bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 5566: Serum Tenascin-C Levels Might be a Novel Biomarker for Predicting Short-term Prognosis of Type B Acute Aortic Dissection
Authors: Nozato Toshihiro , Sato Akira , Hirose Shunsuke , Koura Kenji , Yagishita Atsuhiko , Takahashi Yoshihide , Kamiishi Tetsuo , Hikita Hiroyuki , Takahashi Atsushi , Isobe Mitsuaki , Hiroe Michiaki
Published: April 2024

Backgrounds: Tenascin-C (TN-C), an extracellular matrix glycoprotein, is not normally expressed in adult heart and aorta, but transiently reappears under various pathologic conditions playing important roles in tissue remodeling. The aim of this study is to clarify the clinical implication of serum TN-C levels in patients with type B acute aortic dissection (AD).

Methods: We evaluated 43 patients with type B acute AD between May 2005 and January 2008 and 20 normal controls. We measured serum TN-C levels by ELISA on admission, 7 days, 14 days and 28 days after admission. We compared serum TN-C levels with blood biomarkers, and short-term clinical outcome at a discharge.

Results: Serum TN-C levels (66.4 ng/ml) in AD was significantly higher than normal controls (27.4 ng/ml) at admission (p<0.01). Serum TN-C levels in AD significantly increased at 7 days (82.4 ng/ml), and then gradually decreased to normal range at 28 days (40.4 ng/ml). Serum TN-C levels at 7 days was positively correlated with peak C-reactive protein levels (R=0.45, p=0.02). Three patients of AD died. Serum TN-C levels at 7 days of the death group were significantly higher than those of survival group (79.8±51.5 vs. 168.6±98.6 ng/ml, p=0.01).

Conclusions: We demonstrated that serum TN-C levels are significantly elevated during acute stage after AD, and might be a novel biomarker for prediction of patient prognosis with type B acute AD.
Abstract 5577: Complement Component 3a Receptor (C3aR) Attenuates Atherosclerosis Development
Authors: Rozenberg Izabela , Sluka Susanna H , Wagner Ulrich , Luscher Thomas F , Tanner Felix C
Published: April 2024

Up-to-date molecular biological methods permit efficient identification and characterization of new genes. Using Affimetrix microarrays, we studied gene expression in thoracic aorta of
ApolipoproteinE
−

/

−

(
ApoE
−

/

−
) animals fed either a normal or a high cholesterol diet (HCD) for 4 weeks. Over 200 genes were differentially regulated under these conditions. Interestingly, most components of the complement pathway were upregulated after a HCD, with the complement component 3a receptor (C3aR) exhibiting the highest induction. To study the relevance of the complement pathway in atherogenesis,
ApoE
−

/

−
C3aR
−

/

−

mice and
ApoE
−

/

−

controls were compared after 12 weeks of a HCD.
ApoE
−

/

−
C3aR
−

/

−

animals had on average 14.11%±3.16% of descending aortas occupied by atherosclerotic plaques, which was nearly 2-fold more than controls (7.11%±1.01%). This effect was associated with increased accumulation of leukocytes in atherosclerotic plaques of
ApoE
−

/

−
C3aR
−

/

−

mice. Moreover, production of the proinflammatory cytokines interferon gamma (IFN
γ
), chemokine (C-C) ligand-5 (CCL5), tumor necrosis factor beta (TNF
β
), interleukin-12 (IL-12), and interleukin-18 (IL-18) was higher upon
C3aR
deletion. This study demonstrates that C3aR protects from atherosclerosis development, emphasizing the complex role of the complement cascade in the development of inflammation during atherogenesis.
Abstract 5583: Association Between Newly Onset Oral Anticoagulant Treatment With Phenprocoumon and Serum Levels of the Vitamin-k Dependent Calcification Inhibitor Matrix Gla Protein
Authors: Koos Ralf , Brandenburg Vincent , Kühl Harald Peter , Schurgers Leon
Published: April 2024

Introduction:
Matrix Gla protein (MGP) is a vitamin K-dependent protein. Recently, the uncarboxylated (inactive, uc) form of MGP was associated with vascular tissue calcification. The aim of the present study was to investigate the relationship between a newly initiated oral anticoagulant treatment with the Vitamin-K antagonist phenprocoumon and serum levels of uncarboxylated MGP compared to a control group with aspirine therapy.

Methods:
We performed a prospective study in 43 non-dialyzed patients (mean age 61±10 years, 28 men) with newly initiated oral anticoagulant therapy with phenprocoumon (n=21) or aged-matched subjects with aspirine therapy (n=22). In all patients circulating ucMGP levels (in pM compared to synthetic ucMGP) were measured by a competitive ELISA-based assay (VitaK BV, Maastricht, The Netherlands) at baseline as well as after 1 month and 3 months follow-up.

Results:
At baseline, there were no significant differences in patient characteristics regarding cardiovascular risk factors and the prevalence of coronary artery disease between both groups. In addition, at baseline no significant difference in ucMGP levels between patients with newly initiated oral anticoagulant therapy (1003±526 pM) and control subjects with aspirine therapy (842±214 pM, p= 0.40) was detected. At 1 month follow-up, patients with oral anticoagulation therapy demonstrated significantly increased ucMGP levels (1904±402 pM) compared to the aspirine group (628±223 pM, p<0.001). At 3 months-follow up, a further increase in ucMGP levels in patients with oral anticoagulant treatment (2316±344 pM) compared to the aspirine group (685±378, p<0.001) as well as to study inclusion (1003±526 pM, p=0.003) was observed.

Conclusion:
The present study suggests that newly onset anticoagulation therapy with phenprocoumon is associated with increased serum levels of inactive, uncarboxylated MGP indicating calcification inhibitor deficiency compared to a reference population with aspirine therapy. Long-term follow-up data are needed to evaluate if newly onset oral anticoagulant therapy may be linked to an increased progression of coronary and valvular calcification.
Abstract 5589: Induction of Angiogenesis by RTEF-1 via NF
κ
B-dependent Transcriptional Regulation of HIF-1
α
Authors: Jin Yi , Wu Jiaping , Cully Brittany , Xu Ming , Foo Shiyin , Rosenzweig Anthony , Li Jian
Published: April 2024

Background and Objective
: We previously demonstrated that Related Transcription Enhancer Factor-1 (RTEF-1) plays an important role in hypoxia-induced angiogenesis, in which hypoxia inducible factor-1 (HIF-1) is regulated at the post-translational level to control several angiogenic genes. To further explore the ability of RTEF-1 in the regulation of angiogenesis, this study was designed to assess the potential importance of interrelationships between RTEF-1 and HIF-1 in hypoxic endothelial cells and in an ischemia model of RTEF-1 transgenic mice.

Methods and Results:
Down-regulation of HIF-1
α
mRNA was found in endothelial cells under hypoxic conditions, but it can be recovered by increased expression of RTEF-1. Luciferase assay demonstrated that RTEF-1 increased HIF-1
α
promoter activity by 3.73±0.23 fold. Sequential deletion analyses of the HIF-1
α
promoter identified that an NF
κ
B-related element located between −197 and −188 was essential for RTEF-1 function. By gel shift assay, RTEF-1 was found to interact with the NF
κ
B binding sequence, which was also confirmed by chromatin immunoprecipitation. Wedelolactone, an inhibitor of NF
κ
B, blocked the HIF-1
α
promoter activity in the presence or absence of RTEF-1. In addition, induction of HIF-1
α
promoter activity by RTEF-1 resulted in an increase of HIF-1 protein level and angiogenic processes including endothelial cell proliferation and vascular structure formation. Moreover, over-expression of RTEF-1 in endothelial cells (HMEC-1) led to a significant increase in cell aggregation in matrigel analysis, with further identified increases of connexin 43 expression and promoter activity. In the hindlimb ischemia model of RTEF-1 transgenic mice, increases of HIF-1
α
and vascular endothelial growth factor (VEGF) were detected with an accelerated recovery of blood flow (75% vs. 58%, at day 9) and increased capillary density compared with the wild-type littermate controls.

Conclusion:
RTEF-1 increases angiogenesis via NF
κ
B-dependent transcriptional regulation of HIF-1
α
, and consequently affects endothelial remodeling by increasing cell aggregation via connexin 43. These findings provide a novel mechanistic insight into hypoxia-associated angiogenesis.
Abstract 5594: Human Induced Pluripotent Stem Cells Free of Vector and Transgene Sequences Undergo Cardiogenesis in Defined Conditions
Authors: Zhang Jianhua , Soerens Andrew G , Wilson Gisela F , Yu Junying , Thomson James A , Kamp Timothy J
Published: April 2024

Human induced pluripotent stem (iPS) cells hold great promise for cardiovascular research and therapies, but the lentiviral generated iPS cells contain transgene and vector sequences inserted into the genome that could result in insertional mutagenesis. The recent generation of iPS cells using oriP/EBNA1(Epstein-Barr nuclear antigen-1)-based episomal vectors avoids genomic integrations and brings iPS cells closer to clinical applications. However, the ability of vector-free iPS cells to generate cardiomyocytes (CMs) is unknown. The aim of this study was to test the ability of vector-free iPS cells to undergo cardiac differentiation in embryoid bodies (EBs) as well using directed differentiation in defined medium. Four iPS clones derived from foreskin fibroblasts (iPS-DF6 –9 –9T, iPS-DF6 –9 –12T, iPS-DF19 –9 –7T, iPS-DF19 –9 –11T) were maintained either on mouse embryonic fibroblasts for EB formation, or in feeder-free TeSR medium for directed differentiation. EBs formed contracting outgrowths with variable efficiency among 4 clones (6.1±3.0%, 10.4±3.0%, 0.4±0.2% and 1.6±0.6%, respectively). Likewise, directed differentiation using sequential treatment with Activin A (100 ng/ml) and BMP4 (10 ng/ml) resulted in the formation of contracting CMs, but flow cytometry for cTnT-positive cells demonstrated 10 –15 fold more CMs were obtained using directed differentiation than with the EB method. RT-PCR comparing iPS cell-derived CMs with undifferentiated iPS cells demonstrated strong upregulation of cardiac genes (NKX2–5, TNNT2, MYH6, ACTN2, HPPA, PLN, MYL2 and MYL7) and concomitant downregulation of pluripotency genes (OCT4 and NANOG). Immunolabeling of cTnT, alpha-actinin, MLC2a and MLC2v showed organized sarcomeres in single iPS cell-derived CMs. Sharp microelectrode recordings revealed spontaneous action potentials comparable to those observed in embryonic stem cell-derived CMs (n = 34 cells of 5 microdissected EB outgrowths). Our findings demonstrate that vector-free iPS cells readily undergo cardiogenesis in EBs and that directed differentiation under defined conditions greatly increases cardiogenesis efficiency. Thus, vector-free iPS cells have growing appeal for cardiovascular research and cardiac repair applications.
Abstract 5624: C-Reactive Protein Predicts First Coronary Events but Not Recurrent Stroke: Results From the SPARCL Study
Authors: Ganz Peter , Preston Gregory M , Bao Weihang , Amarenco Pierre , Goldstein Larry , Sillesen Henrik , Welch K M
Published: April 2024

Background:
The inflammatory biomarker C-reactive protein (CRP) predicts the risk for cardiovascular events. The placebo-controlled Stroke Prevention by Aggressive Reduction in Cholesterol Levels (SPARCL) trial evaluated the effect of atorvastatin 80 mg/d on the risk of stroke and major coronary events (COR) in patients with a recent stroke or transient ischemic attack and no known coronary heart disease (CHD). This nested case-control analysis of SPARCL data investigated whether CRP levels at enrolment (1– 6 months after stroke) predicted recurrent stroke or first major coronary events (COR: cardiac death or MI) in placebo-treated patients in this unique patient population.

Methods:
Baseline CRP was measured in 1,243 of 2,366 placebo-treated SPARCL patients, including 278 who had a recurrent stroke, and 94 who had a COR. Baseline characteristics of the sub-study and total SPARCL populations were similar. The predictive value of AHA/CDC recommended cut-points for CRP (<1 [reference group], 1–3, >3 mg/L) was evaluated without and with adjustments for age, gender, smoking, diabetes, entry event, time since entry event, geographic region and race.

Results:
Baseline CRP (median [interquartile range]) was 2.08 [0.96, 4.66] mg/L. Higher baseline CRP predicted COR but not recurrent stroke (Table

). The results were similar using quartiles of CRP.

Conclusion
: In this unique cohort of subjects with cerebrovascular disease but no known CHD, baseline CRP predicted COR, but not recurrent stroke. These results point to potentially important differences in the role of inflammation for first coronary vs. recurrent cerebrovascular events.
Abstract 5625: One-year Mortality in Patients With Troponin I Elevation in an Acute Ischemic Cerebrovascular Event in the Absence of an Acute Coronary Syndrome or Pulmonary Edema
Authors: Purushottam Bhaskar , Amanullah Steven , Kim Jongoh , Amanullah Aman
Published: April 2024

Introduction:
Troponin (TnI) elevation in patients with an acute ischemic cerebrovascular event (AICVE) has a poor prognosis and is often thought to be secondary to cardiac failure {(left ventricular ejection fraction (EF) < 55%} or renal failure (creatinine >1.3 mg/dL). However, the prognosis is not well defined in the absence of cardiac or renal failure.

Hypothesis:
We hypothesize that patients with an elevated TnI (≥0.08 mcg/L) in an AICVE have an increased risk of one-year mortality, even in the absence of cardiac or renal failure.

Methods:
Three hundred consecutive patients who were admitted with an acute ischemic stroke or a transient ischemic attack in the absence of an acute coronary syndrome or pulmonary edema from january 2004 to january 2006 were studied. Hazard ratios for all-cause mortality were determined using multivariate Cox Proportional hazards analysis employing multiple variables between patients with and without TnI elevation.

Results:
The final cohort consisted of 207 patients with a mean age 70 ± 14 years, 62% were females. There was a 33% occurrence of TnI elevation among the entire group of 207 patients (Gr. A) and a 20% occurrence among patients, who had a normal EF(≥ 55%) and a creatinine ≤ 1.3mg/dL (Gr. B, n=91). The one-year mortality was significantly higher in patients with TnI elevation (58% in Gr. A; 42% in Gr. B) as compared to the patients without TnI elevation (20% in Gr. A; 11% in Gr. B).

Conclusions:
The presence of TnI elevation in patients with an AICVE is a significant independent multivariate predictor of one-year mortality in the presence or absence of cardiac or renal failure. It may be plausible that a recent silent myocardial infarction or other neurohormonal mechanisms may cause troponin elevation apart from cardiac or renal failure in the setting of an AICVE.

TnI elevation and one-year mortality
Abstract 5630: Identification of Adventitial Hematopoietic Progenitor Cells in Normal and Atherosclerotic Murine Aortas
Authors: Harbuzariu Adriana , Witt Tyra A , Kleppe Laurel S , Holroyd Eric W , Nesbitt Lisa L , Mueske Cheryl S , Dietz Allan B , Gulati Rajiv , Sandhu Gurpreet S , Simari Robert D
Published: April 2024

Background:
Hematopoietic stem cells and the vasculature share a unique relationship during development, in the adult bone marrow and in the setting of extramedullary hematopoiesis within diseased calcified vessels. We hypothesized that the adult vasculature may contain progenitor cells with the capacity to reconstitute multi-lineage hematopoietic cells.

Methods:
Aortas from normal and ApoE
−/−
adult mice were disaggregated with collagenase and elastase. Disaggregates were cultured in methylcellulose for quantification of short and long term hematopoietic colony forming units (CFU). Flow cytometry assessed various populations of hematopoietic progenitor cells. Aortic disaggregates (or bone marrow or blood) from ROSA26 and GFP mice were transplanted into sublethally irradiated C57BL/6 recipients (n=6) via tail vein injections. Bone marrow cells from the primary recipients were transplanted at 4 months to secondary C57BL/6 recipients (n=6). Mice were sacrificed 3 months later. Blood and bone marrow were analyzed.

Results:
In short term culture in methylcellulose, aortic disaggregates generated erythroid, macrophage, granulocyte and megakaryocyte CFU, with predominance of macrophage colonies, primarily from adventitia (p<0.05) and a greater frequency was found in Apo E
−/−
mice (p<0.05). Long term culture defined that the combined frequency of the hematopoietic precursors approximates the frequency found in bone marrow (1:47,277). Flow cytometry of the aortic disaggregates indicates that among lin-/c-kit+/sca-1+ cells a small population of hematopoietic stem cells (CD150+endoglin+) and a population of multipotent progenitor cells (CD150− endoglin−) exist. Infusion of aortic disaggregates generated bone marrow, blood and spleen chimerism in primary and secondary transplanted mice at 3– 4 months. Flow cytometry of peripheral cells demonstrated multilineage hematopoietic cells.

Conclusions:
For the first time, we have identified a variety of progenitor cell populations in the aorta capable of hematopoiesis. Infusion of these cells into sublethally irradiated mice results in stable chimeras. These data support the presence of novel aortic hematopoietic progenitor cells.
Abstract 5633: Endothelial Progenitor Cells - A Troian Horse in Systemic Endothelial Damage in Patients With Cardiac Parvovirus B19?
Authors: Schmidt-Lucke Caroline , Van Linthout Sophie , Escher Felicitas , Zobel Thomas , Lassner Dirk , Spillmann Frank , Schultheiss Heinz-Peter , Tschope Carsten
Published: April 2024

Detection of the vasculotropic parvovirus B19 (B19V) in endomyocardial biopsies (EMB) is strongly associated with endothelial dysfunction and coronary vasospasms mimicking acute myocardial infarction. B19V replicates primarily in erythroblasts in bone marrow (BM). Endothelial progenitors (EPC) mobilised from BM participate in endothelial cell (EC) regeneration. We, therefore, tested the hypothesis of a systemic EC damage with a direct influence of B19V on EPC and endothelial regeneration. Circulating apoptotic mature endothelial cells (CMAEC; CD45
−
CD146
+
vWF
+
Ann
+
) and CD45
dim
CD34
+
KDR
+
-EPC were quantified from blood by FACS analysis. In 12 patients with B19V (>500 copies/
μ
g DNA) in EMB, numbers of CMAEC (27±12.8/
μ
l) were 6fold higher compared to 9 healthy controls (p<0.05). There were no transcoronary gradients (TCG) of CMAEC, in contrast (p<0.05) to patients with troponin T negative acute coronary syndromes (n=11; +150%; p<0.05), pointing to a systemically induced EC damage. EPC were significantly higher in patients with B19V compared to controls (0.24±0.1%/PMNC vs. 0.008±0.001%/PMNC; p<0.05). In contrast, EPC-function (CFU-Hill) was reduced in these patients (p<0.05). There was a strong trend towards a negative TCG (−33%±40%, p=0.051) of EPC. Intracardiac SDF-1
α
mRNA was 4.6-fold (p<0.05) and CXCR4 mRNA was twofold (p<0.05) increased in B19V+-patients compared to controls, further supporting the hypothesis of a continuous homing. In a second study, CD34
+
KDR
+
-EPC of these patients (n=7) were magnetically separated from BM and blood and B19V DNA with qPCR compared to 5 controls. Compared to controls, only patients had B19V DNA in BM and blood (21399±10378 vs. 16±12 copies/ug DNA in CD34
−
, p<0.05) and documenting replication intermediates of B19V, mRNA was detected in CD34
+
KDR
+
-EPC (4 – 1435 copies cDNA/
μ
g DNA) in BM and blood. Indeed, in one patient with highest copy numbers of B19V mRNA in CD34
+
KDR
+
in blood, B19V mRNA was detected in EMB. Thus, for the first time we present a virally-induced systemic endothelial damage and B19V infection in EPC with replication intermediates. Continuous circulation of BM-derived, B19V-infected EPC could contribute to impaired endothelial regeneration.
Abstract 5636: Time From Onset of Acute Anterior Myocardial Infarction Negatively Influences Bone Marrow Cell Function: Preliminary Data From the NHLBI CCTRN Ancillary Studies
Authors: Cogle Christopher R , Pettway Rodney , Zierold Claudia , Meacham Amy , Madlambayan Gerard J , Carlson Marjorie A , Obodo Udo , Petersen John , Baraniuk Sarah , Moye’ Lemuel A , Traverse Jay , Ellis Stephen , Taylor Doris A , Pepine Carl J
Published: April 2024

Bone marrow (BM) contributes to tissue repair in a time dependent fashion. Impaired BM may lead to decreased response to acute myocardial infarction (AMI) but this hypothesis has been examined only from measurements of peripheral blood (PB) cells among heterogeneous patients.

Methods:

We investigated the association between BM function and time after AMI in a group of patients (mean age 62 years) with recent, initial anterior STEMI, treated by PCI and drug-eluting stenting plus guideline recommended care who had post reperfusion EF ≤ 45% enrolled in CCTRN TIME and Late-TIME trials. BM and PB harvesting were performed at either ≤ 7 days or 14 –21 days after AMI and also from healthy donors. BM and PB endothelial progenitor cells (EPCs) were analyzed by FACS for cell surface phenotyping and by endothelial colony forming cell (ECFC) proliferation.

Results:

Healthy BM generated proliferative ECFC colonies. In contrast, BM from ≤7 day AMI patients showed a trend towards decreased ECFC proliferation and from 14 –21 day AMI patients generated less ECFC colonies (
P
<0.05) with decreased growth rates (
P
<0.05). To relate results to EPC mobilization, we quantified EPCs in BM and PB and found no differences in number of EPCs in BM among all groups. However, in 14 –21 day AMI subjects there was a decrease in PB circulating EPCs (
P
<0.05). Decrease in PB EPCs post-AMI positively correlated with decrease in BM-derived ECFC proliferation (r=0.5515,
P
=0.0268).

Conclusions:

These data are consistent with a time-dependent decrease in EPC proliferative potential after initial anterior reperfused STEMI, suggesting AMI events may preface impaired BM-derived vascular progenitor activity. Another possible explanation for the differences seen is subject differences that guided recruitment into the two studies. These results support the rationale of concentrating angiogenic cells for direct application to sites of myocardial injury.
Abstract 5643 The Erythropoietin Heterodimeric Beta Common Receptor is in a Complex With and Requires VEGFR2 for Downstream Signaling
Authors: Segal Mark , Diao YanPeng , Sautina Laura
Published: April 2024

Background: Endothelial progenitor cells (EPC) circulate in the bloodstream and are capable of carrying out endothelial repair. Erythropoietin (EPO) has been shown to have stimulatory effects on EPC. We have previously demonstrated that EPO effects EPC mobility and increases nitric oxide (NO) levels within cells via the heterodimeric beta-common receptor (BCR) and EPO receptor (EPOR). We now test the hypothesis that not only does EPO induction of NO by the BCR/EPOR complex require vascular endothelial growth factor receptor 2(VEGFR2), but VEGFR2 signaling itself requires the BCR.

Methods: EPC colonies isolated from human or mice were incubated with various kinase or VEGF inhibitors for 30 minutes prior to treating 50 mU/mL EPO and determining the bioavailable nitric oxide (NO) by DAF-FM-Diacetate. In addition we examined the ability of EPO and VEGF to promote an increase in NO in EPC isolated from the BCR −/− mice.

Results: The EPO induced increase in NO could be completely blocked with a VEGFR2 blocking antibody but not by an antibody against VEGF itself. Blocking VEGFR2 also inhibited the EPO mediated increase in transcription of the RNA for endothelial and inducible NO synthase. Interestingly, the increase in NO was not blocked by a JAK-2 kinase inhbitor (Tyrphostin AG-490) confirming that the EPO induced NO effects are independent of the homodimeric EPO receptor-JAK kinase pathway. Most importantly EPC isolated from the BCR−/− mice had no NO stimulation in response to EPO, or VEGF but had a normal NO stimulation in response to bradykinin. We also demonstrate that the VEGFR2 co-immunoprecipitates and colocalizes with the BCR.

Conclusion: We conclude that many of the tissue-protective effects of EPO demonstrated in models of ischemic, traumatic, and inflammatory injuries may be occurring through horizontal signaling of BCR with VEGFR2. In addition VEGFR2 signaling requires the expression of the BCR. These findings have profound implications for the potential side effects of EPO.
Abstract 5649: Human Alanine: Glyoxylate Aminotransferase 2 Lowers ADMA and Protects From ADMA-induced Impairment in Nitric Oxide Production
Authors: Rodionov Roman N , Murry Daryl J , Lentz Steven R
Published: April 2024

Elevated blood concentrations of asymmetric dimethylarginine (ADMA), an endogenous inhibitor of nitric oxide (NO) synthase, are found in association with diabetes, hypertension, congestive heart failure and atherosclerosis. ADMA levels are controlled by dimethylarginine dimethyl-aminohydrolases, cytosolic enzymes which hydrolyze ADMA to citrulline and dimethylamine. ADMA also has been proposed to be regulated through an alternative pathway by alanine-glyoxylate aminotransferase 2 (AGXT2), an aminotransferase expressed primarily in the kidney. Little is known about the intracellular localization or activity of human AGXT2. The goal of this study was to define the subcellular localization of human AGXT2 and test the hypothesis that overexpression of human AGXT2 protects from ADMA-induced inhibition in nitric oxide (NO) production. AGXT2 was cloned from a human kidney cDNA library and overexpressed with a C-terminal FLAG epitope tag in COS-7 cells. Mitochondrial localization of human AGXT2 was demonstrated by confocal microscopy. A 41 amino acid N-terminal mitochondrial cleavage sequence was delineated by N-terminal sequencing of the mature protein. To determine the effect of AGXT2 on NO metabolism, human AGXT2 was overexpressed in MS-1 murine endothelial cells. Overexpression of AGXT2 protected MS-1 cells from ADMA-mediated inhibition of NO production. To determine the effect of AGXT2 on ADMA metabolism
in vivo
, human AGXT2 was overexpressed in the liver of C57BL/6 mice using an adenoviral expression vector. Four days after intravenous injection of the AGXT2 or control adenoviral vectors, mice receiving the AGXT2 vector had significantly lower levels of plasma and tissue ADMA compared with mice receiving the control vector (P<0.05). We conclude that mitochondrially localized human AGXT2 lowers ADMA and protects from ADMA-induced impairment in NO production.
Abstract 5655: Autoantibodies Against Endothelin-1 Receptors Are Involved in the Pathogenesis of Pulmonary Arterial Hypertension
Authors: Dandel Michael , Wallukat Gerd , Lehmkuhl Hans B , Englert Angela , Hetzer Roland
Published: April 2024

Background
In patients with pulmonary arterial hypertension (PAH) production of endothelin-1 (ET-1) is increased and elevated ET-1 plasma levels correlate with PAH severity. After repeated detection of agonistic antibodies against the ET-1 A receptor (ETA) in PAH patients’ sera we assessed their prevalence and properties.

Methods
Using spontaneously beating rat neonatal cardiomyocytes as bioassay we analyzed sera of PAH patients for the presence of autoantibodies (AABs) against ETA receptors. AABs were purified by affinity chromatography.

Results
At the time of AAB testing, the pulmonary vascular resistance (PVR) in the 58 evaluated PAH patients varied between 500 and 1920 dyn • sec • cm
−5
(median 1088 dyn • sec • cm
−5
) and 45 (77.6%) were in functional NYHA/WHO class ≥ III. Of these 58 patients 44 (78.7%) tested positive for AABs against ETA receptors. ET-1 and the AABs against ETA exerted agonistic negative chronotropic effects on the neonatal cardiomyocytes (decrease in pulsation rate), which were blocked by the ETA antagonist BQ610, but not by the ET-1 B receptor blocker BQ788. The AABs induced permanent stimulation without desensitization of the receptor mediated signal cascade and the effect increases with higher AAB concentrations. First attempts to eliminate by immunoadsorption these AABs (which appeared to belong to the IgG2 subclass) in 5 PAH patients showed encouraging results. After AAB removal, which was well tolerated by all patients, at the first controls performed 3 and 6 weeks later all 5 patients showed reduction in right ventricular diameters, lower pulmonary arterial pressure, higher cardiac index and improved exercise tolerance (increase in both VO
2
max and 6 minute walk distance). In one patient the AABs reappeared after 2 months and this AAB reappearance was associated with clinical worsening and PVR increase.

Conclusions
The majority of patients with advanced PAH tested positive for functional serum AABs against ETA These AABs activate the receptors, like ET-1, but prevent the desensitization of the receptor-mediated signal cascade normally seen with ongoing receptor stimulation. Our results suggest that these AABs could be involved in PAH pathogenesis. AAB removal by immunoadsorption might be a possible new therapeutic approach.
Abstract 5671: The Effects of Pharmacological PAI-1 Inhibition in Acute and Chronic Rejection in Murine Cardiac Allografts
Authors: Suzuki Jun-ichi , Ogawa Masahito , Muto Susumu , Yamaguchi Yoichi , Itai Akiko , Hirata Yasunobu , Nagai Ryozo , Isobe Mitsuaki
Published: April 2024

Backgrounds: Acute rejection and graft arterial disease (GAD) in cardiac transplantation are enhanced by inflammation and thrombus formation. However, little is known about the effect of plasminogen activator inhibitor-1 (PAI-1) in heart transplantation. Thus, the objective was to clarify the role of PAI-1 in the progression of rejection.

Methods and Results: Murine hearts were heterotopically transplanted using major mismatch combinations for evaluation of acute rejection and class II mismatch combinations for the GAD. We performed administration of the specific PAI-1 inhibitor (IMD-1622) into murine recipients of cardiac allografts. Nontreated allografts of the major mismatch group were acutely rejected (n=6; 7.3±0.2 days), while the PAI-1 inhibitor prolonged their survival (n=6; 13.7±2.4 days, P < 0.05). Pathologically, severe myocardial cell infiltration (40.8±3.3 %) and fibrosis (44.5±2.8 %) were observed in untreated allografts, the PAI-1 inhibitor attenuated infiltration (25.8±1.9 %, p<0.05) and fibrosis (25.8±3.1 % p<0.05) in the class II mismatch group on day 60. Although severe GAD (luminal occlusion 28.6±6.4 %) was observed in untreated allografts, the PAI-1 inhibitor suppressed the development of neointimal formation (8.1±3.5 %, p<0.05). Immunohistochemically, CD8 and ICAM-1 were enhanced in nontreated allografts, while the PAI-1 inhibitor markedly attenuated expression of these factors. Zymography showed that increased gelatinase activity was observed in the non-treated allograft group, however, the PAI-1 inhibitor decreased the activity. RNAse protection assay revealed that PAI-1 inhibitor suppressed IFN-gamma compared with that of non-treated group.

Conclusion: The PAI-1 inhibitor is potent in the suppression of both allograft rejection and arterial disease because they are critically involved in the development of rejection through the suppression of inflammation.
Abstract 5672: Deletion of Bone Marrow Derived Receptor for Advanced Glycation End-products (RAGE) Does Not Attenuate Plaque Formation but Inhibits Plaque Progression in a Mouse Model of Advanced Atherosclerosis
Authors: Preusch Michael R , Morris-Rosenfeld Sam , Bierhaus Angelika , Albrecht Claudia , Blessing Erwin , Andrassy Martin , Katus Hugo A , Bea Florian
Published: April 2024

Background and Hypothesis: The multiligand receptor for advanced glycation end products (RAGE) of the immunoglobulin superfamily is expressed on multiple cell types implicated in the immune-inflammatory response and in atherosclerosis. We sought to determine the role that bone marrow derived RAGE plays in different stages of atherosclerotic lesion development in apolipoprotein E deficient (apoE
−/−
) mice.

Methods and Results: 7 and 23 week apoE
−/−
mice were lethally irradiated and given bone marrow from RAGE null (RAGE
−/−
//apoE
−/−
) or RAGE
−/−/
/apoE
−/−
mice to create double knock out chimeras. After 16 weeks on a standard chow diet the mice were sacrificed and atherosclerotic lesion development was evaluated. Plaques in the aortic root of the young mice showed no significant difference in maximum plaque size (217 470±17 480
μ
m
2
for the RAGE
−/−
//apoE
−/−
mice compared to 244 764±45 840
μ
m
2
) whereas plaques from the older innominate artery of Rage
−/−
/apoE
−/−
mice had significantly smaller maximum plaque area compared to the Rage
−/−/
apoE
−/−
mice (83 384±17 981
μ
m
2
compared to 136 041±13604
μ
m
2
, p<.05) as well as reduced average necrotic core area (35 582±4 797
μ
m
2
compared to 55 604±6 523
μ
m
2
, p<.05). Reduced plaque size and more stable plaque morphology was associated with significant decreases in the expression of important atherogenic mediators including VCAM-1, ICAM-1, and MCP-1. Accumulation of the RAGE ligand HMGB-1 was also significantly reduced within the lesions of the Rage
−/−
/apoE
−/−
mice.

Conclusions: This study demonstrates that bone marrow derived RAGE is an important factor in the progression of atherosclerotic plaque development by causing increased expression of pro-atherosclerotic mediators. It provides evidence that RAGE could be a potential target for clinical treatment of advanced atherosclerosis.
Abstract 5764: Discharge Without Antiplatelet Therapy After Inhospital Bleeding is an Independent Predictor of 6-Month Mortality Among Patients With Acute Coronary Syndrome: A Pooled Analysis From PURSUIT, PARAGON-A, PARAGON-B and SYNERGY
Authors: Chan Mark , Sun Jie L , Wang Tracy Y , Lopes Renato D , Jolicoeur Marc E , Pieper Karen S , Rao Sunil V , Newby L. K , Mahaffey Kenneth W , Harrington Robert A , Peterson Eric D
Published: April 2024

Bleeding is an important complication of the management of acute coronary syndrome (ACS). Patients (pts) with bleeding are more likely to be discharged without antiplatelet therapy (AT); however, there are limited data on the impact of discharge without AT on longitudinal outcomes in this population.

Methods:
We examined discharge AT (aspirin and/or thienopyridine) use among 8582 pts who experienced inhospital bleeding in 4 large trials of antithrombotic therapy in ACS. After adjusting for the propensity to receive AT, we compared 6-month outcomes among pts discharged with at least one antiplatelet drug vs. those discharged without AT.

Results:
Almost 1 in 10 pts with bleeding were discharged without AT (n=826). These pts had more baseline comorbidities and were more likely to experience other inhospital non-fatal adverse events compared to patients discharged with AT. Adjusted outcomes associated with no AT vs. AT are shown in the Table

. In an interaction analysis, the effect of discharge without AT was worst among pts with more severe bleeding and those treated with PCI. In a treatment effect by treatment intensity analysis, the adjusted hazard ratio for death or myocardial infarction was 1.45 (95% Confidence interval(CI) 1.08–1.94) for pts receiving monotherapy and 1.90 (95% CI 1.27–2.84) for patients receiving no AT, compared to patients receiving dual AT.

Conclusions:
Among patients with bleeding during ACS, discharge without antiplatelet therapy was associated with worse 6-month adverse outcomes. This effect was more pronounced among patients with more severe bleeding and among patients undergoing PCI. A gradient of better outcomes with greater AT intensity was observed. Thus, more research is required to determine the optimal approach to reinitiating AT among patients who bleed yet remain at high risk of recurrent ischemic events after discharge.

Table.

Six-month Outcomes Among Patients Discharged Without Vs. With Antiplatelet Therapy
Abstract 5788: The Incidence and Health Economic Burden of Critical Limb Ischemia and Ischemic Amputation in Minnesota: 2005–2007
Authors: Peacock James M , Keo Hong H , Yu Xinhua , Oldenburg Niki , Duval Sue , Henry Timothy D , Jaff Michael R , Baumgartner Iris , Hirsch Alan T
Published: April 2024

Background:
Critical limb ischemia (CLI) is the most severe manifestation of peripheral artery disease (PAD) and is associated with high rates of morbidity and mortality, including subsequent myocardial infarction, stroke, and amputation. Nevertheless, the annual incidence of CLI in the United States is not known and there is no active national surveillance.

Methods:
We have estimated the incidence of ischemic amputation using all inpatient hospital discharge claims in Minnesota. These claims represent approximately 97% of hospitalizations in the state. Major and minor ischemic amputations were identified through ICD-9 procedure codes for lower limb amputation not due to trauma or cancer. This coding algorithm can be applied to other state-based hospitalization claims databases.

Results:
The age-adjusted incidence of ischemic amputation in 2007 was 18.8 (95% C.I. 17.7–19.9) per 100,000 (n=1,038). Amputations increased significantly with age and were more than twice as high in men as in women (25.1 vs. 11.3 per 100,000). The incidence of amputation did not differ significantly by rural or urban residence. Of these hospitalizations, 45% were for major above ankle leg amputations. The mean total charge for each hospitalization was $53,726, with cumulative charges for ischemic amputation hospitalizations of $56.2 million statewide in 2007. The incidence of ischemic amputation is significant as it represents approximately half of the annual stroke mortality rate (36.2 per 100,000 in 2007) with total charges approximately 20% of those for annual stroke hospitalizations. Applying this incidence and these charges for ischemic amputation in Minnesota to the entire US population in 2008 yields an estimate of more than 57,000 amputations and hospitalization costs of $3.1 billion.

Conclusion:
Ischemic amputation represents only the most severe CLI clinical event and underestimates the total burden of CLI and severe PAD. The incidence of ischemic amputation remains high, and results in major morbidity and health economic costs. These data permit the first population-based estimate of ischemic amputation in the USA in a non-selected population at risk.
Abstract 5792: Lower Ankle-Brachial Index Associated With Lipid-Rich Coronary Plaques in Patient Underwent Elective Coronary Intervention: Integrated Backscatter Ultrasound Analysis
Authors: Uetani Tadayuki , Amano Tetsuya , Shinbo Yusaku , Kunimura Ayako , Kumagai Soichiro , Yokoi Kiminobu , Harada Ken , Yoshida Tomohiro , Kato Bunichi , Kato Masataka , Marui Nobuyuki , Nanki Michio , Izawa Hideo , Matsubara Tatsuaki , Ishii Hideki , Murohara Toyoaki
Published: April 2024

Background
Association between low Ankle Brachial Index (ABI) and cardiovascular events were demonstrated. The aim of this study is to evaluate association between ABI and coronary plaque radiofrequency characterization by integrated backscatter ultrasound (IB-IVUS).

Methods
From Jan.2006 to Dec. 2008 Total of 189 consecutive patients who underwent elective PCI following ABI measurements and IB-IVUS analysis were enrolled. The Color coded maps of tissue characteristics of each 1mm slice of target lesions were constructed and volume of each plaque components (calcification, fibrous and lipid core) was calculated. And percentage of lipid volume in the target lesion (% lipid; volume of lipid core/total plaque volume) was calculated.

Results
ABI significantly correlated with lipid volume of target lesion (r= −0.19, p<0.05), but did not correlate with fibrous volume (r=0.09, P=0.2)(Figure

). ABI (r= −0.18, p<0.01) and HDL-cholesterol (r= −0.14, p<0.05) inversely correlated with %lipid. There was no significant association between %lipid and other clinical factors (age, gender, conventional risk factors, blood pressure, lipid-profile and HbA1c). In multiple regression analysis, ABI significantly associated with %lipid after adjusting for age, gender, prior revascularization of PAD (bypass surgery or angioplasty), systolic BP, LDL-cholesterol, HDL-cholesterol, triglyceride and HbA1c.

Conclusion
Our present findings provide the first evidence that decrease of ABI may be associated with progression of lipid-rich coronary plaques. After adjustment of other clinical factors, ABI independently associated with lipid-rich coronary plaque of coronary stenosis.
Abstract 5793: Lower Extremity Peripheral Vascular Disease is an Independent Predictor of Adverse Outcomes After Percutaneous Coronary Intervention
Authors: Nanayakkara Shane S , Sharma Rahul P , Duffy Stephen J , Ajani Andrew E , Brennan Angela L , Reid Chris M , Andrianopoulos Nick , Shaw James A
Published: April 2024

Background: A significant proportion of patients with coronary artery disease have coexistent peripheral vascular disease (PVD). Whether PVD is a predictor of adverse outcomes in patients undergoing percutaneous coronary intervention (PCI) in the drug eluting stent era is not known.

Methods: Data was analysed from a total of 9110 patients from the Melbourne Intervention Group (MIG) database, a registry of patients undergoing PCI in Melbourne, Australia. Data was collected between April 2004 and July 2007 and analysed for baseline demographic data, traditional risk factors, procedural data and thirty day and twelve month major adverse cardiovascular events (MACE).

Results: Of the 9110 patients undergoing PCI, 632(6.9%) had PVD. Patients with PVD had an average age of 70.6 years, were more frequently diabetic (41.3% vs. 22.6%, p<0.01), were hypertensive (79.7% v 63.1%, p<0.01), and had higher lipid levels (82.5% v 70.6%). In the PVD group patients were more likely to have left main (2.4% vs. 0.7%, p<0.01) and ostial lesions(5.1% vs 3.4%, p=0.02) with no difference in acute procedural success between the 2 groups. 30 day MACE was higher in the PVD group (8.1% vs. 5% p<0.01) with a significant increase in early mortality. 12 month MACE was also more frequent in the PVD group (21% vs. 12.4%, p<0.01), with increased 12 month MI (9.8% vs. 4.3%, p<0.01) and mortality (8.6%v 3.5% p<0.01). On multivariate analysis, PVD independently increased the risk of 12 month MACE (OR 1.456, CI 1.07–1.99 p<0.01).

Conclusions: PVD is an independent predictor of adverse outcomes in patients undergoing PCI even correcting for traditional risk factors. In this study we found a significant increase in 30 day and 12 month mortality and MI in the PVD group, with diabetes and hypertension the most critical modifiable risk factors. Thus in patients undergoing PCI, the presence of peripheral vascular disease should be recognised as an additional factor in determining risk of cardiac events in the short and long term post PCI, with tighter diabetic control and lower blood pressure to be the primary targets of medical therapy in this group.
Abstract 5797: New Biomarkers for Assessing the Activity of Takayasu Arteritis
Authors: Ishihara Takashi , Kamiishi Tetsuo , Haraguchi Go , Isobe Mitsuaki
Published: April 2024

Background - Takayasu arteritis (TA) is a chronic vasculitis, mainly involving the aorta and its main branches. Erythrocyte sedimentation rate (ESR) and C-reactive protein(CRP)have generally been used for monitoring the disease activity. We frequently experience patients who show inflammatory symptoms without an elevation of CRP, even though they are treated with immunosuppressants. Therefore, we sought for useful and sensitive biomarkers to diagnose subtle activity of TA. We hypothesized that pentraxin3 (PTX3) and matrix metalloproteinases (MMPs) could be biomarkers of TA to detect patients who develop recurrence of the disease even in the absence of CRP elevation.

Methods and Results - Consecutive series of 45 patients with TA and 20 age-matched healthy controls were enrolled in this study. According to the clinical symptoms of the patients, 23 patients were in an active phase as evidenced by clinical recurrences within 2 years of blood sampling and 22 patients were in an inactive phase without any inflammation for more than 2 years. Circulating levels of PTX3, MMP-2, MMP-3 and MMP-9 were determined by ELISA. Patients in an active phase had significantly higher level of high sensitive CRP (hsCRP), PTX3, MMP-3 and MMP-9 than those in an inactive patients. Sensitivity (SE), specificity (SP) and area under the ROC curve (AUC) to detect active patients of each biomarker are as follows. hsCRP: SE 53.33%, SP 90.91%, AUC 0.798, PTX3: SE 76.19%, SP 76.19%, AUC 0.871. MMP-3: SE 60.87%, SP 86.36%, AUC 0.831, MMP-9: SE 61.90%, SP 66.67%, AUC 0.728. PTX3 was the best single marker with the highest diagnostic accuracy. Among 21 patients in an active phase, 9 patients showed negative hsCRP level (<4837ng/ml). However, PTX3 level was elevated (>5.27ng/ml) in 7 of the 9 patients. Patients with TA had significantly higher level of MMP-9 (active:360±75ng/ml, inactive:229±48ng/ml) than normal controls (23±5ng/ml) inspective of disease activity. Evaluation of MMP-9 may suggest prior existence of TA even in inactive patients.

Conclusions - PTX3, MMP-3 and MMP-9 could be new and sensitive biomarkers to estimate the activity of TA. We can predict recurrence of TA by elevation of PTX3 even patients whose CRP is negative.
Abstract 5806: SRF Protects Brain Endothelial Cells Against Hypoxic Damage
Authors: Chai Jianyuan
Published: April 2024

Ischemia resulting from brain vascular disorders is the major cause of stroke. Hypoxia triggers both angiogenesis and apoptosis through the induction of HIF1
α
-activated VEGF and the alteration of mitochondrial permeability, respectively. The balance between these two events is crucial to both the progression of vascular disorders and the outcome of injury healing. Our previous study showed that SRF (Serum response factor) is essential to VEGF-induced angiogenesis. This study aimed to determine the roles of SRF in hypoxia-induced brain endothelial cell angiogenesis and apoptosis.

Methods
: Mouse brain endothelial cells (bEND3) were transfected with either an SRF expressing plasmid, SRF shRNA, or the vehicle (control), and cultured under hypoxic condition (5% CO
2
: 94% N
2
: 1% O
2
) for 0, 0.5, 2, 6 and 24 hours. Angiogenesis and apoptosis were assessed by 3-D
in vitro
angiogenesis assay and Annexin V binding assay, respectively.

Results
:

After 30-min hypoxic treatment, SRF mRNA expression was dramatically increased by 54 fold and then gradually declined and returned to normal at 24 hours. SRF protein expression and phosphorylation were also increased by 111±24 % and 74±7 % (both P < 0.01), respectively, after 6-hour hypoxia. Hypoxia also enhanced SRF binding activity to DNA.

Forced overexpression of SRF increased hypoxia-induced angiogenesis by 232±15 % (P < 0.01); and knockdown of SRF completely abolished it.

After 24-hour hypoxia, 29±4 % of the cells underwent apoptosis; while overexpression of SRF reduced this percentage to 5.7±1 %; and knockdown of SRF increased it to 85±4 % (all P < 0.001).

Overexpression of SRF increased Bcl-2/Bax ratio by 23±4 % at baseline and by 167±20 % after 24-hour hypoxia; knockdown of SRF decreased these ratios by 69±3 % and 75±8 %, respectively (all P < 0.01). Knockdown of SRF also increased Bax translocation to mitochondria and Cytochrome C release.

Conclusions
:

Hypoxia triggers SRF activation.

Up-regulation of SRF promotes angiogenesis.

Down-regulation of SRF sensitizes cells to hypoxia-induced apoptosis.

SRF protects against hypoxia-induced apoptosis likely through both direct up-regulation of Bcl-2 and indirect down-regulation of Bax.

This research has received full or partial funding support from the American Heart Association, Western States Affiliate (California, Nevada & Utah).