bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 159: Endothelial Fatty Acid-binding Proteins Contribute To The Pathogenesis Of Pulmonary Hypertension
Authors: Dai Zhiyu , Liu Bin , Yi Dan , Ramirez Karina , Xia Xiaomei , Rafikov Ruslan , Dejesus Vinicio A , Lemay Sarah-Eve S , Bonnet Sebastien , Boucherat Olivier
Published: July 2024

Pulmonary arterial hypertension (PAH) is a devastating disease characterized by obliterative vascular remodeling and persistent increase of vascular resistance, leading to right heart failure. Understanding the cellular and molecular mechanisms will help develop novel therapeutic approaches for PAH patients.

Objectives:
To determine whether upregulation of fatty-acid binding protein 4 and 5 (FABP4/5) is critical in pathogenesis of PAH.

Methods:
FABP4/5 expression was examined in PA endothelial cells (PAECs) and lung tissues from patients with PAH. Plasma proteome analysis was performed in human PAH samples. Echocardiography, hemodynamics, histology, and immunostaining were performed to evaluate the lung and heart PH phenotypes in

Egln1
Tie2Cre

(CKO) mice and

Egln1
Tie2Cre
/Fabp4-5
-/-

(TKO) mice.

Results:
Both FABP4 and FABP5 were highly induced in ECs of
CKO
mice and PAECs from IPAH patients. Plasma levels of FABP4/5 were upregulated in IPAH patients and directly correlated with severity of hemodynamics and biochemical parameters. Genetic deletion of
Fabp4-5
in
CKO
mice caused a reduction of right ventricular systolic pressure (RVSP) and RV hypertrophy, attenuated pulmonary vascular remodeling and prevented the right heart failure.
Fabp4/5
deletion also normalized EC glycolysis, reduced ROS and HIF-2α expression, and decreased aberrant EC proliferation in
CKO
lungs.

Conclusions:
PH causes aberrant expression of FABP4/5 in pulmonary ECs which leads to enhanced EC glycolysis and hyperproliferation, contributing to the accumulation of arterial ECs and vascular remodeling and exacerbating the disease.
Abstract 2027: Enhancing Cell-based Therapies With Peptide-modified Thermoresponsive Citrate-based Biomaterials For Treatment Of Critical Limb Ischemia
Authors: Wang Huifeng , Dang Caitlyn , Rivera-Bolanos Nancy , Tong Ophelia , Wang Xinlong , Riter Rebecca L , Duan Chongwen , Zhang Xiaomin , Haney Chad , Ameer Guillermo A , Jiang Bin
Published: July 2024

Introduction:
Critical limb ischemia (CLI) causes severe morbidity and mortality with limited treatment options. Cell-based therapy has potential to advance treatment outcomes but faces challenges in cell survival and engraftment during delivery. We hypothesize that an antioxidant niche that promotes cell adhesion and spreading will improve cell engraftment and survival rate, with the goal of limb salvage through improved perfusion.

Methods:
Thermoresponsive poly(polyethylene glycol citrate-co-N-isopropylacrylamide) (PPCN) was synthesized and conjugated with laminin-derived (A5G81) or VEGF-mimic (QK) peptides. Human umbilical vein endothelial cells (HUVECs) expressing human sodium iodine symporter (hNIS) were mixed with materials and intramuscularly injected into nude mice hindlimbs and monitored with Single Photon Emission Computed Tomography with Computed Tomography (SPECT/CT) for cell survival. Next, a hindlimb ischemia model was used to assess the efficacy of cell delivery by evaluating limb perfusion, tissue loss, functional impairment and histological examination.

Results:
Cells delivered with PBS or PPCN demonstrated poor cell survival, with over 90% cell loss by day 7 (
Figure 1a
). In contrast, cells delivered with PPCN-A5G81 and PPCN-QK exhibited robust SPECT signal levels for up to four weeks (
Figure 1a,b
). In the hindlimb ischemia model, materials or cells alone did not demonstrate improvement in any of the parameters over the PBS control. However, cells delivered with PPCN-A5G81 and PPCN-QK showed significantly higher blood perfusion (
Figure 1c-e
), with improved tissue salvage and motor functions (
Figure 1f-i
) for both male and female mice.

Conclusion:
PPCN modified with A5G81 and QK promotes cell spreading, proliferation, survival, and tissue regeneration, with potential for more effective cell-based therapies for CLI.
Abstract 2033: Bone Marrow-Derived Macrophage Extracellular Vesicles From Diabetic Mice Suppress Angiogenic Properties In Endothelial Cells
Authors: Nimma Ramesh , Lopez-Diez Raquel , Kumar Vikas , Uribe-Echevarria Zubizarreta Veronica , Yepuri Gautham , Vakil Raveena , Bhallamudi Sangeeta , Erdjument-Bromage Hediye , Neubert Thomas , Ramasamy Ravichandran , Schmidt Ann Marie M
Published: July 2024

Introduction:
Peripheral artery disease (PAD), increased in diabetes, leads to mortality and disability. Diabetes promotes a proinflammatory macrophage phenotype that suppresses repair. Macrophage-derived extracellular vesicles (EVs) have been associated with tissue damage. We hypothesize that diabetes perturbs macrophage EV cargo, thereby contributing to impaired repair responses to PAD/ischemia.

Methods:
Unilateral ligation of the left femoral artery (FAL) was performed to induce hind limb ischemia (HLI) in diabetic Akita (C57BL/6-Ins2
Akita
/J) vs. non-diabetic (NDM, C57BL/6J) male mice. Murine BMDMs were grown in Exo-free medium. Ultracentrifugation was used to isolate EVs. Western blotting, transmission electron microscopy (TEM) and ZetaView nanoparticle tracking analysis were used to characterize EV size, number, concentration, and morphology. We profiled EV microRNA (miRNA) and protein cargos from Akita vs. NDM BMDM mice.

Results:
BMDMs-EV size was similar in Akita and NDM mice (diameter, 20-200nm). TEM showed cup-shaped morphology. Western blot confirmed known positive and negative control exosomal markers. Comparing Akita vs. NDM BMDM EVs, a total of 3277 proteins were identified; 821 proteins were significantly differentially expressed. Enrichment analysis highlighted pathways predominantly related to oxidative stress, insulin signaling, actin cytoskeleton, CLEAR, lipid metabolism, phagosome maturation, RNA binding and DNA replication. A total of 912 EV miRNAs were identified; 39 were significantly up-regulated and 19 were down-regulated. Preliminary analysis links a number of them to endothelial function and angiogenesis.
In vitro
, endothelial tube formation was impaired in human iliac artery endothelial cells treated with diabetic vs. NDM BMDM EVs. BMDM treated with diabetic vs. WT EVs displayed increased expression of inflammatory genes (TNF, IL-6,) and reduced expression of mitochondrial complex I and complex II proteins.

Conclusion:
We identified microRNAs and distinct proteins from diabetic EVs as potential mediators of impaired endothelial function and resolution of inflammation. Targeting macrophage EVs composition might identify therapeutic approaches to prevent and treat PAD in diabetes.
Abstract 2038: Statin Targeted Treatment Against Intimal Hyperplasia Using Unique Chitosan-PLGA Nanoparticles
Authors: Penton Ashley , Poland Gloria Grace , Cabe Maleen , Langert Kelly , Maier Kristopher , Gahtan Vivian
Published: July 2024

Introduction:
Statins have beneficial pleiotropic effects, including reducing intimal hyperplasia (IH) after intervention for peripheral arterial disease. Using a unique nanoparticle (NP) to locally maximize statin’s pleiotropic benefits, we hypothesized chitosan-functionalized polylactic-co-glycolic nanoparticles loaded with simvastatin (SL-cNPs) would: 1) be taken up by endothelial cells (ECs) and vascular smooth muscle cells (VSMCs); 2) affect EC and VSMC function; and 3) reduce IH compared to systemic simvastatin.

Methods:
cNPs were loaded with simvastatin (2%
w/w
) and encapsulated with lipophilic tracer, DiD.
In vitro
, human aortic ECs and VSMCs (passage 3-5) were cultured with 0.2mg SL-cNPs or basal media (30 min). After 24 hours, uptake of SL-cNPs was assessed by immunostaining and flow cytometry. Functional effect of SL-cNPs was determined using qPCR for mRNA expression of RhoA, RhoB and Ras.
In vivo,
male Sprague-Dawley rats (n=6-9/group) underwent carotid artery balloon injury in the following groups: 1) intraluminal therapy using normal saline control, empty cNPs or SL-cNPs; and 2) oral simvastatin (10mg/kg/day, 7 days pre-operative until sacrifice) plus intraluminal saline or SL-cNPs. Rats were sacrificed (day 14), perfusion fixed, arteries harvested, and IH quantified with morphometric analysis. Student’s t-test and one-way ANOVA was performed (p <0.05 for significance).

Results:
In vitro
, SL-cNPs were readily taken up by ECs (79.6%, 38,606/48,501) and VSMCs (46.4%, 25,774/55,549). SL-cNPs induced a compensatory increase in expression of EC RhoA (1.3-fold) and VSMC RhoA (1.7-fold) and RhoB (5.4-fold) (p<0.05).
In vivo,
oral simvastatin plus intraluminal SL-cNPs reduced IH compared to controls (0.44 ± 0.04 vs 0.34 ± 0.03, p<0.05). Although oral simvastatin and SL-cNPs alone trended toward IH reduction, the effect was not significant (0.35 ± 0.03 and 0.377 ± 0.03, respectively). No difference existed between oral simvastatin and SL-cNP therapy.

Conclusion:
cNPs can be used as a novel vehicle to locally deliver statins to vascular cells. Although only the combination of oral simvastatin and SL-cNPs effectively reduced IH, different routes and/or concentration of SL-cNPs may allow for a more robust effect on IH prevention.
Abstract 2039: Limb Flexion Compresses The Anterior Tibial Artery And Changes Its Take-off Angle In A Sex-Dependent Manner
Authors: Pipinos Margarita I , Amato-Hanner Elizabeth A , Jadidi Majid , MacTaggart Jason N , Kamenskiy Alexey V
Published: July 2024

Introduction:
Peripheral Arterial Disease (PAD) operations and reconstructions continue to fail at high rates, particularly below the knee. During limb flexion, distal superficial femoral and popliteal arteries experience significant deformations that are thought to contribute to disease development, but the biomechanics of the tibial arteries below the knee have not been characterized.

Methods:
Perfused human cadavers (n=15, 81±9 years old, range 60-93 years, 9F/6M) were imaged with their limbs in standing (180°), walking (110°), sitting (90°), and fetal/gardening (60°) positions using Computerized Tomography (CT). Three-dimensional geometries of the below-knee arteries were reconstructed in Mimics software and used to assess the take-off angle of the anterior tibial artery (AT) and the distance from the AT take-off to the nearest AT side branch.

Results:
The AT take-off angle relative to the tibioperoneal trunk increased with more acute limb flexion, from 70±19° in standing to 81±24° in walking, 83±24° in sitting, and 89±27° in fetal/gardening postures (p<0.01 for all postures). The AT take-off angle was larger in females than in males (24% in standing and 35% in walking, sitting, and gardening postures, p <0.01), but there were no differences between left and right limbs of either sex (p=0.66-0.99). The distance from the AT take-off to the nearest AT side branch decreased with more acute limb flexion (p< 0.01 for all postures).

Conclusions:
Limb flexion increases the AT take-off angle in a sex-dependent manner and compresses the AT, potentially contributing to the differences in hemodynamics and arterial mechanics and affecting the development and progression of tibial artery pathology. A better understanding of the below-knee artery biomechanics can provide valuable insights for surgical planning and may lead to improvements in the success rate and durability of PAD repairs.
Abstract 2048: Pla2g12b Is Critical For Intestinal Lipid Absorption In Mice
Authors: Prakash Binu , Gangula Bhargavi , Palaia Thomas , Yadav Pradeep K , Valmiki Swati , Hussain M M
Published: July 2024

We previously showed that Pla2g12bhlb218/hlb218 (Pla2g12bmt/mt) mice injected with viruses expressing mutant PCSK9, fed a western diet had significantly lower plasma lipids and fewer atherosclerotic plaques. Here, we aimed to explain mechanisms for resistance to hyperlipidemia and atherosclerosis in these mice. We performed studies in 3-month-old male and female chow fed Pla2g12bmt/mt and control mice. Dual x-ray absorptiometry did not reveal any significant differences in body weight, fat weight and fat percentage in Pla2g12bmt/mt mice compared to controls. Moreover, there were no significant differences in energy expenditure and food intake between mutant and control mice. However, these mice had significantly lower levels of plasma glucose, insulin, triglyceride, cholesterol, HDL-c, LDL-c, apoB48, and apoB100. In oral fat tolerance test Pla2g12bmt/mt mice showed significant defects in triglyceride absorption. Furthermore, these mice excreted more fecal triglyceride, but not cholesterol. Oil Red O, H & E staining, transmission electron microscopy, and quantifications of lipids in the duodenum indicated accumulation of large lipid droplets and dilated endoplasmic reticulum. These studies suggest that Pla2g12bmt/mt mice have significant defect in intestinal fat absorption. These defects were not due to reductions in apoB and MTP expression. RNA-Seq analysis of duodenal samples revealed upregulation of glycerolipid metabolism, arachidonic acid metabolism, oxidative phosphorylation, and thermogenesis. These pathways were likely upregulated to increase cellular catabolism and oxidation of accumulated lipids. Down upregulated pathways were related to pancreatic secretion, protein digestion and absorption. Despite severe defects in fat absorption, these mice maintain normal growth and physiology. These mice may serve as model to understand metabolism in hypolipidemia disorders associated with significant defects in lipid absorption such as abetalipoproteinemia and hypobetalipoproteinemia.
Abstract 2053: Platelet Depletion Alters Plaque Phenotype In Diet-induced Obesity And Glucose Intolerance During Atherosclerosis Regression
Authors: Laskou Maria , Hannemann Carmen , La Forest Maxwell , de Moura Virginio Vitor , Krautter Franziska , Weinstock Ada , Barrett Tessa , Berger Jeffrey , Fisher Edward A
Published: July 2024

Cardiovascular disease treatment of type 2 diabetes (T2D) patients remains challenging. T2D individuals often exhibit hyperglycemia and obesity, leading to an increased rate of atherosclerotic cardiovascular disease (ASCVD). Activated platelets, prevalent in obesity and diabetes, promote monocyte adhesion to atherosclerotic lesions, crucial for ASCVD progression. However, the role of platelets in atherosclerosis regression in T2D and obesity is unclear. In this study,
we hypothesized that platelet activation due to diet-induced obesity and impaired glucose tolerance (iGT) negatively affects atherosclerosis after lipid lowering
.

Ldlr
-

/-
mice fed a 20-week high-fat, high-cholesterol diet, developed obesity, iGT and hypercholesterolemia. Atherosclerosis regression was induced by a 3-week apoB antisense oligonucleotide treatment, lowering lipids from 1,036 to 143.7 mg/dL (n=23-40, p<0.0001). Concurrently, mice were injected with platelet depletion or control antibody every 3 days. Post-treatment, though plasma cholesterol decreased, iGT and insulin resistance persisted. Interestingly, compared to controls, effective platelet depletion during lipid lowering treatment significantly reduced foam cell (CD68+) content (n=11-12, CD68%=11.6±1.5 vs 7.88±0.9, p=0.04) and increased collagen content (n=10-14, Collagen%=2.67±0.08 vs 4.1±0.6, p=0.04) within plaques. Preliminary data from multiplex imaging revealed a reduction in leukocytes, particularly myeloid cells, in the platelet-depleted mice (n=2-4, plaque cell% of MHCII
+
=24.8±1.3 vs 17.06±0.9 p=0.02). In progress are scRNA-seq analyses of plaque leukocytes to assess, in particular, genes in inflammatory pathways of macrophage subsets in platelet-sufficient and depleted mice. In conclusion, these findings indicate a crucial role for platelets in modulating macrophage responses in atherosclerotic plaques in a regression environment. Further investigation of leukocyte populations in plaques of platelet-depleted mice will deepen our understanding of platelet-mediated mechanisms in plaque regression and potentially help identify platelet-related factors that can be identified in T2D and obese individuals with ASCVD.
Abstract 2058: Endothelial Cell Regeneration By Secretome Obtained From P53 Silenced Bone Marrow Derived MSCs (BMSCs).
Authors: Nandula Seshagiri Rao R , Sen Sabyasachi
Published: July 2024

Introduction:
Diabetes is associated with impaired endothelial function. Studies have demonstrated that endothelial progenitor cells (EPCs) and mature endothelial cells (EC) are susceptible to apoptosis in hyperglycemic environment. Hyperglycemia promotes p53 gene over-expression and p53 gene silencing prior to hyperglycemia exposure prevents cellular senescence and helps vascular regeneration. Previously we have shown that p53 silenced human CD34+ve progenitor cell therapy and even associated cell derived secretome, helped tissue regeneration. Our goal in this project was to test whether conditioned media (CM) obtained from p53 silenced human stem cells (other than CD34+ve cells) can also improve endothelial regeneration exposed to hyperglycemia. We compared CM obtained from adipose and bone marrow derived mesenchymal stem cells on endothelial regeneration

Methods:
Ad-human-P53 (TP53)-shRNA was used to silence P53 and Ad-scrambled-null-shRNA was used as control in bone marrow derived mesenchymal stromal cells. To collect conditioned media BM-MSCs were cultured in exosome free FBS media for 5 days, following transduction with Ad-p53sh or Ad-null, as the case may be. After transduction, cells were grown in media containing 2% exosome free FBS for only 48h and the collected supernatant was concentrated 10-fold to obtain the CM. Endothelial regeneration was tested using endothelial wound healing assay kit ( from Cell Biolabs # CBA-120-T). We compared CM obtained from null and p53sh in adipose and bone marrow derived mesenchymal stem cells on human endothelial cells in normal and high glucose conditions

Results:
Proliferation rate of endothelial cells was enhanced when we used CM from bone marrow derived cells as compared to CM from adipose derived mesenchymal stem cells, by nearly two fold. We are currently analyzing the secretome by mass spectroscopy.

Conclusion:
P53 silencing appears to improve endothelial regeneration on using CM obtained from bone marrow derived stem cells. This maybe secondary to improved mitochondrial function post p53 gene silencing. The CM thus obtained are in sufficient quantities and may have therapeutic role in treating diabetes vascular complications such as diabetic wound healing.
Abstract 2091: A Theory And Computational Framework For Fracture Of Blood Clots
Authors: Liu Dongxu , Nguyen Nhung , Pocivavsek Luka
Published: July 2024

Introduction and Research Questions:
The fracture resistance of blood clots is crucial for physiological hemostasis and pathological thrombosis. However, the mechanism of blood clot fracture is still not well understood. Quantifying the fracturemechanisms is essential for diagnosing and treating bleeding disorders and thrombotic diseases.

Goals and Methods:
This work aims to formulate a thermodynamically consistent, multi-physics theoretical framework fordescribing the time-dependent deformation and fracture of blood clots. This theory concurrentlyincorporates fluid transport through porous fibrin networks, non-linear visco-hyperelastic deformationof the solid skeleton, solid/fluid interactions, mechanical degradation of tissues, gradient enhancementof energy, and protein unfolding of fibrin molecules. The constitutive relations of tissue constituents andthe governing equation of fluid transport are derived within the framework of porous media theory byextending non-linear continuum thermodynamics at large strains. A physics-based, compressiblenetwork model is developed to describe the mechanical response of the fibrin network of blood clots.The kinetic equations of the internal variables are formulated according to the principles ofthermodynamics by incorporating the corresponding energy terms into the total free energy densityfunction. An energy-based damage model is developed to predict the damage and fracture of bloodclots.

Results:
The proposed model is experimentally validated, which shows that this model can accurately capturethe experimentally measured deformation and fracture. The fracture of blood clots subjected tomultiple loading conditions is simulated, and the results demonstrate that the tissue subject to a higherloading rate is stiffer. The damage at the crack tip initiates earlier and propagates faster when theloading time approaches the characteristic time of viscoelasticity and fluid transport.

Conclusions:
This work proposes a physics-based theory and computational framework to describe the fracturebehavior of blood clots. The proposed model is validated by comparing the computational results withexperimental data. The mechanisms of blood clots fracture are analyzed systematically.
Abstract 2111: Nano-based Perivascular Intervention Sustains A Nine-month Long-term Suppression Of Intimal Hyperplasia In Vein Grafts
Authors: Huang Yitao , Shirasu Takuro , Urabe Go , Yodsanit Nisakorn , Stratton Matthew , Joseph Matthew , Xie Ruosen , Wang Yuyuan , Marcho Lynn M , Li Jing , Tang Runze , Park Ki Ho , Wang Bowen , Kent K. Craig , Gong Shaoqin , Guo Lian-Wang
Published: July 2024

Introduction:
Open vascular reconstructions (OVR), such as bypass grafts and dialysis access, are commonly performed to treat cardiovascular and renal diseases. Unfortunately, OVR often fail, largely due to intimal hyperplasia (IH), and there are no clinical methods to prevent the failure. OVR provide accessibility to the graft surface, hence an opportunity for perivascular drug delivery to suppress IH. However, drug release generally lasts for limited time whereas long-term efficacy is important for clinical success.

Goals:
We addressed a prominent question in clinical translation: can IH suppression be realistically sustained for a long term (longer than 6 months) via shorter-term perivascular interventions?

Methods:
We applied Pericelle, a nanoparticle+hydrogel system for perivascular delivery of the IH-mitigating drug rapamycin, to a modified rat vein-graft model that exhibits long-term IH progression.

Results:
IH was reduced throughout 3, 6, and 9 months, as indicated by morphometry (115.58±27.89 to 40.34±5.18 at 9 months) and ultrasonography (Fig 1), although rapamycin release was estimated to be around 3 months. Pericelle also mitigated IH in porcine arteriovenous fistulas (Fig 2).

Conclusions:
Suppression of vein-graft IH can be sustained much beyond drug release; Pericelle provides a potential strategy for reducing OVR failure. We thank Dr. Keith Ozaki and Dr. Prabir Roy-Chaudhury for instructions on models.
Abstract 2112: Silent Killer: The Rare And Potentially Lethal Right Coronary Artery Aneurysm
Authors: khodor maya , Alkowatli hamza , Tashwali Sami
Published: July 2024

Giant coronary artery aneurysms, defined as diameters larger than 20 mm, are exceedingly rare and constitute a life-threatening occurrence. This case involves a patient with a giant right coronary artery (RCA) aneurysm and multivessel coronary artery disease, successfully managed by surgical intervention through quintuple coronary artery bypass grafting (CABG).

A 65-year-old male with a history of hypertension presented with acute-onset chest pain. The initial workup revealed an inferolateral ST-segment elevation myocardial infarction (STEMI). The patient underwent emergency coronary angiography revealing a high-grade stenosis in the proximal and mid left anterior descending coronary artery, along with a remarkably tortuous and enlarged mid to distal RCA aneurysm measuring (21mm wide x 20mm long) and featuring significant post aneurysmal stenosis.

Attempts at percutaneous angioplasty were unsuccessful due to extensive thrombosis in the RCA. The patient continued to experience active chest pain, necessitating the placement of an intra-aortic balloon pump in the right common femoral artery, the insertion of a temporary pacemaker wire to address bradycardia, and the initiation of pressor support for hypotension. Additionally, intravenous heparin and antiplatelet therapy were administered. Finally, the patient underwent a quintuple CABG to address the multivessel disease.

Coronary artery aneurysm is an uncommon yet potentially perilous condition. The selection of appropriate management and intervention is crucial for averting fatal consequences associated with coronary artery aneurysms.
Abstract 2123: Evaluation Of Autonomic Neuropathy In Patients With T2DM By Measuring R-R Interval During Rest, Standing And Deep Breathing
Authors: Katayama Katsuyoshi , Uchida Haruhito A , Sakurabu Yoshimasa , Asakawa Tomohiko , Okamoto Shugo , Onishi Yasuhiro , Uchiyama-Matsuoka natsumi , Takeuchi Hidemi , Umebayashi Ryoko , Takemoto Rika , miyamoto satoshi , Wada Jun
Published: July 2024

Objective:
In patients of diabetes mellitus (DM), heart rate variability (HRV), an index of autonomic nervous system activity, tends to be lower than in healthy individuals. This reduction in HRV is associated with an increased risk of cardiovascular diseases. We aimed to evaluate HRV under varying conditions with novel indicators in patients with DM.

Methods:
The patients were recruited from patients with type 2 DM (T2DM) aged 20 to 79 years old from two facilities from January 2020 to December 2023. The control comprised patients without DM. Patients taking oral corticosteroids, adrenergic agonists and antagonists, and those with malignant tumors were excluded. Heart rate (HR) was measured under varying conditions of rest, standing and deep breathing (inhalation and exhalation alternating every 5 seconds). The R-R interval variation was continuously recorded in each segment of 33 beats. Subsequently, the peak HRs for each load were obtained: minimum HR during rest (mHr), maximum HR during standing (MHs), and minimum HR during deep breathing (mHb). Additionally, the SD-RR for each load was calculated.

Results:
We analyzed a total of 117 subjects, including 75 with T2DM and 42 without DM. The participant demographics were as follows: mean age of 55.4±13.9 years, BMI of 25.9±5.6, eGFR of 71.5±26.1 mL/min/1.73m
2
, HbA1c of 7.31±2.26 %, with 46 % of male, and duration of T2DM in patient group at 8.2±8.3 years. The subjects were categorized into three groups: G1 - non DM with CV-RR≧2, G2 - T2DM with CV-RR≧2, and G3 - T2DM with CV-RR<2.mHr was G1: 64.5±7.4, G2: 69.4±7.7, G3: 75.2±13.1 (beats/minute, p<0.001). G3 is markedly higher than G1 and G2. Furthermore, the HR change ratio from mHr to MHs was: G1: 1.36±0.12, G2: 1.31±0.11, G3: 1.22±0.10 (p<0.001). G3 is markedly lower than G1 and G2. The HR change ratio from mHr to mHb was: G1: 0.99±0.04, G2: 0.96±0.05, G3: 0.96±0.05 (p=0.043), with G3 being markedly lower than G1. The SD-RR during rest was: G1: 26.4±18.6, G2: 28.0±12.5, G3: 10.1±4.4 (p<0.001), with G3 being markedly lower than G1 and G2.

Conclusion:
Assessing R-R intervals under varying conditions of rest, standing, and deep breathing can indicate autonomic nervous system dysfunction among patients with T2DM.
Abstract 2131: Bone Marrow-derived Macrophages May Contribute To Elevation Of Et-1 And Pulmonary Hypertension In Female

Bmpr2
+/r899x

Mutant Mice
Authors: Loya Omar , Erewele Ejehi , Castellon Maricela , Marinho Ygor , Oliveira Suellen Darc
Published: July 2024

Introduction:
Mutation that causes loss of function of bone morphogenetic protein receptor 2 (BMPR2) has been implicated in the pathogenesis of Pulmonary Arterial Hypertension (PAH), a disease that currently has no cure and is characterized by elevated right ventricular systolic pressure and right ventricle hypertrophy. Females experience thrice more incidence of PAH. Elevation in the expression of ET-1, Endothelin-1, has been linked to the development of PAH. It is unclear what is contributing to the elevation of ET-1 observed in female mutants.

Hypothesis:
Macrophage activation in the microvasculature contributes to ET-1 elevation and PH pathogenesis in female

BMPR2
+/R899X

mice.

Materials/Methods:
Lung samples from 12-month-old male and female wild-type and

BMPR2
+/R899X

mutants were used for analysis of the ET-1 expression via Western Blot (WB). Accumulation of CD45+ET-1+ cells in the lungs was evaluated by immunohistochemistry (IHC). Finally, bone marrow-derived macrophages (BMDM) isolated from male and female mutant mice underwent a 24-hour treatment with 100 ng/mL Lipopolysaccharide (LPS) and vehicle (control) and ET-1 expression was measured in the cell lysate by WB.

Results/Discussion:
In exploring the source of ET-1, our initial findings indicated an accumulation in inflammatory perivascular cells. A double-blind analysis of the CD45 pan-leukocyte marker revealed a substantial increase in CD45+ cells in female mutants (4.09 +/- 0.68 and 10.46 +/- 1.15 CD45+ cells/microvessel, male and female, respectively; 8 micrographs/animal; 3 mice/group). Colocalization between CD45+ cells and ET-1 suggests these cells may be a source or contribute to ET-1 clearance in the lungs of female mutants. While the identity of CD45+ cells remain unclear, their morphology suggests they are likely macrophages. Indeed, preliminary results indicate a significant rise in ET-1 expression in BMDM from female mutants at baseline and upon LPS exposure.

Conclusion:
Accumulation of CD45+ cells in female mutant lungs, along with colocalization with ET-1, may contribute to PAH development. Additionally, increased ET-1 expression in macrophages from female mutant mice suggests macrophage activation as a potential factor in ET-1 elevation in female mutant lungs.
Abstract 3000: Mir-146a Deficiency Differentially Regulates Sexual Dimorphism Of Abdominal Aortic Aneurysms In Mice
Authors: Ramesh Nithya , Javidan Aida , Downey Eddie , Subramanian Venkateswaran
Published: July 2024

Background and Objective:
Abdominal aortic aneurysms (AAAs) are permanent aortic dilations with 80% mortality after rupture. AAA exhibits sexual dimorphism, with males at higher risk in both mice and humans. In spite of lower incidence, women acquire aggressive aneurysm growth and rupture at smaller sizes, compared to men. Despite this oddity, mechanisms driving the sexual dimorphism of AAA have not been defined. Previously, we demonstrated that Lysyl Oxidase (LOX) inhibition abolishes sexual dimorphism of AAA in mice. Recent clinical studies showed elevated miR-146a levels, an anti-inflammatory miRNA, in plasma and AAA tissues from patients. In our preliminary studies, we found that aortic miR-146a is significantly lower in females compared to male mice. However, the contribution of miR-146a in the sexual dimorphism of AAAs is unknown. In this study, we examined the role of miR-146a in AAA sexual dimorphism in mice by administering β-aminopropionitrile (BAPN), a LOX inhibitor and Angiotensin II (AngII) concurrently.

Methods and Results:
8 -10 week old male and female C57BL/6J mice that were either miR146a wild type (WT) or deficient (KO; n=18-24/per group) were infused with saline or Ang II (1,000 ng/kg/min) via subcutaneous osmotic mini-pump for 28 days. In addition, mice were co-administered with BAPN (1 mg/ml; 0.15g/kg/d in drinking water) from day 0-14. miR-146a deficiency had no effect on body weight and blood pressure in both male and female mice. AngII+BAPN-administration increased AAA in both male (82%;19/23) and female (72%;18/25) mice compared to saline+BAPN controls. Interestingly, miR-146a deficiency profoundly but significantly increased AAA in males (WT=1.5 ± 0.1 versus KO=2.3 ± 0.1 mm, P<0.05), but significantly decreased AAA in females (WT=1.5 ± 0.1 versus KO=1.2 ± 0.1 mm, P<0.05). By RNA sequencing and qPCR, we identified several differentially expressed genes in the absence of miR-146a, with RFLNA upregulated and ELAVL2 downregulated in male KO mice, and SLC38A5, CCND2, DUSP10 genes upregulated in female KO mice. Ongoing studies aim to delineate the role of these genes in miR-146a-mediated sexual dimorphism of AAA.

Conclusion:
These findings demonstrate that miR-146a plays a critical role in mediating sexual dimorphism of AAA in mice.
Abstract 3023: Association Of Lipoprotein(a) With Adverse Events Following Revascularization For Chronic Limb Threatening Ischemia: A Subgroup Analysis Of The BEST-CLI Trial
Authors: Sullivan Alexander , Kundu Suman , Wells Quinn , Linton Macrae F , Freiberg Matthew S , Beckman Joshua A
Published: July 2024

Background:
Elevated lipoprotein(a) (Lp[a]) levels are implicated in the development of peripheral artery disease (PAD) and in major adverse limb events (MALE) across a range of PAD severity. The association between Lp(a) level and adverse events in patients with chronic limb threatening ischemia (CLTI) remains uncertain.

Research Questions:
Does Lp(a) level associate with MALE and death in patients with CLTI undergoing revascularization?

Goals/Aims:
To assess the association of Lp(a) level with MALE and death in patients with CLTI undergoing revascularization.

Methods:
The BEST-CLI trial was a randomized trial assessing the optimal revascularization strategy in patients with CLTI. A subgroup of patients underwent lipid and Lp(a) assessment during the study period and were included. The primary outcome was the first death or MALE event (above-ankle amputation or major reintervention). Hazard ratios were calculated using Cox proportional hazards models, adjusting for age, sex, statin use, and glomerular filtration rate.

Results:
A total of 192 patients (median [IQR] age, 67.5 [62.1, 74.2] years; 139 men [72.4%]) were included in the cohort. Median Lp(a) level was 26.8 [10.8, 65.1] with normal Lp(a) levels (<30 mg/dL) observed in 104 patients, high levels (30-111 [the 90
th
percentile] mg/dL) in 68 patients, and very high levels (>111 mg/dL) in 20 patients. The primary outcome occurred in 75 (39.1%) patients, including 42 (40.4%), 25 (36.8%), and 8 (40.0%) patients with normal, high, and very high Lp(a) levels. There was no association between Lp(a) and the primary outcome (HR 1.00, 95% CI 0.99-1.02, Figure 1). A significant association between Lp(a) and death (HR 1.03, 95% CI 1.01-1.05) was noted. Results did not vary by randomized revascularization procedure.

Conclusion:
In this cohort, Lp(a) predicts death but not limb events in patients with CLTI. Larger studies are needed to understand the role of Lp(a) in patients with CLTI undergoing revascularization.
Abstract 3065: Biomechanical Forces Can Promote SGK-1-dependent Expression Of Pathologic Matrix Markers
Authors: Barksdale Christian , Silverman Jonah , Fannin Ethan , Gedney Ryan , Mattia Victoria , Figueroa Mario , Xiong Ying , Mukherjee Rupak , Jones Jeffrey A , Ruddy Jean Marie
Published: July 2024

Introduction:
Hypertension is a known predictor of cardiovascular morbidity due to dysfunctional matrix remodeling which promotes aortic stiffness, but achieving normotension does not eliminate this risk. Prior work has demonstrated mechanical sensitivity of the serum and glucocorticoid inducible kinase-1 (SGK-1) in vascular disease through inflammatory signaling. We hypothesize that biomechanical signals promote activity of SGK-1 in VSMCs to drive expression of key pathologic matrix markers (PMMs).

Methods:
Aortic rings were harvested from C57BI/6 WT and smooth muscle specific SGK-1 knockout (SMC-SGK-1KO
+/-
) mice. Rings were placed on a myograph in a physiologic salt solution and exposed to experimentally derived optimal tension (baseline OT, 1.5g) or OT+30% (1.95g) for 6 hrs (n=4-6). WT Rings were homogenized to determine SGK-1 and pSGK-1 abundance by immunoblotting. Replicates were treated ±AngII and ±EMD638683 (EMD; a specific SGK-1 inhibitor). RNA extraction enabled QPCR for select PMMs (Interleukin-6 (IL-6), Cathepsin S (CtsS), Cystatin C (CysC), Tenascin C (TNC)). Statistical comparisons utilized ANOVA with significance set at p<0.05.

Results:
There was a significant increase in pSGK-1 abundance and in the pSGK-1/SGK-1 ratio in the WT aortic rings at OT+30%, confirming that mechanical tension promoted SGK-1 activity. IL-6 and CysC expression was augmented in the WT OT+30% (p<0.05 vs WT OT for each), but returned to baseline in the presence of EMD, implicating that SGK-1 may play a causative role for these effects. While expression of all PMMs increased with OT+AngII or OT+30%+AngII, only IL-6 demonstrated a synergistic effect with biomechanical stimulation. In aortic rings from SMC-SGK-1KO
+/-
animals, the response to mechanical stimulation was blunted for all PMMs, further supporting a role for SGK-1 in pathologic mechanical signaling.

Conclusion:
Mechanical tension on the aortic wall can increase SGK-1 activity, correlating to expression of key mediators of pathologic vascular remodeling. These novel findings suggest that SGK-1 may be a druggable target to reduce aortic stiffness in the setting of hypertensive disease.
Abstract 3071: Ras-related Protein 1a-mediated Regulation Of Proprotein Convertase Subtilisin/kexin Type 9
Authors: Tinsley Brea , Ozcan Lale
Published: July 2024

Background & Aims:
Proprotein convertase subtilisin/kexin type 9 (PCSK9) has recently been identified as a therapeutic target to regulate plasma low-density lipoprotein cholesterol (LDL-C) levels. PCSK9 binds to the LDL receptor (LDLR) and targets it for lysosomal degradation, hindering LDL-C uptake from the circulation. Despite the use of anti-PCSK9 therapies, the molecular mechanisms by which PCSK9 gets regulated are not fully understood. In this regard, we recently discovered that activation of hepatic small GTPase, Ras-related protein 1a (Rap1a), decreases PCSK9 levels and subsequently lowers plasma LDL-C. In addition, the most widely used medicine to lower cholesterol, statins, have been shown to increase circulating PCSK9 levels, which limits their efficacy; however, our understanding of the link between statins and plasma PCSK9 is limited.

Hypothesis & Aims:
As statins block the synthesis of geranylgeranyl isoprenoids (GGPP), which are required for Rap1a activation, we explored the hypothesis that Rap1a is involved in statin-induced expression of PCSK9. We further explored the mechanisms of PCSK9 regulation by Rap1a.

Methods:
Mice were treated orally with statins and geranylgeraniol, the precursor of GGPP by daily oral gavage. Rap1a activator, 8-pCPT-2’-OMe-cAMP, was administered via i.p. injection. Hepatic PCSK9, plasma PCSK9, and LDL-C levels were assayed. For in vitro experiments, primary hepatocytes were isolated from wild-type mice and treated with vehicle, simvastatin, or simvastatin in combination with GGPP or 8-pCPT-2’-OMe-cAMP. PCSK9 levels were measured.

Results:
Statin treatment combined with GGOH lowered plasma PCSK9 and LDL-C in mice (174 ± 25 vs 109 ± 16 ng/mL and 24 ± 1.8 vs 19 ± 1.4 mg/dL, respectively, n=4-5, mean ± SEM, p < 0.05). Rap1a activator, 8-pCPT-2’-OMe-cAMP, also lowered statin-induced PCSK9 in vitro. Mechanistically, we found that Rap1a inhibits the downstream RhoA-ROCK pathway and regulates PCSK9 at the post-transcriptional level.

Conclusions:
In conclusion, our data reveal that Rap1a is a novel regulator of PCSK9 protein and blocking Rap1a prenylation through lowering geranylgeranyl levels contributes to statin-mediated induction of PCSK9.
Abstract 3096: Role Of Circulating Phosphatidylserine In The Procoagulant Activity Of Amniotic Fluid
Authors: Yang Chih-Jen , Vu Helen , Shorey-Kendrick Lyndsey , Scottoline Brian , Shatzel Joseph , Puy Cristina , McCarty Owen J , Lo Jamie
Published: July 2024

Background:
Amniotic fluid (AF) embolism is one of the most catastrophic complications during pregnancy. While studies primarily attributed the procoagulability of AF to circulating tissue factor (TF), the exact mechanism remains unclear.

Aims:
Define the procoagulant mechanisms of AF.

Methods:
Clotting time assays were performed using normal human plasma or plasma depleted of factor (F)XII, XI, X, IX, VII, V, or prekallikrein (PK). Chromogenic substrate assays were used to quantify FX activation by the TF/FVIIa complex and prothrombin activation by FXa. Phosphatidylserine (PS) levels were quantified using a fluorometric kit.

Results:
AF shortened the clotting time of normal plasma from 572±83 to 160±2 seconds; AF-induced clotting times were equivalent for FXII, PK, FXI, FX, FIX, or FV-depleted plasma or in the presence of contact pathway inhibitors as compared to normal plasma. In contrast, the procoagulant effects of AF were diminished or absent for FVII-depleted plasma or in the presence of a blocking anti-TF antibody or low-molecular-weight heparin. In a purified system, AF reduced FX activation by the TF/FVIIa complex. The addition of a blocking anti-tissue factor pathway inhibitor (TFPI) antibody reversed the effect of AF on FX activation. Western blotting confirmed the existence of both TF and TFPI in AF. AF enhanced prothrombin activation by FXa. The addition of annexin V, or lactadherin, which bind to PS, or the use of Gla-domainless prothrombin mitigated the effect of AF on prothrombin activation. PS levels in AF and normal plasma were 811±376 and 10.4±3.8 μM, respectively.

Conclusions:
The procoagulability of AF is independent of the contact pathway while being sensitive to both TF and TFPI levels. Furthermore, PS in AF significantly contributes to its procoagulability. This knowledge is important when developing strategies to lessen the procoagulability of AF without compromising hemostasis in the setting of AF embolism-associated coagulopathy.
Abstract 3143: Computational Modeling Of Non-homeostatic Changes Within An Arterial-venous Fistula System Using In Vivo Validation To Understand Mechanical Causes Of Fistula Failure
Authors: Varsanik M. Alyssa , Thaxton Carly , Li Willa , Dardik Alan , Pocivavsek Luka
Published: July 2024

Introduction:
Arteriovenous fistula (AVF) failure is a clinical problem for end stage renal patients seeking durable long term dialysis access. AVF failure is not fully understood, but linked to neointimal hyperplasia (NH). Understanding the hemodynamic factors that may trigger NH response is important for minimizing this clinical problem.

Methods:
A validated murine AVF model defined the geometry and boundary conditions for the computational model. Anesthetized mice underwent laparotomy and aorta-IVC puncture. Ultrasound specified blood flow velocity within the vessels. In vivo CT angiography scans were segmented using 3D software (Simpleware ScanIP) to establish global geometry which was imported into Xflow, a fluid dynamics simulation software. Velocity, static pressure, and turbulence were averaged along the length of the vein above and below the fistula in this model, a stenotic vein model, and a stenotic fistula model.

Results:
Velocity and turbulence significantly increases in the peri-fistula vein and dramatically increases in systole phase. Velocity, turbulence, and static pressure peak in the stenotic vein. When the fistula is stenosed, native flow is restored.

Conclusions:
The presence of AVF dramatically changes the hemodynamics of the native vein. Increased velocity and turbulence in the peri-fistula vein are significant and peak in systole. Venous stenosis substantially strains the system, demonstrated by the high static pressures proximal to the stenosis. The gradients of high turbulence directly correlate with in vivo histology measurements of neointimal hyperplasia thickness changes that have been previously reported. These results may be used to predict how the underlying hemodynamics within a fistula system preclude biological responses from the tissue. We conclude that non-homeostatic hemodynamics, namely increased venous velocity, turbulence, and static pressure may be a potential trigger for NH and ultimately fistula failure.
Tractography from T1-weighted MRI: Empirically exploring the clinical viability of streamline propagation without diffusion MRI
Authors: Cai Leon Y. , Lee Ho Hin , Johnson Graham W. , Newlin Nancy R. , Ramadass Karthik , Kim Michael E. , Archer Derek B. , Hohman Timothy J. , Jefferson Angela L. , Begnoche J. Patrick , Boyd Brian D. , Taylor Warren D. , Morgan Victoria L. , Englot Dario J. , Nath Vishwesh , Chotai Silky , Barquero Laura , D’Archangel Micah , Cutting Laurie E. , Dawant Benoit M. , Rheault François , Moyer Daniel C. , Schilling Kurt G. , Gore John C. , Landman Bennett A.
Published: July 2024

Over the last few decades, diffusion MRI (dMRI) streamline tractography has emerged as the dominant method for in vivo estimation of white matter (WM) pathways in the brain. One key limitation to this technique is that modern tractography implementations require high angular resolution diffusion imaging (HARDI). However, HARDI can be difficult to collect clinically, limiting the reach of tractography analyses to research cohorts and thus limiting many WM investigations to certain populations and pathologies. As such, a clinically viable tractography solution applicable to wider patient populations scanned as a part of routine care would be of key significance in broadening WM analyses to underfunded or rarer diseases and to the clinical setting. Such a solution would require the ability to perform arbitrary tractography analyses, use only clinical imaging for input, and be open source and widely accessible and implementable. Thus, here we evaluate our recently developed, containerized, and open-source, T1-weighted (T1w) MRI-based deep learning model for streamline propagation. We empirically assess its performance against traditional dMRI-based and established atlas-based approaches in a healthy young population, an aging one, and in those with epilepsy, depression, and brain cancer. In the healthy young population, we find slightly increased error compared to traditional tractography with the deep learning model that falls within the bounds attributable to dMRI variability and is considerably less than the atlas-based approach. Further, seeking to replicate previously published dMRI tractography effects in the remaining cohorts as an initial assessment of clinical viability, we find this model successfully does so in some key cases—particularly in applications that rely on long-range streamlines including those not captured by the atlas-based approach—but importantly not all. These results suggest a deep learning-based approach to tractography with T1w MRI demonstrates promise within the limitations of our definition of clinical viability and especially over atlas-based approaches but requires refinement and more robust consideration of out-of-distribution effects prior to widespread clinical use. We also find these results raise additional questions regarding the differences in image content between dMRI and T1w MRI and their relationship to tractography. Further investigation of these questions will improve the field’s understanding of which features of the brain influence measured tractography effects.