bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 005: An IL23-IL22 Axis Regulates Intestinal Microbial Homeostasis to Protect from Diet-induced Atherosclerosis
Authors: Fatkhullina Aliia , Peshkova Iuliia , Koltsova Ekaterina , Trinchieri Giorgio , Dzutsev Amiran
Published: March 2019

Atherosclerosis is lipid-driven, chronic inflammatory disease of the arterial wall. While commensal microbiota is involved in the distal regulation of systemic immune responses, how this distant connection influences the development of atherosclerosis and what are the underlying mechanisms remains largely unknown. In a mouse model of atherosclerosis, we found, unexpectedly, that disease was augmented when expression of the otherwise inflammatory cytokine IL23 was ablated. IL23 and its immediate downstream target IL22 restrict atherosclerosis by preventing outgrowth of pro-atherogenic microbiota, as inactivation of IL23/IL22 signaling led to dysbiosis and expansion of bacteria with pro-atherogenic properties, due to defective production of antimicrobial peptides in the intestine. These pro-atherogenic bacteria contributed to elevated serum levels of several pro-atherogenic metabolites, which in turn induced osteopontin (OPN) expression by aortic macrophages. Microbiota transfer from IL23 deficient mice accelerated atherosclerosis, while microbial depletion or IL22 administration reduced aortic osteopontin expression and ameliorated the disease. Overall, our work uncovers IL23-IL22 signaling axis as a key regulator of atherosclerosis that controls diet-induced expansion of pro-atherogenic microbiota, and argues for informed usage of cytokine blockers with regard to cardiovascular side effects driven by microbiota and inflammation.
Abstract 010: Similar Increases in Arterial Shear-rate Evoked by Handgrip Exercise Activate Autophagy and Enos to a Lesser Extent in Primary Endothelial Cells From Older vs. Adult Subjects
Authors: Park Seul Ki , La Salle D Taylor , Cerbie James , Cho Jae Min , Nelson Ashely , Morgan David E. , Trinity Joel D. , Symons J David
Published: March 2019

We reported that genetic or pharmacological suppression of autophagy in bovine and human arterial endothelial cells (ECs) prevents shear-stress induced p-eNOS
S1177
and NO generation (Bharath et al.,
Arterioscler Thromb Vasc Biol
, 2017). To date, one human study indicates an aging-associated repression of EC autophagy is concurrent with arterial dysfunction (LaRocca et al.,
J Physiol
, 2012). We tested the hypothesis that a physiological elevation of arterial shear-rate activates autophagy and p-eNOS
S1177
to a lesser extent in primary ECs from older vs. adult participants. After familiarization with laboratory procedures, flow-mediated vasodilation (%FMD) was observed to be lower (p<0.05) in 4 older (68±5 yr; 3.9±1.3 %) vs. 5 adult (23±3 yr; 7.9±1.1 %) male subjects. Within 1-week, a catheter was placed into the radial artery (RA). ECs were collected 30-min later via j-wire from the RA. Next, subjects performed rhythmic handgrip exercise (RHE) for 60-min that elevated (p<0.05) arterial shear-rate similarly (2.6 ± 0.2-fold) between groups from pre-exercise values. Shear-rate [8V
mean
/brachial artery (BA) diameter] was obtained by assessing BA diameter, BA blood flow velocity (V
mean
), and BA blood flow [(V
mean
π (vessel diameter / 2)
2
x 60)] at 5-10 min intervals using Doppler ultrasound. Compared to pre-exercise values, RHE did not alter heart rate, stroke volume, cardiac output, or arterial pressure (plethysmography) in either group. Immediately following RHE ECs were collected from the RA. Using quantitative immunofluorescence, primary ECs (75 ECs per endpoint per subject) were identified by positive co-staining for VE-cadherin and DAPI via confocal microscopy. Immortalized human arterial ECs processed and stained in parallel served as fluorescence intensity controls. Relative to pre-exercise, ECs from adult but not old subjects at 60-min displayed increased expression of microtubule associated protein light chain 3 (p<0.04), lysosomal associated membrane protein 2a (p<0.01), autophagy-related gene 3 (p=0.06), and p-eNOS
S1177
(p<0.02), and decreased expression of the adaptor protein p62 (p<0.02). These data indicate aging limits shear-induced EC autophagy and eNOS activation in response to functional hyperemia.
Abstract 018: TET2 Loss of Function-driven Clonal Hematopoiesis Promotes Age-related Vascular and Systemic Inflammation and Metabolic Dysfunction in Mice
Authors: Fuster Jose J , Zuriaga Maria A , Polackal Maya , Walsh Kenneth
Published: March 2019

Background:
Acquired mutations in the hematopoietic system and subsequent clonal hematopoiesis are emerging as potential new drivers of atherosclerotic cardiovascular disease. TET2 was the first gene reported to exhibit acquired mutations in blood cells of cancer-free elderly individuals and we recently reported that the clonal expansion of TET2-mutant hematopoietic cells accelerates atherogenesis in mouse models by exacerbating inflammatory responses in the atherosclerotic plaque. The objective of this study was to examine whether TET2 loss of function-driven clonal hematopoiesis also has systemic effects on age-related inflammation and metabolic function beyond its direct actions on the plaque.

Methods:
An adoptive transfer technique was used to introduce a small percentage of Tet2-/- hematopoietic cells in non-irradiated C57Bl/6J mice, which were monitored for 16 months following transfer. Flow cytometry techniques were used to follow the expansion of Tet2-/- cells in the blood.

Results:
Tet2-/- hematopoietic cells expanded progressively over time after adoptive transfer, with a mild myeloid bias. This was paralleled by increased expression of the pro-inflammatory cytokines IL-1β and IL-6 in the white adipose tissue and the aortic wall, as well as by increased serum levels of IL-6. Tet2-/- expansion also led to exacerbated systemic insulin resistance, as revealed by insulin tolerance tests.

Conclusion:
TET2-deficient hematopoietic cell expansion promotes vascular and systemic inflammation, as well as metabolic dysfunction in aging mice. These results support a causal role for TET2 mutation-driven clonal hematopoiesis in age-related cardiometabolic disease, and suggest that somatic TET2 mutations may contribute to atherosclerosis by promoting inflammation both systemically and locally in the atherosclerotic plaque.
Abstract 023: Clearance of ApoC-III Glycoforms Associated With Triglyceride Metabolism
Authors: Yassine Hussein , Kegulian Natalie , Ramms Bastian , Horton Steven , Trenchevska Olgica , Nedelkov Dobrin , Graham Mark J , Gordts Philip L
Published: March 2019

Background:
We previously reported an association between higher abundance of disialylated (apoC-III
2
) over monosialylated (apoC-III
1
) apoC-III and lower plasma triglyceride (TG) concentrations in three independent studies.

Methods:
The goal of this study was to get mechanistic insights in the metabolism of apoC-III
1
and apoC-III
2
. First, the relative abundances of human triglyceride-rich lipoproteins (TRL)-associated apoC-III glycoforms in plasma were assessed by mass spectrometric immunoassay (MSIA) over time after injecting human TRL in wild-type mice, mice lacking hepatic heparan sulfate proteoglycans (HSPG), and mice deficient for both low-density lipoprotein receptor (LDLR) and LDLR-Related Protein 1 (LRP-1). Secondly, the relative abundance of plasma apoC-III glycoforms was assessed by MSIA in 11 participants randomized to volanesorsen (apoC-III antisense drug; n=11) or placebo (n=6) treatment for 13 weeks.

Results:
Kinetic studies revealed that half of TRL-associated apoC-III (total) was rapidly cleared via LDLR and LRP1 (t
1/2
= 16.5 min), and the remaining half via the much slower but higher capacity HSPGs (t
1/2
= 55.5 min). After injection, a significant increase in the relative abundance of apoC-III
2
was observed in HSPG-deficient mice , while no increase in mice lacking LDLR and LRP-1. Clinically, the relative abundance of apoC-III
2
significantly increased (42% increase compared to placebo, p=0.05) and that of apoC-III
1
significantly decreased (15% decrease compared to placebo, p=0.007) in participants randomized to volanesorsen. The decrease in the relative abundance of apoC-III
1
after volanesorsen treatment was strongly correlated with the decrease in TG levels (r=0.63, p=0.006).

Conclusions:
Our results indicate slower clearance of apoC-III-containing particles through HSPGs, which preferentially clear apoC-III
2.
In contrast, apoC-III
2
is less effectively cleared by LDLR/LRP-1 compared to apoC-III
1
. Clinically, the increase in the apoC-III
2
/apoC-III
1
ratio upon antisense lowering of apoC-III might reflect faster clearance of apoC-III
1
as it associates with improved TG levels.
Abstract 026: Apolipoprotein A-I Helical Registry Modulates Lecithin:cholesterol Acyl Transferase Activity
Authors: Cooke Allison L , Morris Jamie C , Melchior John T , Jerome W. G , Street Scott E , Thompson Thomas B , Smith Loren E , Shah Amy S , Herr Andrew , Segrest Jere P , Heinecke Jay W , Davidson W S
Published: March 2019

The structure of apolipoprotein (APOA1), the major protein of high density lipoproteins (HDL), can modulate protein interactions affecting the cardioprotective functions of HDL particles. APOA1 on nascent discoidal HDL is composed of ten, tandem, α-helical repeats arranged in an anti-parallel, stacked ring-structure that encapsulates a lipid bilayer. Previous chemical cross-linking studies suggested that these APOA1 rings may adopt two different orientations, or registries, with respect to each other. These orientations include one where the bulk of APOA1 orients with the 5
th
helix of one molecule opposing the 5
th
helix of the other (5/5 helical registry), and another orientation adopting a 5/2 helical registry. Engineered HDL particles locked in both 5/5 and 5/2 registries, or rotamers, equally promoted cholesterol efflux from macrophages, indicating functional particles. However, the ability of lecithin:cholesterol acyl transferase (LCAT) to esterify cholesterol was increased over WT in the 5/5 rotamer, and dramatically impaired in the 5/2 rotamer (
p<0.001
), despite surface plasmon resonance studies showing that LCAT bound equally to both. Cross-linking experiments indicated that LCAT activity requires a hybrid epitope composed of helices 5-7 on one APOA1 molecule and 3-4 on the other. Thus, the reciprocating nature of APOA1’s structure may represent a thumbwheel mechanism that influences the activity of HDL remodeling factors in plasma.
Abstract 028: Neutrophils Aged Under Oxidative Stress Impede Vascular Repair
Authors: Okwan-Duodu Derick , Liu Wenxue , xu Kai , Shin Eric , Quintana Raymundo A , Joseph Giji , Hansen Laura , Sellak Hassan , Levit Rebecca , Archer David , Taylor W Robert
Published: March 2019

Objectives:
The link between chronic inflammation and poor cardiovascular outcomes has long been appreciated, but the regulation of immune cellular targets to improve cardiovascular health remains a challenge. Mounting evidence suggests that neutrophils, previously considered acute first responder cells, play critical role in chronic inflammation. Following systemic surveillance of tissues, patrolling neutrophils age in the circulation and traffic to the bone marrow, where they undergo macrophage-mediated clearance. Here, we assessed how chronic inflammation and oxidative stress modify neutrophil trafficking and function during ischemia.

Methods:
Neutrophils from the Townes humanized sickle cell (SS) mice - surrogate for chronic inflammation and oxidative stress - were compared to neutrophils from MerTK deficient mice (MerKO), which age in the circulation owing to defective efferocytosis. Hind limb ischemia (HLI) was used as model of vascular injury. LASER Doppler perfusion imaging (LDPI) measured perfusion recovery after HLI.

Results:
The proportion of CXCR4
Hi
CD62L
lo
aged neutrophils in circulation was markedly elevated in the SS (34 ± 4%) and MerKO mice (38 ± 4%) when compared to wild-type mice (7 ± 1.2%). However, in contrast with MerKO neutrophils, aged neutrophils from SS showed impaired phagocytosis, increased capacity to produce reactive oxygen species, and pronounced release of extracellular traps (NETs). In response to HLI, neutrophils accumulated in the MerKO and SS mice, but perfusion recovery on LDPI was only impaired in the SS mice, suggesting differential role of neutrophils aged by oxidative stress. Importantly, the phenotype of SS mice was reversed by treatment with the anti-oxidant N-acetyl cysteine (NAC), which had no impact on perfusion recovery in MerKO mice.

Conclusion:
Although SS and MerKO mice demonstrated elevated numbers of aged neutrophils, our data suggest that chronic inflammation and oxidative stress modifies the function of aged CXCR4
Hi
CD62L
lo
neutrophils to impede vascular repair after ischemia. This specialized subset of neutrophils may be targeted to improve vascular health without compromising overall anti-microbial host defenses
Abstract 032: Somatic Editing of
Ldlr
with AAV-CRISPR for Atherosclerosis Studies
Authors: Jarrett Kelsey E , Doerfler Alexandria M , Hurley Ayrea , Hsu Rachel H , De Giorgi Marco , Li Ang , Pownall Henry J , Lee Ciaran , Bao Gang , Lagor William R
Published: March 2019

Background:
Current methods to study genetic contributions to atherosclerosis are time-consuming and expensive. The
Ldlr
and
Apoe
knockout (KO) models are currently the gold-standard, but require extensive backcrossing to homozygosity and congenic status on the C57BL6/J background. More rapid and efficient methods are needed to investigate the ever-growing list of candidate genes identified through human genetics.The
C
lustered
R
egularly-
I
nterspaced
S
hort
P
alindromic
R
epeats/Cas9 (CRISPR/Cas9) genome editing system is a powerful new tool for gene disruption.

Hypothesis:
Liver-directed somatic disruption of
Ldlr
using an all-in-one AAV-CRISPR vector can generate atherosclerotic lesions in adult mice, thereby avoiding the need to cross to
Ldlr
KO mice.

Methods:
An Adeno-Associated Viral (AAV) vector based on serotype 8 was used to deliver
Staphylococcus aureus
Cas9 (SaCas9) and small guide RNA (gRNA) targeting the
Ldlr
gene (AAV-CRISPR). This vector was compared head-to-head with AAV-mediated overexpression of a human gain-of-function variant of
PCSK9
, a recent model that mimics
Ldlr
KO through promoting degradation of the Ldlr protein. Adult C57BL6/J mice received either:
1
) saline,
2
) AAV-CRISPR, or
3
) AAV-hPCSK9, versus
4
) germline
Ldlr
KO mice. Animals were placed on a Western diet for twenty weeks and followed for changes in plasma lipids, atherosclerotic lesion burden, and editing efficiency.

Results and Conclusion:
Disruption of
Ldlr
with AAV-CRISPR vector was robust, resulting in severe hypercholesterolemia and atherosclerosis on a standard western diet. AAV-hPCSK9 also produced atherosclerotic lesions as expected from previous reports. Lesion burden was slightly lower with AAV-CRISPR and AAV-hPCSK9 relative to germline
Ldlr
KO mice. Unexpected sexual dimorphism was also observed with AAV-CRISPR, with the greatest effects seen in male mice. In summary, our all-in-one AAV-CRISPR vector is a rapid and efficient alternative to the use of
Ldlr
KO mice to study atherosclerosis.
Abstract 033: Differential Effects of Proprotein Convertase Subtilisin/Kexin Type 9 (PCSK9) Production and Function on Microsomal Triglyceride Transfer Protein
Authors: Huang Cecilia , Miles Joshua , Tavori Hagai , Toth Peter P , Toth Peter P , Fazio Sergio
Published: March 2019

Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a circulating protein that plays a key role in the regulation of plasma low-density lipoprotein (LDL) cholesterol levels. PCSK9 binding to the LDL receptor (LDLR) leads to receptor-mediated endocytosis and lysosomal degradation of LDLR. Prior studies have shown that PCSK9 increases production of triglyceride-rich apoB-lipoproteins via up-regulation of lipogenic genes, and that PCSK9 inhibition reduces plasma triglyceride levels. However, the effect of PCSK9 inhibition on hepatic lipogenic genes expression remain unclear.

Using a human hepatocellular carcinoma cell line (HepG2) we show that overexpression of PCSK9 upregulated the expression of several lipogenic genes, including a 1.5-fold increase in ATP citrate lyase (ACLY), a 1.2-fold increase in Fatty Acid Synthase (FAS), a 1.2-fold increase in HMG-CoA-reductase, and a 1.8-fold increase in microsomal triglyceride transfer protein (MTTP), when compared with untransfected HepG2 cells. Overexpression of a gain-of-function mutant of PCSK9 (PCSK9-D374Y) had stronger effects in the same direction: a 2.3-fold increase in ACLY, a 1.8-fold increase in FAS, a 1.6-fold increase in HMG-CoA-reductase, and a 2.1-fold increase in MTTP. In HepG2 cells overexpressing either normal PCSK9 or PCSK9-D374Y, inhibition of PCSK9 function using a monoclonal antibody blocked PCSK9-mediated degradation of LDLR, and reduced expression of SREBP-dependent genes (ACLY, FAS and HMG-CoA-reductase) to levels of untransfected HepG2 cells. Interestingly, while inhibition of PCSK9 function via monoclonal antibody did not affect MTTP gene expression in these cells, inhibition of PCSK9 production via RNA interference reduced MTTP gene expression by 66%.

We conclude that inhibition of PCSK9 production decreases MTTP gene expression, whereas blockade of PCSK9 function (and cellular re-entry) only decreases SREBP-dependent genes expression, but does not affect MTTP gene expression. This study shows that while PCSK9 exerts an influence on genes related to both lipogenesis and lipoprotein assembly, any inhibition of PCSK9 controls SREPB-dependent genes but only inhibiting PCSK9 production controls MTTP gene expression.
Abstract 039: A Small Fragment Derived from von Willebrand Factor Improves Survival in a Mouse Model of Endotoxemia-Induced Disseminated Intravascular Coagulation
Authors: Valladolid Christian , Cirlos Sonya , Martinez-Vargas Marina , Guillory Bobby , Cruz Miguel A
Published: March 2019

Introduction:
Disseminated intravascular coagulation (DIC) is a life-threatening condition that results in wide-spread clot formation and organ failure. Sepsis-induced DIC is associated with a high mortality rate, prompting the need for an effective therapeutic intervention. Our lab has identified the A2 domain of the von Willebrand factor (VWF) protein as a potent treatment for sepsis-induced DIC. Historically, VWF has only been described as a contributor of clot formation; however, our lab has demonstrated that purified recombinant full length A2 domain of the VWF protein can dampen fibrin-rich microthrombi formation in mice with endotoxin. The mode of action appears to be via the interaction with the cell-surface expressed vimentin on endothelium, inhibiting VWF strings formation and thus, platelet adhesion.

Goal:
Since the VWF protein is naturally cleaved in circulation by the enzyme ADAMTS-13 via the A2 domain, one can argue whether the beneficial effect observed with full len gth A2 domain is terminated by the cleavage of ADAMTS-13. We produced recombinant proteins encompassing the sequences of the two proteolytic products of the A2 domain; the A2Nt and A2Ct proteins. I will identify which fragment retains the mode of action of the full length A2 domain in a mouse model of endotoxin-induced DIC.

Methods:
The recombinant A2 and vimentin proteins were produced using the bacterial expression system.

Protein:
protein interactions were determined by optical interferometry. The endotoxin-induced DIC model was achieved by injecting C57B/6 mice with lipopolysaccharide (LPS). Treatment with A2 or its fragments was started approximately 2 hours after LPS insult.

Results:
The A2N t protein has a higher affinity for vimentin than the A2Ct. Treatment of A2Nt in a LPS-induced DIC mouse model was effective in maintaining survival in males, a comparable finding that mimics treatment of mice with full-length recombinant A2. Interestingly, A2Nt showed less survival than full length A2 in females. These findings highlight the potential differences in therapy responses between genders, an occurrence that some studies have described to also occur in humans.
Abstract 040: Accelerated Atherosclerosis and Thrombosis in Jak2v617f Mice
Authors: Wang Wei , Liu Wenli , Wang Ying , Tang Yang , Woods Brittany , Welch Carrie , Cai Bishuang , Ai Ding , Yang Yong-Guang , Silvestre Carlos , Soehnlein Oliver , Tabas Ira , Levine Ross L , Tall Alan R , Wang Nan
Published: March 2019

The mechanisms driving increased athero-thrombotic risk in individuals with JAK2V617F positive clonal hematopoiesis or myeloproliferative neoplasms are poorly understood. This study was to assess atherosclerosis and thrombosis and associated mechanisms in hypercholesterolemic mice with hematopoietic JAK2V617F expression.

Irradiated

Ldlr
-/-

mice were transplanted with bone marrow from WT or JAK2V617F mice and fed a high fat high cholesterol diet (WD). After 7 weeks of WD
JAK2V617F
mice showed increased hematopoiesis, platelet activation, accelerated thrombosis and increased atherosclerosis. Early atherosclerotic lesions showed increased neutrophils, correlating with lesion size and in association with increased rolling and adhesion of neutrophils on endothelial cells in carotid artery. After 12 weeks of WD JAK2V617F lesions were slightly larger than controls but showed additional complexity, with increased necrotic core area, prominent iron deposition and co-staining of erythrocytes and macrophages suggesting erythrophagocytosis. JAK2V617F erythrocytes were more susceptible to phagocytosis by WT macrophages and showed decreased surface expression of CD47, a “don’t eat me” signal. Human JAK2V617F erythrocytes were also more susceptible to erythrophagocytosis. JAK2V617F macrophages displayed increased expression of pro-inflammatory cytokines and chemokines, increased p38 map kinase signaling and increased cleavage and reduced levels of MerTK, a key molecule mediating phagocytosis of apoptotic cells (efferocytosis) in atherosclerotic lesions. Erythrophagocytosis also suppressed efferocytosis. As a result, JAK2V617F lesional efferocytosis was reduced in advanced lesions.

In conclusion, JAK2V617F promotes early lesion formation, arterial thrombosis and increased complexity in advanced atherosclerosis. In addition to increasing hematopoiesis and neutrophil infiltration in early lesions, JAK2V617F caused cellular defects in erythrocytes and macrophages, leading to increased erythrophagocytosis but defective efferocytosis. These changes appear to promote accumulation of iron and increased necrotic core formation which, together with exacerbated pro-inflammatory responses, may contribute to plaque instability.
Abstract 046: Obesity-induced Oxidative Stress in Hematopoietic Stem and Progenitor Cells Allows a Sustained Myelopoiesis and Persistent Inflammation in Mouse Peripheral Artery Disease
Authors: Barman Pijus , Fang Milie , Fantuzzi Giamila , Koh Timothy J , Urao Norifumi
Published: March 2019

Peripheral artery disease (PAD) is linked with obesity-induced complications. Obesity induced by a high-fat diet (HFD) modulates myelopoiesis of hematopoietic stem and progenitor cells (HSPCs) and innate immune responses in a long-term. Here we show that, in a mouse hindlimb ischemia, normal mice transplanted HSPCs from HFD mice exhibit persistent inflammation and impaired healing using three-dimensional magnetic resonance images, compared to mice transplanted HSPCs from mice fed a normal diet (ND). Bone marrow cell population analysis revealed HFD induces expansion of myeloid progenitor cells in steady state and prolonged activity of more primitive HSPCs following hindlimb ischemia. Ex vivo culture of HSPCs revealed that HFD increases monocyte generation by damage-associated toll-like receptor stimulation, suggesting that HFD promotes myelopoiesis in a HSPC-intrinsic manner. This notion was confirmed by adoptive transfer of HSPCs from HFD mice in a mouse after hindlimb ischemia that contribute to monocyte generation in vivo. Mechanistically, HFD-induced oxidative stress is associated with increased histone-3 lysine-4 trimethylation (H3K4me3), an activating histone methylation, in HSPCs potentially through KDM5 histone demethylase, as a reduction of oxidative stress by cyclosporine A or MitoTEMPO decreased H3K4me3 and increased KDM5 activity in HSPCs. In myeloid progeny of HSPCs from HFD mice, we found enhanced gene expressions of inflammatory Tnfa and Il6, and increased H3K4me3-marks in KDM5 targets, compared to those from ND mice. Our results suggest that HFD-induced oxidative stress in HSPCs modulates their myelopoiesis and phenotype of their progeny via epigenetic reprogramming that, in turn, persistent inflammatory monocyte/macrophage response in an acute ischemic event of PAD.
Abstract 048: Platelets Amplify Monocyte and Macrophage Inflammation Propagating Atherosclerosis Progression
Authors: Barrett Tessa J , Zhou Felix , Gorenchtein Mike , Fisher Edward A , Berger Jeffrey S
Published: March 2019

Objective:
Platelet-leukocyte interactions are recognized to play a central role in the initiation of atherosclerosis and are hypothesized to propagate lesion inflammation. Clinical studies have found that platelet activity is associated with atherosclerosis; however, how they contribute to sustained lesion development remains to be fully elucidated. In this study, we sought to understand the effect of platelets on monocyte and macrophage (Mθ) inflammatory phenotype and trafficking during plaque development.

Methods and Results:

Ldlr
-/-

mice were fed a western diet for 7 weeks prior to platelet depletion (α-CD42b) for 2 weeks. Platelet depletion had no significant effect on monocyte count, yet, resulted in a significant reduction to the: (1) ratio of Ly6
hi
:Ly6C
lo
circulating monocytes, (2) monocyte surface expression of CD11b, and (3) expression of proinflammatory transcripts (
Ccl2
,
iL-1β
,
Cd11b
). Morphometric and immunohistological analysis of plaques demonstrated that platelet deficient mice have significantly reduced lesion area (31% reduction, p=0.03) and reduced Mθ content (38% reduction, p=0.01), compared to platelet competent mice.
In vivo
cell trafficking assays revealed that platelets play a critical role in the recruitment of circulating monocytes to lesions (58% reduction of recruitment in platelet deficient mice, p=0.04), but do not alter plaque macrophage egress. Furthermore, platelets do not alter the proliferative capacity of plaque Mθs, yet, do promote Mθ survival as assessed by cleaved caspase-3 staining, and skew them towards a proinflammatory phenotype (decreased plaque collagen and mannose receptor content, p=0.03 and p=0.05) that have a reduced capacity for efferocytosis.

Conclusion:
This study highlights for the first time novel roles of platelets to immunomodulate monocytes and Mθs, key cell types central to atherogenesis. We find that platelets are critical mediators of circulating monocyte inflammation and subsequent plaque Mθ inflammation and trafficking. The capacity of platelets to skew leukocytes towards a proinflammatory phenotype represents a significant driver of unresolved plaque inflammation, thereby contributing to atherosclerosis progression.
Abstract 053: Differential Effects of LRP1 in AngII-induced Ascending Aortic PathologiesBetween Male and Female Mice: Lack of Association With Elastin Fragmentation
Authors: Wright Bradley C , Sawada Hisashi , Moorleghen Jessica J , Howatt Deborah A , Rateri Debra L , Majesky Mark W , Daugherty Alan
Published: March 2019

Objective:
Low-density lipoprotein receptor-related protein 1 (LRP1), a transmembrane protein, is important in maintaining elastin fiber integrity of the aortic wall. Smooth muscle cells (SMCs) of the ascending aorta are composed of the inner medial layers from the cardiac neural crest and the outer layers from the second heart field (SHF). LRP1 depletion in SMCs of male mice augments angiotensin II (AngII)-induced ascending aortic dilation and rupture attributed specifically to SMCs of SHF origin. The purpose of this study was to determine whether depletion of LRP1 in SHF-derived SMCs (SHF-SMC LRP1) has differential effects on AngII-induced aortic dilation, rupture, and elastin fragmentation between male and female mice.

Methods and Results:
Female LRP1 floxed mice were bred to male LRP1 floxed mice with Mef2c-Cre transgene to generate SHF-SMC LRP1 +/+ and -/- mice. Male and female mice at 12-14 weeks of age were infused with either saline or AngII (1,000 ng/kg/min) for 28 days (n = 12-31). Ascending aortic diameter (AoD) was measured by ultrasound, rupture was determined by necropsy, and elastin fragmentation was assessed by Movat’s staining. In male mice of both genotypes, AoD and elastin breaks were increased by AngII infusion, and AoD was positively correlated with elastin fragmentation (r
2
= 0.48, p < 0.001). SHF-SMC LRP1 deletion augmented aortic dilation, rupture rate, and elastin fragmentation in AngII-infused male mice. In females, AngII infusion increased AoD in SHF-SMC LRP1 -/- mice, but not in their wild type controls. AngII-induced elastin fragmentation did not differ between SHF-SMC LRP1 +/+ and -/- female mice, and no correlation between AoD and elastin fragmentation was detected (r
2
= 0.01, p = 0.88). Despite the extent of AngII-induced elastin fragmentation that was equivalent to that in males, aortic rupture is lower in female than in male LRP1 deleted mice (9 vs 38%, p = 0.02).

Conclusion:
In male mice, AngII infusion promotes dilation and rupture of ascending aorta that is augmented in SHF-SMC LRP1-/- mice and positively associated with elastin fragmentation. However, AngII only promotes ascending aortic dilation in female mice with SHF-SMC LRP1 deletion, despite elastin fragmentation being equivalent between SHF-SMC LRP1 +/+ and -/- mice.
Abstract 095: Comparison of Various Anthropometric Indices for the Identification of a Predictor of Incident Hypertension
Authors: Choi Jung Ran
Published: March 2019

We compared the predictive capability of weight, waist circumference (WC), waist-to-height ratio (WHtR), waist-to-hip ratio (WHR), body mass index (BMI), body roundness index (BRI), and a body shape index (ABSI) to identify incident hypertension, and to determine whether any of these indices may be used as a better single predictor of incident hypertension. A total of 1,718 populations aged 39-72 years were collected in a longitudinal study. Logistic regression models were used to evaluate various anthropometric indices as significant predictors of hypertension. During 2.8 years of follow-up, 185 new cases of hypertension (10.8%) were reported. The BRI and ABSI were significantly higher in the participants who had developed hypertension than in those who had not (4.15±1.01 vs. 3.57±1.03, 0.80±0.04 vs. 0.78±0.05; respectively,
p
<0.001). After adjusting for confounding variables, logistic regression analysis indicated that participants within the highest quartile of WC and WHtR were 4.79 and 4.51 times more likely to have hypertension than those within the lowest quartile (OR 4.79, 95% CI 2.49-9.20 vs. OR 4.51, 95% CI 2.41-8.43, respectively, p<0.0001); in contrast, no such correlation was found for BMI, WHR, BRI, and ABSI. WC (AUC: 0.672) showed a more powerful predictive ability for hypertension (p<0.0001) than BMI (AUC: 0.623), and an equal predictive power for hypertension as WHtR (AUC: 0.662) and BRI (AUC: 0.662) in the general population. We concluded that WC and/or WHtR but not BMI, showed superior prediction capability compared to WHR, BRI and ABSI, for determining the incidence of hypertension in a community-based prospective study.
Abstract 097: Diagnosis of Atherosclerotic Coronary Heart Disease using Plasma Extracellular Vesicle Proteins
Authors: de Kleijn Dominique P , Waissi Farahnaz , Dekker Mirthe , Bank Ingrid , Schoneveld Arjan , Mosterd Arend , Timmers Leo
Published: March 2019

Atherosclerosis is the underlying syndrome of Cardiovascular Disease (CVD). Ischemic Heart Disease and Stroke are cardiovascular events both due to atherosclerosis that are the number 1 and 2 cause of death in the world and expect to increase especially in Asia.

Ischemic heart disease (IHD) comprises 3 entities: stable coronary artery disease (SCAD), unstable angina (UA) and myocardial infarction (MI). Despite its large numbers, diagnosis IHD is challenging, as many patients present with atypical symptoms with women have a different symptom sensation than men. Troponins are the main diagnostic tool for detection of MI. Blood biomarkers for SCAD (typically causing stable angina) and UA, however, are not available. These diagnoses frequently require hospital visits/admissions for time-consuming and costly (non)invasive tests.

Extracellular vesicles are composed of a lipid bilayer containing cytosolic, membrane and nuclear molecules derived from the cell and cell compartments of origin and released by the cell into extracellular biofluids. Using proteomics, we identified a plasma extracellular vesicle protein signature of inflammatory and coagulation proteins in different plasma vesicle subpopulations that can identify UA and SCAD patients. This signature was verified for UA in 60 acute chest pain patients (AUC 0.93), and for SCAD in 60 chest pain patients scheduled for heart perfusion imaging (AUC 0.86).

This shows that plasma extracellular vesicle proteins can be used for early diagnosis of SCAD and UA and underlines the biological role for extracellular vesicles in atherosclerosis.
Abstract 101: Noninvasive Venous Waveform Analysis (NIVA) for Volume Assessment
Authors: McCallister Reid , Alvis Bret , Eagle Susan , Hocking Kyle , Brophy Colleen
Published: March 2019

Background:
Current approaches to determine changes in intravascular volume are invasive, expensive, and of limited accuracy. Non-Invasive Venous Waveform Analysis (NIVA) uses a unique physiologic signal (the venous waveform) to obtain real-time measurement of intravascular blood volume. A prototype NIVA device was developed to assess blood volume in a porcine hemorrhage/resuscitation model.

Methods:
The NIVA device was placed on a branch of the femoral vein of anesthetized pigs. Swan ganz catheters obtained pulmonary capillary wedge pressure (PCWP) and cardiac output (CO). Pigs were hemorrhaged in 100 mL increments to a total blood loss of 400mL, blood was re-infused and crystalloid was administered in 1 L increments to “over” resuscitate. Fast Fourier transformation and a proprietary weighted algorithm was used to derive a NIVA score (LabChart, ADInstruments, Colorado Springs, CO). Statistical analyses (linear regressions) were performed using GraphPad Prism (GraphPad Software Inc, La Jolla, CA).

Results:
NIVA correlated with absolute PCWP in experimental hemorrhage and over resuscitation in the porcine model (R=0.76, n=10, Figure 1). The correlations of
change
in NIVA and
change
in PCWP with volume changes were higher than the correlation of change in cardiac output with volume change (Table 1).

Conclusions:
NIVA correlated with PCWP, and represents a novel physiologic signal to interrogate for non-invasive assessment of volume status, that may be useful for perioperative management. This study provides proof of concept for the NIVA prototype device and algorithm in measurement of intravascular volume.
Abstract 102: Semi-Automatic Measurement of External and Luminal Diameter Predicts the Four-Year Prognosis of Small Abdominal Aortic Aneurysms
Authors: Bogdanovic Marko , Lindquist Liljeqvist Moritz , Hultgren Rebecka , Gasser Christian , Roy Joy
Published: March 2019

Objective:
Predicting which abdominal aortic aneurysms (AAA) will require surgery and which may remain stable remains a challenge. We set out to evaluate whether semi-automatic diameter measurements and finite element analysis (FEA) might predict the four-year prognosis of AAAs more precisely than standard diameter measurements.

Methods:
From a retrospective dataset of 97 patients with aneurysm baseline diameter of 40-50 mm, 39 AAAs remained below 55 mm in diameter for four years whilst 58 expanded beyond 54 mm within four years, or were treated surgically. Standard diameters, measured by radiologists or vascular surgeons, were recorded at baseline and at follow-up. Maximal external and luminal diameters as well as total and luminal volume were semi-automatically re-measured from 3D models based on CT images. FEA, estimating the peak wall rupture index (PWRI), was subsequently performed. Further, in 94 patients from the dataset, standard diameter growth rates between baseline and follow-up were calculated.

Results:
Aneurysms that would require surgery within four years were identified with 100% specificity by semi-automatic diameter, n=13 (22%), luminal diameter, n=14 (24%), PWRI, n=7 (12%), and luminal volume, n=5 (8.6%). Neither standard diameter nor total volume could reach 100% specificity. AAA diameter growth rate correlated with baseline semi-automatic diameter (r=0.39, p=9.2e-5), luminal diameter (r=0.29, p=0.005) and luminal volume (r=0.23, p=0.027) but not with PWRI (r=0.17, p=0.094), total volume (r=0.16, p=0.13), nor standard diameter (r=0.14, p=0.17).

Conclusion:
In AAAs with small diameters, precise, semiautomatic measurement of the maximal external and luminal diameter is able to specifically identify aneurysms requiring surgery within four years.
Abstract 104: Aortic Dilation and Elasticity in Thoracic Aortic Aneurysm in a Marfan Model is Sexually Dimorphic
Authors: Chen Jeff Z , Sawada Hisashi , Moorleghen Jessica , Daugherty Alan
Published: March 2019

Objective:
Meta-analysis of data derived from Marfan patients in the Genetically-Triggered Thoracic Aortic Aneurysms and Cardiovascular Conditions (GenTAC) consortium revealed that thoracic aortic aneurysms occur more frequently in men than in women. To determine the characteristics of sexual dimorphism in experimental thoracic aortic aneurysms of a Marfan model, we determined ascending aortic and aortic root morphology in male and female fibrillin-1 haploinsufficent (FBN1 C1041G/+) mice using ultrasonography.

Method and Results:
Aortas from 4 to 6 week old male and female FBN1 C1041G/+ mice and their wild type littermates were assessed by high frequency ultrasonography. Mice were anesthetized and maintained at a heart rate of 450-550 beats per minute. Ultrasound images were captured using a Vevo 3100 instrument with a 40 MHz tranducer. The maximal luminal diameter of ascending aortic between the sinotubular junction and the innominate artery, and the aortic root diameter at the sinus of valsalva, were measured in mid-systole and end-diastole by two independent observers who were blinded to mice identity. FBN1 C1041G/+ mice demonstrated greater ascending aortic dilation in male mice compared to female mice, whereas no difference in aortic root diameter were detected between male and female wild type littermates. In addition, modest ascending aortic dilation was detected before aortic root dilation in FBN1 C1041G/+ mice. Ascending aortas from FBN1 C1041G/+ mice also demonstrated loss of elasticity as evidenced by reduced expansion in systole. Interestingly, this difference was detected much earlier than expected.

Conclusion:
The fibrillin-1 haploinsufficent model of Marfan syndrome associated thoracic aortic aneurysm demonstrate sexual dimorphism in ascending aortic dilation. Loss of elasticity in the ascending aorta is also an early manifestation in this thoracic aortic aneurysm model.
Abstract 105: Increased Circulating Trimethylamine N-oxide (TMAO) Augments the Incidence of Abdominal Aortic Aneurysm in Low Penetrant C57BL/6J Mice
Authors: Conrad Kelsey , Jones Shannon , Helsley Robert , Schugar Rebecca , Wang Zeneng , Hazen Stanley , Brown Mark , Owens A. Phillip
Published: March 2019

Background:
The gut microbiota is a metabolically active endocrine organ critical to the maintenance of cardiovascular health. Dietary sources of choline are metabolized by microbial enzymes to form trimethylamine (TMA). Metabolism by the host hepatic enzyme flavin-containing monooxygenase 3 (FMO3) converts TMA to the pro-inflammatory molecule trimethylamine N-oxide (TMAO). Human clinical trials have correlated high levels of circulating TMAO to an increased risk of cardiometabolic diseases. However, this meta-organismal pathway has not been evaluated in the context of abdominal aortic aneurysm (AAA). The objective of this study was to determine the effects of a high choline diet on the development of AAA.

Methods:
C57BL/6J male (n=20) and female (n=20) mice were fed either a standard chow control diet (n = 10 each sex) or a choline-rich diet (1%; n = 10 each sex) for 5 weeks. After 1 week of diet, basal abdominal ultrasounds were performed and angiotensin II (AngII; 1,000 ng/kg/min) was infused for 28 days via implantation of osmotic minipumps. Termination ultrasounds were performed on day 27 and mice were sacrificed on day 28. Aortas were harvested for evaluation of aneurysm progression and plasma was analyzed for the metabolites TMA, TMAO, and choline. To determine whether TMAO was elevated in human patients with AAA, plasma samples from participants with fast growing AAAs (n = 85), slow growing AAAs (n = 84), and normal (non-aneurysmal) aortas (n = 115) were analyzed for plasma TMAO levels via liquid chromatography tandem mass spectrometry (LC-MS/MS).

Results:
Administration of a choline-rich diet augmented the incidence (P < 0.02) and aortic diameter (P < 0.001) of AAAs in both male and female mice versus placebo-fed mice. Plasma levels of TMA, TMAO, and choline were significantly elevated in choline-fed mice versus normal chow (P < 0.05). Importantly, circulating levels of plasma TMAO were significantly elevated in a step-wise fashion with the rate of aneurysm growth versus non-aneurysmal control patients (fast growing > slow growing > normal patients; P < 0.001).

Conclusions:
Our results indicate increases in circulating TMAO augments the growth status of aneurysms in human patients and the incidence of AAA in a low penetrant C57BL/6J mouse model.
Abstract 110: Exogenous Vasohibin-2 Does Not Influence Angiotensin II-induced Abdominal Aortic Aneurysms Formation in Either Normolipidemic or Apolipoprotein E-Deficient Mice
Authors: Otaka Nozomu , Uchida Haruhito A , Hada Yoshiko , Takeuchi Hidemi , Kakio Yuki , Okuyama MIchihiro , Umebayashi Ryoko , Tanabe Katsuyuki , Sato Yasufumi , Wada Jun
Published: March 2019

Objective:
Chronic angiotensin II (AngII) infusion promotes both ascending (TAAs) and abdominal aortic aneurysms (AAAs) in mice. Previously, we demonstrated that exogenous vasohibin-2 (VASH-2) exacerbated AngII-induced TAAs in normolipidemic mice. The purpose of this study was to examine whether exogenous VASH-2 influenced AngII-induced AAAs in mice.

Methods and Results:
In the initial study, male C57BL/6J mice (10 weeks old) were injected with VASH2 or LacZ expressing adenovirus (Ad; 7.5 x 109 vp/100 μ L) via tail vein at 2 week intervals. One week after the first injection, subcutaneous infusion of AngII (1,000 ng/kg/min) by mini osmotic pumps was initiated for 3 weeks. Consequently, mice were divided into 2 groups: AngII + Ad VASH2 in C57BL/6J mice (n=22) and AngII + Ad LacZ in C57BL/6J mice (n=21). VASH-2 overexpression had no effect on systolic blood pressure, heart rate, body weight, or serum total cholesterol concentrations. Exogenous VASH-2 did not affect
ex vivo
measurements of maximal diameters of abdominal aortas (AngII + Ad VASH2; 1.36 ± 0.39 mm, AngII + LacZ 1.34 ± 0.24 mm, n.s.) in AngII-infused mice. In a subsequent study, male apolipoprotein E-deficient (apoE-/-) mice (9 to 13 weeks old) were injected with VASH2 or LacZ as described for C57BL/6J mice. Consequently, mice were divided into 2 groups: AngII + Ad VASH2 in apoE-/- mice (n=14) and AngII + Ad LacZ in apoE-/- mice (n=14). Similarly, overexpression of VASH-2 had no effect on systolic blood pressure, heart rate, body weight, or serum total cholesterol concentrations in apoE-/- mice. Furthermore, exogenous VASH-2 did not affect
ex vivo
measurement of maximal diameter of abdominal aorta (AngII + Ad VASH2; 1.70 ± 0.61 mm, AngII + Ad LacZ; 1.61 ± 0.43 mm, n.s.) in AngII-infused apoE-/- mice.

Conclusion:
Despite our previous demonstration of the effects on TAA, exogenous VASH-2 did not influence AngII-induced AAA formation in either normolipidemic or apoE-/- mice.
Abstract 113: Thoracic Aortic Aneurysms Associated With Bicuspid Aortic Valve Have Altered MicroRNA Expression
Authors: Portelli Stefanie S , Robertson Elizabeth N , Padang Ratnasari , Kekic Murat , Lai Donna , Bannon Paul G , Hambly Brett D , Jeremy Richmond W
Published: March 2019

Bicuspid aortic valve (BAV) is the most common congenital heart malformation, affecting 1-2% of the population. BAV is caused by fusion of two of the three aortic valve leaflets in the heart during foetal development. Progressive dilatation of the thoracic aorta, resulting in aneurysm (TAA), develops in approximately 50% of individuals with BAV, and can be fatal. The cause of BAV-TAA is undetermined, however, current understanding suggests that it is a complex interplay between genetic susceptibility and the abnormal haemodynamic stress generated by the malformed valve. Changes in the expression of post-transcriptional genetic modifiers, such as microRNA (miRNA), can alter protein expression and disrupt normal tissue function, contributing to disease. miRNA are also viable therapeutic targets with demonstrated success in disease treatment. We hypothesised that there is altered expression of miRNA in BAV-TAA tissue that contributes to TAA development. Total RNA was isolated from aortic tissue of 11 patients with BAV-TAA (Age 54 ± 12 years; M:F 10:1), and 6 normal controls with no aortic disease (Age 35 ± 18 years; M:F 4:2). RNA was reverse transcribed with primers for 818 different miRNA and qPCR was performed using Taqman® OpenArray® Human miRNA Panels. A significant difference in miRNA expression was defined as a >±2 fold change (p<0.05) compared to control. In BAV-TAA tissue, miR-214, miR-497, miR-22, miR-151-3p and miR-24 were significantly decreased. These miRNA have been previously implicated in the regulation of biological pathways relevant to BAV-TAA, including vascular smooth muscle cell (VSMC) injury and apoptosis, oxidative and shear stress, vascular relaxation, VSMC phenotype and TGF-beta signalling. Therefore, these miRNA may contribute to BAV-TAA pathogenesis, either as part of a compromised genetic background, or in relation to haemodynamic stress arising from BAV. This study has identified a novel expression profile of miRNA in BAV-TAA, and provides promising candidates for further exploration and treatment.