bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 5810: Perivascular Cells Are a Potential Source of Chondrocyte-Like Cells in Atherosclerotic Lesions of Apolipoprotein E Deficient Mice
Authors: Preusch Michael R , Srirangam Kiranmayi , Ieronimakis Nicholas , Callegari Andrea , Reyes Morayma , Rosenfeld Michael E
Published: April 2024

Introduction:
The presence of chondrocyte-like cells is a common phenomenon in mouse models of advanced atherosclerosis. However the source of those cells is still unclear. Since stem cell antigen-1 (Sca-1) positive cells have been found in the adventitia of apoE−/− mice and chondrogenic transformation of mesenchymal stem cell lineages has been described, we hypothesized that a possible source of the chondrocyte-like cells are perivascular Sca-1 positive cells.

Methods:
Innominate arteries of 23 weeks apoE−/− mice were immunostained for Sca-1, CD 45, CD 31 and Mac-2 and analyzed by fluorescence microscopy. The thoracic aortas, aortic arches and innominate arteries of apoE−/− mice and C57BL/6 controls were harvested from 13, 23 and 34 week old mice. The adventitia and attached perivascular tissue and the remaining aorta were both enzyme digested. The isolated cells were stained for Sca-1, CD 45, CD 31, CD 34, CD11b and F4/80 and sorted by flow cytometry using positive selection for Sca-1, CD 34 and negative selection for CD 45, CD 31, CD 11b and F4/80. RNA was isolated from the sorted cells and the expression of Cbfa1/Runx2 was determined by PCR.

Results:
Sca-1 positive, CD 45, CD 31 and Mac-2 negative cells were observed in the vascular wall by immunofluorescence and were located within close proximity to the chondrocyte-like cells within the atherosclerotic lesions. FACS analyses confirmed the presence of these cells within the aorta, aortic adventitia, aortic arch and the innominate artery of the apoE −/− mice and BL6 controls. Furthermore the perivascular cells also express Cbfa1/Runx2.

Conclusions:
Our data suggest that perivascular derived Sca-1 positive cells exist within both atherosclerotic lesions and within arteries from wild type controls of different ages. These cells also express Cbfa1/Runx2, a transcription factor associated with osteogenesis and therefore may be a source of the chondrocyte-like cells found in adavanced lesions of apoE−/− mice.
Abstract 5812: A Novel Role of Insulin Degrading Enzyme (IDE) in Atherosclerosis: Studies Using IDE
−/−
Bone Marrow-Derived Cells Show Increased Lesion Development in a Bone Marrow Transplant LDL Receptor
−/−
Mouse Model
Authors: Caravaggio Justin W , MacLaren Robin , Hasu Mirela , Thabet Mohamed , Whitman Stewart C
Published: April 2024

Insulin degrading enzyme (IDE) hypofunction has been shown to promote Alzheimer’s disease as well as type II diabetes in humans and animal models, but it is unknown if IDE has a role in atherosclerosis. The scavenger receptor-A (SRA) plays a key role in foam cell formation as well as fatty streak development. Using affinity chromatography we previously identified a novel interaction between SRA and IDE from J774 cytosolic lysate. Utilizing a bone marrow transplant model, we tested the hypothesis that IDE knockout in bone marrow derived cells would increase atherosclerotic lesion severity in C57BL-6 mice on an LDLr
−/−
background fed a high fat diet. At 8 weeks of age, male and female LDLr
−/−
recipient mice underwent
γ
-irradiation to eliminate endogenous bone marrow. Recipient mice were repopulated with bone marrow from either wild type or IDE
−/−
, aged matched donor mice. Following 6 weeks of recovery, recipient mice were placed on a high fat diet (21% fat & 0.15% cholesterol) for 8 weeks to generate an atherosclerotic phenotype.
In vivo
results showed a significant increase in aortic arch lesion area in male mice (6-fold increased lesion area in IDE
−/−
(n=8) vs. WT (n=12), p<0.001) and a corresponding increase in aortic root lesion size (2.5-fold increased lesion area in IDE
−/−
(n=12) vs. WT (n=12), p<0.05). Total serum cholesterol was increased in male recipients repopulated with IDE
−/−
vs. WT (p<0.001).
In vitro
mechanistic studies using IDE
−/−
bone marrow-derived macrophages have revealed differences in adhesion (3.2-fold increase in non-adherent cells from IDE
−/−
mice vs. WT, p<0.05) as well as a difference in uptake and esterification of tritium-labeled oleate (
3
H) into cholesterol ester (2-fold increase in acLDL incorporation of
3
H oleate in IDE
−/−
cells vs. WT cells). This suggests IDE plays a key role in macrophage function and foam cell formation during lesion development. To conclude, IDE
−/−
bone marrow-derived cells resulted in significantly greater lesion development when transplanted into LDLr
−/−
male mice. These results indicate a prominent role for IDE in atherosclerotic lesion formation and suggest that IDE may provide a novel target for therapeutic intervention.
Abstract 16639: Acetylation and Stabilization of STAT3 Mediates Cardioprotection by p300 and HDAC Inhibitor Trichostatin A
Authors: Jain Sumit , Wei JianQin , Hilfiker-Kleiner Denise , Bishopric Nanette H
Published: April 2024

Background:
The nuclear acetyltransferase p300 is rapidly induced in the heart during various types of stress, and mice with 2.5x myocyte overexpression of p300 are protected against ischemia/reperfusion (I/R) injury. Transcription factor STAT3 is strongly cardioprotective, and is acetylated and coactivated by p300.

Hypothesis:
p300 mediates cardioprotection against oxidative stress via activation of STAT3.

Methods:
Oxidative stress was induced in neonatal rat cardiac myocytes with doxorubicin (DOX, 1µM). Protein synthesis inhibitor cycloheximide (CY) was used to assess p300 and STAT3 stability. p300 knockdown (p300KD) was achieved with p300-targeting siRNA and compared to non-targeting control (NS). STAT3 fl/fl, α-MHC-Cre+ (KO) and STAT3 wt/wt, α-MHC-Cre+ (WT) littermates were subjected to I/R injury, 1 h after injection of a singe dose of the class I/II HDAC inhibitor TSA or its vehicle.

Results:
p300 protein rapidly increased to high levels after DOX treatment, and remained elevated for ≥ 16 h (3.7±1.9x of control at 8h; p<0.05), entirely through increased p300 stability (5.53±1.87x after 6h CY; p<0.05). Acetyl-STAT3 and STAT3 half-life increased in parallel (1.89±0.45 fold; p<0.05), as did activating phosphorylation of STAT3 at Tyr 705 and Ser 727. Conversely, p300 loss abrogated the effect of DOX on STAT3 content, phosphorylation, acetylation, and half-life (0.68±0.07 fold; p<0.05), accompanied by reduced cell survival and blunted DNA damage response. p300 loss led to downregulation of multiple STAT3-regulated genes (e.g. MMP9, IL1RN, SERPINA3) vs. NS cardiomyocytes. During I/R in vivo, treatment with TSA increased p300-specific HAT activity and p300 protein levels (p<0.001), associated with 32.72±15.73% reduction in infarct volume (P<0.001, n = 4) in mice, however TSA provided no cardioprotection in KO mice (3.25%, p=0.09, n =4).

Conclusion:
Oxidative stress acutely stabilizes p300 in cardiac myocytes, which is required for acetylation, activation and stabilization of STAT3 and expression of STAT3-dependent genes. TSA activates p300 during ischemia-reperfusion and requires STAT3 for its cardioprotective effects. STAT3 is a critical downstream effector for both TSA- and p300-mediated protection against acute oxidative stress.
Abstract 16686: Early Gadolinium Enhancement Represents Area at Risk Not Infarct Size: Timing and Kinetics Are Critical in Interpreting MRI of Acute Myocardial Infarction
Authors: Leung Steve W , Hsu Li-Yueh , Berry Colin , Wilson Joel R , Kellman Peter , Arai Andrew E
Published: April 2024

Background:
Recent papers have raised questions about whether gadolinium over-estimates infarct size and have indicated that early gadolinium enhancement (EGE) corresponds to the area at risk (AAR) rather than infarct size.Objective: To determine kinetics, timing, and size of early gadolinium enhancement as related to measurements of area at risk and infarct size in patients with acute myocardial infarction (MI).

Methods:
MRI was performed on 35 patients within 7 days of acute MI. T2-weighted images were acquired with bright blood methods to measure AAR. EGE images were acquired 1, 3, 5, and 7 minutes post-contrast with single-shot phase sensitive inversion recovery (PSIR). Late gadolinium enhanced (LGE) images were obtained 10-20 minutes post-contrast using segmented PSIR. EGE images were evaluated semi-quantitatively using a 2-SD threshold and with manual planimetry. LGE Infarct size was measured with the previously validated FACT algorithm. Microvascular obstruction was included in the hyperenhanced zone for EGE and LGE.

Results:
Of the 35 patients (26 were male, age 61±12 years old), 14 had subendocardial MI and 21 had transmural MI. For subendocardial MI, the correlation between EGE and T2-weighted AAR was only fair at 1 minute post contrast (r=0.68), but improved to (r=0.86-0.88) from 3 minutes to 7 minutes post-contrast administration. However, there was significant intersubject variability with regard to the optimal timing for EGE as a measure of AAR. For subendocardial MI, the best agreement between EGE and T2 weighted AAR occurred at 3 minutes in 4 subjects, 5 minutes in 4 subjects, and 7 minutes in 5 subjects. For transmural MI (n=21), the best agreement occurred at 7 minutes post-contrast. The maximal area of hyperenhancement on EGE correlated most closely with T2 AAR for subendocardial MI (r=0.91) and was significantly larger than MI size on LGE images (p<0.001 for all MI and for subendocardial MI).

Conclusion:
Myocardial gadolinium enhancement kinetics influence the size of the hyperenhanced zone such that early gadolinium enhancement between 3-7 minutes closely correlates with area at risk, and thus can over-estimate infarct size for non-transmural MI. Timing is critical when quantifying acute infarct size with gadolinium enhanced MRI.
Abstract 16687: The Incidence and Etiology of GI Bleeding in Patients Supported by Continuous Flow Left Ventricular Assist Device
Authors: Uriel Nir , Sharaiha Reem , Murray Lindsay , Yang Jonathan , Devaraj Kalpana M , Colombo Paolo C , Sims Daniel B , Gonzalez-Costello Jose , Takayama Hiroo , Mancini Donna , Naka Yoshifumi , Jorde Ulrich P
Published: April 2024

Introduction
Gastrointestinal (GI) bleeding is a frequent adverse event reported with continuous flow LVADs which may be explained by acquired von Willebrand disease and antithrombotic therapy. The aim of this study was to determine the incidence and etiology of GI bleeding among patients supported by a HeartMate II (HM II) LVAD.

Methods
A retrospective chart review was performed for all HM II patients implanted from 2004 to 2011. GI work-up included gastroscopy, colonoscopy, capsule studies, and small bowel enteroscopy and was reviewed by a gastroenterologist and a pathologist. GI bleeding was defined as occult or obvious blood in the stool with concomitant drop in hemoglobin.

Results
142 patients aged 56.7±14.9 years were implanted with a HM II. 77% of patients were male and the etiology was ischemic cardiomyopathy in 44%. 74% of LVADs were placed as a bridge to transplant. The rate of GI bleeding was 0.36 events / year. A total of 51 events occurred in 28 patients. The indications for GI work-up were melena (46% of patients), hematochezia (14%), heme-positive stool (14%) and iron deficiency anemia (25%). Full GI work-up (gastroscopy, colonoscopy and capsule study) was performed in 7 patients; 12 patients had gastroscopy and colonoscopy only; and 7 underwent only gastroscopy. The source of bleeding was identified in 71% of patients. Duodenal ulcer was the most prevalent identified source and occurred in 21% of patients (see figure). Only 5 patients had therapeutic interventions performed. Gastric biopsy was performed in 7 patients of which 6 had reactive features including mucous depletion, foveolar hyperplasia, fibromuscular hyperplasia of the stroma, and dilated or congested capillaries.

Conclusions
A high incidence of GI bleeding was found among patients supported with a continuous flow device, with gastric ulcers as the most prevalent etiology. Evaluation for H. pylori infection and proton pump inhibitor use should be studied in continuous flow LVAD patients.
Abstract 16688: Human Cardiosphere-Derived Cells Stimulate Cardiomyocyte Proliferation
in vivo
and in Co-Culture
Authors: Xie Yucai , Cheng Ke , Cho Hee Cheol , Malliaras Konstantinos , Ibrahim Ahmed , Sun Baiming , Galang Giselle , Ionta Vittoria , Shen Deliang , Zhang Yiqiang , Marbán Eduardo
Published: April 2024

Background:
Cardiosphere-derived cells (CDCs) are effective in regeneration after myocardial infarction (MI); however, direct differentiation of transplanted cells only accounts for a minuscule fraction of the benefit.

Methods and Results:
SCID mice underwent LAD ligation and were injected with 10
5
human CDCs or vehicle. Two weeks later, hearts were explanted for myocyte isolation by Langendorff perfusion, or for tissue analysis. Immunohistochemistry revealed decreased numbers of apoptotic cells in the MI border zone of CDC-treated animals vs. controls (14.2 ± 2.9% vs. 32.8 ± 5.2% TUNEL+ cells; n=3; P<0.01). Myocytes isolated from CDC-treated hearts demonstrated increased numbers of cycling cardiomyocytes (1.3 ± 0.1% vs. 0.6 ± 0.4% α-sarcomericactinin/Ki67 double positive cells; n=3; P<0.05). To probe the mechanism of the increase in cycling (Ki67+) myocytes, we co-cultured human CDCs or dermal fibroblasts (DFs) with neonatal rat cardiomyocytes (NRCMs). After 3 days, Ki67+ NRCMs were increased in the CDC co-cultures (22.8 ± 0.8% vs. 9.5 ± 4.7% in DF cocultures; n=3; P=0.004). Cycling NRCMs were smaller than non-cycling NRCMs (cross-sectional area 2.3 ± 1.3E+09 vs. 3.4 ± 1.5E+09 nm
2
; n=11; P<0.05), and the number of beating NRCM clusters was higher in CDC co-cultures (56.5 ± 15.7% vs. 28.0 ± 13.0%; n=3; P<0.05). To investigate paracrine effects, we cultured CDCs (or DFs) and NRCMs in a transwell contact-free system. After 3 days of contact-free co-culture, there were more Ki67+ NRCMs in the CDC group (15.6 ± 2.6% vs. 11.5 ± 1.5% in DF controls; n=3; P<0.05), while cycling NRCMs were again smaller (P<0.01) than non-cycling NRCMs. However, the percentage of cycling NRCMs was less than that in mixed co-cultures (15.6 ± 2.6% vs. 22.8 ± 0.8%; n=3; P<0.01).

Conclusions:
CDCs decrease apoptosis and increase the number of cycling myocytes after MI
in vivo
. CDCs stimulate cardiomyocyte proliferation
in vitro
; the enhancement of proliferation is greater in mixed co-culture than with conditioned media. Although soluble paracrine factors are involved, contact-dependent cell-cell interactions may also play an important role.
Abstract 16717: Human Gene Copy Number Spectra Analysis in Congenital Heart Malformations
Authors: Tomita-Mitchell Aoy , Mahnke Donna K , Struble Craig A , Tuffnell Maureen E , Hidestrand Mats , Stoddard Alexander J , Harris Susan , Goetsch Mary A , Lorier Rachel B , Simpson Pippa M , Bick David P , Broeckel Ulrich , Pelech Andrew N , Tweddell James S , Mitchell Michael E
Published: April 2024

Aim:
To visualize copy number associations and quantify relative risk of Copy Number Variants (CNVs) in 100 well-defined congenital heart disease (CHD) risk genes identified from literature sources.

Method:
CNVs in subjects with congenital heart malformations (n=741) and phenotyped in accordance with the European Paediatric Cardiac Code were compared to the frequencies of CNVs present in a control population of subjects with coronary artery disease (n=880). CNV gains (≥ 200 kb) and losses (≥ 100 kb) were determined using the Affymetrix 6.0 Gene Chip and confirmed with TaqMan copy number assays. In addition, each gene was tested using a two proportions z-test approach to determine if the gene frequency was significantly different in CHD vs control. To quantify the effect of observed CNVs in the CHD population, we collapsed the set of rare individual CNVs into a single group to test the collective frequency difference between cases and controls by adapting a previously described Cohort Allelic Sums Test (CAST).

Results:
The following phenotypes showed a significant enrichment of gains and losses in the CHD population: Aortic Stenosis, Coarctation of the Aorta, Ebstein's Anomaly, Tetralogy of Fallot, Truncus Arteriosus and Ventricular Septal Defect. The CNV percentage was increased in CHD subjects in the following genes (p<0.05 with a Bonferroni correction): Losses: FKBP6(-), ELN(s), GTF2IRD1(-), SOX7(-), GATA4(ns), NOTCH1(ns), MYH11(ns), SALL4(s), TBX1(s), CRKL(-), MAPK1(-); Gains: PRKAB2(-), FMO5(-), CHD1L(-), BCL9(-), ACP6(-), GJA5(ns), NPHP3(s), FOXL2(s), SEMA5A(-), NSD1(s), HOXA1(s), TBX20(ns), SOX7(-), GATA4(ns), HRAS(s), GATA6(ns), RUNX1(s), MAPK1(-), BCOR(s), ATRX(s), GPC3(s), ZIC3(ns), FLNA(s) where s is syndromic, ns is non-syndromic and (-) is not available on the CHD WIKI website (
http://www.chearted.eu
). The approach rapidly identified common, cytogenetically visible abnormalities (i.e. TBX1 in 22q11.2 deletion syndrome, n=30, and RUNX1 in Trisomy 21, n=66).

Conclusion:
This is the first study to establish a copy number spectrum in a CHD population. We conclude that the CAST approach shows promise for evaluating qualitative outcomes and statistical precision of prior studies which have focused on CNVs in CHD and other diseases.
Abstract 16718: The Impact of Discontinuation of Dual Antiplatelet Therapy and Stent Fracture or Peri-Stent Contrast Staining on Very Late Stent Thrombosis After Sirolimus-Eluting Stent Implantation
Authors: Fuku Yasushi , Kadota Kazushige , Hasegawa Daiji , Shigemoto Yoshikazu , Habara Seiji , Tada Takeshi , Tanaka Hiroyuki , Oka Naoki , Goto Tsuyoshi , Mitsudo Kazuaki
Published: April 2024

Objectives:
We evaluated the impact of discontinuation of dual antiplatelet therapy (DAPT) and stent fracture (SF) or peri-stent contrast staining (PSS) on very late stent thrombosis (VLST) after sirolimus-eluting stent (SES) implantation.

Methods:
Consecutive patients who had received the first SES implantation and had been evaluated by follow-up angiography within 12 months were analyzed. Stent thrombosis was defined according to the Academic Research Consortium definition. The incidence of VLST beyond 1 year after SES implantation was compared between the 4 groups of patients either with or without a diagnosis of SF or PSS at follow-up angiographic study within 12 months and continuation or discontinuation of DAPT.

Results:
Of 2494 patients who received the first SES implantation between November 2002 and December 2007, 2048 patients met the criteria for inclusion. The median duration of follow-up after the SES implantation was 1680 days (interquartile range 1362-1963 days). SF was observed at follow-up angiography within 12 months in 243 patients (11.9%), PSS in 59 patients (2.9%) and either SF or PSS (SF/PSS) in 271 patients (13.2%). Figure shows that the 5-year cumulative incidence of definite ST in patients stopped DAPT was significantly higher than that in patients continuing DAPT in the SF/PSS (+) group (11.4% vs. 0 %, p<0.001), whereas no differences were observed in patients stopped DAPT versus patients continuing DAPT in the SF/PSS (-) group (0.7% vs. 0.3%, p=0.29). The incidence of definite ST in patients stopped DAPT was also significantly higher than that in patients continuing DAPT in the SF (+) group and PSS (+) group, respectively (9.9% vs. 0%, p=0.001; 19.1% vs. 0%, p=0.041).

Conclusions:
DAPT should be continued when SF or PSS was observed at follow-up angiography after SES implantation. In contrast, it may be possible that DAPT will be discontinued when SF or PSS was not observed.
Abstract 16723: Dopamine D
3
Receptor Decreases NHE3 Expression and Function by Inhibiting the Activity of the De-Ubiquitinylating Enzyme, USP48, in Renal Proximal Tubule Cells
Authors: Armando Ines , Villar Van Anthony M , Wang Xiaoyan , Asico Laureano , Li Hewang , Escano Crisanto S , Felder Robin , Jose Pedro A , Jones John E
Published: April 2024

Disruption of the dopamine D
3
receptor gene in mice (D
3
R
-/-
) results in high blood pressure, salt sensitivity, decreased ability to excrete a sodium load and increased renal expression of sodium/hydrogen exchanger isoform 3 (NHE3) protein (180%, P<0.05) but not mRNA, and D
3
R blockade in wild-type mice increased (2 fold, P<0.04) NHE3 protein expression. We hypothesized that renal D
3
R regulates NHE3 expression/function. In human renal proximal tubular cells (hRPTCs), the colocalization and co-immunoprecipitation of D
3
R and NHE3 are increased by D
3
R stimulation (PD 128907, 1μM), associated to internalization of D
3
R and NHE3 (confocal microscopy) and inhibition (80%, P<0.05) of NHE3 dependent Na
+
transport. D
3
R stimulation also increases NHE3 ubiquitinylation and internalization of the ubiquitinylated NHE3 in the cytoplasm (150%, P<0.05) suggesting that D
3
R activation promotes NHE3 ubiquitinylation and internalization. To determine the mechanism of the D
3
R-mediated NHE3 ubiquitinylation we performed a yeast-two hybrid screening which showed that the 3
rd
intracytoplasmic loop of human D
3
R interacts with human ubiquitin-specific peptidase 48 (USP48), a de-ubiquitinylating enzyme. D
3
R and USP48, and USP48 and NHE3 co-localized and co-immunoprecipitated in hRPTCs. USP48 accounted for ~30% of hRPTCs' total de-ubiquitinylating activity. D
3
R stimulation increased the interaction between D
3
R and USP48 and resulted in a 35% reduction in USP48 de-ubiquitinylating activity, indicating that D
3
R inhibits USP48 activity in hRPTCs. USP48 silencing increased the degradation of NHE3 (vehicle t
1/2
= 6.1 hr; USP48 siRNA t
1/2
= 12.9 hr; P<0.05) while over-expressing USP48 increased NHE3 half -life (t
1/2
= 21.8 hr; P<0.05 vs. vehicle) indicating that USP48 protects NHE3 from degradation via de-ubiquitinylation. Moreover, selective renal USP48 silencing in mice by subcapsular infusion of USP48 siRNA decreases NHE3 expression (25%, P<0.04) and systolic blood pressure measured under anesthesia (vehicle: 102±2 vs. siRNA 78±2 mmHg; P<0.03). Our data demonstrate that agonist activation of D
3
R promotes the degradation of NHE3 via the inhibition of USP48 and thus provide a novel mechanism by which the D
3
R engenders natriuresis and regulates blood pressure.
Abstract 16735: Role of the Autonomic Nervous System in Cardioprotection by Remote Ischemic Preconditioning
Authors: Donato Martin , Buchholz Bruno , Gelpi Ricardo J
Published: April 2024

The mechanisms underlying remote ischemic preconditioning (r-IPC) have not been fully elucidated. In particular, it is unknown how the cardioprotective signal is transmitted from the preconditioned remote organ to the heart. We hypothesize that r-IPC activates a neural pathway, and that the cardioprotective signal reaches the heart via the vagus nerve, preconditioning the heart through the activation of muscarinic receptors. Our aim was to determine the possible role of the vagus nerve and muscarinic receptors in the mechanisms of r-IPC cardioprotection.

Methods:
anesthetized rabbits (n=6, r-IPC) were subjected to 3-cycle hind limb ischemia (5 min) and reperfusion (5 min). The hearts were then excised and perfused (Langendorff technique). Myocardial infarction was induced by 30 min of global ischemia followed by 2 h of reperfusion. In the non-r-IPC group (n=5), animals were subjected to a sham operation and then received the same protocol as group 1. Group 3 (r-IPC with vagal section, n=4) received the same protocol as group 2, but the left and right vagus nerves were sectioned before r-IPC. Group 4 (r-IPC with atropine, n=4) received the same protocol as group 2, but atropine was administered during the r-IPC protocol. In group 5 (vagal stimulation, n-4), the right vagus nerve was sectioned and efferently electrically stimulated. Then, the same protocol as in group 1 was performed.

Results:
In the non-r-IPC group, infarct size was 39.9±4.8%. r-IPC decreased infarct size to 22.7±5.7% (p<0.05). Vagal section and atropine abolished the effect of r-IPC, reaching an infarct size of 53.2±3.5% and 39.8±2.1, respectively (p<0.05). Vagal stimulation decreased infarct size to 18.1±4.7% (p<0.05).

Conclusion:
r-IPC activates a neural afferent pathway and the cardioprotective signal reaches the heart via the vagus nerve (efferent via). Acetylcholine acting on muscarinic receptors thus activates the IPC phenomenon.
Abstract 16736: Medical vs. Surgical Management of Perivalvular Abscess in Infective Endocarditis: A Propensity Analysis From the International Collaboration on Endocarditis Prospective Cohort Study (ICE-PCS)
Authors: Dickerman Stuart A , Barsic Bruno , Chu Vivian H , Fowler V G , Pappas Paul A , Bayer Arnold
Published: April 2024

Introduction
Myocardial abscess is generally regarded as an indication for surgical intervention in infective endocarditis (IE). There is little information, however, comparing outcomes with medical vs. surgical management of such periannular complications of IE. This study used the database of the ICE-PCS to perform a propensity analysis, comparing one year mortality in patients with IE and abscess subjected to either medical or surgical treatment.

Methods
ICE-PCS enrolled 4794 patients between 2000 and 2006, from 64 centers in 28 countries. There were 699 cases of IE with abscess. A statistical model was built to predict surgery in this cohort.158 patients who underwent surgery for abscess were matched to 158 patients who did not. These 316 patients formed the basis of the analysis.

Results
In the unadjusted group of 699 patients, the one-year survival with medical treatment was 45%, vs. 67% with surgical management (p<.0001). Differences favoring surgery were similar whether or not the patients had poor prognostic indicators (S. aureus infection; prosthetic valve involvement; CHF). Patients with strep viridans infection, however, had similar survival with medical or surgical management (70%, 14/20 vs.77%, 65/84). In the propensity analysis, the one-year survival with medical management was 44%, compared to 58% in the surgical group (p=.0021). (See graph).

Conclusion
One-year survival of IE patients with abscess was superior with surgical compared to medical management. These findings support current recommendations that surgical management of perivalvular extension of infection is generally the preferred approach. Patients with strep viridans infection, however, seem to fare equally well with medical or surgical treatment.
Abstract 16737: Maternal Hypertension is More Often Observed Among Hypertensive Patients, and Associates to SNPs of the X Chromosome
Authors: Iaccarino Guido , Uliano Federica , Trimarco Valentina , Izzo Raffaele , Pagnotta Stefano , Tremigliozzi Rosaria , Rivera natalia , de Luca Nicola , Marino marina , Finelli Rosa , Tagliamonte Virginia , Condorelli Gianluigi , Ceccarelli Michele , Trimarco Bruno
Published: April 2024

Family history of hypertension (HTN) is present in as many as 70-80% of all patients with primary HTN. It was recently noted that in some selected pedigrees, HTN is transmitted according to a diaginic pattern. However, it is not known whether the general population, differences can be observed between maternal or paternal familiarity in the transmission of HTN. We analyzed the CampaniaSalute database of the Hypertension Clinic of the Federico II University of Naples in Southern Italy, comprising 12.520 (6.868 males and 5.636 females) cases, which were compared with 6.352 non HTN patients present in database. For both groups we calculated the “odd” value for a family history of HTN. Among the HTN group, family history is present in 75% of cases (odds 3.77), whereas in the Non HTN group, occurs in only 26%, (0.94). The odds ratio (OR) indicates that familiarity increases the risk of developing hypertension 2.91 times (2.67-3.17, p <0.001). We found that in mothers HTN (HTN-M) occurs in 32% of cases [OR 3.01 (2.66-3.41, p <0.001)], in fathers (HTN-F) occurs in 18% [2.31 (2.01-2.68, p <0.001)] and in both parents (HTN-Both) in 15% of cases [3.45 (2.87-4.01, p<0.001)]. Such a pattern is similar between male and female patients. Given the higher prevalence of maternal familiarity, we assessed in 375 male HTNs (which bear the same X chromosome of the mother), the presence of polymorphisms on the sexual chromosome that associate with maternal history of HTN, using the Illumina 300K chip technology. In particular, four polymorphisms (ie, rs2182289, rs1317098, rs2346677, and rs2346678) showed a different distribution with a p value <0.0005, being less frequent in male patients with HTN-M compared with male patients with no HTN in the family (respectively 22.1 vs 36.4%, 16.4 vs 30.6%, 9.0 vs 19.4%, 8.6 vs 19.4%). A similar pattern although less significant was observed in female patients, which bear x chromosomes from both parents. These results suggest that polymorphisms on the X chromosome associate with the maternal transmission of HTN.
Abstract 16785: Molecular Mechanisms of the Crosstalk Between Mitochondrial and NADPH Oxidase Derived Reactive Oxygen Species in White Blood Cells - Implications for Cardiovascular Diseases
Authors: Wenzel Philip , Schuhmacher Swenja , Oelze Matthias , Scholz Alexander , Daub Steffen , Knorr Maike , Xia Ning , Otto Christian , Opitz Benjamin , Schulz Eberhard , Li Huige , Lackner Karl , Münzel Thomas , Daiber Andreas
Published: April 2024

Background:
Many diseases and drug-induced complications are associated or even based on an imbalance between the formation of reactive oxygen and nitrogen species (ROS/RNS) and antioxidant enzymes catalyzing the break-down of these harmful oxidants. Previously we and others have reported on a crosstalk between different sources of oxidative stress. With the present study we sought to determine the underlying mechanism for this crosstalk.

Methods and results:
Human neutrophils were used to study the induction of Nox-dependent oxidative burst by mitochondrial ROS (mtROS) in response to antimycin A or myxothiazol. Activation of Nox2 was measured by extracellular detection of ROS by a peroxidase-coupled chemiluminescence assay. We used different inhibitors for the involved signaling cascade such as cyclosporine A (inhibits the mitochondrial permeability transition pore [mPTP]), chelerythine (inhibitor of protein kinase C) and apocynin (inhibitor of NADPH oxidase). All of these inhibitors suppressed the antimycin A/myxothiazol-triggered extracellular ROS signal, whereas exogenous hydrogen peroxide mimicked the effect of mitochondrial ROS formation and caused extracellular superoxide formation (lucigenin ECL). According to preliminary data, this crosstalk between mtROS and NADPH oxidase was also suppressed in white blood cells from mice deficient of p47phox (an essential subunit for activation of Nox2) or cyclophilin D (a regulatory subunit of the mPTP), whereas this crosstalk was amplified in white blood cells from GPx-1 deficient mice or from MnSOD+/- mice. We also observed that increases in blood pressure, in endothelial dysfunction as well as in NADPH oxidase activity were more pronounced in MnSOD+/- mice as compared to their wild type littermates.

Conclusions:
Previous data have demonstrated that NADPH oxidase derived ROS/RNS stimulate mitochondrial oxidative stress contributing to overall angiotensin-II dependent cellular dysfunction. Our data now show that mitochondrial ROS trigger the activation of NADPH oxidase, which may have severe effects on progression of cardiovascular diseases representing a feedback loop creating a vicious circle of oxidative stress.
Abstract 16807: New Approach to Pressure-Volume Loop Analysis Using Custom Made High Fidelity MRI Compatible Pressure Sensor Tipped Catheters and Real Time MRI
Authors: Dori Yoav , Zviman Menekhem , Miller Frank , Halperin Henry , Rome Jonathan J
Published: April 2024

Background:
Pressure-Volume (PV) loop analysis remains the only load-independent method to assess cardiac function. However, current methods limit the clinical use of PV loops particularly in congenital defects. We developed a novel technique for PV loop analysis using custom made high fidelity MRI compatible pressure sensor tipped catheters (hfMRIc) with real time MRI (rtMRI).

Hypothesis:
Accurate PV loop cardiac function can be obtained using single plane rtMRI imaging and hfMRIc pressure sensor tipped catheters.

Methods:
PV loop analysis was performed in the LV of 4 swine. Instantaneous ventricular volume estimation was obtained with the Simpson method using V = ∑Vi(Li) where Vi(Li) are length volume relations derived from a short axis cine stack and a LA cine plane and Li are lengths obtained using custom made segmentation software and a single rtMRI LA plane (Figure 1(a)). Volume error calculations were performed at two different preload conditions. Pressure measurements were obtained with custom made hfMRIc fiber optic pressure sensor tipped catheters. Pressure measurements were compared to values obtained with a high fidelity Millar catheter positioned in the Aorta.

Results:
At a sampling rate of 20 Khz, the pressure difference between the hfMRIc catheters and the Millar catheter was 1% with a standard deviation (SD) of 0.9%. There was no effect of MRI scanning on pressure recording. The difference between the volume measured with a short axis cine stack and the volume estimated with our methodology using a single LA cine plane was 6.7% with a SD of 4.7%. EDPVR and ESPVR and instantaneous PV loops were obtained by inflating a balloon in the IVC while obtaining rtMRI planes at a temporal resolution of 42 msec (Figure 1(b)).

Conclusions:
Using this new technique, we can accurately assess the instantaneous cardiac function using PV loops. Unlike current technology this new modality is not dependent on ventricular shape or valve function.
Abstract 16819: Intracellular Dyssynchrony in Ca Removal from the Cytosol In Ventricular Cardiac Myocytes
Authors: Hohendanner Felix , McQuaide Niall , Antoons Gudrun , Sipido Karin , Pieske Burkert , Heinzel Frank R
Published: April 2024

In cardiomyocytes, cytosolic Ca removal is mediated by the sarcoplasmic reticulum Ca ATPase (SERCA), sarcolemmal Na/Ca exchanger (NCX), plasmalemmal Ca ATPase and mitochondrial Ca uniporter (MCU). We report, for the first time, spatiotemporal inhomogeneities in cytosolic Ca removal in normal and diseased heart and explore underlying mechanisms. Spatially resolved Ca transients (1 Hz, Fluo-4 AM) were recorded at steady state using confocal microscopy in ventricular cardiomyocytes. The time constant of cytosolic [Ca] decay (tau_loc) of local Ca transients was quantified at 1 µm intervals across the cell. In murine cardiomyocytes tau_loc distribution was inhomogeneous, with a maximal difference between cell regions of 237±9 ms and a variation coefficient (CV_tau) of 14±2% (mean±S.E.M., n=10 cells). tau_loc was not releated to the local amplitude or release kinetics of Ca. Coherent regions of fast (fastCaR, tau_loc < mean tau of whole cell) and slow (slowCaR) Ca removal had similar widths (4.6±0.2 vs. 5.1±0.5 µm). Forskolin accelerated, and SERCA inhibitor cyclopiazonic acid (CPA) inhibited Ca removal significantly more in slowCaR than in fastCaR. In contrast, NCX-inhibitor SEA0400 slowed cytosolic Ca removal similarly in slowCaR and fastCaR. Also, CV_tau was similar in NCX+/- knock-out vs. wild-type mice, suggesting no contribution of NCX to the dyssynchrony of cytosolic Ca removal. tau_loc distribution was also inhomogenous in pig ventricular myocytes. Ca removal from the cytosol was more dyssynchronous in cardiomyocytes from chronic (4 weeks) ischemic pig myocardium (peri-infarct zone) vs. sham-operated pigs (CV_tau 24±1% vs. 20±1%, n=57 cells/group, p<0.05). Simultaneous recording of Ca transients and mitochondrial signal (mitotracker) unveiled a significant correlation between FastCaR regions and mitochondria (n=9 cells, p<0.05). Ru360 (MCU-inhibitor) slowed tau_loc more in FastCaR (to 126±12% vs. 112±10% of baseline in SlowCaR; p<0.05, n=5).

Conclusion:
Cytosolic Ca removal is dyssynchronous in mouse and pig cardiomyocytes. Ca removal in chronically ischemic cardiomyocytes is more dyssynchronous, suggesting a potential new mechanism of cardiomyocyte dysfunction. Mitochondria influence the differences in local Ca reuptake.
Abstract 16853: Endoglin is an Epigenetically Regulated Novel Endothelial Specification Gene
Authors: Banerjee Saswati , Bacanamwo Methode
Published: April 2024

Recent studies indicate that Endothelial cells (EC) are required for the engineering of all tissues/organs and their dysfunction mediates the pathogenesis of most cardiovascular diseases. However, EC production remains inefficient and identifying novel EC specification genes remains a priority. Dnmt1-/- embryonic stem cells (ESC) where found to have a higher propensity to differentiate into EC than wild type EC. In addition, endoglin (Eng), recently implicated in early vascular development and angiogenesis, was methylated and repressed in WT ESC, but was demethylated and expressed in Dnmt1-/- ESC. Interestingly, a CpG site located at a locus predicted to bind HIF1 and Sp1, the two main transcription factors regulating Eng transcription, was not methylated in Dnmt1-/- ESC, but was 80% methylated in WT ESC. We hypothesized that Eng may be an epigenetically regulated EC specification gene. Mouse ESC were treated with recombinant Eng or the Eng ORF, and differentiated in the presence or absence of BMP4. Expression of the mesoderm and EC marker genes, and known mediators of EC specification and their downstream targets was monitored by QRT-PCR, western, immunocytochemistry, and flow cytometry. Functionality of the differentiated EC was assessed by in vitro angiogenesis assay and the induction of ICAM-1 expression in response to TNF-α. The percentage of Tie2-positive cells was increased by recombinant Eng relative to vehicle-treated cells (17.1 ± 2.5 vs. 26.0 ± 1.5, p≤0.01). BMP4 also increased the percentage of Tie2-positive cells relative to controls (17.1 ± 2.5 vs. 26.4 ± 1.3, p≤0.01), and the percentage was further increased by combining BMP4 with recombinant Eng (26.4 ± 1.3 vs. 34.5 ± 2.6, p≤0.01). Similar results were obtained with forced Eng expression and with the other EC markers Flk1, vWF, and CD31. The Eng-induced differentiation was independent of known mediators of EC specification such as Indian Hedgehog and BMP4 or of BMP4/phospho-Smad1/5/8 pathway. The effect of CpG methylation at the HIF1/Sp1 binding site on the binding of these factors and transcriptional regulation of Eng are being investigated. The data indicates that Eng is a novel epigenetically regulated EC specification gene, which induces robust EC production in combination with BMP4.
Abstract 16891: The Apelin Receptor is an Allosteric Inhibitor of the Angiotensin II Type 1 Receptor
Authors: Smith Layton H , Siddiquee Khandaker , Hampton Jessica
Published: April 2024

Apelin and its receptor are emerging as an endogenous counter-regulator of the renin-angiotensin-aldosterone system. In experimental models, apelin opposes angiotensin II (Ang II)-mediated hypertension and cardiovascular fibrosis as well as the acceleration of atherosclerosis and the development of abdominal aortic aneurisms. In vitro studies show that apelin antagonizes Ang II signaling through the Ang II Type 1 receptor (AT1), but the precise molecular mechanism has not been elucidated. In the present study, we sought to test the hypothesis that the apelin receptor is a dominant negative regulator of AT1. Using co-immunoprecipitation and bioluminescence resonance energy transfer (BRET) techniques, we confirm that the apelin receptor (APJ) and AT1 physically interact to form a heterodimeric complex. The interaction between APJ and AT1 was increased by the APJ agonist apelin-13 (Ap13), but not Ang II. Ap13-mediated dimerization between APJ and AT1 eliminated Ang II-mediated recruitment of β-arrestin to AT1 and significantly decreased Ang II-induced Ca
2+
flux. Radioligand binding studies revealed that Ap13 reduced Ang II binding without direct competition. Taken together, these data lead us to conclude that apelin counter-regulates the renin-angiotensin-aldosterone system through the apelin-dependent heterodimerization and APJ-mediated negative allosteric regulation of AT1 function.
Abstract 16907: Modification of Hemeoxygenase-1 Expression Enhances the Therapeutic Potency of CD34+ Cells in Ischemic Tissue Repair
Authors: Tongers Jörn , Jujo Kentaro , Roncalli Jerome G , Renault Marie-Ange , Kamide Christine E , Klyachko Ekaterina , Thorne Tina , Bauersachs Johann , Amrani David L , Motlagh Delara , Losordo Douglas W
Published: April 2024

The potency of cell-based therapy in ischemic tissue repair is limited by low viability and retention of transplanted cells. The hemeoxygenase system catabolizes heme cleavage and has been shown to exert cytoprotection via its anti-oxidative, -apoptotic, -inflammatory properties. HO-1, inducible in various pathologic states, has been shown to influence progenitor cell biology. We hypothesized that HO-1 over-expression enhances the therapeutic potency of CD34+ cells for ischemic tissue repair. CD34+ from healthy donors were enriched from the peripheral blood mononuclear cell fraction of healthy donors after G-CSF mobilization (Isolex 300i). Adenoviral transduction of HO-1/GFP or GFP led to a substantial increase of HO-1 expression (HO-1/18s RNA expression: 62-fold increase, protein expression: 20-fold increase; n=3). After allowing 48 hrs for target gene expression, CD34+ cells were FACS-sorted for GFP followed by overnight recovery in-vitro. A low-therapeutic dose (10^5 cells/kg) of HO-1+/GFP+ (HO-1) or GFP+ (GFP) CD34+ cells was locally injected into the border zone after complete ligation of the left coronary artery in 8-week-old, male nude mice (saline control: 2x 15 μl). At day 28, LV ejection fraction of mice treated with HO-1 over-expressing CD34+ cells was significantly improved (fractional shortening: saline 19±2%, HO-1 30±1%, GFP 24±1%; n=6-8/group, P<0.01). This functional improvement can be explained by a preservation of the border-zone: the number of lectin+ capillaries was increased (lectin staining: saline 73±5/hpf, HO-1 99±4/hpf, GFP 79±7/hpf; n=7-8/group, P<0.01) with the use of HO-1 modified CD34+ cells while lectin+/alpha-smooth-muscle actin+ structures indicative for resistance vessels remained unchanged (P=ns). Apoptosis of resident myocardial cells was attenuated (TUNEL staining: saline 69±5/hpf, HO-1 31±3, GFP 54±9, P<0.05). As a result, the infarct size in mice treated with HO-1 modified cells was reduced (Masson-trichrome staining: saline 39±1%, HO-1 36±2%, GFP 42±1%; n=6-8/group, P<0.05). Ongoing studies are addressing the underlying mechanism(s) on cellular basis as well as the clinical relevance.

Conclusion:
HO-1 gene modification enhances the therapeutic potency of CD34+ in ischemic tissue repair.
Abstract 16966: MicroRNA-92a Differentially Regulates Cell Cycle Proliferation and Angiogenesis in Huvec And Hepg2 Cells
Authors: Adi Nikhil C , Adi Jennipher , Webster Keith A
Published: April 2024

Introduction:
The conserved miR17~92 cluster, is up-regulated in tumors, and its OE promotes tumor angiogenesis and growth. Recent studies indicate that miR92a is anti-angiogenic while the regulation of angiogenesis by miR17/20 is context-dependent.

Hypothesis:
miR92a action is also context and cell type-dependent and differentially regulates cell growth and angiogenesis in tumor vs endothelial cells.

Methods:
HepG2 and HUVECs were transfected with pre and anti miR92a molecules. Cell proliferation was assayed in both cell types using BrdU 48hr after transfection. HUVEC-Matrigel tube formation assays were implemented in parallel to assess angiogenic potential. VEGF was measured in spent media from HepG2 cells and the same media was used in HUVEC tube formation. Protein lysates were collected for WB 48hr after transfections of both cell types.

Results:
Pre-miR92a transfection inhibited proliferation of HUVECs while anti-miR92a, but not pre-miR92a inhibited proliferation of HepG2 cells (all, p<0.05). Pre-miR transfection blocked HUVEC tube formation (p<0.01). Tube formation was supported by spent medium from HepG2 cells ± transfection with pre-miR-92a but the ability of HepG2 spent medium to support tube formation was blocked by transfection with anti miR92a (all, p<0.05). Consistent with the latter result, VEGF levels in spent HepG2 media were decreased by transfection with anti-miR92a. In HepG2 cells transfection of anti-miR92a increased PTEN expression and decreased AKT-P-Thr308 (both p<0.05) PTEN and AKT-P-Thr-308 were not affected by miR92a transfection of HUVECs, however pre-miR-92a significantly decreased eNOS, ITG5α, and CCND1 protein levels (all, p<0.05). These results support our hypothesis that miR92a has opposite effect on the angiogenic properties of HepG2 vs HUVECs.

Conclusions:
MiR92a augments proliferation of HepG2 cells and promotes angiogenesis by inhibiting PTEN, activating Akt and increasing VEGF expression and secretion. Conversely, miR92a inhibits angiogenesis of HUVECs by targeting angiogenic regulators. The differential contextual functions of miR92a explain why this regulation supports tumor growth and angiogenesis while blocking EC function.
Abstract 16967: Preclinical Evaluation of Biopolymer-Delivered Endothelial Progenitor Cells in Hibernating Myocardium
Authors: Giordano Céline , Thorn Stephanie L , Suuronen Erik J , Renaud Jennifer M , Al-Atassi Talal , Boodhwani Munir , Zhang Pingchuan , Ascah Kathy J , DaSilva Jean N , deKemp Rob A , Beanlands Robert , Ruel Marc
Published: April 2024

Background:
Vasculogenic cell-based therapy combined with tissue engineering is a promising revascularization solution for patients with hibernating myocardium (HM), a common clinical condition. However, large animal studies are lacking and we therefore used a clinically relevant pig model of HM to examine the benefits of endothelial progenitor cells (EPCs) and biopolymer therapy in this context. We hypothesized that the delivery of EPCs within a matrix would yield greater improvements in myocardial blood flow (MBF), metabolism, and function than EPCs alone or PBS control.

Methods:
Twenty-three Yorkshire swine underwent placement of an ameroid constrictor on the left circumflex artery (LCx). After 2 weeks, animals underwent echocardiography, a rest and a dipyridamole-induced stress 13N-NH3 PET scan, and a rest 18FDG-PET viability scan. At the same time, EPCs were isolated and cultured for 1 week, and swine were randomized to intramyocardial injections (total: 3 ml) of PBS (n=9), EPCs (∼30 x 106; n=7), or EPCs + a collagen-based matrix (n=7). PET imaging was repeated 4 weeks post treatment, and swine were sacrificed the following week.

Results:
Hibernation was detected in 60% of animals and was specific to the LCx region (p=0.002). Baseline MBF was reduced at rest and stress in the affected region, compared to the remote area (1.13 ± 0.09 vs. 1.57 ± 0.15 ml/min/g at stress, respectively, p=0.01). At follow-up, MBF at stress and myocardial flow reserve (not shown) were higher in the EPC + matrix group compared to EPCs or PBS (EPCs + matrix: +21 ± 13%; EPCs -17 ± 8%; PBS: +5 ± 10%, p<0.05). This was paralleled by improved wall motion abnormalities (EPCs + matrix by 30 ± 9 %; EPCs by 22 ± 21%; PBS by 15 ± 12 %) and improved mismatch scores (EPCs + matrix by 79 ± 10%; EPCs by 55 ± 15%; PBS by 35 ± 19%)(all p<0.05). MBF at stress in the LCx segments positively correlated with smooth muscle actin staining for arterioles (r=0.7; p=0.02).

Conclusions:
This preclinical swine model successfully demonstrated the creation of HM, which was improved by vasculogenic cell therapy. The additional effects conferred by a collagen-based delivery matrix over EPCs alone included improved MBF in the EPC + matrix group, as well as circulatory and metabolic recovery with improvements in myocardial function.