different conditions. Bet hedging has also been described in metazoan cells, primarily in vitro. However, its role in tissue homeostasis has yet to be established. Here, we show that von Willebrand factor (vWF) is expressed in a spatially heterogeneous manner in a small fraction of capillary endothelial cells in the heart, skeletal muscle, lung and brain. Moreover, these mosaic patterns are dynamic, in that vWF expression stochastically toggles ON/OFF over time. By contrast, expression of vWF in the aorta and liver is static in time. In cultured primary endothelial cells, biological noise resulted in mosaic vWF heterogeneity through a promoter-level DNA methylation switch. Finally, vWF-/- mice demonstrated extensive endothelial cell damage in capillaries of the heart and impaired cardiac function, but not kidney or aorta. Taken together, these findings suggest that dynamic mosaicism of vWF expression is functionally relevant and that bet hedging represents a previously unrecognized strategy for adaptive, organ-specific homeostasis.
Abstract 448: Isolation and Characterization of Lesion Specific Cells Harvested in vivo During Percutaneous Coronary Intervention
Authors: Smith Triston B , Pollett Jonathan B , Carothers Wayne J , Lasorda David
Published: July 2021

Background:
Endothelial cells (EC) dysfunction is linked to the development of CAD. Current studies rely on ECs harvested from the umbilical vein or bovine aorta because of the ease of isolation. This fails to account for phenotypic variations and may not adequately represent pathologic changes occurring at the site of specific coronary artery lesion. The need to examine EC phenotypes and cellular processes in the context of existing disease is necessary.

Hypothesis:
Coronary artery lesion specific cells can be harvested and isolated from baloons used during PCI.

Methods:
Coronary balloons used during PCI were immediately placed in endothelial cell media and subjugated to a series of washes, including phosphate buffered saline (PBS) and trypsin. The washes and balloons were centrifuged at 1,700 rpm for 10 minutes and the resulting pellets re-suspended in ammonium-chloride-potassium (ACK)/PBS solution to lyse erythrocytes. These suspensions were centrifuged at 2,500 rpm for 10 minutes to pellet the cells. An aliquot of this final cell harvest was analyzed by fluorescence- activated cell sorting (FACS) with antibodies for endothelial antigens.

Results:
30 samples were analyzed and subpopulations of cells were identified. Between 400-4000 non-hematologic cells per sample were isolated. Cell populations were considered to be endothelial when they were CD31, CD146 double positive and CD45 negative, and populations that were CD222 positive (CD56 CD45 double negative) and CD56 positive (CD222 and CD45 negative) were considered to be fibroblastic and myogenic respectively. Endothelial cells (Figure 1A), myogenic cells (Figure 1B) and fibroblastic cells (Figure 1C) could be visualized.

Conclusions:
Lesion specific cells were successfully harvested and isolated from coronary balloons used during PCI. Proteomic and genetic analyses of these cells may provide invaluable information about pathologic cellular processes during ACS.
Abstract 450: Tumor Necrosis Factor Disrupts Endothelial Surface Glycocalyx Imaged by High-pressure Freezing, Freeze Substitution TEM
Authors: Xu Chang , Austin Joe , Hack Brad , Cunningham Patrick
Published: July 2021

Objective:
The endothelial surface glycocalyx (GCX) is a hydrated mesh formed from a wide variety of endothelial cell membrane-associated macromolecules including glycoproteins bearing sialic acid and proteoglycans. It is thought to be damaged in diseases associated with elevated levels of tumor necrosis factor α (TNF-α) since surface proteoglycan and sialic acid amount decreased after TNF treatment. We imaged in vitro GCX in response to TNF-α using rapid freezing/freeze substitution (RF/FS) transmission electron microscopy (TEM) so that the GCX can be stably preserved in its hydrated configuration.

Methods and Results:
Human glomerular ECs (HGenC) were seeded onto fibronectin- coated PET membrane of the 96-multiwell inserts and allowed to grow to confluence for 4 days. Before experimentation, cells were then treated with TNF (20 ng/ml) or control medium for 24 h. The PET membrane with the cells attached were removed from the Transwell insert by carefully cutting around the membrane edges with a scalpel, and transferred to aluminum planchettes (Ted Pella). The membranes were then subjected to RF/FS-TEM. RF/FS-TEM revealed impressively thick HGenC GCX of ≈13 μm under control condition. In cells treated with TNF, the GCX completely disappeared. Immunoconfocal studies confirmed that in vitro GCX was damaged by TNF.

Conclusions:
New observations by RF/FS-TEM reveal substantial GCX layers on cultured human glomerular EC, which are completely abolished by TNF-α.
Abstract 463: Induction of Smooth Muscle-specific Deletion of Low-Density Lipoprotein Receptor-Related Protein 1 in Adult Mice Augments Angiotensin II Vascular Pathologies
Authors: Rateri Debra L , Balakrishnan Anju , Howatt Deborah A , Moorleghen Jessica J , Cassis Lisa A , Muratoglu Selen C , Strickland Dudley K , Daugherty Alan
Published: July 2021

Objective:
Low-density lipoprotein receptor related protein 1 (LRP1) deletion in smooth muscle cells (SMCs) during embryonic development augments angiotensin (Ang) II induced aneurysm formation in the ascending aorta and superior mesenteric artery. However, these studies have the potential confounding issue of LRP1 deficiency causing structural changes to arteries that may be the basis for this augmented pathology. The purpose of this study was to determine whether AngII-induced vascular changes were influenced when LRP1 deletion occurred in adult mice.

Methods and Results:
To temporally delete LRP1 in SMCs, we used a tamoxifen (tam)-dependent Cre (ERT2) recombinase under the control of a mouse smooth muscle cell actin (SMA) gene. SMA-ERT2-Cre +/0 males containing the ROSA26 reporter were bred to LRP1 floxed female mice to develop SMA-ERT2-Cre 0/0 and +/0 LRP1 x ROSA26 mice. SMC-specific deficiency of LRP1 was induced in mature mice via an injection of an ERT2 ligand, tam (1.5 mg/kg for 5 consecutive days). Non- transgenic littermates were also injected with tam. Tam injections in mice expressing ERT2-Cre led to uniform staining of β-galactosidase in medial SMCs of the aorta and arteries. LRP1 was deleted effectively from SMCs in ERT2-Cre expressing mice as demonstrated by Western blotting of aortic medial extracts. AngII (1,000 ng/kg/min) was infused via mini-osmotic pumps for 28 days into tam-injected mice that were either ERT2-Cre 0/0 or +/0. Consistent with constitutive SMC deletion, SMC LRP1 deletion in mature mice had no effect on dilation of the suprarenal aorta, but augmented expansion of the thoracic aorta and superior mesenteric artery in both male and female mice.

Conclusion:
Consistent with constitutive deletion, SMC LRP1 deletion in mature mice resulted in augmentation of AngII-induced thoracic aortic and superior mesenteric arterial aneurysms. These results demonstrate that undefined ligands acting at LRP1 are responsible for site-specific AngII-induced vascular pathologies.
Abstract 468: HIF-1α Mediates Initiation and Progression of Experimental Abdominal Aortic Aneurysms
Authors: Xu Baohui , Fujimura Naoki , Deng Hongping , Xuan Haojuan , Furusho Yuko , Glover Keith J , Gerritsen Mary , Gurtner Geoffrey C , Michie Sara A , Dalman Ronald L
Published: July 2021

Abdominal aortic aneurysm (AAA) is a life-threatening chronic disease. Although low mural oxygen tension is present in the aneurysmal aorta, with increased hypoxia inducible factor (HIF)-1α expression, the role of HIF-1α in AAA pathogenesis remains uncertain. These experiments separately examined the influences of myeloid cell-specific HIF-1α deletion, as well as pharmacologic HIF-1α inhibition, on experimental AAA progression. Methods: AAAs were created via intra-aortic porcine pancreatic elastase (PPE) infusion in myeloid cell-specific HIF-1α conditional knockout (Lyz2 Cre+/HIF-1α Flox+/+) or control (HIF-1α Flox+/+) mice, as well as C57BL/6J mice treated with HIF-1α inhibitors or vehicle. AAA formation and progression was evaluated in vivo via ultrasonography, and histology at sacrifice. Results: HIF-1α mRNA expression was remarkably elevated in aneurysmal (PPE) as compared to non-aneurysmal (PBS), aortae. Flow cytometry detected nearly 65% of aortic CD11b+ myeloid cells expressed HIF-1α. Myeloid cell HIF-1α deletion reduced time-dependent aneurysmal enlargement noted following PPE infusion. Similar inhibition was observed in WT mice following daily treatment with acriflavine or doxorubicin (1 mg/kg), beginning 3 days prior to PPE infusion. Histologically, both myeloid cell HIF-1α deletion and pharmacologic HIF-1α inhibition attenuated medial elastin degradation and smooth muscle cell depletion, with reduced aortic mural macrophage and neovessel density compared to the corresponding controls. Conclusion: HIF-1α is an essential modulator of AAA pathogenesis, thus representing a novel pharmacologic target for medical disease management.
Abstract 469: Relationship of Heart Rate and Physical Activity with Cardiovascular Risk Factors in a Multi-ethnic Population.
Authors: Grandinetti Andrew , Kaholokula Joseph K
Published: July 2021

Introduction:
Resting heart rate (RHR) has been identified as an independent risk factor for cardiovascular disease and mortality.

Hypothesis:
Physical activity and RHR are associated with hypertension, dyslipidemia and insulin resistance in a multiethnic population.

Methods:
Cross-sectional data from 1,440 participants of Native Hawaiian, Japanese, Filipino, Caucasian, and mixed ethnic ancestries were examined. Body fat was estimated using body mass indices (BMI); fat distribution by waist-hip ratios (WHR); and insulin resistance using the Homeostasis Model (HOMA-IR). Total cholesterol, triglycerides, and HDL-cholesterol levels were assayed from fasting plasma samples. LDL cholesterol was calculated using the Friedewald formula. Blood pressure measurements and medical histories were obtained to determine hypertension status. Leisure time physical activity (PA) was estimated by calculating metabolic equivalents (METS) using the Modifiable Activity Questionnaire. Associations were estimated using logistic regression for dichotomous outcomes (HTN) and general linear models (GLM) for continuous variables.

Results:
Caucasians had lower prevalence of HBP and insulin resistance than all other ethnic groups; there were no statistically significant differences between other ethnic groups on mean RHR. HOMA-IR, RHR, BMI and WHR all differed significantly by ethnic group. Low RHR and high levels of physical activity were inversely associated with insulin resistance. After adjusting for covariates, only RHR remained significantly associated with HBP prevalence. Likewise, only RHR was significantly associated with total cholesterol, LDL and triglycerides. In contrast, HDL cholesterol was positively associated with physical activity but not associated with RHR.

Conclusion:
The relationship of RHR and PA is complex with RHR and PA actions differing for different CVD risk factors. Since low RHR may be an indicator of cardiovascular fitness, these findings suggest fitness, rather than energy expenditure, may be important in preventing most CVD risk factors, while activity level may be more important for increasing HDL levels. A customized approach to activity plans may need to consider specific patient risk profiles.
Abstract 48: Myelopoiesis Following Myocardial Ischemia (MI) Involves Activation of the Nlrp3 Inflammasome by Neutrophil-Derived S100a8/a9
Authors: Nagareddy Prabhakara R , Annabathula Rahul , Ye Shaojing , Klaychkin Yuri M , Abdel-Latif Ahmed , Smyth Susan S
Published: July 2021

Ischemic myocardial damage triggers leukocytosis, particularly the production of monocytes and neutrophils from the bone marrow and spleen (myelopoiesis). These cells infiltrate the evolving myocardial wound, degrade extracellular matrix, and aid in the clearance of dead cardiac myocytes and their debris. Although this inflammatory process is a prerequisite for tissue healing, it is non-specific and often blunt. If unchecked, excessive production of monocytes and neutrophils may result in abnormal ventricular remodeling and heart failure. The myocardial cellular and molecular events that orchestrate with the BM/spleen to regulate myelopoiesis remain unclear. We report here that the number of circulating monocytes and neutrophils peak within 24 hours following coronary artery ligation (LAD) in mice. This is due to expansion and proliferation of hematopoietic stem and multi-potential progenitor cells (HSPC) in the BM as well as extramedullary hematopoiesis in the spleen. MI induced-myelopoiesis was associated with a dramatic increase in the expression of S100a8/a9 (a damage associated molecular pattern), its receptor (Tlr4), the Nlrp3 inflammasome, and pro-IL1β in the heart. Cell separation studies revealed that the infiltrating neutrophils and cardiac fibroblasts are the predominant source of S100a8/a9 and the Nlrp3 inflammasome respectively in the heart. Furthermore, deletion of S100a8/a9 not only reduced MI-induced myelopoiesis but also significantly improved the mortality and cardiac function in mice following LAD. These data supports our hypothesis that neutrophil-derived S100a8/a9 interact with Tlr4 on cardiac fibroblasts to induce the Nlrp3 inflammasome and produce IL1β, which in turn stimulates IL-1R on HSPCs to promote myelopoiesis. Pharmacological strategies aimed at inhibition of S100a8a/9 or the Nlrp3 inflammasome-mediated production of IL1β may be a promising approach to limit inflammation following acute coronary syndrome.
Abstract 483: Non-redundant and Opposing Roles of Group X and Group V Secretory Phospholipase A2s on Pancreatic Beta-cell Function
Authors: Shridas Preetha , Noffsinger Victoria P , Webb Nancy R
Published: July 2021

Background:
Group X and group V secretory phospholipase A2s (GX and GV sPLA2s) potently release arachidonic acid (AA) from the plasma membrane of intact cells. AA is an activator of glucose-stimulated insulin secretion (GSIS) by β-islet cells. However, the AA metabolite prostaglandin E2 (PGE2) is a known inhibitor of GSIS. Both GX and GV sPLA2s are expressed in mouse pancreatic islet cells. We previously demonstrated that GX sPLA2 negatively regulates GSIS by a PGE2-dependent mechanism. In this study we investigated whether GV sPLA2 similarly regulates GSIS.

Methods and Results:
GSIS was measured in pancreatic islet cells isolated from WT and GV sPLA2-deficient (GV KO) mice. To complement these studies, GSIS was also assessed in vitro using MIN6 pancreatic beta cell lines with or without GV sPLA2 overexpression or silencing. In marked contrast to our findings in GX KO mice, GSIS was significantly decreased in islets isolated from GV KO mice compared to WT mice. Similarly, there was a significant decrease in GSIS in MIN6 cells with siRNA-mediated GV sPLA2 suppression. Consistent with these findings, MIN6 cells overexpressing GV sPLA2 (MIN6-GV) showed a significant increase in GSIS compared to control cells. As expected, the amount of AA released into the media by MIN6-GV cells was significantly increased compared to control MIN6 cells. However, unlike MIN6 cells overexpressing GX sPLA2, MIN6-GV cells did not exhibit enhanced PGE2 production or decreased cAMP content compared to control MIN6 cells, despite similar amounts of sPLA2 activity produced by the two cell lines.

Conclusions:
We conclude that GX and GV sPLA2s play opposing and non-redundant roles in pancreatic β-cell function. Whereas GV sPLA2 activates GSIS, GX sPLA2 suppresses this process. This functional difference appears to be due to the extent to which AA generated by the respective sPLA2’s is coupled to PGE2 production.
Abstract 486: Genetic Deletion of Akt2 and AMPK accentuates Insulin Resistance-Induced Cardiac Contractile Anomalies
Authors: Wang Qiurong , Kandadi Machender R , Ren Jun
Published: July 2021

BACKGROUND:
Insulin resistance is associated with high incidence of cardiovascular diseases although the precise mechanisms remain elusive. Heme oxygenase-1 (HO-1) has recently been proven as a proinflammatory involved in the development of metaflammation (metabolic inflammation). This study was designed to examine the role of HO-1 in insulin resistance-associated cardiac dysfunction and potential roles for two pivotal metabolic signaling molecules AMPK and Akt.

Methods:
Whole-body insulin resistance was initiated in wild-type C57BL/6J and AMPK Akt double knockout mice by 8 weeks of sucrose feeding. Contractile and intracellular Ca2+ properties were evaluated in cardiomyocytes using and IonOptix system. Intracellular O2¯ and reactive oxygen species (ROS) were measured.

Results:
Sucrose-fed mice displayed impaired glucose tolerance, enhanced oxidative stress, and normal body weight. Cardiac mechanical properties were compromised in sucrose-fed mice including elevated left ventricular end-systolic and reduced factional shortening. Cardiomyocytes from sucrose-fed mice exhibited depressed peak shortening and maximal velocity of shortening/relengthening, prolonged relengthening duration and impaired intracellular Ca2+ homeostasis, the effects of which were exacerbated in AMPK and Akt double knockout mice. In the sucrose-induced insulin resistant mice, levels of HO-1 and ROS were elevated. Metaflammation related cell signaling markers including TNF-α, NF-κB, IL-6 were upregulated in the sucrose-fed mice, whereas the phosphorylation of Akt and AMPK were downregulated. However, the negative feedback from ROS to Akt and AMPK were absent in AMPK Akt double knockout mice. Levels of HO-1, ROS and metaflammation were enhanced in AMPK and Akt double knockout.

Conclusions:
These findings indicate that metaflammation may play an important role in cardiac contractile dysfunction associated with glucose intolerance and possibly related to proinflammatory HO-1 signaling. A negative feedback loop was present for AMPK/Akt, the deficiency of which led to exacerbated cardiac contractile anomalies under insulin resistance.
Abstract 49: The Transcriptional Repressor MafG Regulates Cholesterol Catabolism
Authors: de Aguiar Vallim Thomas Q , Tarling Elizabeth J , Ahn Hannah , Hagey Lee R , Romanoski Casey E , Lee Richard G , Graham Mark J , Motohashi Hozumi , Yamamoto Masayuki , Edwards Peter A
Published: July 2021

Elevated circulating cholesterol levels is a major risk factor for cardiovascular diseases (CVD), and therefore understanding pathways that affect cholesterol metabolism are important for potential treatment of CVD. The major route for cholesterol excretion is through its catabolism to bile acids. Specific bile acids are also potent signaling molecules that modulate metabolic pathways affecting lipid, glucose and bile acid homeostasis. Bile acids are synthesized from cholesterol in the liver, and the key enzymes involved in bile acid synthesis (
Cyp7a1
,
Cyp8b1
) are regulated transcriptionally by the nuclear receptor FXR. We have identified an FXR-regulated pathway upstream of a transcriptional repressor that controls multiple bile acid metabolism genes. We identify
MafG
as an FXR target gene and show that hepatic MAFG overexpression represses genes of the bile acid synthetic pathway, and modifies the biliary bile acid composition. In contrast, MafG loss-of-function studies cause de-repression of the bile acid genes with concordant changes in biliary bile acid levels. Finally, we identify functional MafG response elements in bile acid metabolism genes using ChIP-Seq analysis. Our studies identify a molecular mechanism for the complex feedback regulation of bile acid synthesis controlled by FXR. The identification of this pathway will likely have important implications in metabolic diseases.
Abstract 495: Body Mass Index is Associated with Progression of Carotid Intima-Media Thickness in Black, but not in White Young Adults: The Bogalusa Heart Study.
Authors: Fernández Alonso Camilo , Barshop Rupert , Galazka Patrycja , Li Shengxu , Chen Wei , Berenson Gerald S
Published: July 2021

Introduction:
Increased Carotid intima-media thickness (CIMT) has been shown to be predictive of future cardiovascular events in middle-age and older adults, with this outcome being detrimentally enhanced by increased body adiposity -as measured by Body Mass Index (BMI).

Hypothesis:
We assessed the hypothesis that the association between BMI and CIMT varies among race (black/white) and gender groups.

Methods:
B-mode ultrasound images of the far walls for both carotid arteries were obtained from 676 community-based subjects, aged 24 to 43 years, enrolled in the Bogalusa Heart Study (70% white and 43% male). CIMT, BMI and traditional cardio metabolic (CM) risk factors were measured at baseline, and a second set of CIMT imaging was performed at follow-up after an average of 6.9 years. Composite CIMT was considered for the analyses. Progression of IMT was defined as an increase of IMT between examinations, that was greater than the upper endpoint of the 95% confidence interval of the mean difference between the baseline samples and their blind duplicates.

Results:
BMI was the only significant predictor of progression of CIMT within the general population (Odds Ratio [OR]= 1.062 p=0.0031), after controlling for age, race, sex and traditional CM risk factors. In race-specific analyses, BMI significantly influenced the odds of CIMT progression only in the black subpopulation (OR= 1.092 p<0.0001). Gender-specific analyses showed independent association of these parameters in females only (OR= 1.063 p=0.0179).

Conclusions:
In conclusion, these findings help support the proposed hypothesis of racial and gender divergences in the association between BMI and progression of CIMT, which suggest that race (black-white) and gender-specific mechanistic differences exist in the impact of increased body adiposity on carotid artery disease. Further, these observations may help strengthen individualized race and gender approaches to cardio-metabolic risk prevention in the community.
Abstract 498: Combinatorial Extracellular Matrix Microarrays for Probing Endothelial Differentiation of Human Induced Pluripotent Stem Cells
Authors: Hou Luqia , Coller John , Natu Vanita , Huang Ngan
Published: July 2021

Human induced pluripotent stem cell (iPSC)-derived endothelial cells (iPSC-ECs) are a promising cell source for vascular regeneration in patients with peripheral arterial disease. However, a critical bottleneck to their clinical translation is the ability to differentiate the cells reproducibly at high yields. Since endothelial cells interact with the basement membrane extracellular matrix (ECM), we sought to examine the role of ECMs on endothelial differentiation using combinatorial ECM microenvironments. ECM microarrays were developed by covalent conjugation of ECMs (gelatin, fibronectin, laminin, heparin sulfate proteoglycans, collagen IV, matrigel) and the multi-component combinations thereof. The pluripotent stem cells attached to the ECMs and subsequently differentiated over the course of 5 days. Endothelial differentiation was semi-quantitatively scored based on the degree of CD31 staining. Our results demonstrated greater levels of CD31staining when cultured on gelatin + matrigel + laminin (G+M+L) or fibronectin + laminin + heparan sulfate (F+L+H), compared to other combinations across three human pluripotent stem cell lines (iPSC-Huf5, iPSC-CON1, and ESC-H1). This enhancement in endothelial differentiation on the microscale was confirmed at larger cell culture platforms in which a marked increase in CD31+ cells was observed in G+M+L modified-dishes (> 5 fold), and F+L+H combination (> 10 fold), compared to gelatin-modified dishes. RT-PCR further confirmed the transcriptional upregulation in endothelial markers for CD31 (> 2 fold) and VE-cadherin (> 4 fold) on G+M+L, compared to gelatin-modified dishes. To elucidate the role of cell-ECM interactions on endothelial differentiation, gene expression of integrin subunits were examined. Gene expression was markedly upregulated in integrins α1 (>10 fold); α4, α5, and αV (>5 fold); and β1, β3 (>50 fold), and β4, when comparing differentiated cells on day14 to undifferentiated cells. The upregulation of integrin subunits was concomitant with upregulation in endothelial genes. Together, this data suggested that combinatorial ECMs differentially promote endothelial differentiation, in part through integrin-mediated pathways.
Abstract 5: Disruption of Semaphorin 7A Confers Protection against the Development of Atherosclerosis
Authors: Hu Shuhong , Yang Fei , Tang Chaojun , Zhu Li
Published: July 2021

Background:
Atherosclerosis is an inflammatory disease of which both innate immunity and adaptive immunity are involved. Increasing evidence suggests that Semaphorin7A (Sema7A), a membrane-associated glycosylphosphatidylinositol (GPI)-linked cell-surface glycoprotein, plays important roles in the immune system and participates in several pathophysiological processes. Whether Sema7A participates in the progress of atherosclerosis is still unclear. Here we investigated the role of Sema7A in the development of atherosclerosis using the Sema7A-deficient mice that have been crossed with atherosclerosis-prone mice. Methods and Results: Atherosclerosis-prone ApoE-/- mice were crossed with Sema7A-/- mice to generate Sema7A-/- ApoE-/- mice and Sema7A+/+ ApoE-/- mice. After 12 weeks on high fat diet, mice were sacrificed and atherosclerotic plaque size was analyzed by en face preparation following Sudan IV Red staining. Although Sema7A deletion did not change mouse blood lipid profile, atherosclerotic lesion size in the whole aorta was decreased by 51.18% (P=00024) in Sema7A-/-ApoE-/- mice (5.01±1.04%, n=13) compared to Sema7A+/+ApoE-/- control mice (10.26±1.15%, n=13). When the aortic root was analyzed using Oil Red O staining, plaque formation was reduced by 45.96% (P=0.0016) in Sema7A-/-ApoE-/- mice (15.80±2.78%, n=7) compared to Sema7A+/+ApoE-/- control mice (31.09±2.56%, n=7). Immunohistochemical studies showed that macrophages within aortic root plaques were significantly reduced by 58.437% (P=0.0010) in ApoE-/- Sema7A-/- mice (24.68±2.767%, n=14) compared with Sema7A+/+ApoE-/- control mice (59.38±4.273%, n=14). Further studies showed a significant decrease in CD4+ cells by 51.31% (P<0.0001) and dendritic cells by 60.4% (P<0.0001), respectively, in the aortic roots in ApoE-/- Sema7A-/- mice compared to ApoE-/- mice. Additionally, Sema7A-/-ApoE-/- mice showed significantly higher collagen content (P<0.0022) and vulnerability index (P <0.0001) than Sema7A+/+ApoE-/- mice. Conclusion: These results demonstrate that genetic deletion of Sema7A delayed the formation of atherosclerotic plaque formation, likely by impairment of the accumulation of macrophages, T cells and dendritic cells in vascular wall.
Abstract 513: Evaluation of Proinflammatory Effects of Low Dose Angiotensin II Infusion in Intact Female Mice and Adiponectin Deficient Mice
Authors: Yu Esther , Wang Xia , Pamidimukkala Jaya
Published: July 2021

Circulating vasoactive peptide Angiotensin II (ANGII) has a well-known role in the development of hypertension and other cardiovascular diseases. In addition it has significant proinflammatory actions in the vascular wall inducing the production of reactive oxygen species, inflammatory cytokines and adhesion molecules. It has been previously shown that estrogen in female mice protects against ANGII mediated hypertension and against proinflammatory effects. It is not clear if the protective effects of estrogen extend to adiponectin deficient mice. Adiponectin is one of the few peptides secreted by fat to have anti-inflammatory properties. The present study evaluates the effect of chronic ANGII infusion on expression of cytokines in female C57BL/6J and adiponectin deficient mice (adipo-/-). Female mice (24- 28wks), were implanted with osmotic pump containing either ANG II (800 ng/Kg /min) or saline. Blood samples and tissue were collected at the end of 14 days. Plasma levels of TNFalpha and IL6 were measured using enzyme linked immunoabsorbent assay. Renal tissue expression of the cytokines were quantified using real-time PCR (Eppendorf Realplex 4 mastercycler) and SYBR Green ROX mastermix. Plasma TNFα levels were similar in saline infused C57Bl/6J(11±1 pg/ml) and adiponectin deficient mice(10± 1 pg/ml). ANGII did not significantly increase TNFα in the control(14±3 pg/ml) or adipo-/- mice(13+1 pg/ml). Plasma IL6 levels were also not significantly different in the two groups. A microarray for mRNA expression of markers of endothelial activation and adhesion molecules was also performed in the renal tissue. Preliminary data show that TNFalpha mRNA expression levels were not increased by ANG II infusion and IL6 expression was undetectable. ANGII also did not alter E-Selectin,VCAM1, Collagen1a1 and eNOS expression. In conclusion, ANGII infusion did not result in a proinflammatory milieu in both female C57BL/6J and Adipo -/- mice.
Abstract 520: High-density Lipoprotein Inhibits Human M1 Macrophage Polarisation through the Redistribution of Caveolin-1
Authors: Lee Man K , Moore Xiao-Lei , Fu Yi , Al-sharea Annas , Dragoljeic Dragana , Fernandez-Rojo Manuel , Parton Robert , Sviridov Dmitri , Murphy Andrew J , Chin-Dusting Jaye P
Published: July 2021

Macrophages play a critical role in the development and progression of atherosclerosis. Depending on their surrounding milieu, macrophages can adopt a wide range of functional phenotypes; pro-inflammatory (M1) and pro-resolving (M2). HDL has many cardio-protective properties including potent anti-inflammatory effects, largely through the removal of cholesterol from cells. It is currently not known if this extends to influencing human macrophage phenotypes. Thus, we aimed to investigate the effect of HDL on human macrophage polarisation. Human blood monocyte-derived macrophages were induced to either an M1-phenotype by incubation with LPS and IFN-γ or to an M2-phenotype with IL-4. Macrophages were differentiated in the presence or absence of human HDL and their phenotypes were characterised using cell surface markers, reactive oxygen species (ROS) production by flow cytometry, and mRNA expression by real-time PCR. Downstream signalling pathways were also explored. We discovered that HDL inhibited the induction to M1 as evidenced by a decrease in cell surface marker expression; CD192 and CD64. This was accompanied by a decreased expression of M1-associated inflammatory genes TNF-α, IL-6 and MCP-1. However, HDL had no effect on induction to the M2 phenotype. Similarly, methyl-beta-cyclodextrin (MβCD), a non-specific cholesterol acceptor was also able to suppress M1 induction suggesting cholesterol efflux is important in this process. Further we found that HDL decreased membrane caveolin-1 in M1 macrophages and redistributed it intracellularly. The requirement of caveolin-1 was revealed as bone marrow-derived macrophages from Cav-1-/- mice continued to differentiate into M1 despite the addition of HDL. Moreover, we demonstrated a decrease in STAT3 and ERK1/2 phosphorylation in M1 macrophages treated with HDL, suggesting cholesterol efflux inhibits the STAT3s and MAPKs during induction to the M1 phenotype. Finally, we found that HDL also inhibited M1 function; with reduced reactive oxygen species (ROS) production. We provide evidence that HDL reduces macrophage induction to the inflammatory M1 phenotype, but not M2, via cellular redistribution of caveolin-1 and inactivation of STAT3 and ERK1/2 signalling pathway.
Abstract 530: A Cluster of Proteins Implicated in Kidney Disease Are Increased in HDL of Hemodialysis Subjects
Authors: Shao Baohai , de Boer Ian , Mayer Philip S , Zelnick Leila , Afkarian Maryam , Heinecke Jay W , Himmelfarb Jonathan
Published: July 2021

Objectives:
Cardiovascular disease (CVD) is the leading cause of death in uremic subjects on dialysis. However, this large increase cannot be completely explained by the conventional risk factors frequently seen in this population. One important factor could be alterations of high-density lipoprotein (HDL) protein composition that impair its cardioprotective functions. We therefore investigated alterations in the HDL proteome of end-stage renal disease (ESRD) patients undergoing hemodialysis.

Methods and Results:
We first used shotgun proteomics to investigate the protein composition of HDL isolated by ultracentrifugation in 20 control and 40 ESRD subjects. This analysis identified 63 proteins in HDL, which were linked to lipid and lipoprotein metabolism, immune response, acute phase response, complement regulation, inhibitors of proteolysis, vitamin binding and transport, and platelet activation/coagulation. We then used targeted proteomics by isotope-dilution MS/MS with selected reaction monitoring (SRM) to provide precise and accurate relative quantification of 37 proteins in HDL. This quantitative approach revealed that 22 proteins (including SAA1, apoA-IV, apoC-II, apoC-III) were significantly enriched and six proteins (including apoA-I, apoA-II, PON1) were significantly decreased in ESRD patients. Moreover, several of these proteins (SAA1, apoC-III, PON1, etc.) have been associated with atherosclerosis. Strikingly, six proteins implicated in renal disease–AMBP, B2M, CFD, CST3, PTGDS and RBP4–were markedly increased in HDL of uremic subjects.

Conclusions:
Our observations indicate that ESRD markedly remodels the HDL proteome. Moreover, our quantitative analysis of the HDL proteome identified a cluster of six proteins that are dramatically enriched in HDL from patients with ESRD. Our observations support the proposal that the protein cargo of HDL can serve as a marker - and perhaps mediator - of renal disease and serve as novel biomarkers for ESRD status. Therefore, quantifying proteins in HDL might help diagnose and perhaps treat human cardiovascular disease in kidney disease.
Abstract 541: SmgGDS as a Crucial Mediator of the Inhibitory Effects of Statins on Cardiac Hypertrophy and Fibrosis -Novel Mechanism of the Pleiotropic Effects of Statins-
Authors: Kudo Shun , Satoh Kimio , Nogi Masamichi , Suzuki Kota , Sunamura Shinichiro , Omura Junichi , Kikuchi Nobuhiro , Kurosawa Ryo , Satoh Taiju , Minami Tatsuro , Ikeda Shohei , Shimokawa Hiroaki
Published: July 2021

Background:
Statins are widely known to exert beneficial pleiotropic effects mediated by anti-oxidative and anti-inflammatory mechanisms, independent of their LDL-cholesterol lowering effect. However, the detailed molecular mechanisms of the pleiotropic effects of statins remain to be fully elucidated. Here, we demonstrate that small GTP-binding protein GDP dissociation stimulator (SmgGDS) is a crucial mediator of the pleiotropic effects of statins.

Methods and Results:
SmgGDS
+/-
and wild-type (WT) mice were treated with continuous infusion of angiotensin II (AngII) for 2 weeks with and without oral treatment with atorvastatin (10 mg/kg/day) or pravastatin (50 mg/kg/day). At 2 weeks, the extents of AngII-induced cardiac hypertrophy and fibrosis were comparable between the 2 genotypes. However, statins significantly attenuated cardiomyocyte hypertrophy, fibrosis and left ventricular diastolic dysfunction in WT mice, but not in SmgGDS
+/-
mice. Since SmgGDS was highly expressed in cardiac fibroblasts (CFs) in the heart, we then examined the role of SmgGDS in cultured CFs from WT and SmgGDS
+/-
mice. In SmgGDS
+/-
CFs, Rac1 expression, ERK1/2 activity, Rho-kinase activity and inflammatory cytokines secretion in response to AngII were significantly increased as compared with WT CFs. Atorvastatin significantly reduced Rac1 expression and oxidative stress in WT CFs, but not in SmgGDS
+/-
CFs. Furthermore, Bio-plex analysis revealed significant up-regulations of inflammatory cytokines/chemokines and growth factors in SmgGDS
+/-
CFs as compared with WT CFs. Importantly, conditioned medium from SmgGDS
+/-
CFs increased BNP expression in neonatal rat cardiomyocytes to a greater extent than that from WT CFs. Furthermore, atorvastatin significantly increased SmgGDS secretion from mouse and human CFs. Finally, treatment with recombinant SmgGDS significantly reduced Rac1 expression in SmgGDS
+/-
CFs.

Conclusion:
These results indicate that both intracellular and extracellular SmgGDS play a crucial role in the inhibitory effects of statins on cardiac hypertrophy and fibrosis, partly through inhibition of Rac1, Rho-kinase and ERK1/2 pathways, demonstrating the novel mechanism of the pleiotropic effects of statins.
Abstract 552: IL-17C-Producing Aortic Smooth Muscle Cells Play a Pro-Atherogenic Role Via the Recruitment of Inflammatory IL-17A+ T Cells
Authors: Butcher Matthew J , Galkina Elena V
Published: July 2021

Atherosclerosis is characterized by frequent communication between infiltrating leukocytes and vascular cells, through chemokine and cytokine networks. Various IL-17 cytokine family members are detected within atherosclerotic plaques; however the involvement of IL-17B-IL-17E has not been thoroughly examined. Aortic IL-17 cytokines were initially profiled and Il17a, Il17f, and Il17c mRNA expression was found to be elevated in Apoe-/- aortas in comparison to C57Bl6 controls. To determine the cellular sources of aortic IL-17C, aortic IL-17C-producing cells were phenotyped by flow cytometry. Surprisingly, these experiments revealed a major population of IL-17C-producing smooth muscle cells within Apoe-/- versus C57Bl6 aortas. To determine the role of IL-17C in atherosclerosis, we generated Il17c-/-Apoe-/- mice and compared 12 week western diet-fed Apoe-/- and Il17c-/-Apoe-/- mice. Atherosclerotic lesions were reduced by 42±2.6% in WD-fed Il17c-/-Apoe-/- aortas in comparison to Apoe-/- controls. Flow cytometric analyses of Il17c-/-Apoe-/- mice revealed a reduction in the number of aortic leukocytes, macrophages, neutrophils, and T cells, which corresponded with smaller atherosclerotic lesions (40±5%). However, the percentage and number of aortic IL-17A+ TCRγδ T cells and Th17 cells were disproportionately reduced within Il17c-/-Apoe-/- aortas (70±5%). We hypothesized that IL-17C might support the recruitment of Th17 cells to the aorta. To study comparative Th17 migration, Apoe-/- T cells were transferred to western diet-fed Il17c-/-Apoe-/- and Apoe-/- recipients and examined for Th1 and Th17 migration 48 hours post transfer. We found diminished accumulation of Th17, but not Th1 T cells, in the aortas of Il17c-/-Apoe-/- versus Apoe-/- recipients. Thus smooth muscle cell-derived IL-17C plays a pro-atherogenic role by supporting the recruitment of Th17 cells to atherosclerotic lesions.
Abstract 560: Association of Female Gender with Arterial Stiffness in Patients With End-Stage Renal Disease
Authors: Guajardo Isabella , Claire Mills , Ganz Peter , Peralta Carmen A , Dubin Ruth
Published: July 2021

Objectives:
Cardiovascular mortality remains disproportionately high in patients with end-stage renal disease (ESRD), and arterial stiffness is a strong risk factor for death in these patients. Recent studies have shown that women are at increased risk for death in this population. We explored whether arterial stiffness differed by gender in a diverse cohort of patients on hemo- or peritoneal dialysis.

Methods:
Stable patients on hemo- or peritoneal dialysis (N=86) underwent arterial stiffness testing by radial tonometry (SphygmoCor, AtCor Medical). Patients were studied on Tuesday or Wednesday mornings (for patients on hemodialysis, the day after the first dialysis session of the week). Central aortic augmentation index (AIx) was calculated at a standardized heart rate of 75 bpm using a generalized transfer function.[[Unable to Display Character:  ]]

Results:
The mean±SD age was 54±12 years, 30% were women, 36% were African American, and 38% had diabetes. The median(IQR) years on dialysis was 4(1.8, 7), 80% were on hemodialysis, and 20% were on peritoneal dialysis. Overall, mean±SD Aix was 24±11%; mean±SD Aix was 29±9% for women vs. 21±9% for men (p<0.001). Clinical factors associated with both higher Aix (worse arterial stiffness) and female gender (p≤0.2) included older age, diabetes, lower albumin, lower hemoglobin and higher Kt/V. After adjustment for these covariates, female gender remained a significant correlate of worse arterial stiffness (β=8%;95%CI [-14, -1.7];p=0.01). Further adjustment for peripheral systolic blood pressure, height and weight (parameters used to calculate Aix) did not attenuate this effect (β=9%;95%CI [-15, -3.1];p=0.004).

Conclusions:
In this ESRD cohort, female gender was associated with higher Aix (worse arterial stiffness) after adjustment for clinical factors. Further studies are needed to explore whether residual confounding explains this association or if hormonal and metabolic factors could lead to worsened arterial stiffness in women with ESRD.
Abstract 59: Disparity in Protection from Thrombotic Risk Depends on Platelet Thrombin Receptor PAR4
Authors: Tourdot Benjamin , Holinstat Michael
Published: July 2021

Aberrant platelet activation contributes to the mortality associated with coronary heart disease (CHD). Our lab group has recently shown that platelets from black donors were hyper-responsive to activation of the thrombin receptor Protease-Activated Receptor-4 (PAR4) compared to platelets from white donors. The racial difference observed through PAR4 persists in platelets treated ex vivo with dual antiplatelet inhibitors (COX-1 inhibitor Aspirin and P2Y12 receptor inhibitor Clopidogrel), a standard of care for many CHD patients. The racial difference in PAR4 signaling is largely attributable to one PAR4 variant more frequently expressed in blacks than whites, PAR4 T120A. Independent of race, T120 is associated with greater PAR4-mediated platelet reactivity compared to the A120 variant. The role PAR4 variation plays in regulating thrombin-mediated platelet activation in CHD has not been elucidated to date. We hypothesize that patients homozygous for the T120 will have an increase in PAR4-mediated platelet reactivity compared to patients homozygous for A120. To investigate this hypothesis, we studied PAR4 signaling in platelets obtained from healthy donors and patients with CHD on dual antiplatelet therapy. Healthy donors taking either a COX-1 or P2Y12 inhibitor who are homozygous for threonine at amino acid 120, have an increase in platelet reactivity as measured by aggregation, αIIbβ3 activation and granule secretion compared to individuals who are homozygous for alanine at amino acid 120. Interestingly, subjects heterozygous at 120 exhibit platelet reactivity which is in between what is observed for people homozygous for alanine or threonine at this amino acid. This study reinforces the personalized medicine approach to therapeutic intervention and challenges the one size fits all approach which often leaves at risk populations without adequate protection from thrombotic events and stroke.