bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 419: Wall Thickness Influence on Computational Wall Stress of Arteries
Authors: Shang Eric K , Xu Chun , Nathan Derek P , Fairman Ronald M , Gorman Robert C , Gorman Joseph H , Vigmostad Sarah C , Jackson Benjamin M
Published: March 2022

Objective:
Biomechanical analysis can predict mechanical stresses in cardiovascular structures. Prior models often assume uniform arterial wall thickness (WT). We hypothesize that including local WT in finite element analysis (FEA) of the carotid artery will effect wall stress (WS) distribution and maxima.

Methods:
CTAs in patients with carotid occlusive disease (n=5) were analyzed with custom algorithms to segment luminal and adventitial surfaces, providing local WT. A control model was defined with variable wall thickness (VWT) and hyperelastic material properties (MP). Experimental models tested the effects of: (1) uniform wall thickness (UWT), (2) soft and calcified plaque, and (3) linear MP. Commercial FEA software was used to load each model with 80 mmHg pulse pressure and compute Von Mises WS.

Results:
The mean (across n=5 patients) peak WS of the VWT model was 162±29 kPa. Peak WS decreased with the imposition of UWT (112±19 kPa, P=0.01) and with linear MP (105±24 kPa, P=0.01). UWT decreased both the average WS over the entire carotid bifurcation (31±4 kPa, P=0.04) compared to VWT control (42±9 kPa) and the mean normalized nodal variance (a measure of inhomogeneity) of the WS over the bifurcation (0.34±0.03 vs 0.30±0.04, P=0.03).

Conclusion:
Incorporation of local WT can significantly increase peak WS and cause less uniform WS distribution. Aortic wall thickness should be incorporated in FEA of aneurysms to accurately predict rupture risk and in other bioengineering analyses.
Abstract 423: High-Risk Plaque in the Carotid and Femoral Arteries of Men and Women with Peripheral Arterial Disease: Prevalence and Clinical Correlations
Authors: McDermott Mary , Liu Kiang , Carroll Timothy J , Yuan Chun , Xu Dongxiang , Kramer Christopher , Ferrucci Luigi , Guralnik Jack M , Berry Jarett , Polonsky Tammy , Tao Huimin , Tian Lu , Carr James C , Zhao Lihui , Kibbe Melina , Criqui Michael H
Published: March 2022

OBJECTIVES.
It is currently unknown whether high-risk atherosclerotic plaque develops simultaneously in multiple arterial beds or whether it is an isolated phenomenon. We used direct plaque imaging with magnetic resonance imaging (MRI) in participants with lower extremity peripheral arterial disease (PAD) to determine whether individuals with lipid rich necrotic core (LRNC) in the femoral arteries have a higher prevalence of LRNC in the carotid arteries than individuals without LRNC in the femoral arteries.

METHODS AND RESULTS.
Twenty-four participants with PAD underwent direct plaque imaging of both their carotid and superficial femoral arteries. Four participants (16.7%) had LRNC in the femoral artery and 18 (75%) had LRNC in the carotid artery (P<0.001). The prevalence of LRNC in the carotid arteries was 75% among participants with LRNC in the femoral arteries vs. 75% among participants without LRNC in the femoral arteries (P=1.00). Mean ages were 69.0+8.7, 61.9+9.6, and 63.7 +3.8 among participants with LRNC in neither the carotid or femoral arterial beds, in only one of the arterial beds, and in both the arterial beds, respectively, (P=0.27). Among participants with LRNC in neither arterial bed, only one arterial bed, and both arterial beds, mean ABI values were 0.62+0.08, 0.76+ 0.12, and 0.57 + 0.14 (P=0.02), prevalences of cigarette smoking were 20.00%, 56.25%, and 66.67% (P=0.30), and prevalences of males were 80.0%, 93.75%, and 100%, respectively.

CONCLUSION.
Among people with PAD, the prevalence of plaque containing LRNC is higher in the carotid than in the femoral arteries. The prevalence of femoral artery LRNC is similar between participants with vs. without carotid artery LRNC.
Abstract 426: Epidermal Growth Factor Receptor Increases MMP-9 Expression in Carotid Plaques of Symptomatic Patients with Carotid Stenosis
Authors: Rao Velidi H , Kansal Vikash , Sur Swastika , Tyndall Steve H , Agrawal Devendra K
Published: March 2022

Unstable carotid atherosclerotic plaques are characterized by rupture of their fibrous cap, leading to thromboembolism and stroke. The role of MMPs in plaque disruption is not well understood. The control of MMP transcription is complex but recently, EGFR expression has been implicated in the up-regulation of MMP-9. The aim of this study was to determine the MMP-9 mRNA expression as well as other MMPs (MMP-3, -7, and -14/MT1-MMP) expression in carotid plaques as well as in isolated SMCs from asymptomatic and symptomatic patients with carotid stenosis. The MMP-2 and MMP-9 activities were determined by gelatin zymography. The mRNA transcripts of MMP-9, -3, -7, -14 and EGFR were quantified by qPCR. The activity of MMP-9 on gelatin zymograms and the MMP-9 mRNA transcripts were significantly increased in symptomatic carotid plaques and isolated SMCs compared to those from plaques of asymptomatic patients. There was a parallel increase in the mRNA transcripts of MMP-7, -3 and -14. The mRNA transcripts for EGFR were also up-regulated. These results suggest that MMP-9 may be activated by MMP-7 and -3 and thus destabilizes the symptomatic plaques of patients compared to those from asymptomatic plaques and this effect may be mediated by EGFR.
Abstract 431: Impact of Liver-Specific Transgenic Overexpression of Drosophila and Human MTP on Diet-Induced Hepatic Steatosis and Lipoprotein Secretion
Authors: MacArthur Philip S , Fabritius Melissa A , Cheng Dongmei , Hussain M M , Shelness Gregory S
Published: March 2022

Microsomal triglyceride transfer protein (MTP) is essential for the formation of apolipoprotein B (apoB)-containing lipoproteins. Previous results established that invertebrate forms of MTP (e.g.,
Drosophila
) are capable of phospholipid (PL) transfer, whereas vertebrate forms of MTP (e.g., human) engage in both PL and triglyceride (TG) transfer. To determine,
in vivo,
whether the PL and TG transfer activities of MTP play different roles in lipoprotein formation and protection from hepatic steatosis, we created B6D2F1 mice that express human MTP (hMTP) or
Drosophila
MTP (dMTP) under the control of the apoE 5’ proximal promoter and 3’ hepatic control region in the plasmid pLIV11. Immunoblot analysis confirmed that transgenic expression was limited to the liver and that expression of the transgenes did not impact the abundance of endogenous mouse MTP. To assess the ability of MTP to protect mice from hepatic steatosis, dMTP transgenic mice, hMTP transgenic, and wild type (WT) littermates were placed on a high fat (45% from lard) and cholesterol (0.2%) diet for eight weeks. Mice displayed no significant differences in plasma TG or total plasma cholesterol; however, hMTP mice displayed a trend toward reduced liver TG content, indicating that chronic MTP overexpression may protect the liver from hepatic lipid accumulation. Surprisingly, however, a 2-fold increase in liver TG was observed in two separate founder lines of dMTP transgenic mice. H&E staining confirmed an increase in neutral lipid accumulation in mice expressing the dMTP transgene, relative to both WT and hMTP mice. Quantitative PCR analysis did not reveal changes in expression of genes responsible for hepatic lipogenesis or lipid oxidation. These data complement earlier studies using adenovirus-mediated expression, indicating that acute hMTP and dMTP expression in liver-specific MTP knockout mice can reverse hepatic steatosis caused by MTP deficiency. In contrast, chronic hepatic overexpression of dMTP in a wild type background appears to interfere with endogenous lipoprotein formation and secretion. Understanding the basis for this phenotype may provide insights into how the specific lipid transfer and other activities of MTP contribute to apoB lipoprotein formation.
Abstract 437: Simvastatin/Ezetimibe 20/10 mg Versus Atorvastatin 20 mg in Patients with Type 2 Diabetes: Results of Single-Center, Randomized Controlled Trial
Authors: Lee Ju-Hee , Park Kyung Woo , Kim Hyung-Kwan , Cho Hyun-Jai , Choi Eue-Keun , Kang Hyun-Jae , Kim Yong-Jin , Koo Bon-Kwon , Oh Seil , Kim Hyo-Soo , Sohn Dae-Won , Oh Byung-Hee , Park Young-Bae
Published: March 2022

Introduction
Though the efficacy on traditional lipid parameters of simvastatin/ezetimibe and atorvastatin has been studied extensively, ApoB/ApoA1 ratio which has better predictive value for cardiovascular event has not been compared as a primary endpoint in these two treatment groups before, especially in diabetes.

Methods
This study was a single center, open label, randomized phase 4 study to compare the efficacy and safety of simvastatin/ezetimibe 20/10mg versus atorvastatin 20mg once daily in subjects with type 2 diabetes. One hundred thirty-two well controlled diabetic patients (HbA1C <8.5%) with high LDL cholesterol levels (≥ 100mg/dL) were randomized to two groups. The primary endpoint was the difference in percent change of ApoB/ApoA1 ratio at 12 weeks and secondary endpoints were changes in conventional lipid profiles and apolipoproteins.

Results
In total, 132 patients (66 for each group; 60 males; mean age, 64.6±7.6 years) were enrolled and 125 patients were included in the full analysis set (62 for simvastatin/ezetimibe; 63 for atorvastatin). Baseline characteristics including lipid profiles were comparable between two groups. After 12 weeks of treatment ApoB/ApoA1 ratio was significantly reduced in both groups, however the differences of % changes between two groups were statistically not significant (-38.1±17.9% vs. -34.2±16.2%,
p
=0.279) and the changes in conventional lipid profiles including total cholesterol, LDL cholesterol and triglyceride also showed no significant difference. However, HDL cholesterol and ApoA1 levels increased significantly in the simvastatin/ezetimibe group, whereas decreased in the atorvastatin group (HDL cholesterol 4.2±12.7% vs. -0.2±14.8% and ApoA1 2.8±10.0% vs. -1.8±9.8%,
p
=0.041 and 0.002). In per-protocol analysis, improvement of ApoB/ApoA1 was significantly greater in the simvastatin/ezetimibe group (-42.3±11.9% vs. -36.5±14.1%,
p
=0.031). The changes of HOMA index and hsCRP were comparable between two groups in full analysis set. But in per-protocol analysis, HOMA index and hsCRP decreased in the simvastatin/ezetimibe group whereas increased in the atorvastatin group although there was no statistic significance. The frequency of adverse reaction was comparable between two groups.

Conclusion
Based on the results of this randomized controlled trial, simvastatin/ezetimibe 20/10mg might be preferable to atorvastatin 20mg for management of dyslipidemia in the patients with type 2 diabetes.
Abstract 439: Dietary Sphingomyelin Lowers Hepatic Lipids by Inhibiting Intestinal Cholesterol Absorption in High-Fat-Fed Mice
Authors: Chung Rosanna W S , Kamili Alvin , Tandy Sally , Rye Kerry-Anne , Cohn Jeffrey S
Published: March 2022

Background:
Accumulating evidence suggests that tissue sphingolipids such as sphingomyelin (SM) and ceramide contribute to the onset and development of atherosclerosis. Dietary sphingolipids may not however be pathogenic and we have shown that a phospholipid-rich extract from dairy milk that contains SM has potent liver lipid-lowering properties.

Objective:
To investigate the effect of pure dietary egg SM on hepatic lipid metabolism in high-fat fed mice.

Methods:
Four groups of male C57BL/6 mice were given a high-fat diet containing 21% w/w butterfat and 0.15% w/w cholesterol (HF) or the HF diet supplemented with egg SM (0.3, 0.6 or 1.2% w/w). After 4 weeks, livers were analysed for lipid and mRNA. Intestinal cholesterol absorption was measured with [14C]cholesterol and [3H]sitostanol in mice fed 0.6% w/w SM for three weeks.

Results:
All animals weighed similarly. Liver weight was significantly lower in 1.2%-fed mice (HF vs SM: 1.05 ± 0.03 v 0.86 ± 0.03 g, P < 0.05 by ANOVA). Total liver lipid (mg/organ) was significantly less in 0.6%SM- and 1.2%SM-fed mice (-33% and -40% vs HF resp., P < 0.001). This was due to significantly reduced levels of both triglyceride and cholesterol. Mice given 0.6%SM for 3 weeks were found to have significantly increased faecal lipid and cholesterol output and reduced liver accumulation of gastrically-administered [14C]cholesterol. Intestinal cholesterol absorption was significantly reduced by 0.6%SM supplementation (HF vs SM: 52.4 ± 2.6 vs 36.2 ± 3.5%, P < 0.01).

Conclusion:
Dietary egg sphingomyelin has pronounced lipid-lowering properties, suggesting it may have atheroprotective rather than atherogenic properties.
Abstract 442: Metformin Mitigates the Apoptosis Pathway in Ischemic Myocardium
Authors: Elmadhun Nassrene Y
Published: March 2022

OBJECTIVE:
Epidemiologic data has shown that metformin confers a survival advantage in patients with cardiovascular disease. Although the underlying cardioprotective mechanism is unclear, it appears to be independent of metformin’s insulin-sensitizing effect. The purpose of this study is to evaluate the effect of metformin on the apoptosis pathway in the ischemic cardiac tissue in a swine model of metabolic syndrome.

METHODS:
Ossabaw miniswine were fed either a regular diet (OC, n=8), or a high-cholesterol diet (OHC, n=9) or a high-cholesterol diet supplemented with metformin (OHCM n=8). After three weeks, all animals underwent placement of an ameroid constrictor to the left circumflex coronary artery to induce chronic ischemia. Seven weeks after ameroid placement, animals underwent cardiac harvest.

RESULTS:
Western blot analysis of protein expression in the ischemic territory is reported as fold change ± standard error of the mean (FC ± SEM) with respect to OC. See table. FoxO3a is a pro-apoptotic protein that is inactivated with phosphorylation. Caspase 3 is a pro-apoptotic protein that is activated only when cleaved. P38 and ERK1/2 are activated when phosphorylated and are involved in the apoptosis/survival pathway.

CONCLUSIONS:
Meformin selectively alters the apoptosis pathway by inhibiting FoxO3 and decreasing the active form of caspase 3, cleaved caspase 3. Metformin also up-regulates mitogen-activated kinase proteins p38 and ERK1/2, which are considered cardioprotective during ischemic preconditioning. Perhaps the altered activation of the apoptosis pathway in ischemic myocardium is one mechanism by which metformin is cardioprotective.
Abstract 443: Inhibition of TGFβ Attenuates Angiotensin II Effects to Increase Vascular Biglycan and Atherosclerosis
Authors: Tang Tao , Wilson Patricia G , Thompson Joel C , Nelson Christina , Hamilton Meghan , Tannock Lisa
Published: March 2022

Background:
Proteoglycans, a major component of extracellular matrix in the vascular wall, play a critical role in the development of atherosclerosis due to their ability to bind and retain atherogenic lipoproteins. Of all the vascular proteoglycans, biglycan has been shown to be the one most closely associated with apolipoprotein B. Our previous studies showed that AngII increases vascular biglycan content and predisposes to diet-induced atherosclerosis in
Ldlr
null mice. Moreover, we demonstrated that TGFβ is required for AngII stimulation of vascular biglycan content
in vitro
. In this study, we determined whether TGFβ is required for AngII-induced biglycan accumulation and atherosclerosis predisposition
in vivo
.

Methods and Results:
Ldlr
null mice fed a normal chow diet were infused with AngII or saline for 28 days. TGFβ1 levels in AngII-infused mice peaked at 24h then returned to baseline levels by day 5. A single injection of TGFβ neutralizing antibody 1D11 (2mg/kg) not only prevented the induction of TGFβ1 levels by AngII, but also attenuated AngII-induced accumulation of aortic biglycan content, whereas there was no attenuation of AngII-induced increase in TGFβ or biglycan by a control antibody. To study atherosclerosis, the experiment was repeated but after 28 days of AngII or saline infusions the pumps were removed and the mice were fed Western diet for 6 weeks. TGFβ neutralizing antibody injection had no effect on systolic blood pressure or plasma lipids (triglyceride and cholesterol). However, AngII-infused mice that received TGFβ neutralizing antibody injection showed reduced atherosclerotic lesion area (aorta root, en face and the brachiocephalic artery) compared to AngII-infused mice that received a control antibody injection. Therefore, our study showed that TGFβ inhibition attenuates AngII stimulation of vascular biglycan content
in vivo
and attenuates AngII-accelerated atherosclerosis.
Abstract 448: Tetrahydrobiopterin Rescues Mitochondrial Dysfunction by Regulating Mitochondrial Biogenesis and Oxidative Phosphorylation
Authors: Han Jin , Kim Hyoung Kyu , Kim Nari , Ko Kyung Soo , Lee Byoung Doo
Published: March 2022

The multifunctional cofactor tetrahydrobiopterin (BH
4
) has antioxidant effects, and low levels of BH
4
expression are implicated in various cardiovascular diseases involving mitochondrial dysfunction[1]. We investigated the role of BH
4
in the regulation of cardiac mitochondrial function using sepiapterin reductase knockout (
Spr–
) mice as a model of BH
4
deficiency[2]. Based on systematic data-integrative analysis of the mitochondrial proteome, we found that BH
4
deficiency resulted in significant remodeling of oxidative phosphorylation, which impaired mitochondrial membrane potential, oxygen consumption, and ATP production, and induced oxidative stress in mitochondria DNA, leading to cardiac contractile dysfunction. Moreover, the expression of major regulators of mitochondrial biogenesis and respiration, such as PGC1-α and
mt
TFA, was reduced by BH
4
deficiency. Finally, we confirmed that BH
4
supplementation could rescue cardiac mitochondrial biogenesis and oxidative phosphorylation, which demonstrates the therapeutic potential of BH
4
for the treatment of cardiovascular diseases involving mitochondrial dysfunction.

Reference

[1] Kim, H.K., Ha, S.H., and Han, J. (2010). Potential therapeutic applications of tetrahydrobiopterin: from inherited hyperphenylalaninemia to mitochondrial diseases. Ann N Y Acad Sci
1201
, 177-182.

[2] Yang, S., Lee, Y.J., Kim, J.M., Park, S., Peris, J., Laipis, P., Park, Y.S., Chung, J.H., and Oh, S.P. (2006). A murine model for human sepiapterin-reductase deficiency. Am J Hum Genet
78
, 575-587.
Abstract 452: Cyclic Strain Induces Osteopontin Expression in Vascular Smooth Muscle via a Hydrogen Peroxide--Dependent Mechanism
Authors: Caesar Christa , Lyle Alicia N , Taylor W Robert
Published: March 2022

Introduction:
Vascular inflammation, a hallmark of hypertension, is associated with increased cyclic strain on the vessel wall. Reactive oxygen species (ROS), such as superoxide and hydrogen peroxide, are known to play an important role in the progression of cardiovascular inflammation. However, the underlying molecular mechanisms of ROS modulation of vascular inflammation through increased cyclic strain remain largely unknown.

Objective:
In this study, we sought to determine whether cyclical strain induces an increase in osteopontin (OPN), a pro-inflammatory protein, in a hydrogen peroxide dependent manner using an in vitro cell system.

Methods:
Rat aortic smooth muscle cells were seeded on collagen-I coated, flexible bottomed six-well plates and cyclically stretched at 10% elongation in a biaxial stretch bioreactor. Following stretch, cells were harvested and protein, mRNA, and hydrogen peroxide levels were measured via Western Blotting, quantitative real-time PCR, and Amplex Red assay, respectively.

Results:
Preliminary results indicate that cyclic strain promotes a 16% ± 6 increase in hydrogen peroxide levels compared to non-stretched cells at 8 hours. In addition, elevated expression of OPN protein was observed with 24 hours of stretch (195% ± 16). Increased OPN mRNA was also observed with 12 hours of stretch (39% ± 15). Finally, OPN expression was blunted when cells were simultaneously stretched for 24 hours with 200U/mL PEG-Catalase (a H2O2 scavenger) suggesting that the stretch induced increase in OPN expression is mediated by hydrogen peroxide (49%±14 decrease in cells stretched with PEG-Catalase versus non- PEG-Catalase treated but stretched cells)

Conclusion:
These data support that increased cyclical mechanical stretch, as experienced by the vascular wall under hypertensive conditions, increases osteopontin expression, via a hydrogen peroxide-mediated pathway.
Abstract 457: Constitutive Expression of Fibroblast Growth Factor 1 in Endothelial Cells Alters Adipose Tissue Development
Authors: Keeley Tyler , Kirov Aleksandr , Prudovsky Igor , Small Deena J
Published: March 2022

The development of healthy adipose tissue is dependent on both adipogenesis and angiogenesis. Resident endothelial and adipogenic cells within the growing depot produce angiogenic factors that stimulate angiogenesis in order to provide vascular support for the metabolic demands of the tissue. Since endothelial cells also produce adipokines that promote fat cell maturation, depots with higher vascular density tend to have more numerous, but smaller adipocytes than those with fewer vessels. Although the risk for the development of insulin resistance syndromes and cardiovascular disease is significantly higher in individuals with hypertrophic obesity - a condition characterized by the presence of enlarged and poorly vascularized adipocytes - the molecular mechanisms governing communication between adipogenic and endothelial cell populations within an adipose depot are poorly characterized. Acidic Fibroblast Growth Factor (FGF1) is an angiokine secreted by both adipocytes and endothelial cells and has also been reported to stimulate the production of adipocytes. To test the role of FGF1 in adipose tissue development, we analyzed weight gain, blood glucose, adipogenic hormone levels and fat tissue morphology in control and genetically-manipulated mice constitutively expressing an endothelial-specific FGF1 transgene. For these studies, mice were fed a high-fat, “Western” diet over the course of fourteen weeks. We found that a subpopulation of the FGF1-transgenic animals displayed significant differences in weight gain, fat cell size and hormone levels in comparison to control animals. Interestingly, the fat depots from the leanest animals also appeared to contain more “brown fat” than those taken from the more obese individuals. These results suggest that FGF1 may be a critical mediator in maintaining the balance between adipocyte number, size and vessel density within adipose depots. Since obesity is a health crisis in the United States, these studies may contribute to the development of therapeutic agents used to treat pathological conditions associated with abnormal weight gain.
Abstract 463: Dietary Nitrite Plays a Role in Wound Healing in Diabetic Mice
Authors: Madigan Michael , Gladwin Mark , Tarpey Margaret , Zuckerbraun Brian , Tzeng Edith
Published: March 2022

Intro:
Diabetic wounds heal poorly leading to major morbidity, mortality, and financial cost. Nitric oxide (NO), produced in wounds by NO synthases (NOSs), is essential for normal wound healing and is reduced in diabetic wounds, contributing to poor healing. It is known that NO can also be regenerated from nitrite through xanthine oxidoreductase (XOR). Thus, we hypothesize that dietary nitrite may be an important source of NO in diabetic wound healing.

Methods:
Diabetic Db/Db mice were fed normal chow, nitrite depleted chow, or nitrite depleted chow with 1.5g/L nitrite supplemented water starting 1 week prior to wounding. Excisional wounds were created on each mouse and wound areas were measured every other day. Skin and wound samples from a separate group of Db/Db mice were obtained prior to and 5 days after wounding. Tissues were used for Western blots and XOR activity assay. Plasma was used for nitrite measurements.

Results:
Nitrite depletion lowered plasma nitrite by 50-60% vs mice on chow or on nitrite water. XOR was highly expressed in the skin, less so at the wound edge, and minimally in granulation tissue by Western. This was confirmed by XOR enzyme assay showing 5 fold more activity in the skin compared to the wound tissues. Wound healing was delayed in nitrite depleted mice as compared to the chow fed mice (Figure). Nitrite supplementation showed a trend in improved healing rates vs nitrite depleted mice.

Conclusion:
Diabetic skin and wound edge expresses XOR. Nitrite does contribute to wound healing in Db/Db mice. Dietary nitrite increased plasma nitrite levels and increased the wound healing rate. This suggests a potential therapeutic role for dietary nitrite in diabetic wound care.
Abstract 465: ETC-1002, a Novel Dicarboxylic Fatty Acid Analog, Inhibits Inflammatory Response in Primary Human Monocyte-Derived Macrophages as Well as in Adipose Tissue of Insulin-Resistant Mice via AMPK-Dependent Mechanisms
Authors: Filippov Sergey , Lister Richard J , Pinkosky Stephen L , Bradshaw Cheryl D , Hanselman Jeffrey C , Hurley Tim R , Spahr Mark A , Srivastava Rai Ajit , Newton Roger S
Published: March 2022

ETC-1002, a small molecule regulator of imbalances in lipid and carbohydrate metabolism, is an investigational drug currently in Phase 2 development to treat dyslipidemia and other cardiometabolic risk factors. In hyperlipidemic LDL receptor-deficient mice, robust antiatherosclerotic activities of ETC-1002 coincided with reduced levels of inflammatory markers in mouse atheroma. To further investigate anti-inflammatory properties of ETC-1002, human monocyte-derived macrophages (MDMs) differentiated in autologous serum were stimulated with 100 ng/ml of LPS in the absence or presence of the 10 μM and 30 μM of ETC-1002. TLR4-mediated activation of downstream kinases as well as the production of pro-inflammatory mediators were assessed with phosphokinase and protein arrays. Lower levels of JNK, cJUN, p38 and ERK phosphorylation in cells treated with ETC-1002 were consistent with reduced production of pro-inflammatory cytokines (TNF-α, IL-6, IL-8 and MIP1α) and chemokines (CXCL10, CXCL1, CCL2 and CCL5). ETC-1002 at 30 mg/kg dose largely diminished thioglycolate-induced homing of neutrophils and macrophages into the mouse peritoneal cavity, supporting the inhibitory effect of ETC-1002 on leukocyte chemotactic and inflammatory activity. Furthermore, in a mouse model of diet-induced obesity and insulin resistance, epididymal fat pad mass and IL-6 release by inflamed adipose tissue were significantly attenuated (by 32% and 80% respectively) in animals treated with ETC-1002. Importantly, enhanced levels of AMPK phosphorylation, changes in intracellular energy charge coupled with reduced basal rates of sterol and fatty acid synthesis by human MDMs strongly supported AMPK-dependent anti-inflammatory effects of ETC-1002. Thus, our data suggest that ETC-1002, via stimulation of AMPK activity, may provide additional clinical benefits for patients with metabolic syndrome by reducing systemic inflammation and other cardiometabolic abnormalities linked to vascular disease.
Abstract 476: TLR4 Activation in Microvascular Endothelial Cells Triggers a Robust Inflammatory Response and Crosstalk with Mononuclear Cells
Authors: Lu Zhongyang , Li Yanchun , Jin Junfei , Zhang Xiaoming , Lopes-Virella Maria F , Huang Yan
Published: March 2022

Although coronary artery disease is a macrovascular disease, microvessels are developed in atherosclerotic plaques from the vasa vasorum and contribute to inflammation and intraplaque hemorrhage. As the major types of cells in vascular system, both macrovascular endothelial cells (MAC ECs), which cover the luminal surface of atherosclerotic plaques, and microvascular endothelial cells (MIC ECs), which line small size vessels and capillaries inside lesions, are important players in atherosclerosis. In recent years, a large number of studies have provided evidence that toll-like receptor (TLR)4, an innate immune receptor, plays an essential role in atherosclerosis. However, the inflammatory responses by MIC and MAC ECs to TLR4 activation have not been fully characterized and compared. In this study, ELISA, real-time PCR and gene expression arrays showed that TLR4 activation by lipopolysaccharide (LPS) stimulated a much more robust expression of pro-inflammatory genes including cytokines, chemokines, growth factors and adhesion molecules in human dermal MIC ECs than human aortic MAC ECs. For example, TLR4 activation increased interleukin (IL)-6 secretion from MIC and MAC ECs by 185- and 17-fold of that from control cells (150,916 vs 814 pg/ml for MIC ECs and 501 vs 29 pg/ml for MAC ECs), respectively. Also, TLR4 activation increased chemokine CXCL10 secretion from MIC and MAC ECs by 5,405- and 12-folds of that from control cells, respectively. Furthermore, TLR4 activation in MIC ECs triggered a crosstalk with U937 mononuclear cells through MIC EC-released IL-6 to upregulate matrix metalloproteinase (MMP)-1 expression in U937 cells. In contrast, TLR4 activation in MAC ECs did not trigger the crosstalk with mononuclear cells. To explore the mechanisms underlying the different degrees of responsiveness to TLR4 activation by MIC and MAC ECs, we found that TLR4 and CD14 expression in MIC ECs was significantly higher than that in MAC ECs. Finally, we found that the nuclear factor kappa B (NFκB) activity in response to TLR4 activation increased by10- and 3-fold, respectively, in MIC and MAC ECs. Taken together, this study showed that TLR4 activation in MIC ECs triggered a more robust inflammatory response than that in MAC ECs and facilitated MIC ECs to crosstalk with mononuclear cells. Given the major contribution of cytokines and MMPs to atherosclerotic plaque destabilization, our study indicates that MIC ECs may play a major role in plaque vulnerability through a TLR4-dependent mechanism.
Abstract 48: Inhibition of Interleukin 27 Attenuates Atherosclerosis
Authors: Ramos Ornélia H , de Jager Saskia C , van Santbrink Peter J , ter Borg Mariette , van der Sluis Ronald , Kuiper Johan
Published: March 2022

Interleukin (IL) 27 is an interesting cytokine as it has both pro- and anti-inflammatory properties. IL-27 binds to cytokine receptor WSX-1 to activate cells. In this experiment we determined the effect of IL-27 on atherosclerosis and blocked the function of IL-27
in vivo.

This was achieved by injecting a lentivirus encoding the WSX-1 receptor lacking the intracellular domain with a human IgG2 tag (sWSX-1) in female LDL receptor deficient mice. The sWSX-1 protein was continuously released into the blood for at least 10 weeks and the responders (n=11) had a high serum hIgG level in comparison to the mice which received an empty virus (n=10), whereas the non-responders (n=9) did not show an increased hIgG level. After 10 weeks of atherogenic diet, mice were sacrificed .

In vitro
sWSX-1 was able to inhibit the IL-27-mediated IL-2 secretion in spleen cells. The
in vivo
blockade of IL-27 via the soluble receptor sWSX-1 resulted in a 31 % reduction in aortic lesion size in the responder group (2,7 x 10
5
μm
2
± 2,2 x 10
4
) compared to the control group (3,7 x 10
5
μm
2
± 3,5 x 10
4
). No change was measured in the non-responders (3,4 x 10
5
μm
2
± 2,5 x 10
4
) The lesion size negatively correlated with the concentration of the soluble receptor in the serum. sWSX-1 production did not affect the collagen content of the lesion, while there was a trend towards an increased macrophage content. FACS analysis showed that blockade of IL-27 increased the circulating T helper cell and cytotoxic T cell population by 2-fold. Surprisingly, no changes were detected in the Th1 and Th17 populations. There was a 5-fold increase in the percentage circulating regulatory T cells in the presence of sWSX-1. Furthermore, CCR2 expression on circulating Ly6C
low
monocytes was reduced by 44 %. The proliferation of spleen cells showed no change between the control and mice with overexpression of sWSX-1.

Taken together these results indicate that blockade of interleukin 27 reduces lesion formation by inducing a less inflammatory immune status.
Abstract 485: Potential Neuronal-Specific Influence on AT1a Receptor--Mediated Hyperplasia in the Ascending Aorta
Authors: Poduri Aruna , Rateri Debra L , Howatt Deborah A , Moorleghen Jessica J , Cassis Lisa A , Daugherty Alan
Published: March 2022

Introduction and Objective:
Angiotensin II (AngII) infusion stimulates AT1a receptors to promote medial thickening throughout the aorta. Medial thickening is due to hyperplasia in the ascending aorta and hypertrophy in the rest of the aorta. The cell type(s) expressing AT1a receptors that promote medial thickening have not been defined. We have previously demonstrated the surprising result that deletion of AT1a receptors in either endothelial or smooth muscle cells had no effect on medial thickening. Therefore, the purpose of this study was to determine the role of AT1a receptors in neuronal cells on AngII-induced aortic medial thickening.

Methods and Results:
We generated male neuronal cell-specific AT1aR deficient mice by breeding AT1aR floxed mice with transgenic mice expressing Cre driven by enolase 2 promoter (AT1aR x Eno2 Cre +/0). Male non-transgenic littermates (AT1aR x Eno2 Cre 0/0) were used to compare AT1aR x Eno2 Cre +/0 mice. Mice were fed a normal laboratory diet and implanted with mini-osmotic pumps to infuse either saline or AngII (1,000 ng/kg/min) for 28 days. The expression of Cre had no effect on body weight or blood pressure. At termination, mice were perfusion-fixed at physiologic pressure and ascending and descending thoracic regions were sectioned serially. Aortic sections were stained with H&E and propodium iodide. The medial thickness (>5 per region) and nuclei counts (>2 per region) of each aortic tissue section was determined by image analysis. Cre expression under the control of Eno2 led to a significant reduction in medial thickness and number of nuclei in the ascending aorta (p<0.05). Conversely, there was no effect on medial thickening in the rest of the aorta. To determine specificity of Cre expression under control of the Eno2 promoter, Eno2 Cre+/0 mice were bred to ROSA26 mice which have repressed beta-galactosidase activity. In addition to the expected expression in neuronal cells, positive staining was also noted in the adventitia adjacent to the region of attenuated medial thickening.

Conclusion:
AngII-induced ascending aortic hyperplasia was attenuated in mice in which AT1a receptors were deleted using Cre driven by the Eno2 promoter. However, the non-neuronal expression of Eno2 may be the basis for this effect.
Abstract 507: Effects of Arteriole Tortuosity on Platelet Activation and Collision in Thrombosis
Authors: Chesnutt Jennifer K , Han Hai-Chao
Published: March 2022

Introduction:
Thrombosis and artery tortuosity are major contributors to cardiovascular disease. A fundamental gap exists in understanding how tortuosity regulates thrombosis through modification of blood flow. Accordingly, the objective of this research was to elucidate the physical mechanisms of artery tortuosity in thrombus formation.

Hypothesis:
An increase in arteriole tortuosity promotes platelet adhesion to the convex surface of the endothelium by increasing platelet collisions and shear-induced platelet activation.

Methods:
Platelet-mediated thrombosis was computationally simulated in 100-μm diameter vessels with sinusoidal shapes. The model used a discrete element algorithm for transport, collision, shear-induced activation, and receptor-ligand adhesion and aggregation of individual platelets. For a given simulation, platelets were set to be either (1) unactivated, (2) pre-activated, or (3) activated by shear stress. Tortuosity values (ratio of amplitude to wavelength) were set from 0 to 0.23, average velocity was 0.1 cm s
-1
, and Reynolds number was 0.1.

Results:
For cases with platelets subject to shear-induced activation, increased tortuosity increased platelet activation, and platelets adhered to the convex endothelial wall, where shear stress was highest, while negligible numbers of platelets contacted the concave wall. For cases in which all platelets were previously activated, however, considerable platelet adhesion occurred on the concave wall. For cases with unactivated platelets in tortuous arterioles, as tortuosity increased, platelet contacts with the concave wall decreased until reaching the same value as the convex wall. Additionally, for cases with unactivated platelets, collisions between platelets was highest for the straight vessel, and collisions with walls throughout the entire arteriole were relatively negligible for straight and highly tortuous arterioles, but much larger for arterioles with intermediate tortuosity.

Conclusions:
Shear-induced activation due to tortuosity generates platelet adhesion to the convex wall of arterioles. Increased tortuosity increases platelet activation, though the relationship between tortuosity and platelet collisions is less evident.
Abstract 517: Diabetic Status Is More Predictive of Necrotic Core Burden in Acute Coronary Syndrome Patients Versus Stable Patients Undergoing Percutaneous Coronary Intervention as Quantified by Intravascular Ultrasound Virtual Histology
Authors: Nathan Sandeep , Jagadeesan Vikrant , Gajjar Mark , Lee Linda
Published: March 2022

Background:
ACS plaque is thought to differ from non-ACS plaque by manifesting more inflammation / cellular debris and lower plaque volume. We used intravascular ultrasound-Virtual Histology (IVUS-VH) to validate reported plaque compositional differences pre- PCI in ACS vs. non-ACS patients (pts).

Methods:
2,579 IVUS-VH frames were acquired in 36 PCI pts (ACS: n = 15, 6 diabetic (DM); non-ACS: n = 21, 9 DM). Automated IVUS-VH pullbacks were performed using Volcano EagleEye Platinum 20 MHz probes with VH frames traced and blindly adjudicated. VH data was divided into Fibrous (FI), Fibro-Fatty (FF), Necrotic Core (NC), and Dense Calcium (DC). Statistical analyses were performed in groups stratified by ACS and DM status.

Results:
ACS pts overall had slightly higher plaque area/frame (Fig 1) with lower proportional NC (16% vs. 21%, p <0.001) and absolute NC amount (1.6 vs. 1.9 mm
2
, p<0.0001) than non-ACS pts. NC/DC ratio, a known VH correlate of lesion instability, was higher however (3.2 vs. 1.9, P<0.0001) in ACS pts. DM status conferred opposite NC content signals when pts were divided by ACS status, with DM/ACS plaque having the greatest NC content of all. (Fig.1)

Conclusions:
ACS plaque does not uniformly contain greater NC but does evidence greater NC/DC ratio suggesting increased overall plaque vulnerability. DM status is more predictive of absolute NC burden in ACS patients than non-ACS patients. This raises the question of whether the impact of DM on NC content eclipses the impact of ACS and merits further study.
Abstract 522: Effect of Allopurinol on Endothelial Dysfunction: A Meta-analysis of Randomized Controlled Trials
Authors: Dahal Khagendra , Pandey Kaushal , Patel Mitkumar , Johnstone Michael , Garcia Lawrence A
Published: March 2022

Introduction:
Endothelial dysfunction is a precursor of atherosclerosis, and has been found to be present in patients with diabetes, hypertension, hypercholesterolemia, smoking habit, obstructive sleep apnea, hyperuricemia and chronic kidney disease. Measurement of endothelial function by either flow-mediated dilatation (FMD) or forearm venous-occlusion plethysmography (FVOP) of brachial artery has been previously validated. Allopurinol is a xanthine oxidase inhibitor that has been shown to correct endothelial dysfunction in coronary artery disease, congestive heart failure and hypertension by different mechanisms. We conducted a meta-analysis of all available randomized controlled trials (RCTs) that studied the effect of allopurinol in endothelial dysfunction as measured by FMD or FVOP of brachial artery.

Methods:
We performed literature search with predefined criteria with "allopurinol" and "endothelial function" or "endothelial dysfunction" in PubMed, Cochrane Database, EMBASE and proceedings from various society meetings through December 15, 2011. Only the RCTs that compared the effect of allopurinol with placebo on endothelial dysfunction in patients with proven risk factors for endothelial dysfunction were included in the analysis. Random Effects Model was used for analysis of the data using software MetaAnalyst Beta 3.13 (Tufts, Boston, USA).

Results:
A total of 10 RCTs with 519 patients were included in the analysis.When compared to placebo, allopurinol improved the endothelial dysfunction as measured by FMD or FVOP (pooled mean difference 2.335, 95% CI = 1.46 - 3.21, p <0.05, I2 = 98%). The heterogeneity of data was due to different disease conditions, dose and duration of allopurinol and the methods of measurement of endothelial dysfunction.

Conclusion:
We conclude that allopurinol improves endothelial dysfunction in patients with various cardiovascular risk factors. Allopurinol may have a role in preventing atherosclerosis in patients with various cardiovascular risk factors for endothelial dysfunction.
Abstract 540: Plasma Cell Infiltration and Mucoid Degeneration in the Media of Ascending Aorta in Patients with Coronary Artery Disease
Authors: Nagareda Tomofumi , Hirata Nobuaki , Ohsawa Masahiko , Kosakai Yoshio
Published: March 2022

Introduction
Atherosclerosis results in inflammatory changes in the aortic intima, but little is known regarding medial changes. The inflammatory diseases of the aorta and large artery display characteristic features of inflammatory cell infiltration in the media, and they are usually diagnosed by medial changes.

Aims
Atherosclerosis of the ascending aorta coexists with coronary artery disease. The aim of this study was to investigate the atherosclerotic changes in 44 biopsy specimens of media of the ascending aorta associated with coronary artery disease.

Methods
We compared plasma cells, and matrix metalloproteinase (MMP)-2-, -9- and -12-positive cells immunohistochemically, and we also compared mucoid degeneration and fibrosis determined by special staining using a point-counting method, for groups with a variable number of coronary stenotic (≥75 %) lesions.

Results
and Conclusion To investigate the appearance of plasma cells, smooth muscle cells, and mucoid degeneration in the media of the atherosclerotic aorta, we examined three cadavers with known atherosclerosis. CD38-positive plasma cells are pleomorphous in the aortic media. Desmin-positive smooth muscle cells appeared fibrous or solitary spindle cells. Mucoid degeneration was shown in all three cadaver cases. In patients with one to three coronary stenotic lesions, plasma cells and mucoid degeneration were low in the aortic media. With four to five lesions, both plasma cells and mucoid degeneration increased significantly compared with those in the group with one to three lesions, and MMP-12-positive cells significantly decreased. In patients with six to nine lesions, the number of plasma cells was significantly lower than in patients with four or five lesions, whereas mucoid degeneration significantly increased. There was no change in fibrosis. In conclusion, both the infiltration of short lived plasma cells and the exudation of proteoglycan are known to be regarded as characteristic features in the autoimmune disease. The delayed stage in our study is suggested to be caused by autoimmune reaction. If the cause of the delayed medial atherosclerosis stage is found, then more effective therapies to suppress its development can be introduced.
Abstract 543: The Janus-Faced Transcription Factor Sister-of-Mammalian Grainyhead: 2 Isoforms with Opposing Effects in Endothelial Cells and in Vivo
Authors: Mlynek Arne , Lukosz Margarete , Köhrer Karl , Deenen René , Jakob Sascha , Güttler Christopher , Graf Martin , Winkler Christoph , Haendeler Judith , Altschmied Joachim
Published: March 2022

Maintenance of apoptosis protection and migratory capacity are absolutely required for proper endothelial function. Several pathways are described, which are necessary for these processes including activation of the protein kinase B (Akt) and the endothelial nitric oxide synthase (eNOS). Recently, we identified a new player, the transcription factor sister-of-mammalian grainyhead (SOM)/Grainyhead-like 3, which demonstrated anti-apoptotic and pro-migratory properties in endothelial cells (EC).

In humans, three isoforms of SOM are expressed, two of which, SOM1 and SOM3, are derived from an alternatively spliced transcript. SOM3 lacks exon 2, described to contain the putative transactivation domain, and thus, has been suggested to be a repressor. SOM1 and SOM3 are co-expressed in numerous tissues, however, their exact cellular functions have not been investigated so far.

Here we demonstrate that SOM1 and SOM3 are expressed in primary human EC. In contrast to published data, which were based on a yeast two hybrid analysis with isolated domains, we could show that both full length proteins are transcriptional activators. Moreover, they have opposing functions in EC and in a whole animal model. Overexpression of SOM1 inhibited apoptosis and induced migration of EC. The latter effect could be due to the observed activation of eNOS and its upstream regulator Akt. On the contrary, SOM3 has no effect on apoptosis and significantly inhibits migration and eNOS activation. To assess the in vivo relevance of the opposing effects of SOM1 and SOM3, we made use of zebrafish as a whole animal model. SOM3, but not SOM1, induced severe malformations in embryos and reduced the number of normally developed embryos significantly. Microarray analyses showed that SOM1 and SOM3 have some overlapping, but an even larger number of distinct target genes.

Our data suggest that the opposing effects of the two SOM isoforms are mediated by transcriptome alterations and that a switch in isoform expression could change the cellular fate with dramatic consequences for a whole organism.
Abstract 56: Thrombin Induces Protease-Activated Receptor 1 and PAR4 Heterodimers
Authors: de la Fuente Maria , Nieman Marvin
Published: March 2022

Thrombin is a potent platelet agonist. Thrombin activates platelets and other cells of the cardiovascular system by cleaving its receptors, protease activated receptor 1 (PAR1), PAR4 or both. PARs are G-protein coupled receptors that activate cellular signaling through Gq and G12/13. Our previous studies have determined that PAR4 forms homodimers and have mapped the homodimer interface to specific residues in transmembrane helix 4 (TM4). We have also shown that coexpression of PAR1 with PAR4 lowers the threshold for PAR4 activation by thrombin ∼10-fold. The purpose of the current study is to examine the physical interaction between PAR1 and PAR4 and how these interactions influence PAR1’s ability to enhance PAR4 activation. The PAR1-PAR4 heterodimers were examined by bioluminescence resonance energy transfer (BRET). PAR1-PAR4 heterodimers were not detected under basal conditions. However, when the cells were stimulated with 10 nM thrombin, we were able to detect a strong interaction between PAR1 and PAR4. In contrast, stimulating PAR1, PAR4, or both with agonist peptides TFLLRN (100 microM) or AYPGKF (500 microM), respectively did not induce heterodimers. Further, point mutations that prevent PAR1 or PAR4 cleavage did not from heterodimers when stimulated with thrombin. Other proteases that activate PAR1 or PAR4 such as activated protein C (APC) and plasmin do not induce PAR1-PAR4 heterodimers. Finally, we have introduced point mutations in transmembrane helix 4 of PAR1 and PAR4 that prevent heterodimer formation when stimulated with thrombin. These data demonstrate that PAR1 and PAR4 require allosteric changes induced by receptor cleavage by thrombin to mediate heterodimer formation and have determined the PAR1-PAR4 heterodimer interface. Taken together, these data suggest that PAR1 and PAR4 have a dynamic interaction depending on the context of their expression. Since PAR1 is an attractive antiplatelet target, the molecular interactions of this receptor on the cells surface must be taken into account when developing and characterizing these antagonists.