bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 5092: Effects of Intensive Lipid Therapy on Atherosclerotic Plaque Burden and Time-course: A Prospective, Randomized, Double Blinded Study With Magnetic Resonance Imaging (MRI)
Authors: Dong Li , Yuan Chun , Hatsukami Tom S , Neradilek Moni B , Chu Baocheng , Underhill Hunter R , Moore Andrew , Monick Duane A , Lee Colin D , Okawa Minako , Zhao Xihai , Zhao Xue-Qiao
Published: April 2024

Background:
Intensive lipid lowering therapy has been associated with regression of atherosclerotic disease burden. However, the time-course for response to therapy has not been established. In this study, we examined the effects of intensive lipid-lowering therapy on atherosclerosis burden and its time course over 3 yrs in the carotid artery.

Methods:
Subjects (
N
=112) with coronary or carotid artery disease, apoB >= 120 mg/dl, received intensive lipid lowering therapy for 3 yrs. Subjects underwent carotid MRI scans at baseline and annually for 3 yrs. Expert reviewers, blinded to clinical information and scan time sequence, analyzed the atherosclerosis burden (percent wall volume (PWV) =100% × wall volume/[lumen volume + wall volume]). The burden change, expressed as ΔPWV, between pre- and post-therapy and the time course of changes over 3 yrs were compared. We also modeled (ΔPWV) as a function of the extent of maximum wall thickness (MaxWT) to assess association between the degree of disease in selected patients and ΔPWV.

Results:
84 subjects completed 3 yr follow-up scans with acceptable image quality. After 3 yrs of therapy, PWV significantly regressed (−3.8%, p<0.001). The time course over 3 yrs showed that PWV did not change significantly after the 1
st
yr (−0.3 %, p=0.7), but showed significant regression during the 2nd yr (−3.6 %, p<0.001). In this sample population, an additional year of therapy did not result in significant change from the reduction that occurred during the second year (+0.1 %, p=0.9). Average MaxWT at baseline was 2.27 mm (range: 1.0 to 5.6 mm). More thickened arteries showed significantly more reduction in atherosclerosis burden over 3 yrs compared to thinner arteries (p<0.001 for MWT slope, fitted linear regression model for ΔPWV: mean ΔPWV =0.46 – 0.41*MWT).

Conclusions:
Maximal effect of intensive lipid-lowering therapy on atherosclerotic burden was observed after 2 years of therapy. This regression was mainly demonstrated in thickened arteries, but not in relatively normal (thinner) arteries. Our findings suggest that efficacy of lipid-lowering therapy on carotid plaque burden may be best evaluated in a population with pre-existing carotid atherosclerotic disease and 2 years of therapy.
Abstract 5101: ACS in Patients With and Without Spontaneous Dissections of Coronary Arteries: The Discovery Study
Authors: Pagliani Leopoldo , La Vecchi Luigi , Cabianca Ester , Bonanno Carlo , Benettin Alessandra , Desideri Alessandro , Santarelli Alessandro , Manari Andrea , Benassi Alberto , Fontanelli Alessandro
Published: April 2024

Spontaneous coronary dissection ( SCAD) is an unusual, life threatening cause of acute myocardial ischemia. Over 200 cases of SCAD have been reported in the literature but, most of the available clinical information about SCAD derives from case reports and prospective data are lacking. The purpose of DISCOVERY study (DISsection of COronary Arteries: Veneto and Emilia RegistrY) was to prospectively assess clinical characteristics, treatment and outcome of SCAD pts. w/o spontaneous dissections with comparable pts. and to attempt to assess the role of SCAD in the pathogenesis of Acute Coronary Syndromes (ACS) and to determine whether peculiar clinical characteristics may help in defining a diagnostic pattern for SCAD-ACS. From October ’05 to December ’07, 42 consecutive pts ( mean age 51 yrs) were enrolled in the Cardiology Units of the Veneto and Emilia regions. Only cases of angiographically-proven SCAD of major coronary branches were included. Concomitantly, the first incoming ACS pts following a SCAD pts of comparable age and sex was enrolled. Insodoing, a control group of non-SCAD ACS was obtained. Analysis report data on two groups (ACS and SCAD-ACS) in order to analyze presentation, treatment strategies and outcome. In the SCAD-ACS group, no case was associated with peripartum, nor any showed associated carotid disease or lens subluxation. Baseline characteristics were similar in SCAD and non-SCAD ACS. However, SCAD pts had fewer risk factors (r.f.) for coronary disease (p=0.0016), The LAD coronary artery was more commonly involved. Also, presenting ecg pattern was more frequently ST elevation in SCAD compared to non-SCAD (58% vs 33%). Treatment were significantly different in the 2 groups: 54 % of SCAD population had conservative treatment vs 31% in the control group (p=0.002). In revascularized pts, PCI was by far the preferred treatment option. All pts were discharged alive in both groups. During a 18 month f.u., incidence of MACE was 2.3% in SCAD vs 27% in non-SCAD (p<0.001). Our data show how SCAD-ACS pts have fewer coronary r.f. compared to non-SCAD-ACS of comparable age, are less prone to undergo revascularization and exibit a much better long-term prognosis. Some features of SCAD-ACS traditionally considered in SCAD do not appear to be such in our series.
Abstract 5102: Patients Developing STEMI In-hospital Have Delayed Reperfusion and Increased 1-year Mortality Compared to STEMI Patients Presenting to the Emergency Department
Authors: Haryani Anand , Richmond Tyler , Haw Janet M , Melgoza Norma , Purim-Shem-Tov Yaninia , Calvin James E , Schaer Gary L
Published: April 2024

BACKGROUND: Despite recent advances in ST-elevation myocardial infarction (STEMI) systems of care, there has been little focus on patients who develop STEMI in-hospital. We hypothesized that inpatients who develop STEMI receive less timely care and have worse clinical outcome compared to patients who present to the emergency department (ED) with STEMI.

METHODS: One hundred and twenty-six consecutive cases of STEMI treated with primary percutaneous coronary intervention (PCI) from May 2004 to May 2009 were included in the analysis. A retrospective medical record review was utilized to identify ECG to balloon times, intraaortic balloon pump (IABP) utilization, pre-discharge left ventricular ejection fraction, and duration of hospital stay following STEMI. In-hospital, 30-day, and 1-year mortality data was collected using the National Death Registry.

RESULTS: Twenty-nine inpatients and 97 ED patients diagnosed with STEMI and undergoing PCI were included in this analysis. ECG to balloon time (357±419 min vs 163±188 min, p <0.001), one-year mortality (24.1% vs 5%, p <0.006), and duration of hospital stay (9±12.6 d vs 4.1±4.5 d, p <0.001) were significantly increased in inpatients compared to patients admitted through the ED. Inpatients trended toward increased need for IABP (28% vs. 14.4%, p =0.09) and increased 30-day mortality (13.8% vs 5%, p=0.12). There were no differences in LVEF (45% vs 50%, p=0.22) or in-hospital mortality (10% vs 5%, p=0.27) between the two cohorts.

CONCLUSION: Inpatients who develop STEMI have delayed reperfusion, increased length of stay, and greater 1-year mortality compared to STEMI patients admitted through the ED. Further study is needed to determine if improved identification and more rapid PCI could improve the clinical outcome of patients who develop STEMI in-hospital.
Abstract 5103: Perivascular Adipose Tissue in Angiotensin II Dependent Hypertension
Authors: Guzik Tomasz J , McCann Louise , Lob Heinrich , Mikolajczyk Tomasz , Budzyn Klaudia , Naruszewicz Marek , Harrison David G
Published: April 2024

Recent studies have suggested that perivascular adipose tissue (pvAT) is not just a mechanical scaffold for the vasculature but that it plays a critical role in vascular disease. The effects of mediators such as angiotensin II on perivascular adipose tissue however remain unknown. Accordingly, we examined the effects of chronic (14 day) angiotensin II (490ng/kg/min) infusion in mice on perivascular adipose tissue in comparison to effects on visceral (VAT) and subcutaneous (SAT) adipose tissue. Real time PCR and Western blotting showed that both AT1R and AT2R angiotensin II receptors were expressed in all studied compartments of AT but were greater in pvAT (485±90 vs. 45±18 in VAT and 3±0.7 in SAT U/ug RNA for AT1R mRNA; p<0.001). Angiotensin II infusion decreased epidydymal fat pad weight (413±45 mg vs 181±23mg; p<0.01) with concomitant increase of perivascular fat measured histologically (18±3 vs. 30±6 AU per aortic lumen; p<0.01). Interestingly, the cellularity of both pvAT and VAT increased (pvAT>>VAT) significantly but remained unchanged in SAT upon Angiotensin II infusion. Angiotensin II significantly altered adipocytokine expression in pvAT and VAT but not in SAT. Resistin levels increased in pvAT in response to angiotensin II, while leptin, visfatin and adiponectin decreased. The most significant changes were observed in inflammatory cytokines and chemokines. IL-6, TNF-a, IL-17A and were induced in pvAT to a greater degree than in VAT and were not changed by angiotensin II in SAT. Angiotensin II increased RANTES mRNA 6-fold in pvAT but not in VAT or SAT. Immunohistochemistry confirmed an increase in RANTES expression selectively in pvAT. These changes of cytokine and chemokine expression were accompanied by increased infiltration of pvAT with T lymphocytes expressing the RANTES receptor CCR5. These data indicate that angiotensin II affects adipocytokine and cytokine expression in adipose tissue and these effects are most pronounced in pvAT leading to T cell infiltration. These data indicate that the pvAT is a unique compartment of fat predisposed to angiotensin II-induced inflammation and a potential target for therapeutic intervention.
Abstract 5105: Increased Glucose Uptake in Visceral versus Subcutaneous Adipose Tissue Revealed by PET Imaging
Authors: Christen Thomas , Rocha Viviane , Sheikine Yuri , Goldfine Allison , Di Carli Marcelo , Libby Peter
Published: April 2024

Background
: The amount of visceral and subcutaneous adipose tissue (VAT and SAT, respectively) differentially predicts cardiovascular risk. Beyond volumetric measurement, glucose uptake in adipose tissue might yield additional information for risk stratification. This study explored differences in glucose utilization between VAT and SAT and potential mechanisms for this finding.

Methods and Results
: Retrospective analysis of whole body FDG-PET scans from 59 patients showed that VAT exhibited higher FDG uptake than SAT (Figure A

, p<0.0001) independent of age, gender, body mass index, co-morbidities, or medications. To investigate the mechanisms underlying this observation, we then studied glucose uptake in stromal vascular cells (SVCs), a fraction of the adipose tissue which is rich in inflammatory cells. SVCs from VAT of diet-induced obese C57BL/6 mice exhibited higher glucose uptake than those from SAT (Figure B

, n=6, p=0.01). Evaluation of the expression of various genes involved in cellular glucose metabolism such as glucose transporters (GLUT 1, 3 and 4) and hexokinases (HK-1 and 2) showed increased expression of HK-1 in VAT-compared to SAT-derived SVCs (Figure C

, n=10, p=0.005). This finding illustrates one mechanism that may explain the above results.

Conclusions:
This study showed non-invasively with FDG PET imaging that VAT has increased glucose uptake compared to SAT in human subjects, and showed experimentally that inflammatory cells may contribute to this difference. These findings add to our understanding of the nexus between adiposity, inflammation, and cardiovascular risk.
Abstract 5110: The Inhibition of Poly(ADP-ribose) Polymerase-1 (PARP-1) Significantly Accelerates the Vasculogenesis in a Model of Hind Limb Ischemia
Authors: Elmageed Zakaria Y Abd , Amin Ali H , Nair Devika , Megan Partyka , Belmadani Souad , Matrougui Khalid
Published: April 2024

Background: Cardiovascular and metabolic diseases are associated with impaired vasculogenesis in response to ischemia. PARP-1, an abundant nuclear enzyme involved in DNA repair and transcriptional regulation and recognized as a key regulator of cell survival and cell death in response to noxious stimuli in various forms of cardiovascular and metabolic disease. Our hypothesis is to determine whether PARP-1 has a key role in vasculogensis in response to ischemia.

Methods and Results: Hind limb ischemia was performed in C57/Bl6 mice for 15 days with and without PARP-1 inhibition using TIQ (PARP-1 inhibitor) or local injection of lentivirus-PARP-1-siRNA. We also used microvascular cultured endothelial cells stimulated with hypoxia with and without PARP-1 inhibition (TIQ and lentivirus-PARP-1-siRNA) and determined the expression of VEGF. Mice treated with TIQ significantly accelerated the recovery of blood flow, as determined with MoorLDI-Laser and X-ray, by 50 – 60% the first week and complete recovery at the second week compared to control. Interestingly, mice locally injected with lentivirus-PARP-1-siRNA significantly accelerated blood flow recovery compared to control. Western blot analysis revealed an increase in VEGF level in muscle from mice treated with TIQ or injected with lentivirus-PARP-1-siRNA compared to control. VEGF release in the conditioned media was increased when cultured endothelial cells were stimulated with hypoxia. In addition, endothelial cells pre-treated with TIQ or transfected with lentivirus-PARP-1-siRNA and then stimulated with hypoxia significantly increased VEGF release in the conditioned media compared control and to cells stimulated with hypoxia.

Conclusion: Our novel data provide evidence that PARP-1 plays a key role in the regulation of vasculogenesis in response to ischemia. We demonstrated that the inhibition of PARP-1 enhances earlier the formation of new vessels in response to ischemia through the increase of VEGF release. The inhibition of PARP-1 would be interesting in model of altered vasculogenesis such cardiovascular and metabolic diseases.
Abstract 5113: Ablation of Lectin-like Receptor for Oxidized Low-density Lipoproteins (LOX-1) Resulted in Suppression of Ischemia-Induced Angiogenesis
Authors: Shiraki Takeru , Aoyama Takuma , Yokoyama Chiharu , Kazuhiko Nishigaki , Takemura Genzou , Fujiwara Takako , Fujiwara Hisayoshi , Sawamura Tatsuya , Minatoguchi Shinya
Published: April 2024

Background: We previously identified LOX-1 as a receptor for oxidized low-density lipoproteins. Recent studies show LOX-1 contributes to capillary formation induced by Angiotensin II or oxLDL, suggesting the potential effects of LOX-1 on angiogenesis. Therefore, we elucidated whether LOX-1 might be involved in angiogenesis in ischemic hindlimb tissue.

Methods and Results: Ischemia was surgically produced by femoral artery ligature in both wild-type mice (WT) and LOX-1 deletion mice (LOX-1 KO). The ratio of ischemic/nonischemic leg blood flow measured by laser Doppler perfusion imaging was significantly smaller in LOX-1 KO than WT on Day 14, 21 and 28 (p<0.01). In agreement, histological capillary density (vWF immunostaining) was significantly lower in LOX-1 KO than in WT on Day 28 (115±28 vs. 177±19/HPF, p<0.01). Next, we analyzed phosphorylation of p38 involved in LOX-1 signaling. Phosphorylation of p38 was suppressed in ischemic hindlimb tissue of LOX-1 KO mice compared with WT mice. Western blot analyses also showed remarkable induction of VEGF in hindlimb of WT on Day 3 after ischemia. On the other hand, ischemic tissues of LOX-1 KO contained very low VEGF, resulting in suppressed phospholylation of Akt and eNOS, which are downstream signaling of VEGFR2. Considering the fact that inflammation is important for ischemia-induced angiogenesis, we examined whether infiltrated macrophages (F4/80 immunochemistry) release VEGF in ischemic hindlimb using two-color immunofluorescence histochemical staining. Fewer numbers of macrophages infiltrated into the ischemic tissues in LOX-1 KO than in WT (64±24 vs. 118±25/HPF, p<0.01), and immunohistochemical analysis revealed the number of infiltrated macrophages expressing VEGF was also decreased in LOX-1 KO (LOX-1: 34±19 vs. WT: 50±14/HPF, p<0.01).

Conclusions: This study demonstrates that LOX-1 deletion negatively modulates ischemia-induced angiogenesis through impairment of early inflammatory mononuclear cell infiltration, which releases VEGF. LOX-1 would become a novel target molecule for modulating angiogenesis in ischemic tissue.
Abstract 5117: Sustained Release of Granulocyte Colony-Stimulating Factor Using Biodegradable Gelatin Hydrogel Microspheres Persistently Improves Blood Flow of Ischemic Limbs in Mice
Authors: Kawamura Itta , Takemura Genzou , Kanamori Hiromitsu , Li Longhu , Okada Hideshi , Esaki Masayasu , Kobayashi Hiroyuki , Tsujimoto Akiko , Goto Kazuko , Maruyama Rumi , Takeyama Toshiaki , Kawaguchi Tomonori , Watanabe Takatomo , Hattori Arihiro , Ushikoshi Hiroaki , Aoyama Takuma , Kawasaki Masanori , Nishigaki Kazuhiko , Fujiwara Hisayoshi , Tabata Yasuhiko , Minatoguchi Shinya
Published: April 2024

Granulocyte colony-stimulating factor (G-CSF) is a potent angiogenetic factor, but its use is limited because of the unfavorable systemic effects. We hypothesized that G-CSF immersed gelatin hydrogel microspheres (GHM) injected into the ischemic limb might continuously release a small amount of G-CSF to locally stimulate angiogenesis without adverse effects. We ligated the right femoral artery of BALB/c mice and randomly assigned mice into 5 groups: the saline group with a single intramuscular injection of saline into the ischemic limb; the G-CSF sc group with a subcutaneus injection of G-CSF (100
μ
g/kg) for 5 days; the G-CSF im group with an intramuscular injection of G-CSF (20
μ
g/kg) into the ischemic limb for 5 days: the GHM group with a single intramuscular injection of GHM alone into the ischemic limb; and the G-CSF-GHM group with a single intramuscular injection of G-CSF (100
μ
g/kg) immersed GHM into the ischemic limb. The blood flow ratio of the ischemic/non-ischemic limb assessed by a laser Doppler perfusion imager became significantly improved 6 weeks later in the G-CSF-GHM group compared with the other groups and lasted until 8 weeks (saline; 0.63±0.10, G-CSF sc; 0.70±0.14, G-CSF im; 0.61±0.15, GHM; 0.59±0.12, G-CSF-GHM; 0.88±0.20). Both in the G-CSF sc and im groups, the blood flow ratio was significantly improved at the early time (1 and 2 weeks later in the G-CSF sc group, 3days later in the G-CSF im group), but was only transient. Not only capillaries but also arterioles were significantly increased in the G-CSF-GHM group where granulocyte and endothelial progenitor cell counts were not elevated but the Akt/endothelial nitric oxide synthase pathway in the ischemic limb was significantly activated. The present drug delivery system is suggested to have potential as a novel noninvasive therapy in patients with peripheral artery disease because of not only its efficacy but also less invasiveness and less unfavorable systemic effect.
Abstract 5118: Cardiac
β
1-Adrenergic Receptor Blockade Promotes Angiogenesis in the Post-Ischemic Failing Heart via Activation of VEGF- and Akt-dependent Signalling Pathways
Authors: Zincarelli Carmela , Rengo Giuseppe , Golino Luca , Lymperopoulos Anastasios , Altobelli Giovanna Giuseppina , Cimini Vincenzo , Marchese Massimo , De Lucia Claudio , Koch Walter J , Leosco Dario
Published: April 2024

Impaired angiogenesis in the post-myocardial infarction (MI) heart, contributes to the progression to heart failure (HF). The inhibition of vascular endothelial growth factor (VEGF) signaling has been shown to induce the transition from compensatory hypertrophy to failure. In the present study, we investigated whether treatment with
β
1- adrenergic receptor (
β
-AR) blockade Bisoprolol (B) may reactivate VEGF-dependent angiogenesis and counteract maladaptive remodeling in HF. 4 weeks from surgically-induced MI, 40 male Sprague Dawley rats, were randomized to receive B (10 mg/Kg/d, for 10 weeks) or vehicle. 10 sham-operated rats were also included in the study. Echocardiography revealed reduced cardiac LV diameters in B-treated compared to vehicle-treated rats. Moreover, B treatment was associated with increased LV capillary and arteriolar density, and with increased
in vivo
coronary perfusion and reserve. Importantly, 10 days after B treatment was started, increased cardiac VEGF expression and Akt and eNOS activation were observed by comparing B-treated to drug-untreated hearts. The concomitant administration of B and the mTOR inhibitor Rapamycin (R), which inhibits Akt activation, was associated with increased LV dilatation and dysfunction, and decreased myocardial capillary and arteriolar density compared to B treatment alone. Finally, to test whether the pro-angiogenic effects of B act via activation of VEGF expression, rats were intravenously injected with adenoviral vector encoding a decoy VEGF receptor (Ad-Flk) or a control adenovirus (Ad-C), at the start of the treatment. After 10 weeks from the start of the treatments, histological analysis revealed reduced capillary and arteriolar density, and coronary perfusion in B-treated+Ad-Flk compared to B-treated+Ad-C. Moreover, VEGF inhibition counteracted the positive effects of B on cardiac remodeling and function. In conclusion,
β
1-AR selective blockade promotes angiogenesis in HF hearts via activation of VEGF- and Akt-dependent signaling pathways. The
β
-blocker-induced enhancement of vascular bed prevents progression from adaptive to maladaptive remodeling in the failing myocardium, thus counteracting the progressive dysfunction of the failing heart.

This research has received full or partial funding support from the American Heart Association, Great Rivers Affiliate (Delaware, Kentucky, Ohio, Pennsylvania & West Virginia).
Abstract 5123: Assessment of Myocardial Microvascular Abnormalities in Chronic Kidney Disease
Authors: Kuliszewski Michael A , Rudenko Dmitriy , Connelly Kim A , Yuen Darren , Trogadis Judy , Gilbert Richard E , Leong-Poi Howard
Published: April 2024

Recent data has shown that chronic kidney disease (CKD) has been linked to the development of cardiovascular disease and been associated with an adverse prognosis. It has been hypothesized that effects on the coronary microcirculation play a critical role in the exacerbation of disease. Thus, we assessed the myocardial microvasculature in CKD, using fluorescent microangiography (FMA) and myocardial contrast echocardiography (MCE).

Methods:
8 week old male Fischer 344 rats underwent subtotal nephrectomy (n=10) (SNX), or sham surgery (n=10). Systolic blood pressure (SBP), proteinuria, left ventricular (LV) ejection fraction (EF), end-diastolic pressure volume relationship (EDPVR) and Tau were assessed at baseline and 8 weeks following surgery. MCE was performed to assess regional myocardial perfusion, and rats were subjected to FMA (10% solution of 0.02
μ
m fluorescent microspheres). Thick (200
μ
m) sections of fixed tissue can then be visualized under confocal microscopy. Stacks of images are collected and 3D reconstruction and visualization carried out in order to quantify the density, size and shape of blood vessels. An automated tracing algorithm was used to create a solid 3D model.

Results
: SNX animals showed increased in SBP and urinary protein confirming CKD. LVEF was preserved (68±16 versus 53±14 % in controls), while diastolic function was impaired in both the energy-dependent early (Tau) (16.7±3.1 versus 11.0±1.2 msec; p<0.05) and late passive EDPVR, a measure of LV compliance (0.052±0.026 versus 0.018±0.009 mmHg/ul in controls; p<0.01). FMA analysis illustrated a reduction in average vessel diameter (3.84±0.08 versus 4.68±0.16
μ
M in controls; p < 0.05), and a reduction in vascular density (2.81E-03±1.72E-04 versus 3.63E-03±1.56E-04
μ
M in controls; p < 0.01). MCE-derived endocardial-epicardial blood flow ratio was reduced in CKD as compared to sham controls (0.67±0.14 versus 1.12±0.35 in controls; p<0.05).

Conclusion:
Myocardial microvascular loss occurs in CKD, and is associated with a reduction in endocardial blood flow and impaired diastolic function, with preserved LV systolic function.
Abstract 5125: Elevated Serum Brain Natriuretic Peptide Level Predicts the Presence of the Left Atrial Appendage Dysfunction in Patients With Acute Ischemic Stroke
Authors: Tamura Harutoshi , Watanabe Tetsu , Sasaki Shintaro , Nishiyama Satoshi , Takahashi Hiroki , Arimoto Takanori , Shishido Tetsuro , Miyashita Takehiko , Miyamoto Takuya , Nitobe Joji , Kubota Isao
Published: April 2024

Background:
It is well known that the left atrial appendage (LAA) dysfunction plays an important role in the occurrence of the cardioembolic stroke. It has been reported that the atrium is the main source of brain natriuretic peptide (BNP) in atrial fibrillation patients without heart failure. Therefore, we hypothesized that serum BNP levels are sensitive predictor for LAA dysfunction and thrombus formation in patients with acute ischemic stroke.

Methods:
We performed trans-thoracic and esophageal echocardiography within 7 days after the onset (male, n=64, 69±11 years old; female, n=38, 68±15 years old) and measured serum hemostatic markers and BNP in 103 patients with ischemic stroke. No patients had history of heart failure.

Results:
LAA thrombus was detected in 8 patients (36%) among 22 patients with atrial fibrillation. Even in 3 patients (4%) among 80 patients with sinus rhythm, LAA thrombus was detected. There were no significant differences in left ventricular end-diastolic dimension (LVDd), and the ratio of the early diastolic trans-mitral inflow velocity (E) to mitral annular velocity (E′) between patients with and without LAA thrombus (LVDd, 49±6 vs. 47±5 mm; E/E′, 8.6±1.3 vs. 7.2±0.9). Serum BNP levels were markedly higher in patients with LAA thrombus formation than in those without it (191 (128 –278) vs. 39 (13–100) pg/ml, P<0.01). Furthermore, in patients with sinus rhythm (n=80), the BNP levels were significantly correlated with LAA peak emptying flow velocity (R=−0.34, P<0.01). The multivariate logistic regression analysis revealed that BNP >120 was an independent predictor of LAA thrombus formation (odds ratio, 13.50; 95% confidence interval, 2.482–73.40; P=0.0026).

Conclusions:
The BNP levels were correlated with LAA peak emptying flow velocity. The elevated BNP level could be used as a non-invasive surrogate maker to predict LAA dysfunction and thrombus formation in patients with acute ischemic stroke.
Abstract 5137: Visualization of Accumulating Reactive Oxygen Species During Myocardial Ischemia by Real-time Two-photon Imaging of Perfused Rat Hearts
Authors: Iguchi Moritake , Akao Masaharu , Kato Masashi , Matsumoto-Ida Madoka , Kimura Takeshi , Kita Toru
Published: April 2024

Background: During myocardial ischemia/reperfusion injury, a burst of reactive oxygen species (ROS) is considered to occur at the onset of reperfusion and cause cardiomyocyte death through the loss of mitochondrial membrane potential (ΔΨ
m
), but there are evidences against this concept. We aimed to visualize the dynamic changes of ROS during ischemia/reperfusion in intact rat hearts using two-photon laser scanning microscopy.

Methods and Results: Langendorff-perfused rat hearts were loaded with 5-(and -6)-chloromethyl-2′,7′-dichlorodihydrofluorescein diacetate acetyl-ester and tetramethylrhodamine ethyl-ester, fluorescent indicators of ROS and ΔΨ
m
, respectively. Under the two-photon excitation, spatio-temporal changes of ROS and ΔΨ
m
in response to ischemia/reperfusion were simultaneously monitored at cellular level. As soon as ischemia started, ROS level of each cell began to increase, despite maintained ΔΨ
m
. Importantly, the rate of ROS accumulation during the early phase of ischemia was variable from cell to cell, and irreversible ΔΨ
m
loss occurred in an all-or-none manner depending on cellular ROS level, with a clear cut-off value. Upon reperfusion, we were unable to detect a burst of ROS, but there were more cells undergoing similar ΔΨ
m
loss after the cell-to-cell specific period of latency. Importantly, there was a significant inverse correlation between the level of ROS accumulation during ischemia and the latency until which the cell underwent ΔΨ
m
loss (r=−0.89). Ischemic preconditioning delayed the increase of ROS during ischemia and protected against ΔΨ
m
loss. An iron chelator deferoxamine, which reduces the generation of hydroxyl radical through inhibiting the iron-mediated Fenton reaction, neither attenuated the ROS generation nor protected against ΔΨ
m
loss. A free radical scavenger N-(2-mercaptopropionyl)-glycine attenuated the ROS accumulation, but it did not protect against ΔΨ
m
loss.

Conclusion: In myocardial ischemia/reperfusion, ROS are mainly generated during ischemia, but not after reperfusion, and play a critical role in mitochondrial collapse and cardiomyocyte death. However, reducing the ROS accumulation during ischemia may not suffice to prevent cell death.
Abstract 5138: Secretoneurin Therapy Improves Cardiac Function in a Model of Myocardial Infarction in-vivo and Induces Angiogenesis and VEGF Expression in Cardiac Cells in-vitro
Authors: Schgoer Wilfried , Albrecht Karin , Theurl Markus , Beer Arno , Tancevski Ivan , Vasiljevic Danijela , Voelckl Jakob , Bonaros Nikolaos , Kocher Alfred , Wiedemann Dominik , Aumayr Florian , Oeberseder Gregor , Steger Christina , Schratzberger Peter , Patsch Josef , Kirchmair Rudolf
Published: April 2024

Introduction
The angiogenic neuropeptide Secretoneurin (SN) is upregulated by hypoxia in several tissues like neurones and skeletal muscle cells and SN gene therapy restores tissue integrity, function and perfusion in the mouse hind-limb ischemia model by induction of angiogenesis, arteriogenesis and vasculogenesis. We therefore hypothesized that this cytokine might also affect function of cardiac cells and exert beneficial effects in a rat model of myocardial infarction (MI).

Results
To investigate the effect of SN
in vitro
, a matrigel assay using human coronary artery endothelial cells (HCAEC) in the absence or presence of different concentrations of SN was performed. SN dose dependently induced angiogenesis
in vitro
with a maximum effect at 1ng/ml (rel. capillary tube formation 2.06+/−0.15; n=4; p<0.001 vs. Ctr.). A specific SN antibody abolished the observed effect (rel. cap. tube form. 1.05+/−0.14; n=4; p<0.01 vs. SN). Western Blot analysis of HCAEC extracts revealed stimulation of ERK1/2 by SN 10ng/ml indicating activation of this signal transduction pathway. PCR analysis of hypoxic human cardiac myocytes showed no effect on the expression of SN (in contrast to skeletal myocytes) but stimulation of this cell line with SN 100ng/ml lead to an upregulation of VEGF mRNA after 12 hours. In an acute MI model in rats, SN gene therapy was performed by injecting 500ug plasmid intramyocardially 10 min after LAD ligation. GFP plasmid was used as control. Left ventricular function was evaluated post-operatively and on day 14 and 28 after LAD ligation by echocardiography (ejection fraction, EF) and fractional area change (FAC) was measured using magnetic resonance imaging. On day 28 SN treated rats showed a significant better outcome with improved EF (75% for SN vs. 44% for Ctr, n=5; p<0.05) and improved FAC (0.59 for SN and 0.46 for Ctr, n=5; p<0.05).

Conclusion
In vitro
data for SN on cardiac cells suggest induction of angiogenesis and upregulation of VEGF.
In vivo
, SN gene therapy significantly improved left ventricular function in the rat MI model. Future studies will have to clarify if beneficial effects of SN
in vivo
are mediated by VEGF. These findings suggest that SN gene therapy might be a promising new therapeutic agent to improve ischemic cardiac dysfunction.
Abstract 5144: The Obligatory Role of Heat Shock Protein 90 in iNOS Induction in Postischemic Hearts
Authors: Luo Suxin , Wang Tingting , Chen Lucy L , Xia Yong
Published: April 2024

Inducible nitric oxide synthase (iNOS) plays important roles in the process of myocardial infarction, heart failure, and cardiac preconditioning. The expression of iNOS is mainly induced by inflammatory substances such as endotoxins and cytokines. NF-
κ
B and STAT-1 are the two transcriptional factors that mediate iNOS expression in stimulated cells. Previous studies showed that heat shock protein 90 (Hsp90) associates with iNOS and this association enhances iNOS activity. Here we show that Hsp90 is also essential for iNOS induction. To determine the role of Hsp90 in iNOS induction, we treated mouse macrophages (RAW 264.7) with lipopolysaccharide (LPS, 1
μ
g/ml) and interferon-
γ
(IFN-
γ
, 100 units/ml) in the absence and presence of Hsp90 inhibitor geldanamycin (1
μ
M). Hsp90 inhibition completely prevented iNOS protein expression. RT-PCR experiments showed that iNOS mRNA transcription was blocked by Hsp90 inhibition. Radicicol (10
μ
M), another Hsp90 inhibitor whose structure is different from that of geldanamycin, also blocked iNOS mRNA transcription. Further experiments showed that Hsp90 inhibition prevented NF-
κ
B activation in LPS-treated cells. In IFN-
γ
-stimulated cells, STAT-1 activation was also abolished by Hsp90 inhibition. To explore whether these findings from cultured cells occurred in diseases, we performed
in vivo
myocardium infarction on mice. While no iNOS expression was detected in the heart of mice in sham group, iNOS mRNA and proteins were observed in the infarcted areas of the hearts subjected to 1-hr coronary artery (LAD) occlusion and 24-hr reperfusion. Geldanamycin (4 mg/kg, i.p.) completely prevented iNOS mRNA and protein expression in the infarcted myocardium (
P
<0.001, n=5). Taken together, these results demonstrate that besides acting as an allosteric enhancer, Hsp90 is also required for iNOS gene transactivation. Intervening Hsp90 may represent a novel approach to modulate iNOS expression in myocardial infarction and cardiac preconditioning.
Abstract 5147: GLP-1 Improves Insulin-Stimulated Nitric Oxide-Dependent Vasodilator Responsiveness in Patients With Metabolic Syndrome
Authors: Tesauro Manfredi , Schinzari Francesca , Rovella Valentina , Mores Nadia , Pitocco Dario , Ghirlanda Giovanni , Lauro Renato , Cardillo Carmine
Published: April 2024

Our previous work has shown that patients with metabolic syndrome (MetS) have impaired insulin-stimulated vasodilation, which may be involved in the development of hypertension and contribute to insulin resistance by affecting the glucose uptake. Glucagon-like peptide-1 (GLP-1), an amino acid gut hormone that stimulates both insulin secretion and sensitivity, is also provided of cardioprotective and vasoactive properties. We therefore examined the effects of exogenous GLP-1 on insulin-stimulated, nitric oxide (NO)-dependent vasodilation in patients with obesity-related MetS by use of the forearm perfusion technique. Forearm blood flow (FBF) responses to intra-arterial infusion of graded doses of acetylcholine (ACh; stimulus for endothelial release of NO) and sodium nitroprusside (SNP; exogenous NO donor) were assessed in 10 patients with MetS during coinfusion of either saline or regular insulin (0.2 mU/Kg/min). Thereafter, patients underwent to intra-arterial infusion of GLP-1 (30 ng/min for 45 minutes) and responses to ACh and SNP were then reassessed. In the absence of hyperinsulinemia, GLP-1 did not impact unstimulated FBF (from 3.5±0.2 [mean±SEM] to 3.6±0.3 ml/min/dl; P=0.59); similarly, exogenous GLP-1 did not affect vascular responses to both ACh (from 8.8±1.1 to 8.7±1.2 ml/min/dl at the highest dose; P=0.97) and SNP (from 10.7±0.6 to 9.6±0.7 ml/min/dl; P=0.68). In the presence of hyperinsulinemia, GLP-1 infusion was associated with mild increase in basal FBF (from 4.7±0.4 to 5.7±0.6 ml/min/dl; P=0.25); under those conditions, however, exogenous GLP-1 resulted in a significant improvement in vascular reactivity to both ACh (from 13.7±0.7 to 17.4±1.5 ml/min/dl; P=0.03) and SNP (from 12.7±1.0 to 15.9±1.2; P=0.02). In conclusion, exogenous GLP-1 improves both endothelium-dependent and -independent vasodilator responsiveness during hyperinsulinemia in patients with MetS. This suggests that GLP-1 may exert a facilitatory effect on insulin signaling to enhance NO-dependent reactivity in vascular smooth muscle. As a consequence, strategies targeting the GLP-1 system for treatment of type 2 diabetes might beneficially affect both metabolic and hemodynamic abnormalities associated with insulin resistance.
Abstract 5157: Atherosclerotic Plaque Development Depends on T-cell Priming Within the Secondary Lymphatic Organs
Authors: Luchtefeld Maren M , Grothusen Christina , Pabst Oliver , Förster Reinhold , Bernhard Schieffer
Published: April 2024

Background
: The chemokine receptor 7 (CCR7) is required for the migration of dendritic cells and T-cells to the secondary lymphoid organs (SLO) in order to promote DC-dependent T-cell priming. Here we investigated the influence of CCR7-deficiency on T-cell distribution in animals on cholesterol rich diet.
In vitro
, we analyzed whether oxLDL induced the expansion of a T-cell carrying a specific T-cell receptor subset and whether this T-cell receptor could be found in animals fed with a high cholesterol diet. Finally we investigated whether atherosclerosis can be reconstituted when
ex vivo
primed T-cells were given to CCR7-deficient mice while feeding with a cholesterol rich diet.

Methods and Results:
T-cell number within the aorta was significantly increased, whereas T-cells within the SLO were reduced in
ccr7
−/−
/ldlr
−/−
mice compared to
ldlr
−/−
-mice as analyzed by CD3 staining of serial sections (immunhistochemistry, n=12–17). OxLDL (10
μ
g/mL) but not nLDL (10
μ
g/mL) promoted DC maturation
in vitro
(CCR7 and MHC-II expression, n=4) and
in vivo
(CD80 and CD86 expression, n=4) as monitored by FACS-analysis. To characterize an oxLDL specific T-cell, we co-cultivated nLDL and oxLDL stimulated DCs with T-cells and analyzed T-cell activation (BrdU, n=5), and the expression of an oxLDL specific T-cell receptor subset (real-time PCR, n=4). OxLDL but not nLDL stimulated DCs induced T-cell proliferation, which was accompanied with the expression of the T-cell receptor chains V
α
10 and V
β
6. These T-cells could also be found within the aorta and the SLO of
ldlr
−/−
-mice following feeding and was found to be significantly reduced in
ccr7
−/−
/ldlr
−/−
mice as analyzed by T-cell receptor expression (real-time PCR and FACS-analysis, n=4–5). Finally, to investigate whether this specific T-cell is responsible for atherosclerotic plaque development, we injected oxLDL expanded T-ells into
ccr7
−/−
/ldlr
−/−
mice while feeding with a cholesterol rich diet (1×10
7
cells) and observed a complete reconstitution of the atherosclerotic phenotype (
en face
preparation of oil red stained aortas, n=5–14),

Conclusion:
Our results indicate that atherosclerotic plaque development is driven by an antigenic process induced by oxLDL and promoted by a specific T-cell.
Abstract 5169: Endovascular Injury Induces Rapid Phenotypic Changes in Perivascular Adipose Tissue
Authors: Takaoka Minoru , Nagai Ryozo , Sata Masataka
Published: April 2024

Background: It was reported that the serum levels of circulating proinflammatory cytokines are increased in overweight people with enhanced accumulation of visceral fat. In contrast to visceral fat, little attention has been paid to the role of perivascular fat in the pathogenesis of obesity-associated diseases. Here, we investigated the effect of endovascular injury on the phenotype of perivascular fat.

Methods and Results: First, endovascular injury significantly down-regulated adiponectin and up-regulated proinflammatory adipocytokines, TNF-alpha, IL-6, PAI-1 and MCP-1 within perivascular fat tissue in models of murine femoral artery wire injury and rat iliac artery balloon injury (p<0.05). Second, diet-induced obesity augmented endovascular injury-induced up-regulation of proinflammatory adipocytokines within perivascular fat tissue (p<0.05) and neointimal hyperplasia (p<0.05) compared with standard chow diet in wild-type mice. Third, up-regulation of proinflammatory adipocytokines within perivascular fat by endovascular injury was significantly blunted in TNF-alpha deficient mice (p<0.05). Neointima formation was attenuated in TNF-alpha deficient mice compared with wild-type mice (p<0.05). Furthermore, perivascular delivery of TNF-alpha to the femoral artery using osmotic minipump enhanced endovascular injury-induced up-regulation of proinflammatory adipocy-tokines within perivascular fat (p<0.05) and neointimal hyperplasia (p<0.05) compared with contralateral PBS delivered artery in TNF-alpha deficient mice. Fourth, conditioned medium from the coculture of 3T3L1-derived adipocyte and RAW264 macrophage stimulated rat vascular smooth muscle cells proliferation relative to the control medium from 3T3L1-derived adipocyte (p<0.05). Blockade of TNF-alpha in the co-culture using an anti-TNF-alpha neutralizing antibody reduced the stimulatory activity of the condition medium (p<0.05).

Conclusions: Our findings indicate that endovascular injury induces rapid and marked changes in perivascular adipose tissue, mainly mediated by TNF-alpha. It is suggested that the phenotypic changes in perivascular adipose tissue may have a role in the pathogenesis of neointimal hyperplasia after angioplasty.
Abstract 5170: Atrial Endothelial Impairment Induced by Toll-Like Receptor-4 Signaling May Cause Atrial Thrombogenesis
Authors: Katoh Shigehiko , Watanabe Tetsu , Suzuki Satoshi , Takahashi Hiroki , Arimoto Takanori , Shishido Tetsuro , Miyashita Takehiko , Miyamoto Takuya , Nitobe Joji , Kubota Isao
Published: April 2024

Background
Recently atrial endothelial dysfunction has gained more attention as an important cause for atrial thrombogenesis in patients with atrial fibrillation (AF). It was reported that AF patients with left ventricular hypertrophy or heart failure (HF) have higher thromboembolic risks than those without. Several reports showed that angiotensin II receptor blockers prevented occurrences of AF and thrombogenesis by inhibiting inflammation. However, little is known how AF causes atrial inflammation and thrombogenesis. We examined whether atrial inflammation through toll-like receptor (TLR)-4 signaling causes atrial thrombogenesis.

Methods and result
First, we examined serum and atrial tissue heat shock protein 60 (HSP60) levels, which have been known as a TLR-4 ligand, in patients with and without AF. Atrial HSP60 increased in HF patients with AF compared to those without AF whereas serum HSP60 level was lower in HF patients with AF compared to those without AF (NYHA class I and II). Then, we created thoracic transverse aortic constriction (TAC) model in TLR-4 knockout (KO) mice (n=15) and wild-type (WT) mice (n=20). We performed echocardiography at 4 weeks after TAC surgery. Then, the atrium was excised, and vascular cell adhesion molecule (VCAM)-1, endothelial nitric oxide synthase (eNOS), extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK) and mitogen activated protein kinase p38 (p38) were analyzed by Western blotting. We observed less frequent occurrence of atrial thrombosis in TLR-4 KO-TAC mice (4/15) than WT-TAC mice (16/20). Decrease in eNOS phosphorylation and increase in VCAM-1 expression in WT-TAC mice were significantly attenuated in TLR-4 KO-TAC mice (P<0.05). Nuclear factor-kappa B (NFkB) was attenuated in TLR-4 KO-TAC mice compared to WT-TAC mice by electrophoretic mobility shift assay analysis. Phosphorylation of p38, but ERK and JNK was also attenuated in TLR-4 KO-TAC mice (P<0.05). These results suggested that activation of VCAM-1 and decreased eNOS through TLR-4/p38 and NFkB signaling may be associated with atrial thrombogenesis in AF.

Conclusion
Atrial inflammation caused by TLR-4 signaling may play a role in atrial thrombogenesis in heart failure mice model.
Abstract 5201: Low-dose Tetrahydrobiopterin Supplementation Reduces Atherosclerosis Progression in Apolipoprotein E-Knockout Mice via Effects on Vascular T cell Infiltration
Authors: Schmidt Tim S , McNeill Eileen , Douglas Gillian , Crabtree Mark J , Hale Ashley B , O’Neill Charles A , Channon Keith M , Alp Nicholas J
Published: April 2024

Tetrahydrobiopterin (BH4) supplementation improves endothelial function in models of vascular disease by maintaining endothelial nitric oxide synthase coupling and nitric oxide bioavailability. Recent studies using pharmacological and transgenic endothelial-specific BH4 supplementation highlighted the therapeutic potential of BH4 in native atherosclerosis and vascular injury and suggested a potential role for BH4 as regulator of inflammation and vascular remodeling. However, the cellular mechanisms through which this leads to reduced atherosclerosis remain unclear. We have used a pharmaceutical BH4 formulation to address the hypothesis that improved endothelial function resulting from exogenous BH4 supplementation leads to reduced atherosclerosis through effects on vascular inflammation. Single oral dose pharmacokinetic studies revealed rapid BH4 uptake from the gastrointestinal tract into plasma and organs. While in most organs BH4 levels returned to baseline by 8 hours after dosing, BH4 levels in aorta remained markedly increased at 24 hours, suggesting an active mechanism for BH4 retention. Daily oral BH4 supplementation in 8 week old chow-fed ApoE-KO mice for 8 or 12 weeks had no effect on plasma lipids or hemodynamic parameters, but resulted in a significant 45% relative reduction in aortic root atherosclerosis only at 10 mg/kg BH4, compared with placebo controls (0.049 vs. 0.088 mm2, P<0.05 after 8 weeks; 0.142 vs. 0.259 mm2, P<0.05 after 12 weeks). BH4 treatment led to a striking reduction in T cell infiltration, both within plaques (0.001886 vs. 0.006653 mm2; P<0.05) and in the surrounding adventitia (0.007741 vs. 0.020133 mm2; P<0.05). Importantly, BH4 treatment had no effect on circulating leukocyte numbers, and furthermore no evidence for BH4 uptake by peripheral blood mononuclear cells from plasma after single oral supplementation was found. Experiments using endothelium-targeted GTP-cyclohydrolase 1 transgenic mice, with a specific increase in endothelial BH4 levels, revealed a similar reduction in vascular T cell infiltration, suggesting that BH4 supplementation reduces atherosclerosis through a reduction in vascular T cell infiltration mediated by salutary effects on endothelial nitric oxide bioavailability.
Abstract 5207: Role of Endothelial Nitric Oxide Synthase in Endothelial Cell Mitochondrial Biogenesis and Stress Adaptation
Authors: Li Chunying , Keaney John F
Published: April 2024

Activation of metabolic stress pathways is known to confer stress adaptation and promote longevity in lower organisms. We have recently found that chronic stimulation of AMP-activated protein kinase (AMPK) in endothelial cells promotes both mitochondrial biogenesis and stress adaptation that prevent hypertension-induced endothelial dysfunction. However, the precise mechanisms for endothelial cell mitochondrial biogenesis and stress adaptation are not well defined. To this end, we chronically activated AMPK with 5-aminoimidazole-4-carboxamide riboside (AICAR) and observed enhanced mitochondrial biogenesis in wild-type mouse endothelium as determined by mitochondrial-specific fluorescence (Mitotracker green) and protein expression (cytochrome c and SOD2). In contrast, endothelium from eNOS-null mice exhibited no mitochondrial biogenesis in response to AICAR and this defect was rescued by reintroduction of eNOS by adenoviral-mediated gene transfer. Moreover, eNOS appeared critical for stress adaptation as chronic AMPK stimulation attenuated H
2
O
2
-induced cell death in wild type, but not eNOS-null endothelium. To test this pathway
in vivo
, we used 20-hydroxy-eicosatetraenoic acid (20-HETE) administration to induce endothelial dysfunction measured as prostacyclin-dependent, acetylcholine-induced arterial relaxation. We found that 20-HETE produced equivalent endothelial dysfunction in both wild-type and eNOS-null mice. However, chronic AICAR administration was able to prevent 20-HETE-induced dysfunction in wild-type, but not eNOS-null mice. Thus, these data indicate that eNOS is involved in AICAR-induced mitochondrial biogenesis and stress adaptation in endothelial cells. These results have implications for targeting stress adaptation as a means to prevent endothelial dysfunction and potentially ameliorate vascular disease.