bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 196: Impact of Cholesteryl Ester Transfer Protein Inhibition With TA-8995 on High Density Lipoprotein Function
Authors: Shrestha Sudichhya , Jessup Wendy , Tabet Fatiha , Barter Philip J , Rye Kerry-Anne
Published: June 2021

Background:
The effect of inhibiting cholesteryl ester transfer protein (CETP) activity on high density lipoprotein (HDL) function is poorly understood.

Aim:
To determine whether the CETP inhibitor, TA-8995, influences the cholesterol efflux capacity and anti-inflammatory properties of HDLs.

Methods:
Subjects were treated with TA-8995 (2.5 mg/day, n=14 or 10 mg/day, n=13), or placebo (n=14) for 12 weeks. Blood was collected prior to (Visit 2), and after TA-8995 treatment (Visit 5). The composition and size of ultracentrifugally isolated HDLs was determined. Serum cholesterol efflux was quantified in J774 macrophages and CHO cells. Inhibition of vascular cell adhesion molecule (VCAM)-1 and intercellular adhesion molecule (ICAM)-1 expression was determined by flow cytometry in tumor necrosis factor (TNF)-alpha activated human coronary artery endothelial cells (HCAECs) after pre-incubation with isolated HDLs.

Results:
Plasma HDL-cholesterol (HDL-C) levels, HDL composition and size were comparable at Visit 2 and Visit 5 in the placebo group. Treatment with 2.5 and 10 mg TA-8995 increased HDL-C levels by 113±0.6% and 153±0.5%, respectively. HDL composition in the placebo and treatment groups was comparable at Visit 2. At Visit 5 the apoA-I/apoA-II molar ratio, and wt% phospholipid, cholesteryl ester and unesterified cholesterol increased and the wt% triglyceride and total protein decreased in both treatment groups. TA-8995 treatment also increased HDL size.

At Visit 5 versus Visit 2 cAMP-specific cholesterol efflux from J774 macrophages increased by 24±0.3% and 21±0.3%, respectively, in the 2.5 mg and 10 mg TA-8995 treatment groups while ABCA1-specific cholesterol efflux increased by 10±0.1% and 20±0.1%, respectively, and ABCG1-specific cholesterol efflux increased by 20±0.2% and 7±0.2%, respectively. Cholesterol efflux was comparable at Visit 2 and Visit 5 in the placebo group.

Pre-incubation of HCAECs with HDLs from the placebo and both TA-8995 treatment groups inhibited TNF-alpha-induced ICAM-1 and VCAM-1 expression comparably at Visits 2 and 5.

Conclusion:
Inhibition of CETP with TA-8995 changes HDL composition. It also increases HDL size and cholesterol efflux and conserves the anti-inflammatory properties of HDLs.
Abstract 20: Nogo-B Controls Inflammation by Regulating Autophagy in Macrophage
Authors: Tao Bo , Yu Jun
Published: June 2021

Objective:
Autophagy, initiated from endoplasmic reticulum (ER), plays a housekeeping role that controls inflammatory response in cell. However, the precise mechanisms underlying this process are not yet fully understood. The ER is believed to be one of the major membrane sources and the scaffold for autophagosome formation. ER remodeling participates in autophagy initiation. Nogo-B is a ubiquitously expressed reticulon family protein that mainly localized to ER membrane. Nogo-B has been shown to regulate ER membrane remodeling and trafficking. Herein, we hypothesize that Nogo-B may have a critical role in balancing autophagy, inflammation and cell homeostasis.

Approach and Results:
In this study, we have been able to show that genetic loss of Nogo-B (Nogo-B
-/-
) protected against cecal ligation and puncture or lipopolysaccharide (LPS)-mediated sepsis in vivo. Nogo-B deficiency increased autophagic flux in the lung and, decreased proinflammatory cytokine, IL-1beta, production in Bronchoalveolar lavage fluid and blood
in vivo
. Macrophage specific Nogo-B deletion mice phenocopied the global Nogo-B
-/-
mice, suggests that Nogo-B in macrophage serves a critical role in controlling the process of sepsis. Furthermore, loss of Nogo-B in monocytes and macrophages enhanced starvation and LPS-induced autophagy and ameliorated IL-1β production, which was not due to the alteration of LPS-mediated NFκB signaling or MAPK signaling, but through enhanced inflammasome degradation, in vitro. Mechanistically, Nogo-B associates with Bcl2 and stables Bcl2:Beclin1 complex in ER and in turn negatively regulates autophagosome formation.

Conclusions:
Taken together, our findings define a novel mechanism controlling autophagy by a ER membrane protein, Nogo-B. Nogo-B is a negative regulator of the autophagy initiation and, facilitates inflammation. The results from this study deepen our understanding on the mechanisms of autophagosome formation and its role in inflammation regulation.
Abstract 205: Impact of PLPP3 Gene Deficiency on Lipid Metabolism and Signaling
Authors: Halder Suchismita
Published: June 2021

Genome wide association studies identify heritable variants within the final intron of the PLPP3 gene that associate with coronary artery disease risk. The risk associated alleles of these variants alter activity of an enhancer sequence that acts in cis to upregulate expression of the gene in response to inflammatory and atherogenic stimuli (for example oxidized low density lipoprotein) and may have longer distance effects on expression of other genes. PLPP3 encodes lipid phosphate phosphatase 3 (LPP3) which is an integral membrane enzyme that dephosphorylates lipid phosphate esters. These substrates are bioactive signaling molecules at the cell surface including lysophosphatidic acid (LPA) and sphingosine 1 phosphate (S1P) and intracellular intermediates in pathways of lipid metabolism and signaling, for example ceramide 1 phosphate and phosphatidic acid. PLPP3 is essential for vascular development in mice. We have generated mice with inducible post-natal and tissue specific inactivation of this gene which are viable and exhibit accelerated atherosclerosis on a low density lipoprotein receptor knockout background after feeding atherogenic diets. Analysis using targeted and untargeted approaches using multistage and high resolution mass spectrometry reveal that LPP3 deficiency results in significant differences in levels of some LPP3 substrates and products in cells, tissues and biological fluids from these animals. These observations support the hypothesis that alterations in levels of these LPP3 substrates or their dephosphorylation products account for or contribute to the mechanisms by which alterations in LPP3 expression influence cardiovascular disease risk.
Abstract 227: The Role of Fatty Acid Desaturase 1 in Inflammation Initiation and Resolution in Atherosclerosis
Authors: Gromovsky Anthony , Schugar Rebecca , Brown Amanda , Burrows Amy , Ferguson Daniel , Sansbury Brian , Graham Mark , Crooke Rosanne , Lee Richard , Hazen Stanley , Parks John , Spite Matthew , Brown J. Mark
Published: June 2021

Studies suggest that dietary enrichment of polyunsaturated fatty acids (PUFAs) may improve CVD outcomes, however the mechanisms underlying these observations remain elusive. Additionally, genome-wide association studies have linked single nucleotide polymorphisms (SNPs) in genes encoding essential PUFA metabolizing enzymes, the Fatty Acid Desaturases 1, 2, and 3 (
FADS1-2-3
), to dyslipidemia and other metabolic risk factors known to be linked to CVD. Since enzymatic oxidation of specific PUFAs, commonly arachidonic acid (AA), eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA), generate signaling mediators that coordinate both the initiation and resolution of inflammation, understanding the interplay of PUFA metabolism and vascular inflammation has broad implications in cardiometabolic drug discovery. Here we have altered the abundances of distinct dietary and endogenously-synthesized PUFAs, and associated lipid mediators, in hyperlipidemic Ldlr-/- mice to understand the signaling mechanisms underlying the PUFA-CVD link. We diversified PUFA profiles by feeding diets enriched in either ω-6 or ω-3 PUFAs, and inhibited endogenous production of AA and EPA by antisense oligonucleotide (ASO)-mediated knockdown of the Δ5 desaturase Fads1. Fads1 knockdown results in accumulation of its substrate fatty acids (dihomo-γ-linolenic acid and eicosatrienoic acid) and marked reduction in its enzymatic products (AA and EPA). Metabololipidomic profiling reveals that Fads1 knockdown alters select AA-, EPA-, and DHA-derived lipid mediators in a diet-specific manner. Interestingly, Fads1 knockdown protects Ldlr-/- mice from obesity and insulin resistance regardless of diet, yet alters increases circulating monocyte subsets and promotes dyslipidemia in a highly diet-specific manner. How these phenotypes link to diet-induced atherosclerosis burden in Fads1 knockdown mice will also be presented. Collectively, our studies provide new information in regards to the relative contribution of endogenous (FADS1-derived) and exogenous (diet-derived) AA and EPA in driving cardiometabolic phenotypes. Furthermore, our studies provide the first mechanistic understanding behind the genetic link of
FADS1-2-3
SNPs and atherosclerosis.
Abstract 23: Microrna302-367 Sphingosine 1 Phosphate Receptor 1 Pathway Prevents Tumor Growth via Restricting Angiogenesis and Enhancing Vascular Stability
Authors: Pi Jinjiang , Tao Ting , Zhuang Tao , Sun Huimin , Chen Xiaoli , Shen Yixin , Yu Zuoren , Zhu Helen H , Gao Wei-qiang , Suo Yuanzhen , Wei Xunbin , Zheng Xiangjian , Tian Ying , Morrisey Edward , Zhang Lin , Zhang Yuzhen
Published: June 2021

Angiogenic hypersprouting and leaky immature vessels of pathological angiogenesis are essential for tumor growth. MicroRNAs have unique therapeutic advantages by targeting multiple pathways of tumor-associated angiogenesis, but the function of individual miRNAs in angiogenesis and tumors has not yet been fully evaluated. Here, we show that miR302-367 elevation in endothelial cells reduces retina sprouting angiogenesis and promotes vascular stability in vivo, ex vivo and in vitro. Erk1/2 are identified as direct targets of miR302-367, and down-regulation of Erk1/2 upon miR302-367 elevation in endothelial cells increases the expression of Klf2 and in turn S1pr1 and its downstream target VE-cadherin, suppressing angiogenesis and improving vascular stability. Conversely, both pharmacological blockade and genetic deletion of S1pr1 in endothelial cells reverse the anti-angiogenic and vascular stabilizing effect of miR302-367 in mice. Pathological angiogenesis in tumors shares features of developmental angiogenesis, and endothelial specific elevation of miR302-367 reduces tumor growth by restricting sprout angiogenesis and decreasing vascular permeability via the same Erk1/2-Klf2-S1pr1 pathways. In conclusion, miR302-367 regulation of an Erk1/2-Klf2-S1pr1 pathway in the endothelium advances our understanding of angiogenesis, meanwhile also provides opportunities for therapeutic intervention of tumor growth.
Abstract 235: Prevalence of Lipoprotein(a) Measurements and Extent of Lipoprotein(a) Elevations in 2,266 Patients with Aortic Stenosis: Results from an Academic Echocardiography Laboratory Practice Setting
Authors: Wilkinson Michael J , Ma Gary S , Yeang Calvin , Strachan Monet , Wilson Joel , DeMaria Anthony N , Tsimikas Sotirios , Cotter Bruno
Published: June 2021

Background:
The
LPA
gene causes elevated lipoprotein(a) (Lp(a)) levels and causally mediates calcific aortic valve stenosis (AS). Elevated Lp(a) and its associated oxidized phospholipids (OxPL-apoB) predict the progression of pre-existing AS and need for aortic valve replacement and are targets of therapy.

Methods:
We determined the prevalence and the extent of Lp(a) elevation in patients with AS diagnosed by echocardiography performed at the University of California San Diego between 2010-2015. Severity of AS was classified as critical, severe, moderate, mild, or trace. Lp(a) levels were organized as Lp(a) <30 mg/dL, 30-50 mg/dL, 50-100 mg/dL and >100 mg/dL.

Results:
2,266 patients with AS were found, with 130 critical, 333 severe, 477 moderate, 1318 mild, and 8 cases of trace AS. Mean age was 75.0 and range 18-106 years. 51% of patients were male. Prevalence of any Lp(a) measurement was 159/2,266 patients (7.02%). The number (%) of patients with an Lp(a) level was: 1) critical (n=4, 3.1%), 2) severe (n=28, 8.4%), 3) moderate (n=56, 11.7%), 4) mild (n=71, 5.4%), and 5) trace (n=0). The extent of Lp(a) elevation within each AS category is in Table 1: 55/159 (34%), 35/159 (22%) and 19/159 (12%) of patients had Lp(a) >30 mg/dL, >50 mg/dL and >100 mg/dL, respectively.

Conclusion:
Lp(a) was measured in only 7.0% of patients with AS in an academic setting. Given the ongoing development of therapies to lower Lp(a) in patients with AS, educational efforts are needed to raise awareness of Lp(a) as a causal risk factor for AS.
Abstract 240: T Cell Specific CD40L Promotes to Atherosclerosis
Authors: Bürger Christina , Winkels Holger , Spitz Charlotte , Meiler Svenja , Reim Sigrid , Weber Christian , Lutgens Esther , Gerdes Norbert
Published: June 2021

Background:
Adaptive immunity and co-stimulatory signals are pivotal to all stages of atherosclerosis. CD40-CD40L (CD154) co-stimulatory molecules are central mediators of disease progression. However, it remains undetermined whether platelet- or T cell-specific CD40L contributes to its pro-atherogenic function.

Method:
We generated mice carrying a mutated allele of the CD40L gene flanked by loxp-sites sensitive to cre-mediated inactivation. We crossed these to
Apoe
-/-
-deficient mice which also expressed Cre specifically in T cells (
Cd4Cre
tg
) or platelets
(Pf4Cre
tg
) to generate
Apoe
-/-
Cd40lg
fl/fl
-Cd4Cre
tg
,
-Cd4Cre
wt
,
-Pf4Cre
tg
, or
-Pf4Cre
wt
mice. After consuming chow diet for 28 weeks the immune status and atherosclerosis was assessed by flow cytometry, RNA expression analysis and histology of the ascending aorta, blood, draining lymph nodes, and spleen.

Results:
Specific deletion of CD40L on T cells leads to decreased atherosclerotic lesion size in the ascending aorta while plaque size in
Apoe
-/-
Cd40lg
fl/fl
-Pf4Cre
tg
or
-Pf4Cre
wt
mice did not differ. Furthermore, platelet-specific ablation of CD40L did not results in changes of immune cells distribution. In contrast, cytokine and RNA analysis data pointed to a less inflammatory phenotype in
Apoe
-/-
Cd40lg
fl/fl
Cd4Cre
tg
mice compared to
Apoe
-/-
Cd40lg
fl/fl
Cd4Cre
wt
controls. Interestingly, T cell-specific inactivation of CD40L also caused a systemic decrease of Tregs in draining lymph nodes, spleen, blood, and thymus.

Conclusion:
Taken together our data confirm that CD40L expressed on T cells but not on platelets contributes to the disease-aggravating role of the CD40/CD40L dyad in atherosclerosis. Thus, T cell-expressed CD40L may provide a promising candidate for further investigation in atherosclerosis and other inflammatory diseases.
Abstract 244: Myocardial Infarction Accelerates Atherosclerosis in Low Density Lipoprotein Receptor Knockout Mice
Authors: Covarrubias Roman , Wilhelm Ashley , Chepurko Elena , Novitskaya Tatiana , Moore Jared L , Rhoads Jillian P , Major Amy S , Gumina Richard J
Published: June 2021

Objective:
To elucidate the molecular mechanisms leading to accelerated atherosclerosis after a myocardial infarction (MI).

Approach and Results:
Human studies suggest that atherosclerosis is accelerated following MI. Although, this has been recapitulated in apolipoprotein E deficient (apoE
-/-
) mice, the mechanisms by which MI-accelerated atherosclerosis have not been elucidated. Here, we demonstrate that, similar to apoE
-/-
mice, MI accelerates atherosclerosis in high-fat diet-fed LDLr
-/-
mice. We fed high-fat diet (21% saturated fat, 0.15% cholesterol) to LDLr
-/-
mice for 1 week prior to coronary artery ligation (CAL) or sham surgery. After surgery, LDLr
-/-
mice were maintained on high-fat diet and sacrificed after 3 weeks. Total serum cholesterol (Sham: 1349.0 ± 50.09 and CAL: 1355 ± 20.09, p = 0.19) and triglycerides (Sham: 394.6 ± 32.4 and MI 421.1 ± 6.27, p =0.48) were comparable between sham and CAL mice. Examination of aortic sinus by oil red O-stained cross-sections revealed an increase in atherosclerotic lesion area in LDLr
-/-
that received an MI (CAL: 78,465 ± 15,239 microm
2
/section and sham 42,697 ± 4757 microm
2
/section, p= 0.03). Additionally, we found a statistically significant increase in the number (sham: 17,943 ± 7762, n=8 and CAL: 58,129 ± 13,444, n=6, p = 0.017) and percentages (sham: 2.2 ± 0.41, n=8, CAL: 5.32 ± 1.105, n=6, p = 0.01) of Ly6G
+
neutrophil infiltrate in para-aortic lymph nodes in LDLr
-/-
mice that received MI versus sham surgery. Analysis of cellular composition in atherosclerotic lesions is ongoing.

Conclusion:
Atherosclerosis is accelerated following an MI in high-fat fed LDLr
-/-
mice.
Abstract 246: Adiporon, an Adiponectin Receptor Agonist, Inhibits P70s6kinase Signaling in Vascular Smooth Muscle Cells: Implications Toward Suppression of Injury-induced Neointima Formation in Mice
Authors: Fairaq Arwa , Segar Lakshman
Published: June 2021

Background:
Hypoadiponectinemia is closely correlated with insulin resistance and coronary artery disease. The use of exogenous adiponectin as a rational therapy to reduce cardiovascular disease risk has been hampered in a clinical setting due to its large molecular size and short plasma half-life. Recent studies demonstrate that Adiporon, a small molecule adiponectin receptor agonist, improves insulin resistance and glycemic control in type 2 diabetic mice. The present study is aimed at examining the likely regulatory effects of Adiporon on vascular smooth muscle cell (VSMC) phenotype.

Methods:
Using human aortic VSMCs, we examined the effects of AdipoRon (5 to 100 μM) on platelet-derived growth factor (PDGF, 30 ng/ml)-induced proliferative signaling events. In addition, we examined the effects of orally-administered AdipoRon (50 mg/kg/d) on injury-induced neointima formation in CJ57BL6 mice. Morphometric analysis of injured femoral artery was performed using H&E and EVG-stained sections.

Results:
Exposure of VSMCs to AdipoRon resulted in a significant decrease in PDGF-induced proliferation by ~63.8% and cyclin D1 expression by ~92.5% (n = 3). In addition, AdipoRon treatment led to significant diminutions in PDGF-induced phosphorylation of Akt, p70S6K, and its downstream targets that include 4E-BP1 and ribosomal protein S6, as revealed by immunoblot analysis (n = 3). Importantly, Adiporon administration (50mg/kg/d, PO) for three weeks led to a significant attenuation of neointima formation by ~63.7% in the injured femoral artery (n = 5-8).

Conclusion:
The present findings suggest that Adiporon (a small molecule adiponectin receptor agonist) may inhibit neointima formation after arterial injury by targeting p70S6K signaling in VSMCs. Future studies should investigate the likely vasoprotective effects of Adiporon in the mouse models of obesity and atherosclerosis.
Abstract 248: Stat4 Promotes Atherosclerosis by a Reduction in Cd8
+
Tregs, Supporting Tfh and Plasma B Cell Development, T Cell Activation and Cd11b+ Cell Accumulation in the Aortic Wall
Authors: Taghavie-Moghadam Paresa , Gjurich Breanne , Nadler Jerry , Galkina Elena
Published: June 2021

Atherosclerosis is an inflammatory disease of medium and large size arteries. Signal transducer and activator of transcription 4 (STAT4) is a transcription factor that supports generation of cells with features of both follicular helper T cells (Tfh) and T helper 1 (Th1) cells. To date, the role of STAT4 in atherogenesis and its effects on the induction of Tfh cells is an important area of active research. To investigate the role of STAT4 in atherogenesis, we generated STAT4-deficient, low-density lipoprotein-deficient (

Stat4
-/-
Ldlr
-/-

) mice. STAT4 deficiency resulted in a significant reduction in plaque burden within the aortas of

Stat4
-/-
Ldlr
-/-

mice fed a high fat/high carbohydrate diet with added cholesterol (DDC) for 16 weeks compared to age and diet-matched

Ldlr
-/-

mice (p<0.01). STAT4 deficiency resulted in the reduction of CD19
+
B cells (p<0.0001) CD8
+
T cells (p<0.01) and CD3
+
T cells (p<0.05) in spleens and peripheral lymph nodes of

Stat4
-/-
Ldlr
-/

vs

Ldlr
-/

mice. Additionally, the number of splenic CXCR5
+
Tfh cells and CD138
+
plasma B cells was also diminished (p<0.003), suggesting reduced follicular germinal center activities under conditions of STAT4 deficiency. CD8
+
Tregs are suppressors for autoreactive CD4
+
T cells and also target Tfh cells. Importantly, CD8
+
Tregs were elevated in spleens of

Stat4
-/-
Ldlr
-/-

compared to

Ldlr
-/-

mice (p<0.05), suggesting that decreased Tfh cell numbers are due to increased CD8
+
Tregs in

Stat4
-/-
Ldlr
-/-

mice. While STAT4 deficiency had no effects on M1/M2 macrophage polarization, Stat4-deficient M1 macrophages showed reduced MHCII expression and diminished secretion of pro-inflammatory cytokines upon stimulation in vitro. Furthermore, STAT4 deficiency led to a reduced MHCII expression by antigen-presenting cells (APC) and decreased content of CD11b
+
CD11c
-
cells in the aortas of

Stat4
-/-
Ldlr
-/-

vs

Ldlr
-/-

mice. Together, these data demonstrate a pro-atherogenic role of STAT4 in multiple cell types via the regulation of CD8
+
Treg-dependent germinal center formation, modulation of levels of APC activation, and alteration of the immune response within the aortic wall.

This work is supported by NHLBI HL112605 (J.N., E.G.), HL112605 supplemental grant 02S1 (J.N., P.T-M), and by HL107522 (E.G.).
Abstract 256: Global and Hepatocyte-specific Ablation of Bmal1 Induces Hyperlipidemia and Enhances Atherosclerosis
Authors: Pan Xiaoyue , Bradfield Christopher A. , Hussain M. Mahmood
Published: June 2021

Circadian rhythms controlled by clock genes affect plasma lipids, known risk factors for atherosclerosis. We observed that global ablation of a critical clock gene,
Bmal1
, in

Apoe
–/–

and

Ldlr
–/–

mice, and liver-specific ablation of Bmal1 in

Apoe
–/–

(L

-Bmal1
–/–
Apoe
–/–

) mice increases hyperlipidemia and atherosclerosis. In contrast, overexpression of BMAL1 in

L-Bmal1
–/–
Apoe
–/–

mice decreases hyperlipidemia and atherosclerosis. Physiologic studies showed that Bmal1 deficiency augments hepatic lipoprotein secretion and diminishes cholesterol excretion to the bile. Molecular studies identified that Bmal1 deficiency reduces expression of Shp, a repressor, and Gata4, an activator. Reductions in Shp increase Mtp expression and lipoprotein production, whereas reductions in Gata4 diminish Abcg5/Abcg8 expression and cholesterol excretion to the bile. SHP expression normalized lipoprotein secretion with no effect on cholesterol excretion to the bile, while GATA4 expression increased cholesterol excretion to the bile and reduced plasma lipids in

L-Bmal1
–/–
Apoe
–/–

mice. Taken together, our data indicate that Bmal1 modulates lipoprotein secretion to the plasma and cholesterol excretion to the bile by regulating the expression of Mtp and Abcg5/Abcg8 via Shp and Gata4, and that the loss of Bmal1 promotes hyperlipidemia and atherosclerosis.
Abstract 266: Role of Marine Long-chain Monounsaturated Fatty Acid in Cardiovascular Disease Prevention and Possible Mechanism to Reduce the Atherosclerosis
Authors: Yang Zhi-Hong , Vaisman Boris , Pryor Milton , Bando Masahiro , Fukuda Daiju , Sata Masataka , Sakaue Hiroshi , Miyahara Hiroko , Takeo Jiro , Remaley Alan
Published: June 2021

Numerous studies have shown cardiovascular benefits of fish oil, and most of these favorable effects have been attributed to omega-3 fatty acids. Fish oils, however, also contain varying amounts of other unusual types of fatty acids, such as long-chain monounsaturated fatty acids (LCMUFA; i.e., C20:1 and C22:1 isomers combined). Compared with well-studied omega-3, limited information is available on the role of LCMUFA on cardiovascular disease. In the current study, we examined the effect of saury fish oil-derived LCMUFA concentrate on the pathogenesis of atherosclerosis in ApoE-deficient and LDLR-deficient mice. Male ApoE-deficient mice were fed a western diet supplemented with 5% LCMUFA concentrate or not (control) for 12 weeks.
En face
Sudan IV staining of the aorta revealed that LCMUFA significantly suppressed the development of atherosclerotic lesions, and down-regulated inflammatory gene expression. In LDLR-deficient female mice, we also observed that 12-week supplementation of 2% LCMUFA on a western diet significantly decreased atherosclerosis lesion areas and accumulation of macrophages, compared with western diet (control) or western diet supplemented with 2% olive oil enriched in shorter-chain MUFA oleic acid (C18:1). There were no differences, however, in plasma or hepatic lipid profiles between the three groups. LCMUFA, but not olive oil, also significantly suppressed several plasma inflammatory cytokine levels. RNA sequencing and subsequent qPCR analyses revealed that LCMUFA upregulated PPAR signaling pathways in liver. In cell culture studies, apoB-depleted plasma from the LCMUFA group promoted cholesterol efflux from macrophage-like THP-1 cells and ABCA1-overexpressing BHK cells. LCMUFA also suppressed LPS-induced inflammatory cytokine production by macrophages. In conclusion, our research showed for the first time that LCMUFA consumption could protects against atherosclerosis, possibly by upregulating the PPAR signaling pathway. Omega-3 fatty acids are generally considered the major active components in fish lipids, but our findings provide new insights into potential health effect of LCMUFA-rich fish oil and suggest that LCMUFA-rich fish oil may also be beneficial for reducing cardiovascular disease risk.
Abstract 319: Cardiac Deficiency of AMPK Exacerbates Heart Hypertrophy by Pressure Overload
Authors: Wang Jinli , Quan Nanhu , Wang Lin , Sun Wanqing , Cates Courtney A , Chu Dongyang , Chen Xu , Rezaie Alireza , Li Ji
Published: June 2021

Introduction:
Heart hypertrophy caused by pressure overload is characterized by the activation of adenosine monophosphate-activated protein kinase (AMPK), which is a major energy sensor in the heart. However, the anti-inflammatory effect of AMPK has not been determined in the hypertrophy model. Activated protein C (APC) is a vitamin-K dependent plasma serine protease which inhibits blood clotting, and APC is an endogenous AMPK agonist. This study was designed to examine the role of AMPK in hypertrophy and the underlying mechanisms by which APC inhibits heart hypotrophy by pressure overload.

Hypothesis:
AMPK play a role in preventing heart from hypertrophy induced by pressure overload. APC could inhibit high blood pressure-induced hypertrophy via activation of AMPK signaling pathway.

Methods:
Wild-type (WT) and AMPK-kinase dead (KD) transgenic mice were subjected to transverse aortic constriction (TAC) surgery. Echocardiography was performed to evaluate the heart function, and histology staining revealed the morphological changes. Real-time PCR and western blotting were used to determine the signaling changes of mRNA and protein expression levels.

Results:
There is no phenotype difference between WT and AMPK-KD mice under normal physiological conditions. However after 4 weeks of TAC surgery, AMPK-KD mice demonstrated significantly bigger heart than WT mice (p<0.05), and the cardiac functions measured by echocardiography in AMPK-KD hearts were significantly impaired as compared with WT hearts (p<0.05). The immunohistochemical staining showed that the increased macrophage infiltration and reactive oxygen species (ROS) including activated p66shc, 4-Hydroxynonenal accumulation and phosphorylation of extracellular signal-regulated kinase (ERK) were observed in the AMPK-KD hearts after 4 weeks of TAC surgery (all p<0.05 versus WT hearts). APC administration significantly attenuated hypertrophy and fibrosis caused by pressure overload, and macrophage infiltration and p66shc activation were also inhibited by APC treatment.

Conclusions:
Cardiac AMPK deficiency aggravates hypertrophy caused by pressure overload. AMPK activator APC could be a therapeutic drug for treatment of hypertrophy by high blood pressure.
Abstract 32: Analysis of Serum Clopidogrel Active Metabolite Concentration Identifies Novel Genetic Variants Associated With Clopidogrel Pharmacokinetics
Authors: Backman Joshua D , Lewis Joshua P , Horenstein Richard B , Pacanowski Michael A , Figg William D , Shuldiner Alan R , O'Connell Jeffrey R , Yerges-Armstrong Laura M
Published: June 2021

Clopidogrel with aspirin is standard of care for the secondary prevention of ischemic events in patients with acute coronary syndromes. However, substantial inter-individual variation in clopidogrel response results in sub-optimal therapy in some patients and an increased risk of recurrent events. Up to 73% of the variation in clopidogrel efficacy is found to be heritable, and while key genetic variants such as
CYP2C19
*2 have been identified, a large portion of the genetic heritability remains unaccounted for. To date, few studies have used clopidogrel active metabolite to evaluate clopidogrel pharmacogenomics. The evaluation of this biomarker may be particularly informative as it enables us to focus almost exclusively on clopidogrel pharmacokinetics and may allow new discoveries leading to a more thorough understanding of clopidogrel activation.

Serum clopidogrel active metabolite concentration was measured by HPLC-MS/MS in 506 healthy, Amish participants of the Pharmacogenomics of Anti-Platelet Intervention (PAPI) Study after a 7-day 75-mg/day clopidogrel intervention. Genome-wide genotyping was performed using Affymetrix 5.0 and 6.0 arrays, and SNPs were imputed to the 1000 Genomes cosmopolitan panel. Mixed models analysis was conducted under an additive model and adjusted for age, sex, BMI, and relatedness among study participants with MMAP (http://edn.som.umaryland.edu/mmap).

The most significant signal of our GWAS was the previously identified
CYP2C19
/
CYP2C9
locus (P= 9.2x10
-15
), and was fully accounted for by the
CYP2C19
*2 variant. In addition to this signal, novel genome-wide significant signals were identified in loci on Chr. 3 (rs187941554, P=4.7x10
-11
), Chr. 7 (rs73407739, p=3.6x10
-8
), and Chr. 17 (rs80343429, P=3.7x10
-8
).

In this study we conducted the first ever GWAS of serum clopidogrel active metabolite concentration. Novel associations for clopidogrel efficacy were identified, and further study is required to determine the impact of these variants on clinical care Continual analysis of clopidogrel active metabolite concentration will contribute to a better understanding of the clopidogrel metabolic pathway, and potentially lay the groundwork for improved anti-platelet treatment for CVD patients in the future.
Abstract 323: Acute Reduction of Ambient Air Pressure Lowers Systemic Vascular Resistance and Improves Cardiac Output in Mice
in-vivo
Authors: Shahid Anmol , Patel Vaibhav B , Oudit Gavin Y , McMurtry Michael S
Published: June 2021

Background:
Humans have lower risk for myocardial infarction and stroke at higher altitudes. The mechanism is unknown. We previously demonstrated that acute reductions in ambient air pressure result in enhanced arterial vasodilation
ex vivo
using a myograph model of murine resistance arteries. Enhanced vasodilation might increase effective arterial compliance
in vivo
, thus we further hypothesized a reduction in systemic vascular resistance
in vivo
as a result of lowered ambient air pressure.

Methods:
In-vivo
Pressure Volume Loops: A novel hypobaric chamber was designed and constructed with the capacity to hold a single anaesthetized mouse and simulate a variety of lowered air pressures within the chamber. The design of the chamber allowed for monitoring of murine heart rate, respiratory rate, & blood oxygen saturation at all times. Using inhaled isoflurane to anesthetize the animal, a closed-chest left-ventricular catheterization procedure was performed within the chamber on free-breathing adult male C57-WT mice (n=8) during consecutive acute exposures to the three air pressure steps: 754 mmHg (control or room atmospheric pressure), 714 mmHg (p1) and 674 mmHg (p2).

Results:
In-vivo
: Total systemic vascular resistance was reduced by acute exposure to lower air pressure (10.09±0.15 mmHg*min/μL at control vs. 8.11±1.45 and 8.18±1.24 mmHg*min/μL at p1 and p2, respectively; p<0.05). Cardiac output was significantly improved by lowering atmospheric pressure below room pressure (7.36±0.15 mL/min at control versus 9.71±1.45 mL/min and 9.59±1.77 mL/min at p1 and p2, respectively; p<0.05). Significant increases in stroke volume with a reduction in air pressure were also observed (p<0.05 for p1 and p2 vs. control). We did not note significant changes in heart rate, mean arterial pressure, or contractility of the mice with lowered atmospheric pressure.

Conclusion:
Acute reductions in ambient air pressure lower systemic vascular resistance, and increase cardiac output and stroke volumes
in vivo
.
Abstract 330: Testicular Vein Thrombosis Etiology and Outcomes
Authors: Lenz Charles J , Saadiq Rayya , Simmons Benjamin , Cohoon Kevin , McBane Robert , Daniels Paul , Wysokinski Waldemar
Published: June 2021

Background:
Testicular vein thrombosis (TVT) is not currently well-described in the literature. A better understanding of the natural history of TVT will allow for development of optimal management strategies for this little known entity.

Methods:
The goal of this project was to compare TVT with ovarian vein thrombosis (OVT) and lower extremity deep vein thrombosis (DVT). Patients with TVT, OVT, or DVT between 1995 and 2015 were identified and a chart review was performed.

Results:
A total of 39 patients with TVT occurring between 1995 and 2015 were identified and compared with 35 patients with OVT and 114 randomly chosen DVT patients. Mean duration of follow-up was 2.9 years (range 0-16.1). Patient demographics can be seen in table 1. Cancer-related (p-value=<0.0001) and idiopathic thrombosis (p-value=0.01) rates were significantly different among the groups. Recurrence rates were similar between TVT and OVT (4.2 vs. 3 per 100 patient-years, p-value = 0.9) as well as TVT and DVT (4.2 vs 2.2 recurrences per 100 patient-years, p-value= 0.37). There were 25 deaths (22%) in DVT patients, 9 (26%) in OVT patients, and 12 (31%) in TVT patients (p-value= 0.541). There was one major bleeding event in both an OVT patient and a TVT patient.

Conclusion:
Etiology of TVT differs from that of OVT and DVT, specifically in cancer-related and idiopathic thrombosis rate. Our data supports a similar rate of recurrent venous thromboembolism and death in TVT as compared with DVT and OVT.
Abstract 332: Characteristics of Patients with Submassive Pulmonary Embolism with Early Re-hospitalization After tPA Administration
Authors: Valadri Ravinder , LeYu Naing , Desai Pramthesh , Thomas Linda H , Pancholy Samir B , Singhania Namrata , English Richard , William Dempsey , Litchman Maureen
Published: June 2021

Background:
Evidence for clinical predictors of early re-hospitalization after tissue plasminogen activator (tPA) therapy in patients with submassive pulmonary embolism (SMPE) is currently lacking. We sought to explore the characteristics of these patients in a retrospective cohort study at our hospital

Methods:
Clinical, biochemical, hemodynamic, echocardiographic characteristics and clinical course of consecutive patients with SMPE [as defined by Acute PE without systemic hypotension (systolic blood pressure >90 mm Hg) but with either Right Ventricular strain/dysfunction demonstrated on echocardiography obtained within 24 hours of presentation or myocardial necrosis as evidenced by elevated cardiac troponins and/or elevated brain natriuretic peptide levels)] were studied in a retrospective cohort study. Unplanned all cause re-hospitalizations after the index hospitalization for SMPE were captured

Results:
A total of 145 (in past 5 years) patients with pulmonary embolism diagnosed with spiral CT angiography or ventilation perfusion scan were screened for patients with SMPE. 17 patients with SMPE were identified in whom tPA was administered. No patient had major bleeding, one patient suffered gross hematuria during the tPA administration. After a mean 2.1 years of follow up 5 (29%) patients suffered unplanned all cause re-hospitalizations among which 4 were within one year from the index hospitalization for SMPE. Baseline characteristics are shown in table 1. No patients had recurrent PE after the index hospitalization

Conclusion:
A significant proportion of patients were re-hospitalized within one year after tPA administration in patients with SMPE. Further studies in larger cohorts are needed to identify independent clinical predictors of re-hospitalization in patients with submassive pulmonary embolism
Abstract 337: Plasma Advanced-oxidized Protein Products Promote Platelet-endothelial Crosstalk and Endothelial Tissue Factor Expression
Authors: Pasterk Lisa , Lemesch Sandra , Leber Bettina , Trieb Markus , Curcic Sanja , Stadlbauer Vanessa , Schuligoi Rufina , Schicho Rudolf , Heinemann Akos , Marsche Gunther
Published: June 2021

Background/Aims:
End-stage renal disease (ESRD) is associated with a prothrombotic phenotype and substantial activation of platelets can occur in the course of hemodialysis. However, the underlying mechanisms for increased platelet reactivity remain unclear. Increased levels of oxidized albumin, termed “advanced oxidation protein products (AOPPs)” accumulate in subjects with renal disease.

Methods:
Platelet aggregation was assessed with platelet aggregometry. Platelet activation, formation of reactive oxygen species and tissue factor expression on the surface of platelets and endothelial cells was measured by flow cytometry. Platelet adherence was assessed under flow using the Cellix system. The oxidative status of albumin isolated from ESRD patients was determined by photometric analysis. Tissue factor levels in serum samples were determined using a commercially available ELISA kit.

Results:
Albumin isolated from hemodialysis patients as well as in vitro generated AOPP-albumin promoted platelet activation via CD36, dependent on the AOPP-content of albumin. AOPP-albumin mediated platelet activation was prevented by scavenging superoxide anions and inhibitors of phospholipase C and protein kinase C. Furthermore, AOPP and serum tissue factor levels were considerably increased in ESRD patients on hemodialysis and a significant correlation of AOPP and serum tissue factor was found.

Conclusion:
Interaction of platelet CD36 with endogenous oxidized albumin may link oxidative stress with a prothrombotic phenotype in ESRD.
Abstract 345: Syzygium Jambos: A Potential Therapeutic Approach to Treat the Vascular and Hematological Complications of Sickle Cell Disease
Authors: Santiago-Perez Analaura , Inostroza-Nieves Yaritza , Gil de la Madrid Daniel , Alicea Isamar , Vega Christopher , Prado Gregory N , Rivera Alicia , Romero Jose , Valentin Shirley
Published: June 2021

Protein disulfide isomerase (PDI) is an oxidoreductase that mediates thiol/disulfide interchange reactions and has been reported to play a critical role in thrombus formation following vascular injury. PDI has also been shown to regulate leukocyte adherence to the endothelium and nitric oxide delivery. We recently reported that PDI is present at high levels and regulates erythrocyte homeostasis and Gardos Channel activity in humans with Sickle Cell Disease (SCD). Thus, PDI inhibition has been proposed as a promising therapeutic approach to ameliorate both the vascular and hematological complications of SCD. Syzygium jambos (S. jambos) is purported to have anti-inflammatory and antioxidant properties. However, the regulation of PDI activity by S. jambos has not been studied. We studied in vitro PDI activity in the presence of the S. jambos aqueous leaf extract using a PDI insulin turbidity assay. We observed significant reductions in PDI activity at 25 μg/mL (66.0 ± 9.7%, p<0.01, n=3), 50 μg/mL (83.3 ± 6.0%, p<0.01, n=3), and 100 μg/mL (91.6 ± 11.5%, p<0.01, n=3). S. jambos extract showed a dose-dependent anti-PDI activity with an IC50 of 14.40 μg/mL. We then tested the effects of S. jambos on endothelin-1 (ET-1)-stimulated PDI activity in human endothelial cells. Using a fluorescence based PDI activity assay, we observed that ET-1 increased PDI activity (1.7 ± 0.7 folds, n=3) that was dose-dependently blocked by S. jambos extract. In addition, we observed that ET-1 stimulated ex vivo human polymorphic nucleated (PMN) leukocyte migration toward the endothelial cells that was likewise dose-dependently blocked by S. jambos extract. (p<0.01, n=3). We also quantified the levels of reactive oxygen species (ROS) production in ET-1 treated endothelial cells. ET-1 stimulation significantly increased ROS levels [3 fold] when compared to vehicle treatment (p<0.05, n=3). S. jambos extract reduced ET-1 stimulated ROS to baseline levels (p<0.05, n=3). Our results suggest that S. Jambos may represent a novel pharmacological approach to treat complications of SCD.
Abstract 352: Real-time Intravital Optical Imaging Reflects the Dynamic Changes of Oxidative Stress Induced by Cigarette Smoking in Vasculatures
Authors: Kim Ji Bak , Ryu Jiheun , Song Joon Woo , Oh Dong Joo , Gweon DaeGab , Yoo Hongki , Kim Jin Won
Published: June 2021

Background:
Reactive oxygen species (ROS) play a central role in cigarette smoking-induced atherogenesis. The present study aims to assess the smoking-induced acute oxidative stress within vasculatures, and evaluates whether the resveratrol, a natural polyphenol antioxidant, can counteract this ROS production, using a customized, high resolution intravital optical imaging in real-time.

Methods and Results:
20-week-old male C57BL/6 mice were divided into four groups according to the preceding administration of resveratrol (R) (25mg/kg via gavage, for 7 days) and exposure to cigarette smoke (CS). To in vivo assess acute oxidative stress in blood vessels, dihydroethidium, which forms a red fluorescence (ethidium, excitation/emission: 520nm/610nm) upon reaction with ROS, was injected intraperitoneally. During CS exposure, temporal changes of fluorescence signals from the mouse cremaster muscle including vasculatures were assessed by intravital optical imaging for 15 minutes. Fluorescence signals were much more pronounced in CS exposed mice than controls (p<0.001). Resveratrol p.o. significantly reduced the CS-induced ROS signals compared to the non-treated group (fluorescence signal to noise ratio, SNR, 2.51±0.09 vs. 12.52±2.116, p=0.0002) (Figure A). Without CS exposure, fluorescence signals in targeted vasculatures were very low showing no difference between groups (SNR, 1.65±0.19 vs. 1.53±0.07, p=0.80) (Figure A). Lipid peroxidation was increased in CS group and significantly attenuated in resveratrol-treated mice (Figure B). Fluorescence microscopy and immunostainings corroborated the in vivo findings.

Conclusions:
The intravital optical imaging was able to in vivo estimate the dynamic changes of ROS production by CS exposure. Our data demonstrated that even a brief exposure to CS increased oxidative stress in vasculatures promptly, and the resveratrol exerts protective effects against the CS-induced acute oxidative stress.
Abstract 355: Progenitor Cells and Extracellular Vesicles as Biomarkers for Vasculitis
Authors: Mizus Marisa C , Sreih Antoine G , Rogers Wade T , Zhang Lifeng , Schretzenmair Richard , Moore Jonni S , Merkel Peter A , Mohler Emile R
Published: June 2021

Background:
Patients with vasculitis (VS) have complex organ- and life-threatening disease manifestations due to autoimmune-mediated inflammatory vascular damage and an increased risk of cardiovascular disease (CVD). Accurate biomarkers do not exist for VS. Cells release extracellular vesicles (EVs) in the setting of activation, injury, or death. In patients with diabetes mellitus and CVD, the effects of the inflammatory cascade, platelets and endothelium, positively correlate with levels of EVs bearing receptors for CD3, CD41, or CD105, and negatively correlate with levels of circulating endothelial progenitor cells (CPCs). Because VS is characterized by systemic inflammation, platelet activation and endothelial damage, this group of diseases may have unique effects on EVs and CPCs. This study evaluated a high dimensional flow cytometry approach to profiling EVs and CPCs in plasma to discern a biomarker pattern in VS.

Methods:
Levels of CPCs and EVs were measured with a panel of cell surface markers in 10 VS patients and 5 age-similar healthy controls (HC). Data were acquired on a modified Becton Dickinson FACSCanto.

Results:
Although no significant increase in CPCs in VS compared with HC was detected, the standard deviation of CPC levels in VS was more than double that in HC (SD=574 vs. 165, p=0.081), suggesting heterogeneity in the VS group. EV subsets with lower concentrations in VS, compared with HC, included: Annexin
+
CD3
+
double-positive T Lymphocyte (p=0.026) and CD105
+
CD64
+
double-positive activated Macrophage (p=0.030). CD144
+
CD105
+
double-positive endothelial EVs were only detected in a subset of VS and not in HC. CD31
+
CD41a
+
double-positive platelet EVs were increased in VS compared with HC (894 (580) vs. 416 (128), p=0.004).

Conclusion:
This proof of concept study discovered a distinct EV pattern that correlates with a diagnosis of VS, suggesting that cell- and vesicle-based assays could be useful biomarkers in evaluating VS. This pattern is different from that found in patients with diabetes mellitus and CVD. Larger studies could seek to elucidate correlations between CPCs and disease activity and identify a unique EV signature for VS with increased risk for CVD.
Abstract 358: Myeloid-specific Il-4 Receptor α Knockout Alters Cardiac Remodeling Post-myocardial Infarction
Authors: Song Jianrui , Vigil Thomas , Chung Yutein , Frieler Ryan , Goonewardena Sascha , Mortensen Richard
Published: June 2021

Introduction:
Cardiac remodeling post myocardial infarction (MI) can be a critical determinant of outcome for patients with MI. Well-contained inflammation results in successful infarct healing while excessive inflammation cause adverse remodeling which leads to heart failure. Macrophages are important participants in inflammation, helping resolve pro-inflammatory reactions and performing reparative processes. Reprogramming macrophages towards a resolving and reparative phenotype is a potential therapeutic approach. We hypothesized that IL4/IL13-induced, alternatively activated macrophages (M2) have an important role in cardiac remodeling post-MI, and we tested this hypothesis in a mouse model of MI using myeloid-specific IL4 receptor α knockout mice (MyIL4RaKO).

Methods:
MyIL4RaKO mice were generated using IL4Ra
flox/flox
;LysM-Cre. MI was induced by ligating the left anterior descending coronary artery. Hearts were cut into 1mm sections, and then stained by tetrazolium chloride for infarct size measurement. Evenly spaced radians were taken through the infarct with the center of left ventricle in 5μm heart sections, and the average infarct thickness was calculated. qPCR was used to determine gene expression. Echocardiography was performed at baseline and 3 weeks post MI.

Results:
Initial infarct size was not affected by IL4Ra knockout but at 1-week post MI, infarct size of MyIL4RaKO mice (16.54 ± 2.433, n=11) was shown significantly smaller than that of FC mice (24.96 ± 2.005, n=15) showing changes in remodeling (p= 0.0129). Changes in remodeling continued and at 3-week post MI, infarct thickness of MyIL4RaKO mice (0.2171 ± 0.01053, n=6) was significantly increased, compared with that of FC mice (0.3508 ± 0.03629, n=8, p= 0.0094). These changes were accompanied by MyIL4RaKO mice also showed lower level of fibrosis markers: Col1A1 and Plod2. A significantly lower ejection fraction was observed in MyIL4RaKO mice (25.46 ± 3.749, n=5) compared with FC mice (37.90 ± 2.309, n=5) at 3 weeks (p= 0.0223).

Conclusions:
Myeloid-specific IL4Ra knockout results in alteration of remodeling, altered fibrosis and decreased cardiac function post MI, although the cardiac hypertrophy did not show significant change.