bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 479: Juvenile Idiopathic Arthritis Does Not Impair ABCA-1 and SR-B1-Mediated Cholesterol Efflux Capacity
Authors: Mani Preethi , Uno Kiyoko , Duong MyNgan , Wolski Kathy , Husni Elaine , Spalding Steven , Nicholls Stephen
Published: March 2022

Background:
Autoimmune disease in adults significantly increase atherosclerotic burden and cardiovascular risk, potentially secondary to impaired high density lipoprotein (HDL) function. It is unknown whether adolescents with autoimmune diseases such as juvenile idiopathic arthritis (JIA) demonstrate changes in HDL function. We hypothesized that patients with JIA have intact HDL functionality as measured by ATP binding cassette A1 (ABCA-1) and scavenger receptor B1 (SR-B1) mediated cholesterol efflux.

Methods:
22 patients with JIA and 14 healthy controls underwent physical examination and fasting lipid panel. Subjects with significant cardiovascular risk factors, personal or family history of early cardiovascular events, or other chronic inflammatory disease were excluded. ABCA-1 and SR-B1 mediated efflux of serum were studied using J774 macrophages and Fu5AH hepatoma cells, respectively.

Results:
While JIA patients were younger than controls (14.8±4.7 y vs 21.1±6.5 y, p= 0.002), there were no other differences in risk factors, lipids, or lipoproteins. JIA patients demonstrated less SR-B1 mediated efflux (9.8±2.5 % vs 12.1±3.3%, p=0.007) than controls. However, after controlling efflux for ApoA1 level, no difference was noted (0.08±0.01% vs 0.08±0.02%, p=0.60). In contrast, no difference was observed in ABCA-1 mediated efflux (11.5±3.6% vs 11.4±4.9%, p=0.96). Analysis of JIA patients showed that presence of higher active joint counts had no effect on SR-B1 or ABCA-1 mediated efflux. In the entire cohort, HDL-C and ApoA1 levels correlated with SR-B1 mediated efflux (r=0.74 and r=0.77, respectively; both p<0.0001,), but not with ABCA-1 mediated efflux.

Conclusions:
Our results suggest that the capacity for SR-B1 mediated efflux is determined by the number of carriers present, while ABCA-1 mediated efflux is independent of carrier levels. In addition, the results show that patients with JIA have intact capacity for cholesterol efflux via ABCA-1 and SR-B1. Our study is the first to demonstrate that, despite ongoing systemic inflammation, the lipid transporting ability of HDL in JIA patients is not impaired.
Abstract 49: The Neuroimmune Guidance Molecule Semaphorin 3e Is Expressed in Macrophages of Advanced Atherosclerotic Plaques and Induces Macrophage Chemostasis
Authors: Seibert Tara , Wanschel Amarylis C , Hewing Bernd , Ramkhelawon Bhama , van Gils Janine M , Rayner Katey J , Feig Jonathan , O’Brien Edward , Fisher Edward A , Moore Kathryn J
Published: March 2022

Objective:
The persistence of myeloid-derived cells in the artery wall is a characteristic of advanced atherosclerotic plaques. However, the mechanisms by which these cells are retained in plaques are poorly understood. Recently, we reported the involvement of Netrin-1, a neuronal guidance molecule, in promoting chronic inflammation in atherosclerosis by retaining macrophages in plaque. Semaphorins, another class of neuronal guidance molecules, play a critical role in vascular patterning and development, yet little is known about their role in atherosclerosis. The present study evaluates the expression of Sema3E in the setting of atherosclerosis and its contribution to macrophage accumulation in the atherosclerotic plaque.

Methods and Results:
Immunofluorescence staining of aortic root plaques from ApoE
-/-
mice fed a western diet showed abundant expression of Sema3E, as well as its ligand binding receptor PlexinD1, in macrophage foam cells. In vitro, incubation of bone marrow derived macrophages with physiological drivers of plaque inflammation, OxLDL and hypoxia, induced a 3 and 6 fold up-regulation of Sema3E mRNA levels respectively. In addition, Sema3E was also found to be upregulated 4-fold in inflammatory M1 macrophages compared to their more reparative counterpart, M2 macrophages. To understand how this guidance molecule affects macrophage behavior, we treated macrophages with recombinant (r)Sema3E in the presence/absence of CCL19, a chemokine implicated in the egress of macrophages from atherosclerotic plaques. (r)Sema3E blocked actin polymerization and macrophage migration stimulated by CCL19, suggesting that it may act to immobilize these cells in the plaque. Notably, in two different mouse models of atherosclerosis regression, Sema3E mRNA was highly downregulated in macrophages isolated from regressing plaques compared to progressing plaques. Furthermore, the decrease in Sema3E mRNA and protein in regressing plaques coincided with a reduction in plaque macrophage content and an enrichment in markers of M2 macrophages.

Conclusion:
Together, these findings suggest a role for Sema3E in promoting macrophage retention in atherosclerosis and highlight the emerging role for negative regulators of leukocyte migration in promoting chronic inflammation.
Abstract 532: Periadventitial Adenoviral Ang-1/2 Gene Transfer Interferes with Atherosclerotic Plaque Progression and Angiogenesis in the Carotid Artery of LDLr
-/-
ApoB100/100 Mice
Authors: Lappalainen Jari P , Theelen Thomas , Verheyen Fons , Biessen Erik A , Aavik Einari , Anisimov Andrey , Alitalo Kari , Daemen Mat J , Sluimer Judith C , Ylä-Herttuala Seppo
Published: March 2022

Accumulation of macrophages, red blood cells and lipids in atherosclerotic plaques is associated with plaque rupture. It has been shown that macrophages and erythrocytes can enter the plaque also via plaque microvasculature originating from the adventitia. Furthermore, it has been reported that in advanced or ruptured human coronary lesions the microvessel density is increased and these microvessels are leaky. The leaky phenotype is characterized by poor pericyte coverage and dysfunctional inter-endothelial junctions.

As angiopoietins are involved in angiogenic growth and maturation, we hypothesized that periadventitial Ang-1 or Ang-2 gene transfer would reduce atherogenesis and affect microvasculature in mouse carotid artery.

To study this, atherosclerosis was induced by placing a perivascular silastic collar around the carotid artery of LDLr
-/-
ApoB100/100 mice fed a high cholesterol (0,15 %) diet for 3 weeks. Simultaneous to surgery adenoviral gene transfer (5μl, 5 x 107 pfu/ml) of either Ang-1, Ang-2 or LacZ as a control was applied to the carotid artery (20 mice per group). After 1 or 5 weeks follow-up on high cholesterol diet mice were sacrificed. Plaque size and microvessel density were analyzed with morphometry software (Leica Qwin) using cross-sections stained with hematoxylin and eosin or CD31 respectively. Microvessel ultrastructure was studied using electron microscopy.

Ang-2 gene transfer led to significantly smaller plaque area and plaque volume 5 weeks after collar placement compared to LacZ group (2,7 fold, p<0,001 and 3,9 fold, p<0,01 respectively). Also the microvessel density in intima, media and adventitia was lower in the Ang-2 group compared to LacZ (4,5 fold, p<0,05; 7,5 fold p<0,05 and 11,8 fold p<0,05 respectively). Ang-1 gene transfer led to a non-significant trend towards smaller lesions and lower microvessel density compared to LacZ gene transfer. Plaque phenotype was quite fibrous in all groups at 5 weeks, although lesions in Ang-2 group were most human atheroma-like. Electron microscopy showed intact pericyte coverage and endothelial junctions of microvessels in all groups. Endothelial cell morphology after Ang-1 gene transfer showed less filopodia and cytoplasmic vacuoles compared to Ang-2 and LacZ, suggestive of a quiescent phenotype.

In conclusion:
Ang-2 over expression in mouse carotid arteries led to smaller plaque size and lower microvessel density, suggesting the involvement of angiopoietin signaling in angiogenesis of the atherosclerotic vessel wall.
Abstract 535: Stem Cell--Mediated Atherosclerosis Plaque Repair
Authors: Liu Zhao-Jun , Tan Yurong , Tian Runxia , Li Yan , Beecham Gary W , Seo David M , Vazquez-Padron Roberto I , Pericak-Vance Margaret , Vance Jeffery M , Goldschmidt-Clermont Pascal J , Livingstone Alan S , Velazquez Omaida C
Published: March 2022

Objectives:
Notch Signaling is known to regulate the biology of stem cells and the embryonic development of the aorta, but its role in atherosclerosis has received limited investigation. We recently reported that several Notch pathway components are activated in atherosclerotic plaques and result in endothelial inflammation and senescence. We investigate if differential Notch expression plays a role in the balance between plaque repair versus progression as a putative determinant of overall plaque burden.

Methods:
We assessed Notch expression in Bone Marrow Stem Cells of atherosclerosis-predisposed ApoE
-/-
mice (N=5) by PCRArray. We queried our data base of 1179 human blood samples (patients phenotyped by cardiac catherization and genotyped by Genome-Wide Human SNP Array) to determine if circulating mononuclear cells in patients with severe coronary artery disease (CAD) show variant genetic patterns within the family of Notch genes.

Results:
The Notch pathway was significantly activated in bone marrow-derived Endothelial Progenitor Cells (EPC) and Mesenchymal Stem Cells (MSC) from young ApoE
-/-
mice before initiation of high-fat diet and before aortic development of atherosclerosis plaque (some components such as
Hes-1
and
Hey-1
increased by up to 6.6- and 8.2-fold, respectively
P
<0.001). Genetic association analysis detected 4 SNPs with statistically significant linkage to severe CAD.

Conclusions:
Notch signaling may be linked to CAD risk by modulating the biology of bone marrow-derived progenitor/stem cell pools. These findings, together with our prior findings that Notch signaling is activated in atherosclerotic plaques and results in endothelial inflammation and senescence, strongly implicate the involvement of Notch in atherosclerosis pathogenesis.
Abstract 55: Spatiotemporal Heterogeneity of Platelet Activation and Thrombus Architecture Regulates Plasma Protein Accessibility During Thrombus Formation in Vivo
Authors: Stalker Timothy J , Traxler Elizabeth A , Cermignano Samantha , Voronov Roman , Diamond Scott L , Brass Lawrence F
Published: March 2022

While often described as a linear process in which platelets react in a uniform manner, recent in vivo studies demonstrate that the platelet response to vascular injury is quite heterogeneous. Here, we utilized confocal fluorescence intravital microscopy to provide a detailed description of the heterogeneous nature of thrombi formed in vivo while asking the following questions: 1) does platelet accumulation and activation during thrombus formation follow specific spatio-temporal patterns that result in subpopulations of platelets with different activation states; 2) does the architecture of the hemostatic mass depend on the mechanism of injury; and 3) does heterogeneity within the platelet mass result in discrete regions of a thrombus that are more or less accessible to plasma and its constituents? We found that regardless of the mode of injury, stable thrombi were composed of two distinct and contiguous populations of platelets defined both by their activation state and their packing density. Closest to the site of injury was a population of fully activated, stably adherent, degranulated platelets, which was covered by multiple layers of loosely adherent platelets that had not undergone granule release. Increased packing density of platelets within the thrombus core was associated with reduced plasma volume in this region as well as limited accessibility of large plasma borne tracer molecules and proteins. Taken together, these findings clearly demonstrate the heterogeneous nature of thrombus formation at the molecular and structural levels. Further, the finding of reduced plasma volume and protein accessibility to the thrombus core has implications for delivery of pro- and anti-thrombotic blood factors and therapeutic agents to this region of a thrombus.
Abstract 74: Endothelial PPARγ Protects Against Vascular Thrombosis by Downregulating P-Selectin-Mediated Leukocyte-Endothelial Interactions
Authors: Jin Hong , Gebska Milena A , Chauhan Anil K , Keen Henry L , Sigmund Curt D , Lentz Steven R
Published: March 2022

Background and Hypothesis:
Tissue-specific effects of peroxisome proliferator-activated receptor-gamma (PPARγ) on vascular thrombosis are not well understood. Using a novel murine model of selective interference with endothelial PPARγ, we tested the hypothesis that endothelial PPARγ protects against vascular thrombosis.

Methods:
Transgenic mice expressing a dominant negative PPARγ mutant (E-V290M) selectively in endothelium were compared with non-transgenic (non-Tg) littermates. Susceptibility to occlusive thrombosis of the carotid artery was assessed following chemical (7% FeCl3) or photochemical (rose bengal and green laser) injury. Leukocyte rolling on mesenteric venules was measured by intravital microscopy. P-selectin mRNA levels were assayed by real-time PCR and P-selectin protein levels were measured by ELISA and flow cytometry. To assess the role of P-selectin and leukocytes in thrombosis, P-selectin blocking antibodies or leukocyte-depleting antibodies were infused before carotid artery injury.

Results:
The time to occlusion of the carotid artery was significantly shortened in E-V290M mice compared with non-Tg mice after either chemical (5.1±0.2 vs. 10.1±3.3 vs. minutes; P=0.01) or photochemical injury (48±9 vs. 74±9 minutes; P=0.04). Both plasma P-selectin levels and P-selection mRNA levels in the carotid artery were significantly higher in E-V290M mice than in non-Tg mice (P<0.05), but no differences in platelet surface P-selectin were observed. Increased functional endothelial P-selectin activity was demonstrated by enhanced leukocyte rolling on mesenteric venules in E-V290M mice compared with non-Tg mice (53±8 vs. 25±7 per minute; P=0.025). The shortened occlusion times in E-V290M mice were reversed by P-selectin blocking antibodies or leukocyte-depleting antibodies (P=0.04, and P=0.02 respectively). Gene set enrichment analysis of a gene expression microarray demonstrated significant upregulation of NF-kB target genes, including P-selectin, in endothelial cells from E-V290M mice (P<0.001).

Conclusions:
We conclude that endothelial PPARγ protects from thrombosis through a mechanism that involves downregulation of NF-kB-mediated P-selectin expression and leukocyte-endothelial interactions.
Abstract 97: Spatiotemporal Expression Of Matrix Metalloproteinases (mmps) Is Regulated By Ca2+-signal Transducer S100a4 In The Thoracic Aortic Aneurysm Pathogenesis
Authors: Cao Jiumei , Geng Liang , Wu Qihong , Lu Lin , Chen Ying , Shen Weifeng
Published: March 2022

Purpose:
Ca
2+
signaling pathways play critical roles in a wide range of physiological functions including control and regulation of smooth muscle cell performance and vascular tone. Previous studies have indicated that Ca
2+
signaling transducer S100A4 could increase the secretion of MMPs from endothelial cells and fibroblasts and is greatly expressed in smooth muscle cells in atherosclerotic lesions. This study investigated whether S100A4 has a potential role in the formation of TAA.

Methods:
Thoracic aorta of male Sprague-Dawley rats was exposed to 0.5 M CaCl
2
or normal saline (NaCl). Animals were euthanized at specified time points (2, 4, 10 weeks post-TAA induction). The treated aortic segments were harvested, and mRNA levels, protein expressions and immunohistochemistry regarding MMP-2, MMP-9 and S100A4 were analyzed. The A7r5 cell lines were used as in vitro study. Experiments were also performed using human TAA samples for comparison.

Results:
Localized aneurysmal dilation was observed in the CaCl
2
-treated segments, with disruption of elastic lamellae and decrease of intima-media thickness (minimum at 2 weeks). The transcription levels of S100A4 and MMPs were elevated in CaCl
2
-treated segments versus controls (most prominent at 2 weeks), and a significant correlation between S100A4 and MMPs expression was observed across the time points (MMP-2, r=0.652, p<0.05; MMP-9, r=0.762, p<0.05). Immunohistochemistry studies have revealed similar expression pattern of S100A4 and MMPs proteins, as well as co-localization of S100A4 with the cell lineage markers (αSMA and CD68) and inflammatory markers (MMPs and NF-κB P65 subunit). Proliferation ability of A7r5 cells after transfection with S100A4 siRNA was suppressed and down regulation of S100A4 inhibited MMP-2 expression
in vitro
. Increased expression of S100A4 was observed in the entire layers of aorta wall in human TAA specimens. And serum concentrations of S100A4 determined by ELISA were found significantly increased in TAA patients.

Conclusion:
This study established an important role of S100A4 with MMPs in the development of TAA. S100A4 may serve as a biomarker for TAA diagnosis and a potential target for therapy.
Abstract 105: Timely Removal of Lipid Oxidation Products is Mediated by Macrophages.
Authors: Kim Young-Woong , West Xiaoxia , Byzova Tatiana
Published: March 2022

Reciprocity of inflammation and oxidative stress is now significantly considered as an important mechanism underlying numerous diseases and cancers. As a mechanistic link connecting them, we have shown that one of the end products of lipid oxidation, 2-(ω-carboxyethyl)pyrrole (CEP) is generated during inflammation and wound healing and recognized by Toll-like receptor 2 (TLR2) on endothelial cells. It leads to angiogenesis, which is independent of vascular endothelial growth factor. Importantly, the levels of CEP are reduced back to normal when the wound is healed, but remain constitutively high in pathological conditions, including aging and cancers. Thus, for most tissue repair processes, it appears critical not only to generate CEP, but also to provide its timely removal. Accordingly, our hypothesis is that in order to avoid tissue damage by inflammation-induced oxidative products, CEP needs to be cleared from injured tissues when its role is complete. We have sought to determine a cellular and molecular mechanism of CEP clearance from injured tissues. At first, through CEP binding and internalization assay, comparing with other cell types, we verified that macrophage is the responsible cell type of CEP clearance. Specifically, resident, F4/80
hi
, and M2 macrophages are much more efficient for CEP clearance rather than thioglycolate-elicited, F4/80
lo
, and M1 macrophages, respectively. Secondly, by using several knockout mouse lines, we identified that both TLR2 and CD36 are largely involved in CEP clearance. To examine the
in vivo
relevance of CEP clearance besides skin injury model, we used ApoE deficient mice as an atherosclerotic model because atherosclerosis is known to be intimately associated with inflammation and oxidative stress. Using a CEP-specific antibody, we have been screening the CEP appearance and disappearance in early-to-late stages of atherosclerosis. Recently, we examined that CEP is strongly stained only in and adjacent to the atherosclerotic lesion of ApoE deficient mice fed with western diet for 20 weeks, but barely stained in 30 weeks. Moreover, in the plaque, macrophage-enriched region exhibits weak signals of CEP staining. Therefore, we suggest that timely removal of CEP is mediated by macrophages
ex vivo
and
in vivo
.
Abstract 110: Novel Apoc-II Mimetic Peptide Activates LPL and Decreases Serum Triglycerides in Mice.
Authors: Amar Marcelo J , Freeman Lita , Sviridov Denis , Ashan Lusana , Stonik John , Demosky Stephen , Remaley Alan
Published: March 2022

BACKGROUND

Elevated triglycerides (TG) over 1000 mg/dL in patients with severe dyslipidemias can lead to pancreatitis. ApoA-I and its mimetic peptides, which are currently being explored as therapeutic agents for acute coronary syndrome, can also markedly increase serum TG.

OBJECTIVE

To examine the mechanism behind hypertriglyceridemia caused by apoA-I mimetic peptides and to develop novel peptides based on apoC-II for activating LPL and decreasing serum TG.

METHODS

The 18A-CII peptide was synthesized, using standard solid phase FMOC chemistry, and contains the 18A amphipathic helix (DWLKAFYDKVAEKLKEAF) linked to the last helix of apoC-II (AMSTYTGIFTDQVLSVLKGEE) with an intervening Pro residue.

RESULTS

High doses of the 5A apoA-I mimetic peptide (>120mg/kg) raised serum TG between 1000-1200mg/dL in non-fasting C57B/6 mice after a single bolus. By FPLC analysis, most of the TG were observed in VLDL. By in vitro lipolysis studies, 5A was found to be a direct inhibitor of LPL, whereas 18A-CII activated LPL. An inactive analogue of 18A-CII, containing 4 Ala substitutions for the residues (18A-P-AMSTATGAFTAAVLSVLKGEE) shown to bind to LPL, inhibited LPL activity. Intravenous injection of 30mg/kg of 18A-CII in C57BL/6 mice showed a 52% of reduction of LDL-C. In ApoE KO and LDLR KO mice, treatment with 18A-CII led to a marked TG reduction at 4 h of 85% and 53%, respectively, when compared to sham injected mice. Co-administration of 5A along with 18A-CII in mice still resulted in an increase in HDL-C observed with the 5A peptide but with no increase in serum TG. In order to test the effect of 18A-CII in humans, plasma from an ApoC-II deficient patient was treated with LPL. Addition of 18A-CII or apoC-II was similarly effective in restoring normal level of lipolysis after the addition of LPL.

CONCLUSION

In summary, 18A-CII is able to activate LPL both in vitro and in vivo and reduces serum TG levels and can overcome the inhibition of LPL observed with 5A. These results suggest that 18A-CII may be of value either in combination with an apoA-I mimetic peptide and or can be possibly be used to treat apoC-II deficiency.
Abstract 121: Role of Leukocyte Gpr120 in Atherosclerosis Development in Polyunsaturated Fatty Acid Diet Induced Atheroprotection in LDLrKO Mice
Authors: Shewale Swapnil V , Boudyguina Elena , Bi Xin , Zhu Xuewei , MIshra Nilamadhab , Oh Da Young , Olefsky Jorrold , Parks John
Published: March 2022

Introduction

Diets enriched in polyunsaturated n-3 and n-6 fatty acids (PUFAs) are atheroprotective. FO, which is enriched in n-3 PUFAs, reduces plasma lipids and has anti-inflammatory effects. Recent studies have shown that GPR120 is the physiological n-3 PUFA receptor and its ligation by n-3 PUFAs blunts pro-inflammatory signaling pathways. However, the role of GRP120 in atherogenesis is unknown.

Hypothesis

We hypothesized that leukocyte GPR120 contributes to the atheroprotective effects of n-3, but not n-6, PUFUs by ligation of GRP120, resulting in decreased leukocyte inflammation.

Methods

Female LDLr-/- mice were transplanted with bone marrow from male wild type or GPR120 knockout mice. After recovery from transplantation, recipient LDLr-/- mice were fed atherogenic diets containing 0.2% cholesterol and enriched in saturated/monounsaturated fat (palm oil, PO), n-6 PUFA (borage, BO), a botanical n-3 PUFA (echium oil, EO) or FO for 16 weeks. Periodic blood samples were taken for lipid and lipoprotein measurements and aortas were harvested after 16 weeks of diet feeding to quantify atherosclerosis.

Results

Compared to the PO group, mice fed BO, EO and FO had significantly lower plasma total and very low density lipoprotein (VLDL) cholesterol concentrations. EO and FO groups also had lower plasma triglyceride concentrations compare to PO and BO fed mice. Compared to the PO group, mice fed BO and EO had significantly reduced aortic CE, comparable to the FO group. However, lack of leukocyte GPR120 did not significantly affect plasma lipids or atherosclerosis severity within any diet group.

Conclusion

Atherogenic diets enriched in n-6 and n-3 PUFAs derived from botanical oils (BO and EO) were as atheroprotective as FO compared to a diet containing saturated/monounsaturated fat (PO). Furthermore, lack of leukocyte GPR120 had minimal effect on plasma lipid/lipoprotein profile and atherosclerosis, suggesting that n-3 as well as n-6 PUFA associated atheroprotection is primarily due to reduction of VLDL-cholesterol in LDLr-/- mice.
Abstract 126: Identification of an Individual With Extremely High HDL-C Who is Homozygous for a Loss-of-function Mutation in Scavenger Receptor BI (SR-BI) and Functional Characterization of the Mutation
Authors: Zanoni Paolo , Larach Dan , Peloso Gina , Rodrigues Amrith , Tohyama Junichiro , Billheimer Jeffrey T , Kathiresan Sekar , Cuchel Marina , Rader Daniel J
Published: March 2022

Background

The role of SR-BI in human HDL metabolism and physiology has yet to be fully clarified and no humans with homozygous SR-BI deficiency have been reported to date. We report the identification of an individual with HDL-C > 160 mg/dL who is homozygous for a Pro376Leu mutation in the SCARB1 gene.

Methods

In silico tests were performed to evaluate the effect of the variant on protein function (Polyphen 2 and Condel) and on protein secondary structure (Raptor X and NetSurfP). HDL binding, cholesteryl-ether (CE) uptake and the effect on cholesterol efflux were assessed in vitro in COS7 cells. Serum efflux was measured using J774 macrophages. Subcellular distribution of the mutant protein was assessed through biotinylation of cell surface proteins and trypsin sensitivity.

Results

HDL-C was 166mg/dl and apoA-I was and 275 mg/dl in the homozygous subject. HDL particles were much larger and enriched in cholesterol and triglycerides as compared with a control subject matched for HDL-C levels. In silico analysis predicted that the variant would cause secondary structure modifications and be deleterious for function. In vitro studies in COS7 cells transfected with the P376L mutation showed a strongly diminished HDL binding (-69%) and CE uptake (-79%) as compared to cells transfected with the WT form of SR-BI. Efflux was also impaired (-59%). The biotinylation and trypsin sensitivity assays showed a defect in the delivery of the mutant protein to the cell surface.

Conclusions

We report the first identification of a homozygote for a loss-of-function mutation in SR-BI; the phenotype includes extremely elevated HDL-C levels. The P376L mutation in SCARB1 gene results in marked loss of SR-BI function at least in part by impairing the transport of the protein to the cell surface.
Abstract 133: Hemodynamic Environment-dependent Effects of Human HDL and oxHDL on Vessel Wall Inflammation
Authors: Feaver Ryan E , Turk Jim , Collado Sol , Walton Sandi , Day Nathan , Manka David , Wamhoff Brian , Qualls Charles W , Blackman Brett
Published: March 2022

High density lipoprotein (HDL) concentration is inversely correlated with cardiovascular disease and exerts atheroprotective effects on endothelial (EC) and smooth muscle cells (SMC). Oxidation of LDL (oxLDL) is a well-appreciated driver of atherosclerosis. What is less well known is that HDL can become oxidized (oxHDL) by myeloperoxidase, which may negate its beneficial effects. Further, previous mechanistic studies of HDL were performed on static culture ECs or SMCs that do not take into consideration in vivo hemodynamics; a critical regulator of vascular phenotype. Regional hemodynamics prime EC/SMCs towards either an atheroprotective phenotype or, in regions such as bifurcations, towards an atheroprone, pro-inflammatory phenotype. We assessed the role of the hemodynamic environment in modulating the differential response of vascular cells to human HDL or oxHDL from normal donors. To test this, HDL was added to EC/SMC cocultures primed by atheroprotective or atheroprone shear stresses derived from the human carotid bifurcation or an “Advanced Inflammatory” condition that combines atheroprone shear stress with additional atherogenic factors such as oxLDL and TNFα. In the presence of HDL, ECs under Advanced Inflammatory conditions reduced pro-inflammatory (NFkB Activity, IL6, COX2), and oxidative stress (SOD2), but did not have an effect in baseline atheroprotective and atheroprone conditions. Many of the anti-inflammatory responses of endothelial cells were retained when HDL was oxidized. Conversely, while HDL had minimal effect on SMC gene expression, oxHDL was found to reduced the atheroprotective phenotype by potently inducing oxidative and inflammatory cytokine genes/secreted proteins (IL1B, IL6). Overall, this study illustrates the importance of reproducing relevant in vivo environments in order to understand HDL-mediated mechanisms in vascular biology in vitro. This study supports that oxidation dysfunctionally transforms HDL from an atheroprotective to an inflammatory and intrinsically oxidative particle and might damage otherwise healthy regions of the vasculature. Thus, measurement of circulating HDL without consideration of HDL composition and oxidation may limit its predictive value for cardiovascular risk.
Abstract 139: Post-Translational Modifications of Connexins in Neointimal Formation
Authors: Johnstone Scott R , Martin Patricia E , Lampe Paul D , Isakson Brant E
Published: March 2022

Intro

Connexins are integrally involved in the process of cell division and atherosclerosis. Recently we identified that connexin 43 (Cx43) could significantly alter smooth muscle cell proliferation in vitro and in vivo. We now investigate the involvement of Cx43 in human neointimal formation and the mechanisms through which it controls cell cycle.

Methods

Protein interactions between Cx43 were identified in smooth muscle cells through co-immunoprecipitation, immuno-transmission electron microscopy and proximity ligation assays. Novel mice with alanine substitutions at MAPK phosphorylated serines (Cx43-MK4A) were investigated. Samples from diseased and non-diseased human coronary arteries were assessed for level of Cx43 phosphorylation and interactions. Ex vivo saphenous vein explant cultures were used to assess the contribution of Cx43 phosphorylation to human neointimal formation. In vitro studies were performed in human coronary artery smooth muscle cells and in mouse aortic smooth muscle cells from wild type and knockout mice.

Results

We demonstrate that levels of Cx43 are increased in human atherosclerotic plaques and that neointimal formation in human tissues corresponds to a significant increase in Cx43 phosphorylation and cyclin E interactions. In vitro, Cx43 facilitates trafficking of cyclin E to the nucleus in a manner that is dependent on its MAPK phosphorylation.

Discussion

Our studies suggest that in human vascular disease and neointimal formation Cx43 interactions with cyclin E are elevated. Further, Cx43 facilitates the transport of the cell cycle promoter cyclin E to the nucleus suggesting that this is a critical component of VSMC proliferation and therefore could be a novel target in the prevention of neointimal formation in human vascular disease.
Abstract 153: High Ambient Temperature During Pregnancy Predicts Vascular Disease in the Offspring as Adult
Authors: Teji Jagjit S , Eldeirawi Kamal
Published: March 2022

Introduction and Purpose

It well known that being small at birth is a risk factor for hypertension and obesity and diabetes related disorders of adulthood. Animal studies have demonstrated retarded placental and fetal growth when exposed to higher temperature during pregnancy.

Purpose of this study

Is high ambient temperature during pregnancy associated with low birthweight, LBW.

Methods

The data from vital statistics of birth certificates in USA 1995 through 2002. Analyzed only full term babies. Weather data from (NOAA) website; Logistical regression was performed with LBW as the dependent variable with all the other dichotomized variables gender, race, plurality, maternal education, maternal age as confounding and where independent variables latitude and/or temperature with respect to race using STATA 9.0 SE.

Results

About 25 million records from over 32 million births were used for analysis. It was noted that average annual temperature>55 F and lower latitude in the USA had a higher probability of LBW; Odds ratio (OR) 1.228029, P<0.0005; 95% CL-1.22-1.23).[Figure]. Greatest impact of LBW was for non-hispanic white, NHW and lowest for, hispanic, H, OR 1.22 vs. 1.003. Other variables had no impact on the probability of LBW when inversely related to temperature and/or lower latitude of birth.

Conclusions

1. LBW at birth is directly associated to higher environmental temperature during pregnancy. 2. Increased incidence of chronic diseases later in life, such as diabetes, obesity, and cardiovascular are associated with LBW.
Abstract 156: Statins are Effective in Lowering LDL-C in High-Risk Patients with Mitochondrial Disease
Authors: Hannah-Shmouni Fady , Sirrs Sandra , Mattman Andre
Published: March 2022

Background

Mitochondrial dysfunction affects multiple organs with varying severity and can be either primary or secondary to other diseases (e.g.: Alzheimer’s disease) or medications. Statins may be beneficial in patients with mitochondrial disease (mitoD) who have a higher prevalence of cardiac risk factors including diabetes and sedentary lifestyle. Therefore, it would be useful to know if statins are effective in patients with mitoD for prevention of cardiovascular disease.

Methods

We identified adult patients with confirmed mitoD in our clinic. Clinical and laboratory parameters were compared between the statin and non-statin groups.

Results

56 patients had confirmed mitoD, 8 of which were on statins (average of 2 year, equivalent 20 mg daily atorvastatin) for secondary prevention. The statin group had a higher risk of cardiovascular disease (5 High, 3 Moderate Framingham risk score), including higher age (mean, years = 67 (SD 15) vs. 53 (SD 15), p<0.05) and comorbidities, such as coronary heart disease (50 vs. 15%, p<0.019), obesity (33 vs. 2%, p<0.008), diabetes mellitus (38 vs. 15%, p<0.11) and hypertension (50 vs. 40%, p<0.58). At two years, the mean reduction in LDL-C was 39.8% (SD 17%, measured in 7/8). None had myopathy with an elevated CK more than x10 the upper limit of normal (ULN), or AST more than x3 ULN, either at baseline or after the initiation of statins.

Conclusion

Our study suggests that high-risk patients with mitoD may benefit from statins, if indicated for the prevention of cardiovascular disease. The fact that the lowering of LDL-C in mitoD patients was of the same magnitude as expected in the general population suggests that there is no difference in statin metabolism in these individuals.
Abstract 158: The Impact of the Mediterranean Diet on Inflammatory Markers in Men and Premenopausal Women: Do Sex Differences Exist?
Authors: Bédard Alexandra , Lamarche Benoît , Tchernof André , Corneau Louise , Dodin Sylvie , Lemieux Simone
Published: March 2022

Introduction

Little is known about the impact of the Mediterranean diet (MedDiet) on inflammatory markers.

Objective

To investigate the effects of a 4-week controlled isoenergetic MedDiet on adiponectin and high-sensitivity C-reactive protein (hs-CRP) concentrations in premenopausal women and age-matched men.

Methods

Participants were 37 men and 32 premenopausal women aged 24-53 years with a slightly elevated plasma LDL-C (3.4-4.9 mmol/L) or total cholesterol to HDL-C ratio ≥ 5.0, i.e. individuals for whom the adoption of healthy dietary habits is recommended in
CVD
prevention. During the experimental MedDiet, all foods and drinks were provided to participants. Adiponectin and hs-CRP concentrations were measured before and after the MedDiet by ELISA. This controlled 4-week experimental MedDiet has been previously shown to improve lipid and lipoprotein profiles and blood pressure in both men and women.

Results

Adiponectin concentration decreased in response to the MedDiet (time effect, P<0.0001), with a tendency for a greater decrease in men (-17.3%, P=0.0001) than in women (-10.7%, P=0.27; sex by time interaction effect P=0.075). A higher adiponectin concentration before the MedDiet was associated with a greater decrease in response to the MedDiet (r=-0.36 and P=0.003). The MedDiet had no impact on hs-CRP concentration in both men and women (value before the MedDiet: 1.53±0.25 mg/L in men and 2.32±0.31 mg/L in women; time effect, P=0.50; time by sex interaction, P=0.36). hs-CRP concentration before the MedDiet was negatively associated with variations in hs-CRP concentrations in response to the MedDiet (r=-0.39 and P=0.002).

Conclusions

Our results suggest that short term consumption of MedDiet in isocaloric conditions, which has overall beneficial effects on traditional risk factors, leads to a decrease in adiponectin concentration, an adipokine known for its anti-inflammatory properties, and has no impact on hs-CRP concentration. This decrease in adiponectin concentration may be more
pronounced
in men than in premenopausal women. This unexpected impact of the MedDiet on adiponectin concentration, especially in men, needs further investigation.
Abstract 192: Depletion of Inflammatory Monocytes Reduces Endothelial Nitric Oxide Synthase Uncoupling and iNOS Derived Nitrosative Stress in Angiotensin II-induced Vascular Dysfunction
Authors: Hu Hanhan , Kossmann Sabine , Schwenk Melanie , Knorr Maike , Brand Moritz , Schönfelder Tanja , Karbach Susanne H , Becker Christian , Oelze Matthias , Münzel Thomas , Daiber Andreas , Wenzel Philip
Published: March 2022

Background

Endothelial nitric oxide synthase (eNOS) uncoupling occurs in disease states of vascular dysfunction and amplifies vascular oxidative stress and loss of NO bioactivity. Inflammatory myelomonocytic cells as a major source of superoxide are critical for angiotensin-II induced vascular dysfunction; however their role in mediating eNOS uncopuling has not been defined yet.

Methods and results

Angiotensin II (1mg/kg/d, 7d) increased the number of CD11b
+
Gr-1
low
iNOS
+
monocytes and macrophages in mouse aorta (verified by flow cytometry) in paralell to increasing nox4 and heme oxygenase 1 expression (assessed by Western blot analysis) and inducing eNOS uncoupling, measured by differential NOS dependent superoxide formation in the endothelial layer and glutathionylation of eNOS. Toxin mediated ablation of macrophages in LysM
iDTR
transgenic mice restored all of these parameters. Conversely, ablation of LysM
+
cells reduced iNOS expression and normalized vascular levels of tetrahydrobiopterin (measured by high-performance liquid chromatography) and expression of the GTP cyclohydrolase-1, essential for tetrahydrobiopterin synthesis, all of which were increased by angiotensin II in control mice.

Conclusion

Depletion of LysM
+
monocytes reduces angiotensin II induced eNOS uncopuling in vascular dysfunction, and simultaneously protects from iNOS mediated nitrosative stress: These findings highlight the role of vascular inflammatory cells in disturbed NO-bioactivity and vascular function
Abstract 21: Loss of Abcg1 Drives the Site-selective Expansion of B-1 B Cells and Secretion of Atheroprotective Natural Antibodies, in Response to Local Accumulation of Specific Oxidized Lipids
Authors: Tarling Elizabeth J , Gonen Ayelet , Que Xuchu , Diehl Cody , Witztum Joseph L , Baldan Angel , Edwards Peter A
Published: March 2022

The oxidation of LDL, a key event in the pathogenesis of atherosclerosis, generates multiple oxidation-specific neo-epitopes (OSE). These epitopes are recognized by natural germ line IgM antibodies that are secreted by B-1 B cells. Mice lacking the ATP binding cassette (ABC) transporter, ABCG1, develop chronic inflammation in the lungs, associated with the accumulation of lipid (cholesterol, cholesterol ester, phospholipid)-filled cells and cholesterol crystal deposition, which are also characteristic of atherosclerotic lesions. However, hyperlipidemic mice lacking ABCG1 develop smaller atherosclerotic lesions compared to controls. We previously attributed this decrease to increased apoptosis of macrophages within the atherosclerotic lesions of mice lacking ABCG1. We now demonstrate that the lungs of Abcg1-/- mice have increased levels of specific oxidized phospholipids species. Here we show that there is a significant increase in B cell number in the lungs, but not the spleens, of Abcg1-/- mice. Subsequent comparison of B cell subtypes revealed that B-1 B cells, which secrete natural antibodies (NAbs), are significantly expanded in the pleural cavity of Abcg1-/- mice, compared to wild type mice. In addition, we show that the lungs and plasma of Abcg1-/- mice have increased titers of NAbs to OSE, and increased mRNA transcript levels for the known hypervariable (VH) CDR3 region of the atheroprotective EO6/T15 NAb. These data are consistent with an antigen-specific enhanced local production of NAbs in the pleural cavity. These findings suggest that the accumulation of lipids resulting from loss of ABCG1 induces the specific expansion of B-1 B cells, which secrete NAbs that may help protect against the development of atherosclerosis. These data also suggest that Abcg1-/- mice may represent a new model in which to study the protective functions of B-1 B cells/NAbs, and may provide novel targets for pharmacologic intervention and treatment of disease.
Abstract 227: Adipose Stromal Vascular Fraction (SVF) Vasculogenesis Demonstrates a Dose Response Through the Assembly of Endothelial Networks
Authors: Maijub John G , Boyd Nolan L , Hoying James B , Williams Stuart K , Morris Marvin E
Published: March 2022

Objective

Critical limb ischemia (CLI) is a challenging clinical entity complicated by chronic microvascular disease. Stromal vascular fraction (SVF), a heterogeneous population of cells derived from adipose tissue, has garnered interest as a means of an autologous cell-based approach to vasculogenesis in these patients. The SVF concentration needed to achieve this goal is still unknown. Our goal was to determine if SVF vasculogenesis exhibits a dose-response using in vitro and murine in vivo models.

Methods

SVF was isolated from adult male Sprague Dawley transgenic-GFP+ Rat epididymal fat pads and serially diluted using SVF growth media and then added to a respectively labeled matrigel-coated plates. Wells were then labeled with Griffonia simplicifolia-1 (GS-1) rhodamine lectin. Phase and fluorescent microscopy images of each well were then taken. The same SVF concentrations were suspended in collagen gels and subcutaneously implanted into the flanks of 6 Rag-1 deficient mice and imaged with confocal microscopy 2 weeks later to determine if a dose-response could be seen in-vivo. Tubule length was calculated using Metamorph software.

Results

Vasculogenic networks showed monolayering at concentrations of 5.0 x 10
5
and 1.0 x 10
6
cells/mL, where as the intermediate dose exhibited greater complexity to its network. Total tubule length, measured in μm, peaked with the 2.50 x 10
5
cells/mL concentration. Tubule length increased significantly up to this peak, and then gradually decreased at higher concentrations. GS-1 staining was positive when viewing the tubules under fluorescence microscopy. Preliminary results from the in-vivo model showed increasing vasculogenesis up to a maximal tubule length for the 1 x 10
6
cells/mL concentration.

Conclusion

SVF vasculogenesis exhibits a dose-response. Tubule formation is of endothelial phenotype and tubule length is directly proportional to cell concentration up to a peak concentration of 2.5 x 10
5
cells/mL. Preliminary in-vivo data also suggests that a dose-reponse exists. We are encouraged by these results that suggest that cell based therapy may be tailored for appropriate dosing in CLI patients.
Abstract 232: Delayed Inhaled Carbon Monoxide Improves Diabetic Wound Healing
Authors: Washington Christopher B , Shukla Ankur S , Madigan Michael C , Tzeng Edith
Published: March 2022

Background

Nonhealing wounds are a major source of morbidity in diabetic patients. These wounds persist due to a persistent inflammatory state and poor resistance to infection. Carbon monoxide (CO) is a naturally occurring byproduct of heme degradation that possesses both anti-inflammatory and pro-angiogenic properties. Thus, we hypothesize that inhaled CO will improve cutaneous wound healing in the setting of diabetes.

Methods

Excisional wounds (1.5 x 1.5 cm) were created in db/db mice. The mice were treated with inhaled CO at 250 ppm for 1 hr daily starting immediately after wounding or starting on day 8. Control mice remained in room air. Wounds were measured by planimetry. At day 20, mice were sacrificed and the residual wounds were collected for immunohistochemistry.

Results

Inhaled CO initiated immediately after wounding did not improve healing with less granulation tissue formation. However, when CO treatment was delayed until day 8, wound healing was greatly improved compared to controls (residual wounds measuring 27.2 ± 5.3% of original vs 52.2 ± 5.8% on day 20; N=10-14 per group; p < 0.005). CD31 staining showed increased angiogenesis in the delayed CO treated animals (Figure 1).

Conclusions

Early inhaled CO was detrimental to wound healing while delaying CO treatment accelerated healing, potentially through increased angiogenesis. Early administration of CO likely inhibits inflammatory responses and cellular recruitment essential for wound healing. Delayed CO may inhibit the persistent inflammation that leads to chronic non-healing wounds, allowing for improved wound healing in diabetes.
Abstract 285: The Effects of Hypercholesterolemia and Lipoprotein-Cholesterol Concentrations on Angiotensin II-Induced Abdominal Aortic Aneurysms
Authors: Liu Jing , Lu Hong , Howatt Deborah , Balakrishnan Anju , Cassis Lisa , Daugherty Alan
Published: March 2022

Background and objective

Angiotensin II (AngII) infusion promotes abdominal aortic aneurysms (AAA) in both normo- and hypercholesterolemic mice, but the latter have a greater susceptibility, implicating a role of hypercholesterolemia in AngII-induced AAA. A recent publication demonstrated that high-density lipoprotein (HDL) administration prevented AngII-induced AAA in apoE -/- mice without affecting total plasma cholesterol concentrations. The purpose of this study was to determine the relationship between concentrations and characteristics of plasma cholesterol and lipoproteins and AngII-induced AAA.

Methods and Results

Three strains of male mice were studied: C57BL/6, LDL receptor -/-, and apoE -/-. These mice were fed either a normal diet or a saturated fat- enriched diet. AngII (1 μg/kg/min) was infused for 4 weeks through osmotic minipumps implanted subcutaneously. Total plasma cholesterol concentrations were measured using an enzymatic kit. Lipoproteins were resolved by size exclusion gel chromatography and non-HDL and HDL analyzed using Peakfit software. AAA was quantified by measuring maximal outer width of suprarenal aortas. In C57BL/6 mice, the fat-enriched diet modestly increased concentrations of total plasma cholesterol (114 ± 4 vs 99 ± 6 mg/dl; P=0.03) and HDL-C (98 ± 4 vs 81 ± 7 mg/dl; P=0.02), while did not change non-HDL-C/HDL-C ratio (0.21 ± 0.03 vs 0.24 ± 0.03; P=0.5) and AAA (1.1 ± 0.1 vs 1.1 ± 0.1 mm; P=0.6). In LDL receptor -/- mice, the fat-enriched diet feeding increased concentrations of total plasma cholesterol (1590 ± 58 vs 276 ± 9 mg/dl; P< 0.001), HDL-C (132 ± 5 vs 90 ± 3 mg/dl; P<0.001), non-HDL-C/HDL-C ratio (10.8 ± 0.3 vs 2.1 ± 0.1; P<0.001), and AAA (1.6 ± 0.1 vs 1.2 ± 0.1 mm; P=0.002), compared to LDL receptor -/- mice fed the normal diet. In apoE -/- mice, the fat-enriched diet increased concentrations of total plasma cholesterol (1261 ± 121 vs 348 ± 38 mg/dl; P<0.001), had no effect on HDL-C, increased non-HDL-C/HDL-C ratio (53.1 ± 5.9 vs 10.5 ± 2.0; P=0.016), and had no effect on AAA development (1.5 ± 0.2 vs 1.6 ± 0.2 mm; P=0.4).

Conclusion

Hypercholesterolemia promotes AngII-induced AAA, while AAA has no clear relationship with either concentrations of total plasma cholesterol or HDL-C.