bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 193: First Demonstration that Hepatic Apob100 and Intestinal Apob48 Colocalize with Macrophages in Human Carotid Atherosclerotic Plaques
Authors: Vazquez-Figueroa Jesus G , Sharma Abhinav , Rinehart Sarah , Qian Zhen , Bhatt Kunal , Joshi Parag , Pryor Andrew , Blackman Brett , Teramoto Tamio , Matsushima Teruhiko , Kinoshita Makoto , Voros Szilard
Published: July 2021

Background:
We previously demonstrated the presence and localization of hepatic apoB100 and intestinal apoB48 in human carotid atherosclerotic plaques by immunoperoxidase, immunofluorescence (IF), immunoelectron microscopy, and western blot; however, co-localization of apoB48 and apoB100 with macrophages has not been shown.

Methods:
In 3 patients undergoing carotid endarterectomy (CEA), plaques were stained for apoB100/CD68 and apoB48/CD68 dual stain (DS)-IF. CD68 was used to localize macrophages. Slides were processed for antigen retrieval and incubated with primary antibodies(ApoB100: ab50069-100 rabbit polyclonal antibody recognizing the C-terminal; apoB48:Shibayagi AKHB48E goat monoclonal antibody recognizing the C-terminal; CD68: Santa Cruz Biotechnologies SC-20060 mouse monoclonal antibody) and species-specific secondary antibodies (Invitrogen;A21447,A21235,A21244, or
A11010
).Confocal microscopy was used to visualize DS-IF.

Results:
There is intense CD68 staining throughout the plaques, demonstrating the presence of macrophages(B,E). As previously demonstrated, both apoB100 and apoB48 are present in plaques(A,D). Importantly, while CD68 staining is present throughout the plaque and in areas independent of apoB100 and apoB48, both apoB100 and apoB48 are always co-localized with CD68(C,F).

Conclusions:
This is the first demonstration that both hepatic apoB100 and intestinal apoB48 co-localize with macrophages in human carotid atherosclerotic plaques.
Abstract 269: Collagen Topographical Patterning Modulates Endothelial Cell Morphology, Gene Expression and Function
Authors: Huang Ngan F , Ribeiro Alexandre , Lai Edwina , Fuller Gerald G , Pruitt Beth , Cooke John P
Published: July 2021

Endothelial cells (ECs) exposed to laminar shear stress align in the direction of blood flow and express low levels of adhesion molecules. However, it is unknown whether these biological responses result from shear stress per se, or if they can be recapitulated using topography cues from the underlying extracellular matrix (ECM). We hypothesized that collagen topographical patterning can regulate EC assembly and function in the absence of shear stress. We fabricated collagen-coated polydimethylsiloxane microchannels (30μm wide) to induce EC alignment. When grown on microchannels, the ECs underwent striking re-organization of their actin cytoskeleton (aligned within 10 degrees of the microchannel direction) and their focal adhesions (Fig A). Mimicking the reported EC responses to laminar shear stress, aligned ECs were more quiescent (reduced Ki67 expression), and less adhesive for monocytes (with 50% reductions in the expression of intercellular adhesion molecule 1, and in monocyte adhesion in a functional binding assay). DNA microarrays revealed a transcriptional signature of 600 genes that were differentially expressed by ECs cultured on the patterned substrates (Fig B), including genes previously not associated with EC function such as histones (HIST1H2AH, HIST2H2BF) and heat shock proteins (HSPB7, HSPB9). These results demonstrate that topographical cues from the underlying ECM are potent regulators of EC morphology and function and mimic the effects of laminar shear-stress. This work highlights the importance of cell-ECM interactions in maintenance of EC phenotype and has implications in the design of vascular conduits to minimize atherogenesis.
Abstract 272: Impaired Arteriogenesis in Cystathionine Gamma-Lyase (CSE) and Endothelial Nitric Oxide Synthase (eNOS) Mutant Mice
Authors: Manrique Alvaro F , Bir Shyamal C , Peter Elvis , Wang Rui , Kevil Christopher G
Published: July 2021

Introduction:
Arteriogenesis is an important process involving remodeling of collateral vasculature during chronic or intermittent tissue ischemia. CSE and eNOS are the two principle enzymes for hydrogen sulfide (H
2
S) and Nitric Oxide (NO) generation. Although role of NO during arteriogenesis is partially known, the role of CSE dependent H
2
S production during arteriogenesis remains unknown.

Hypothesis/objective:
We sought to determine the role of CSE generated H
2
S on arteriogenesis activity of CSE knockout (CSEKO) mice compared to eNOS knock out (eNOSKO) and wild type (WT) mice during hind limb ischemia.

Method:
Permanent unilateral hind limb ischemia was induced in 12-week-old CSE KO, eNOS KO, and WT mice and studied until day 21. Tissue perfusion was measured using indicator dye blush rates with the SPY Imager system over time along with laser doppler flowmetry. Temporal development of arterial remodeling was also observed using SPY imaging, and vessel number, length and branch morphometry determined over time. Microfil vascular casting was performed to further examine the number of collaterals in ischemic muscles at the end of the study. Histological arteriogenesis was evaluated by vascular smooth muscle actin (SMA) positive vessel density with dual fluorescence staining of anti-CD31 (endothelial positive cell) and anti-α-SMA antibodies.

Results:
Tissue perfusion was significantly impaired in CSE KO and eNOS KO mice compared to WT mice at different time points throughout the study period. Both CSE KO and eNOS KO mice showed significantly less blush rate than WT mice. SMA positive vessel density was also significantly less in CSE KO and eNOS KO mice than that in WT mice.

Conclusion:
CSE generated H
2
S and eNOS derived NO play important roles in regulating ischemic arterial remodeling as demonstrated in our studies. The mechanism of impaired arteriogenesis in both mutant mice and the interaction between these two gasotransmitters has yet to be elucidated.
Abstract 274: In Vivo Assessment of Adventitial Vasa Vasorum in Patients with Symptomatic Carotid Plaques: A Dynamic Contrast-Enhanced MR Imaging Study
Authors: Sun Jie , Song Yan , Chen Huijun , Kerwin William , Dong Li , Hippe Daniel , Chen Min , Zhou Cheng , Hatsukami Thomas , Yuan Chun
Published: July 2021

Objectives:
To characterize adventitial vasa vasorum (VV) in unilaterally symptomatic patients and evaluate its association with intraplaque hemorrhage (IPH) and symptom status.

Background:
Adventitial VV is thought to be a primary source of IPH and a potential marker for plaque instability. Dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) has been shown capable of quantifying adventitial VV in the carotid artery.

Methods:
Patients with ischemic cerebrovascular symptoms within the past 6 months and ipsilateral carotid plaques underwent MR imaging examination, which included a multi-contrast protocol for detecting IPH and a DCE-MRI protocol for measuring adventitial VV. Both the symptomatic and contralateral asymptomatic arteries were studied. Adventitial VV was measured at consecutive slices as adventitial Ktrans from kinetic modeling. Maximum adventitial Ktrans across all slices were calculated for each artery.

Results:
From the 27 patients (22 men; 69±10 years) recruited, 50 arteries had adventitial Ktrans measurements. IPH arteries (0.142±0.042 vs. 0.112±0.029 min-1, p<0.001) and symptomatic arteries (0.134±0.038 vs. 0.107±0.027 min-1, p=0.001) were shown to have higher adventitial Ktrans compared to their counterparts. In multivariate analysis, IPH, symptom status and male sex were independently associated with adventitial Ktrans.

Conclusions:
Adventitial VV demonstrated significant associations with IPH and symptom status. In vivo assessment of adventitial VV by DCE-MRI may be useful in studying plaque pathogenesis or risk stratification, but further examination through prospective studies is needed.
Abstract 284: Cholesterol Handling in Bone Marrow--Derived Macrophages from Apoe-Deficient AKR and DBA/2 Mice
Authors: Robinet Peggy , Ritchey Brian , Smith Jonathan D
Published: July 2021

It has been demonstrated in humans that about 50% of the susceptibility to atherosclerosis is heritable. Therefore, it is not surprising that mice from distinct genetic backgrounds present different susceptibility to atherosclerosis. For example, apoE
-/-
DBA/2 mice have aortic root atherosclerosis lesion areas >10-fold larger than apoE
-/-
AKR mice. In order to better understand the mechanisms underlying this difference, we studied cholesterol metabolism in bone marrow-derived macrophages from these two strains. In the unloaded state, macrophages from both strains had equivalent amounts of total, esterified, and free cholesterol (TC, CE, and FC, respectively). Cells were loaded with acetylated low density lipoproteins (AcLDL) for 48h and DBA/2 macrophages had 36% higher TC (p<0.0001) vs. AKR macrophages, mainly due to 72% higher CE accumulation (p<0.0001). In contrast the loaded DBA/2 macrophages had 6% lower FC than the AKR macrophages (p<0.05). No difference was seen in AcLDL uptake or acetyl-coenzyme A acyltransferase (ACAT) activity between the two strains. When cells were loaded with AcLDL, the DBA/2 cells released cholesterol less efficiently than the AKR cells to apoAI (18% lower, p=0.01) or HDL (25% lower, p=0.001). However when the loading was performed in presence of ACAT inhibitor, blocking the formation of CE, the efflux from the two strains was equivalent suggesting a blockage of CE hydrolysis in DBA/2 cells. In loaded cells, when a 24h chase period was done in presence of ACAT inhibitor to prevent cholesterol re-esterification, the CE was reduce by 69% in AKR cells (t
1/2
= 47h) compared to chase without inhibitor while it was only reduced by 42% in DBA/2 cells (t
1/2
= 101h). Furthermore, when lalistat, an inhibitor of lysosomal acid lipase (LAL), and ACAT inhibitor were added during the chase, CE levels were equivalent to the one observed in chase without any inhibitors, suggesting that CE hydrolysis occurs primarily in the lysosome. In conclusion, our data support the hypothesis that DBA/2 cells accumulate more CE than AKR cells due to a defect in CE hydrolysis by LAL. Autophagy has been recently described as the pathway in macrophages by which lipid droplets can be delivered to the lysosome, and we are investigating the role of this pathway in our cells.
Abstract 294: The A2b Adenosine Receptor and Newly Identified Opposing Effects of cAMP on Insulin Secretion
Authors: Johnston-Cox Hillary A , Corkey Barbara , Ravid Katya
Published: July 2021

The A2b adenosine receptor (A2bAR) is a G-protein coupled receptor that, upon binding of adenosine, activates adenylyl cyclase and mediates downstream effects through secondary messengers, including cyclic 3’5’ AMP (cAMP) and Ca
++
. We have previously demonstrated that A2bAR knockout (KO) KO mice, post-high fat diet (HFD) develop a type 2 diabetic (T2D) phenotype, evidenced by elevated plasma insulin and glucose. Pancreatic islets from A2bAR KO mice demonstrated insulin hypersecretion post-4 weeks HFD, and high glucose challenge. To further understand the underlying mechanism, we focused on the contribution of the pancreatic A2bAR to this phenomnena. cAMP has been demonstrated to be a significant amplifier of glucose-stimulated insulin secretion. Through the use of A2bAR KO islets and diet-induced stress, we identified a new dual role for cAMP in mediating insulin secretion, dependent on cAMP level and duration. Short exposure to elevated cAMP indeed causes insulin hypersecretion. cAMP has, however, also a downregulating effect on the expression of GLUT2 mRNA and protein, which has the potential to inhibit insulin secretion. Thus, A2bAR short and long-term signaling in the pancreas play an important role in insulin homeostasis.
Abstract 295: Microsomal Prostaglandin E Synthase 1 Deficiency Attenuates Diet-Induced Obesity and Promotes the Formation of Brite Adipose Tissue
Authors: Moretz Natalie J , Hatch Nicholas , Srodulski Sarah , King Victoria L
Published: July 2021

Mice deficient in adipocyte specific phospholipases A2 have a marked reduction in prostaglandin E2 (PGE2) levels and are resistant to the development of diet-induced obesity. Clinical data suggest that obesity is a chronic low grade inflammatory disease, characterized by the influx of inflammatory cells into the adipose tissue. During a chronic inflammatory state, microsomal prostaglandin E synthase-1 (mPGES-1) is the primary source of PGE2. We have previously demonstrated that mice deficient in mPGES-1 (KO) have a marked reduction in body weight gain and adiposity compared to littermate controls (WT) fed a high fat (HF) diet with a concomitant reduction in urinary PGE2 concentrations and an increase in urinary PGI2 concentrations. The reduction in weight gain is not for accounted by alterations in food intake or locomotor activity. However, resting metabolic rate, measured by indirect calorimetry, was increased in KO mice compared to WT fed a HF diet. Moreover, body temperature was also increased in KO mice compared to WT mice (37.0 ± 0.2 vs 35.8 ± 0.2; P < 0.05) fed a HF diet. Taken together these data suggest that mPGES-1 deficiency increases energy expenditure in response to feeding a HF diet. Analysis of white adipose tissue (WAT) depots demonstrated an increase in number of smaller adipocytes per unit area in the KO mice compared to WT mice. The WAT from KO mice also had a marked decrease in triglyceride content, F4/80 staining and CD86 staining with a concomitant increase in CD206 staining suggesting an attenuation in macrophage recruitment into the WAT as well as an M2 phenotype. Additionally, COX-2 and UCP-1 and PPAR-γ expression were increased in WAT depots with a concomitant localization of multi-locular adipocytes in WAT depots, demonstrating the presence of brown adipocytes in WAT depots in KO mice fed a HF diet. These data suggest that the reduction in body weight gain in the KO mice may be due an increase in thermogenesis mediated by the formation of brite adipose tissue in WAT depots.
Abstract 296: Decreased Hepatic Avpr1a Gene Expression and Elevated Serum Bile Acid in Mouse Model of Metabolic Syndrome
Authors: Ma Xiaoya , Bodary Peter F
Published: July 2021

The KK/HIJ mouse has been demonstrated to have polygenic obesity and insulin resistance and serve as a model of metabolic syndrome. From microarray studies in this mouse strain, we observed hepatic gene expression of arginine vasopressin receptor 1A (Avpr1a) as the most differentially elevated gene in the liver following 3-week of voluntary wheel running activity. Subsequent studies validated that hepatic Avpr1a gene expression is significantly upregulated following voluntary activity and also highly suppressed (4 to 8-fold) in 2 independent models of insulin resistance (including obesity and lipoatrophic models). Although Avpr1a is highly abundant in the liver, its physiologic role is not well described. One proposed role for hepatic Avpr1a is mediation of vasopressin-induced bile acid secretion. To further evaluate the relationship between hepatic Avpr1a and bile acid homeostasis, we determined the age-related change in these variables in female KK mice. To investigate, adipose tissue, liver and serum were collected from female KK mice from before sexual maturity (PRE: 4.5 weeks old, n=9) and after sexual maturation (POST: 6 to 30 weeks old, n=12). Consistent with previous studies using this obesity-prone strain, we observed a robust increase in adiposity with age despite a standard rodent chow diet. RT-PCR studies of hepatic gene expression revealed a 53% lower Avpr1a in POST compared to PRE mice (p<0.00001). In parallel with the drop in hepatic Avpr1a gene expression was an increase in serum bile acids (PRE: 26.56± 9.98μmol/L; POST: 39.40± 9.63μmol/L; p<0.01). A negative correlation was evident between hepatic Avpr1a gene expression and serum bile acid level (R= -0.51). The change in Avpr1a and bile acids was pronounced at the age of onset of estrous cycling. In conclusion, female KK mice have a significant increase in fat mass with age in parallel with an elevation of serum bile acids and downregulation of hepatic Avpr1a gene expression. We propose that suppression of hepatic Avpr1a increases hydrophobic bile acids in the liver and serum and promotes hepatic inflammation, contributing to symptoms of the metabolic syndrome.
Abstract 299: Intestinal Apolipoprotein A-IV Expression Conserves Visceral Adiposity in Mice Subjected to Cyclical Feeding and Fasting
Authors: Simon Trang , Weinberg Richard B
Published: July 2021

Background.
We recently observed that although C57BL6 and apo A-IV knockout (A4KO) mice had similar weight gain on rodent chow, C56BL6 mice gained more weight and had greater adiposity when fed a high fat diet for 12 weeks (J Lipid Res 52:1984-1994, 2011). As food intake and intestinal fat absorption were the same in both groups, these data suggest that intestinal apo A-IV expression increases the efficiency of dietary energy storage. Here we have investigated whether apo A-IV could conserve adiposity in the setting of energy deprivation.

Methods.
Four month old male C56BL6 and A4KO mice (n=5) weighing ∼30-35 gm were subjected to repeated 3 day cycles of free access to a high fat (40%) diet alternating with access to only water. Intra-abdominal fat, fat free mass (FFM), and hepatic fat were measured at baseline and after 8 weeks by CT scanning. At the end of the study, livers and epidydimal fat pads were collected and weighed.

Results.
After 8 weeks on the feed-fast regimen, C57BL6 and A4KO mice had lost similar amounts of total weight, 4.4 ± 0.7 vs 5.2 ± 0.5 g (mean ± SE) and fat free mass, 3.0 ± 0.5 vs 3.0 ± 0.4 g; however, C57BL6 mice had lost less fat than A4KO mice, 1.4 ± 0.3 vs 2.2 ± 0.2 g (p=0.05). C57BL6 mice displayed a smaller decrease in adiposity than A4KO mice, 13.8 ± 1.1 to 10.9 ± 1.1 % vs 13.1 ± 0.5 to 7.9 ± 0.8 % (p=0.02 for the difference in the decrease between groups). There was a trend towards lower hepatic fat content (estimated from CT Hounsfield units) in C57BL6 mice, but liver (1.4 ± 0.2 vs 1.7 ± 0.2 g) and fat pad (1.3 ± 0.7 vs 1.1 ± 0.7 g) weights were not significantly different between the strains, suggesting that cyclical fasting affected mainly visceral fat.

Conclusions.
These data establish that intestinal apo A-IV expression conserves visceral adipose tissue during cyclical fasting. Because apo A-IV is a uniquely mammalian apolipoprotein which appeared at the time of the divergence of mammals from the avian-reptile lineages, and as the mammalian survival paradigm requires that sufficient stores of lipid energy are available to support thermogenesis, internal gestation, and lactation, these data suggest that a major role of apo A-IV is to promote energy storage during times of dietary energy surfeit and conserve lipid energy stores during periods of prolonged energy deprivation.
Abstract 302: Presence of Oxidized Low-Density Lipoprotein in the Left Ventricular Blood of Subjects with Cardiovascular Diseases
Authors: Aluganti Narasimhulu Chandrakala , Litvinov Dmitry , Jones Danielle , Sai-Sudhakar Chittoor , Fristenberg Michael , Sun Benjamin , Parthasarathy Sampath
Published: July 2021

Hypothesis:
Oxidized low density lipoprotein (Ox-LDL) has properties that profoundly affect cardiovascular function. We hypothesized that Ox-LDL is likely to be formed in the left ventricular blood (LVB) when the heart is subjected to ischemic conditions and the ejection fraction (EF) is low. We speculated whether “stagnation” of LDL in the LV could result in increased formation of Ox-LDL.

Objective:
We studied whether there is an increased level of Ox-LDL in the LVB as opposed to peripheral blood (PB), and whether its presence correlated with the EF. Also we examined whether a higher level of Ox-LDL negatively correlated with the activity of paraoxonase 1 (PON 1).

Methods:
Following the Institutional Review Board (IRB) approval, 62 HF patients were enrolled in the study. All patients underwent pre-operative transthoracic echocardiographic assessment of ventricular function. Left ventricular ejection fractions were determined using the Simpsons bi-plane technique. 2ml of LVB and 5ml of PB samples were taken before coronary artery bypass surgery, or a surgery with replacement of mitral, aortic or tricuspid valve. Blood level of Ox-LDL was determined by ELISA (Mercodia), and PON 1 activity was determined by the rate of conversion of its substrate p-nitrophenyl acetate into p-nitrophenol.

Results:
The result showed significant increase in Ox-LDL in LVB as compared to PB (p=0.032) in HF subjects even when EF was near normal. There was no significant increase in subjects with lower EF. In contrast, Ox-LDL levels increased in the PB of subjects with lower EF and reached those of LVB. We also noticed that there was a statistically significant negative correlation between EF and Ox-LDL levels in both LVB and PB (p < 0.05). The activity of PON1, an antioxidant enzyme that protects LDL from oxidation showed decreased levels both in LV blood as well as in PB with decreased EF. It was observed that there was a statistically significant difference in PON1 levels between LV and PB of subjects having EF>60% (p = 0.03). Conclusions: In conclusion the results suggest that there might be oxidative stress associated with LVB even when the EF is not compromised. In contrast, the increase in PB Ox-LDL with poor EF might suggest that the low blood flow to peripheral tissues and end organs also might contribute to increased oxidative stress. The results also might suggest that persistent oxidative stress could have affected the clearance mechanisms of Ox-LDL.
Abstract 317: Mast Cells Do Not Contribute to a Reduction in Vein Graft Neointima in the C-kit Mutant KitW-sh/w-sh Mice
Authors: Wu Junxi , Maffia Pasquale , Kennedy Simon , Lawrence Catherine , Wadsworth Roger M
Published: July 2021

Background:
Recent evidence suggests mast cells are involved in the development of vascular diseases such as atherosclerosis. The presence of mast cells in vein grafts led us to hypothesise that mast cells may regulate the extent of vein graft neointima hyperplasia. In this study, we employed the Kit
W-sh/W-sh
mouse strain, a widely accepted mast cell deficient mouse model induced by a spontaneous inversion mutation in the transcriptional regulatory elements of the c-kit gene.

Methods:
The vein graft surgery was performed by inserting the inferior thoracic vena cava from the donor mouse into the common carotid artery of the littermate recipient. Vein grafts were harvested at 4 weeks and fixed with 4% formaldehyde for histology. In some groups, Kit
W-sh/W-sh
mice were reconstituted with cultured mast cells either systemically or locally 9 weeks before vein graft surgery. For primary cell culture studies, aortic endothelial cells and smooth muscle cells were isolated by collagenase digestion method and cultured in standard conditions.

Results:
Kit
w-sh/w-sh
vein grafts had a significant reduction (33%; p<0.05) in neointima area, compared to the wild type controls. The cellular composition of the Kit
w-sh/w-sh
neointima was similar to the wild type controls, most cells being smooth muscle alpha-actin positive. Neither the systemic nor local reconstitution of mast cells restored the neointima area to the wild type level. In the primary cell culture, Kit
w-sh/w-sh
vascular cells showed less migrating activity in the scratch wound healing test, which may account for the decrease in neointima formation.

Conlusion:
This study suggests that it is not the deficiency of mast cells in the Kit
W-sh/W-sh
mice which accounts for the reduction in neointima formation in vein grafts. Other cell types affected by the c-kit mutation may be responsible for the altered neointima formation in Kit
W-sh/W-sh
mice.
Abstract 32: EBP50 and NF-κB: A Feed-Forward Regulation of Vascular Inflammation
Authors: Leslie Kristen , Song GyunJee , Barrick Stacey , Bisello Alessandro
Published: July 2021

Inflammation plays a fundamental role in the development of cardiovascular diseases such as atherosclerosis and restenosis. The vascular response to inflammation in these diseases requires communication between multiple cell types such as vascular smooth muscle cells (VSMC) and macrophages. We have identified a scaffolding protein, Ezrin Binding Protein 50 (EBP50, also known as NHERF-1), which is expressed at very low levels in normal vessels but is up-regulated following vascular injury. EBP50 promotes VSMC proliferation and neointima formation after wire injury in mouse femoral arteries. In the current study, we hypothesized that EBP50 functions as a central mediator linking macrophage activation and the response of vessels to inflammation. Similar to the observations in vessels, treatment of primary VSMC and macrophages with LPS or TNFα increased EBP50 expression. This increase was dependent on NF-κB signaling because pharmacological inhibition of NF-κB and expression of a dominant negative IκBα abolished the induction of EBP50. Interestingly, LPS-induced activation of NF-κB (determined by phosphorylation of IKKα/β, degradation of IκBα, and p65 phosphorylation) was impaired in EBP50 null VSMC and macrophages. Moreover, nuclear translocation of the p65 subunit of NF-κB was also reduced in EBP50
-/-
VSMC. Consistent with the role of EBP50 on NF-κB, LPS- and TNFα-induced macrophage activation (determined by the expression of the inflammatory markers iNOS, TNFα, and IL-1β) was reduced in EBP50
-/-
macrophages and mice. Similarly, the expression of iNOS, VCAM-1, and ICAM-1 in VSMC was decreased in EBP50
-/-
VSMC and mice. Collectively, these observations indicate that EBP50 and NF-κB participate in a positive feedback loop leading to increased macrophage activation and enhanced response of VSMC to inflammation, suggesting its involvement in vascular remodeling.
Abstract 322: Effect of Atorvastatin on Very-Low-Density Lipoprotein Triglyceride Kinetics in Obese Men
Authors: Ooi Esther M , Ng Theodore W , Watts Gerald F , Chan Dick C , Barrett Hugh R
Published: July 2021

Background
Hypertriglyceridemia is a risk factor for cardiovascular disease. Atorvastatin effectively decreases plasma and VLDL triglyceride concentrations in humans, but the mechanism of action is unknown. This study examined the effect of atorvastatin on VLDL triglyceride (VLDL-TG) metabolism in obese men.

Hypothesis:
Atorvastatin decreases VLDL-TG concentrations by increasing VLDL-TG catabolism.

Methods:
25 obese men (mean ± SEM: age 52 ± 3 years, BMI 34.5 ± 1.4 kg/m
2
, plasma triglyceride 1.9 ± 0.1 mmol/L, HDL cholesterol 1.00 ± 0.05 mmol/L) were studied in a two-arm parallel group study design. Eligible subjects entered a 3 week run-in dietary stabilizing period at the end of which they were randomized to a 6 week treatment period of either atorvastatin 40 mg/day (ATV) or placebo. VLDL-TG kinetics were examined using stable isotope methods and compartmental modelling.

Results:
ATV decreased plasma TG, VLDL apoB and VLDL apoC-III concentrations compared with placebo (p<0.05). Compared with placebo, ATV decreased VLDL-TG concentration ( ATV vs. placebo) (-40% vs. +2%, p<0.01) and increased VLDL-TG fractional catabolic rate (FCR; +54 % vs. -7%, p<0.01). ATV did not alter VLDL-TG production rate. Change in VLDL-TG concentration was positively correlated with change in VLDL apoB (r = 0.81, p<0.01) and VLDL apoC-III (r = 0.65, p=0.02) concentrations, and negatively correlated with VLDL-TG FCR (r = -0.54, p=0.05). Change in VLDL-TG FCR was negatively correlated with VLDL apoC-III, but this failed to reach statistical significance (r = -0.53, p=0.06).

Conclusions:
In obese men, ATV decreased plasma and VLDL-TG concentrations chiefly by increasing VLDL-TG catabolism, with no appreciable effect on VLDL-TG synthesis. Our data suggest that reduction in apoC-III concentration with ATV may explain, in part, the increase in VLDL-TG catabolism.
Abstract 371: High On-Treatment Residual Platelet Reactivity Post--Percutaneous Coronary Intervention in Patients Loaded with High-Dose Clopidogrel: Impact of Diabetes, Age, African-American Race and Presentation Acuity on Antiplatelet Hyporesponse
Authors: Nathan Sandeep , Karol Janet , Sweiss Auddie , Jagadeesan Vikrant , Saha Narayan , Gajjar Mark , Lee Linda , Weiss Sandra
Published: July 2021

Background:
High residual platelet reactivity (HRPR) on clopidogrel is variably defined and linked with factors such as age, race, genetics and diabetes (DM). We investigated the prevalence of HRPR and validated its link to reported risk factors in a high-risk PCI population using published definitions.

Methods:
127 PCI pts had prospective platelet inhibition testing via the Accumetrics VerifyNow P2Y12 platform, 24 hrs post-600 mg clopidogrel po. Inhibition of platelet aggregation (IPA) and residual platelet reactivity (P2Y12-reactivity units, PRU) were analyzed on continuous and dichotomous scales with varying data-supported cutoff values: PRU<230, <208; IPA<10%. Predictors of HRPR were analyzed.

Results:
In 127 pts (age 64 ± 11, DM 50.4%, male 63.7%) mean PRU was 198.0 ± 107 (39.3% ± 30.2% IPA). Using cutoffs of PRU > 230, PRU > 208 and IPA < 10%, HRPR was seen in 42.5%, 48.0% and 24.4% of pts respectively. (Fig 1) DM (220 +/-114.3 vs 177+/-97, P=0.022) and age > 65 yrs (177+ 107 vs 219 + 106, P=0.028) were predictive of high residual platelet reactivity (HRPR) and increasing age was continuously correlated with HRPR (r=0.184, p=0.038). 70.1% of pts were African American (AA) and 34.6% presented with STEMI/ACS but neither factor correlated with HRPR.

Conclusions:
HRPR on clopidogrel was highly prevalent post-PCI but with great variability based on definition. DM and older age were linked with HRPR but several other risk factors. These data suggest limited predictability of HRPR based on published metrics of risk.
Abstract 385: Amelioration of Heart Transplantation-Induced Ischemia-Reperfusion Injury by Melatonin and Ghrelin in Rats
Authors: Alamran Fadhil
Published: July 2021

Background:
Global myocardial ischemia reperfusion (I/R), which is often obligatory during cardiac surgery, induce an inflammatory response in the transplanted heart. Inflammatory reactions in the graft have a pivotal influence on acute as well as long term graft function.

Objective:
This study was undertaken to investigate the potential role of melatonin and ghrelin in amelioration of I/R injury in the transplanted heart in rat model.

Materials and Methods:
The rats were randomized into 4 equal groups. Group 1 sham group, rat underwent the same anesthetic and surgical procedure as the control group except for heterotopic heart transplantation, group 2 control group, rats underwent heterotopic heart transplantation and subjected to global ischemia for 30 min and reperfusion for 2 hours, group 3 donor and recipient rats received melatonin 10 mg/kg i.p. 30 min before the transplant procedure and the same dose at the reperfusion, group 4 donor and recipient rats received ghrelin 10
-8
mol/kg i.p. ( 2 doses) 13 & 1 hour before the transplant procedure, the heterotopic heart transplantation is done using the cuff technique in the neck ( the aorta of the donor is anastomosed with the carotid artery of the recipient and the pulmonary artery of the donor to the jugular vein of the recipient). At the end of experiment (2 hr of reperfusion), blood samples were collected from the heart for measurement of plasma level of cardiac troponin I(cTn I). The heart were harvested, the apical side was fixed in 10% formalin for histological examination and the basal side was homogenized for the measurement of tissue tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) and intracellular adhesion molecule-1 (ICAM-1).

Results:
Compared with the sham group, levels of myocardial TNF-α & IL-1β, and ICAM-1; plasma cTn I were increased (p<0.05), Histologically, all induced untreated rats showed significant myocardial injury (
P
< 0.05) Both melatonin and ghrelin significantly counteract the increase in myocardium level of TNF-α, IL-1, & ICAM-1 (
P
< 0.05). histological analysis revealed that both melatonin and ghrelin markedly reduced (
P
< 0.05) the severity of heart injury in the rats underwent transplant procedure.

Conclusion:
The results of the present study reveal that melatonin and ghrelin may ameliorate I/R injury in the grafted heart in rats via interfering with inflammatory reactions which induced by I/R injury . These findings suggested that both melatonin and ghrelin have a promising graft protective effect against transplant induced global I/R injury.
Abstract 401: Experimental Aneurysm in Rats Excluded by Stent Graft: A New Model to Assess the Impact of Endovascular Aneurysm Repair on the Pathophysiology of Abdominal Aortic Aneurysm.
Authors: Rouer Martin , Rouer Martin , Alsac Jean-Marc , Michel Jean-Baptiste
Published: July 2021

Introduction Biological study of the impact of endovascular aortic repair (EVAR) on pathophysiology of aortic abdominal aneurysms (AAA) can only be performed indirectly in humans, by imaging or search for peripheral biomarkers in the circulating blood. Therefore biological mechanism’s modifications into the aneurismal wall related to its endovascular exclusion are still to be elucidated, and small animal models should bring a valuable help in this field. We describe a new experimental model of stentgraft implantation for the exclusion of AAA in rats.

Methods Aneurysms were induced as previously described by intra-aortic elastase injection in Wistar rats, or by aortic decellularized xenograft transplantation in Lewis rats. At least 15 days later, the midline laparotomy was reopened, and 3mm covered stentgraft were inserted and deployed in the AAA to obtain its exclusion. The patency of the graft and the AAA exclusion could be assessed by a global arteriogram through the carotid artery. After closure of the laparotomy, the rats were awakened and returned to a normal diet.

Results This experimental model of AAA exclusion by a stentgraft allows many in vivo and ex vivo studies of the pathophysiology of AAA after EVAR. Histological modifications of the aortic wall and the intra-luminal thrombus could be assessed. The impact of EVAR on the adventitial immuno-inflammatory activity could be studied by different imaging such as MRI, scintigraphy or PET-scan. In situ biological and enzymatic activities could be evaluated to better understand the local mechanisms leading to AAA shrinkage or expansion after EVAR.

Conclusion Exclusion by stentgraft of experimental AAA in rats is the first described model of EVAR in small animals. It is feasible and reproducible for both elastase and xenograft experimental AAA models. This model will definitely help to a better analysis and understanding of the impact of stentgrafting on biological mechanisms in the aneurismal wall, that lead to EVAR success with shrinkage of aneurismal sac or EVAR failure with its continuing expansion.
Abstract 415: Oxidized High-Density Lipoprotein Reduces Blood Clot Firmness
Authors: Talens Simone , Leebeek Frank W , Rijken Dick C
Published: July 2021

Low high-density lipoprotein (HDL) levels are a risk factor for cardiovascular disease. The best established anti-atherogenic property of HDL is its role in reverse cholesterol transport. Unlike HDL, low-density lipoprotein (LDL) has atherogenic properties and these are enhanced when LDL is oxidatively modified. Oxidized lipids can be transferred from oxidized LDL (oxLDL) to HDL and then be transported to the liver. However, other properties of circulating oxidized HDL (oxHDL) are not extensively investigated. Therefore we studied the role of native HDL (nHDL) and oxHDL in hemostasis. We collected citrated blood from healthy volunteers (n=5) and examined the effects of 4 mg/ml added nHDL and oxHDL on tissue factor-induced clot formation and clot firmness using thromboelastography (ROTEM). The effects of nLDL and oxLDL were measured as a control. Lipoproteins were isolated from plasma with density-gradient ultracentrifugation and oxidized with copper sulphate. The effect on fibrin degradation was examined by thromboelastography in the presence of cytochalasin D (2.5 μg/ml) and by measuring fibrin degradation products in the serum after thromboelastography using an ELISA. The ROTEM parameters clotting time, clot formation time and α-angle, were not affected by the addition of nHDL or oxHDL. The clot firmness, measured as amplitude, reduced during the thromboelastography by the addition of oxHDL. The amplitude (±SD) at 30 minutes, at 45 minutes, and at 60 minutes decreased from 100 (±0)%, 96 (±1.8)% and 91 (±2.9)% to 63 (±24.8)%, 51 (±24.3)% and 45 (±23.5)%, respectively (p<0.05) by the addition of oxHDL but did not change by the addition of nHDL. Fibrin breakdown, as measured by clot firmness in the presence of cytochalasin D was not affected by added oxHDL. In addition, the amount of fibrin degradation products after thromboelastography was comparable between control blood and blood with added oxHDL. No effects on clot firmness were observed with nLDL and oxLDL.

In conclusion, oxHDL, but not nHDL, reduces blood clot firmness in a time-dependent manner without interfering with the clot formation. This effect does not reflect fibrinolysis but most likely involves platelets. The precise mechanism by which oxHDL reduces clot firmness has yet to be elucidated.
Abstract 438: Global Gene Expression Analysis of Human Perivascular Adipocytes Reveals Reduced Expression of Insulin and Wnt Signaling Genes: Implications for Inflammatory Crosstalk
Authors: Chatterjee Tapan K , Kuhel David G , Hui David Y , Weintraub Neal L
Published: July 2021

Inflammatory crosstalk between PV adipose tissue and the blood vessel wall has been proposed to contribute to the pathogenesis of atherosclerosis. We reported that PV adipocytes exhibit a pro-inflammatory phenotype, reduced state of differentiation, and altered expression of developmental genes as compared with subcutaneous (SQ) adipocytes derived from the same human subjects. To define global differences in gene expression patterns between PV and SQ adipocytes, genome-wide microarray studies were performed in three sets of
in vitro
differentiated SQ and PV adipocytes derived from unrelated human subjects. Insulin-regulated and Wnt signaling genes were markedly down-regulated in PV adipocytes. Validation of microarray data by qPCR demonstrated reductions in expression of C/EBPα, PPARγ, FABP4, adiponectin, lipoprotein lipase, hormone sensitive lipase and perilipin in PV compared to SQ adipocytes. We further observed that insulin-induced Akt ser-473 phosphorylation and glucose uptake were markedly reduced (∼ 3 fold and 4 fold, respectively) in differentiated PV adipocytes compared to SQ adipocytes. The mRNA levels of insulin and insulin-like growth factor receptors, however, were similar in adipocytes differentiated from these two depots. Regarding the Wnt pathway, PV adipocytes exhibited dramatically elevated expression of Wnt inhibitor DKK1 (2864%) and reduced expression of Wnt 5A (50%), FDZ4 (38%), and LRP5 (38%). Further evaluation revealed that these Wnt signaling pathway genes, like those of the insulin signaling pathway, correlated with the extent of adipogenic differentiation. We propose that dysregulation of Wnt 5A/FDZ4 and insulin signaling pathways contributes to impaired adipogenic differentiation and insulin resistance in PV adipocytes. This, in turn, may contribute to heightened inflammatory crosstalk between PV adipose tissue and the vascular wall in the setting of atherosclerosis.
Abstract 449: Novel Role of Bim in the Regulation of Allogeneic T-Cell Activation and Development of Transplant Vasculopathy
Authors: von Rossum Anna , Enns Winnie , Shi Yu P , Choy Jonathan C
Published: July 2021

Transplant vasculopathy (TV) is an arteriosclerotic disease characterized by intimal thickening of allograft arteries and is a leading cause of heart transplant rejection. T cell responses towards allograft arteries are responsible for the development of TV and understanding the regulatory pathways controlling T cell activation in allograft arteries provides opportunities for the therapeutic attenuation of TV as well as other arteriosclerotic diseases. Bim is a pro-apoptotic Bcl-2 protein known to down-regulate immune responses after viral infections by inducing cell death of effector T cells but its role in regulating allogeneic T cell responses is not known. We compared cell death and alloantigen-driven activation of T cells from Bim
+/+
(wild-type), Bim
+/-
and Bim
-/-
mice as well as the development of TV in these mice. Bim was required for cell death of both CD4 and CD8 T cells in response to cytokine deprivation
in vitro
. Unexpectedly, Bim was also required for alloantigen-induced proliferation of both CD4 and CD8 T cells as well as for IL-2 production. When TV was examined in aortic interposition grafts implanted into complete major histocompatibility complex-mismatched mice, intimal thickening was significantly reduced in Bim
+/-
but not Bim
-/-
recipients as compared to Bim
+/+
counterparts. There was signficantly less CD4 T cell accumulation in the intima of arteries from Bim
+/-
as compared to Bim
+/+
recipients but this effect was not observed in Bim
-/-
recipients. The accumulation of CD8 T cells in allograft arteries was not affected by differences in Bim expression. Taken together, our data support a novel role for Bim in driving T cell activation in response to allogeneic stimuli and indicate that the effects of this Bcl-2 protein in the pathogenesis of TV likely depends on its dual role in supporting T cell activation and death.
Abstract 451: HPODE Upregulates the Expression of Secretory Phospholipase A2 Group IIA in Rat Smooth Muscle Cells. Potential Contribution to the Progression of Atherosclerosis
Authors: Yang Zhaohui , Aluganti Narasimhulu Chandrakala , Jiang Xueting , Parthasarathy Sampath
Published: July 2021

Introduction
Peroxidized lipids have been suggested to contribute to atherosclerosis in a number of ways. Accumulating evidence indicates that Group IIA secretory phospholipase A2 (sPLA2-IIA) may contribute to the atherogenic process and serve as markers for cardiovascular diseases. However, the relationship between lipid peroxides and sPLA2-IIA remain poorly identified.

Hypothesis
Peroxidized lipids such as HPODEs might induce the expression of sPLA2-IIA and enhance the uptake of LDL, thereby contributing to lipid accumulation.

Methods
Different concentrations of HPODE (0 μM, 12.5μM, 25μM) or oxidized -LDL (ox-LDL) (25μg/ml) were incubated with SMCs at 37°C. Real-time PCR was used to assay gene expression. The degradation of HPODE and ox-LDL was also monitored by the loss of peroxides and conjugated dienes. The formation of HPODE decomposition products was followed by electrospray mass spectrometry.

Results
Incubation of SMCs with HPODE resulted in an up-regulation of sPLA2-IIA mRNA expression in a concentration dependent manner. However, ox-LDL had limited or no effects in the expression of sPLA2. In addition, SMCs and peritoneal mouse macrophages pretreated with HPODE for 8hrs followed by DiI-LDL (20μg/ml) showed an increase in accumulation of labeled LDL as compared to control. We also noted that HPODE was decomposed by SMCs in hours as compared to no cell incubations. However, incubation of peroxidized lipids in the form of ox-LDL resulted in poor decomposition of the peroxides.

Conclusion
SMCs may break down the peroxidized fatty acids leading to the formation of simple compounds such as aldehydes and carboxylic acids. HPODE or its decomposition products may induce the expression of sPLA2-II and enhance foam cell formation.
Abstract 454: Impact of Uremia on Human Adipose Phenotype
Authors: Ho Karen J , Xue Hui , Mauro Christine R , Nguyen Binh T , Yu Peng , Tao Ming , Brunelli Steve , Ozaki C Keith
Published: July 2021

Background:
Recognition of adipose-related signaling participation in vascular processes is increasing, though direct interrogation of clinically relevant human adipose has been sparse. Few scenarios rival uremia for health impact. To advance understanding of the spectrum/determinants of human adipose biology, we compared fresh extremity adipose from patients with/without uremia.

Methods:
Subcutaneous adipose and clinical data was collected from patients undergoing open elective surgery. Protein was isolated, and mediator levels determined via multiplex assays.

Results:
In the 71 patient cohort, mean age=63.4y; 63.3% had diabetes, 49.2% had hyperlipidemia and 53.5% had coronary disease. Compared to non-uremic patients, uremic patients had 1/10th the levels of leptin, 1/3rd the levels of adiponectin, and 3-fold higher levels of resistin (Table 1). Females had 6-fold higher leptin, 1.5-fold higher adiponectin and 2-fold higher TNF but equivalent resistin. There were no differences in mediator levels when stratified by age. In both the obese/non-obese strata, we observed a concordant pattern of association (magnitude/significance) of uremia and leptin/adiponectin/resistin. No differentials in other mediators emerged upon BMI stratification. Multiple regression analysis for leptin/adiponectin/resistin (with age/gender/uremia as independent variables) showed uremia as the highest independent predictor of all three mediators, followed by gender, whereas age had no effect.

Conclusions:
Based on direct biopsy of living human adipose, uremia is associated with adiposopathy, including perturbations in the adipose derived hormones leptin, adiponectin, and resistin. Compared with other parameters including BMI, uremia overall dominated in determining adipose phenotype.
Abstract 455: Extracellular Matrix Incorporation of Thrombospondins Depends on Intermolecular Recruitment by Their L-lectin Domains: A Novel Process in ECM Organization
Authors: Christofidou Elena D , Stenina Olga , Keene Douglas R , Adams Josephine C
Published: July 2021

Thrombospondins (TSPs) are conserved, multi-domain, extracellular glycoproteins, that participate in many biological processes. TSP1 is present in low levels in the extracellular matrix (ECM) of healthy vessel walls but is up-regulated during neointima and plaque formation. TSP1 is responsible for inducing vascular smooth muscle cell (VSMC) proliferation and migration upon vascular injury or during atherosclerosis. We have examined the organisation of elastic fibril layers in the aortae of 10 to 12-week old wild-type and Thbs1
-/-
female and male mice. The total width of elastic layers was measured from medial cross sections of the ascending arch and thoracic aorta. 1055 measurements were made from Thbs1
-/-
and 671 from wild-type sections. The total width of layers is increased in Thbs1
-/-
mice (p<0.05) and organisation of layers is more irregular. Similar perturbations were identified in aortae from 12-week old transgenic mice that specifically over-express TSP1 in arterial smooth muscle cells (based on 878 and 1503 measurements of TSP1 transgenic mice and mice expressing cre in SMC as control, respectively). Thus, ECM organisation in the aortic wall is calibrated to the physiological level of TSP1: either increasing or decreasing TSP1 perturbs the aortic wall ECM. The mechanisms by which TSP1 is deposited into ECM are poorly understood. Previous research from this laboratory identified a conserved site on the outer face of the C-terminal L-lectin domain that is necessary for accumulation of TSP-1 or TSP-5 into insoluble ECM. We present data from ECM assays, western blotting and immunofluorescence that identify a novel mechanism of trans inter-molecular interactions between TSP molecules within ECM. Trans-interactions can be homophilic (TSP1/TSP1 or TSP5/TSP5) or heterophilic (TSP1/TSP5). The mechanism is L-lectin domain specific and is also supported by glycosaminoglycan-dependent interactions that alter the patterning and rate of accumulation of TSP1 in ECM. Concentration-dependent matrix trapping of TSPs may explain how a low abundance component, TSP1, can impact on large-scale ECM organisation, as in the wall of the aorta. This mechanism could provide a new starting point for controlling TSP deposition in ECM under normal or pathological conditions.