tbuckley/cabot-search huggingface dataset:
Abstract 5592: Role of Afadin in Vascular Endothelial Growth Factor- and Sphingosine 1-Phosphate-Induced Angiogenesis
Authors: Hideto Tawa, Yoshiyuki Rikitake, Motonori Takahashi, Hisayuki Amano, Mitsuo Kinugasa, Yuichi Nagamatsu, Takashi Majima, Hisakazu Ogita, Jun Miyoshi, Yoshimi Takai, Ken-ichi Hirata
Journal: Circulation (2009)

[Background] Better understanding of mechanism of angiogenesis and angiogenic signaling of pro-angiogenic factors such as vascular endothelial growth factor (VEGF) and sphingosine 1-phosphate (S1P) will aid in developing therapeutic angiogenesis for ischemic heart disease and peripheral arterial disease. Recently, Rap1 small G protein was identified as a key angiogenic signaling molecule. However, mechanisms involved in the Rap1 regulation of angiogenesis and downstream effectors of Rap1 are not known. Afadin is an adherens junction protein that connects immunoglobulin-like adhesion molecule nectins to the actin cytoskeleton and binds activated Rap1. Recently, we identified that, besides the role in cell-cell adhesion formation in epithelial cells, afadin plays a role in the regulation of directional cell movement of fibroblasts. We therefore attempted to investigate the role of endothelial afadin in angiogenesis and clarify the underlying molecular mechanism. [Method and Results] Knockdown of Rap1 or afadin by siRNA inhibited the VEGF- and S1P-induced tubulogenesis, migration and proliferation, and conversely, increased the serum deprivation-induced apoptosis of human umbilical vein endothelial cells (HUVECs). Immunoprecipitation and immunofluorescence studies revealed that Rap1 regulated the interaction between afadin and phosphatidyl-inositol 3-kinase (PI3K) and recruitment of the afadin-PI3K complex to the leading edge, suggesting that Rap1 and afadin regulates the PI3K signaling. Indeed, knockdown of Rap1 or afadin by siRNA inhibited the VEGF- and S1P-induced phosphorylation of Akt. Thus, afadin regulated the PI3K-Akt signaling. To examine the role of afadin in angiogenesis in vivo, we developed endothelial cell-specific afadin-knockout mice using the Cre/loxP system. Majority (96%) of afadin-knockout mice were embryonically lethal, suggesting an essential role of endothelial afadin in embryonic vascular development and angiogenesis. We performed Matrigel plug assay in born adult afadin-knockout mice and observed that VEGF- and S1P-induced angiogenesis was impaired in afadin-knockout mice. [Conclusion] These results demonstrate that afadin regulates VEGF- and S1P-induced angiogenesis downstream of Rap1.
Abstract 5589: Induction of Angiogenesis by RTEF-1 via NF <i>κ</i> B-dependent Transcriptional Regulation of HIF-1 <i>α</i>
Authors: Yi Jin, Jiaping Wu, Brittany Cully, Ming Xu, Shiyin Foo, Anthony Rosenzweig, Jian Li
Journal: Circulation (2009)

Background and Objective : We previously demonstrated that Related Transcription Enhancer Factor-1 (RTEF-1) plays an important role in hypoxia-induced angiogenesis, in which hypoxia inducible factor-1 (HIF-1) is regulated at the post-translational level to control several angiogenic genes. To further explore the ability of RTEF-1 in the regulation of angiogenesis, this study was designed to assess the potential importance of interrelationships between RTEF-1 and HIF-1 in hypoxic endothelial cells and in an ischemia model of RTEF-1 transgenic mice. Methods and Results: Down-regulation of HIF-1 α mRNA was found in endothelial cells under hypoxic conditions, but it can be recovered by increased expression of RTEF-1. Luciferase assay demonstrated that RTEF-1 increased HIF-1 α promoter activity by 3.73±0.23 fold. Sequential deletion analyses of the HIF-1 α promoter identified that an NF κ B-related element located between −197 and −188 was essential for RTEF-1 function. By gel shift assay, RTEF-1 was found to interact with the NF κ B binding sequence, which was also confirmed by chromatin immunoprecipitation. Wedelolactone, an inhibitor of NF κ B, blocked the HIF-1 α promoter activity in the presence or absence of RTEF-1. In addition, induction of HIF-1 α promoter activity by RTEF-1 resulted in an increase of HIF-1 protein level and angiogenic processes including endothelial cell proliferation and vascular structure formation. Moreover, over-expression of RTEF-1 in endothelial cells (HMEC-1) led to a significant increase in cell aggregation in matrigel analysis, with further identified increases of connexin 43 expression and promoter activity. In the hindlimb ischemia model of RTEF-1 transgenic mice, increases of HIF-1 α and vascular endothelial growth factor (VEGF) were detected with an accelerated recovery of blood flow (75% vs. 58%, at day 9) and increased capillary density compared with the wild-type littermate controls. Conclusion: RTEF-1 increases angiogenesis via NF κ B-dependent transcriptional regulation of HIF-1 α , and consequently affects endothelial remodeling by increasing cell aggregation via connexin 43. These findings provide a novel mechanistic insight into hypoxia-associated angiogenesis.
Abstract 5595: Regulated Expression and Function of c-Myb During Cardiovascular-directed Differentiation of Mouse Embryonic Stem Cells
Authors: Masayoshi Ishida, Karolina M Kolodziejska, Steven Kattman, Gordon Keller, Mansoor Husain
Journal: Circulation (2009)

Background: c-Myb plays a pivotal role in hematopoiesis and in proliferation of vascular smooth muscle cells (VSMC). We showed that c-myb null (KO) mouse embryonic stem cells (ESC) were unable to form contractile VSMC, but formed contracting cardiomyocytes (CM) in embryoid bodies (EB) with seemingly greater efficiency. This suggested lineage-defining effects of c-Myb during CM and VSMC differentiation. Methods &amp; Results: To investigate underlying mechanisms, we first studied c-Myb levels in wild-type (WT) ESC. In serum-conditioned spontaneously differentiating WT-ESC, Western blot revealed high c-Myb levels on days 0 –2, rapid/complete loss of c-Myb on days 2.5 to 3, and subsequent re-expression during days 4 – 6 of differentiation. The loss of c-Myb between days 2.5–3.0 did not occur with any change in mRNA, but was completely blocked by proteosome inhibitor MG132. We next studied ESC-derived cardiovascular progenitor populations by flow cytometry using VEGFR2 and PDGFR α makers. By day 3, VEGFR2+/PDGFR α - cells were significantly fewer, while VEGFR2-/PDGFR α + and VEGFR2+/PDGFR α + cells were more frequent, in KO vs. WT EB. In serum-free cardiac-directed differentiation cultures of WT ESC, Western blot revealed the highest levels of c-Myb expression at day 2, with rapid loss by day 3 in VEGFR2+/PDGFR α + cells. Of interest, VEGFR2+/PDGFR α - cells showed the highest levels of c-Myb, while VEGFR2+/PDGFR α + cells expressed the lowest. Subsequently, c-Myb was not detectable in beating CM populations and differentiating CM that emerged under these conditions. Lentiviral constructs enabling inducible expression of a shRNA against c-Myb were used to temporally knock down c-Myb expression and observe corresponding effects on cardiovascular differentiation. Finally, co-IP and mass spectrometry identified TIF1 β , a known negative regulator of c-Myb, as a putative c-Myb partner proteins affecting c-Myb-modulated cardiovascular lineage specification. Summary: These data show that c-Myb is tightly regulated by proteosomal degradation and protein-protein interactions during ESC differentiation, and that it plays a role in the differentiation of VEGFR2+/PDGFR α - cells, which may represent the source of the contractile VSMC observed in EBs.
Abstract 5624: C-Reactive Protein Predicts First Coronary Events but Not Recurrent Stroke: Results From the SPARCL Study
Authors: Peter Ganz, Gregory M Preston, Weihang Bao, Pierre Amarenco, Larry Goldstein, Henrik Sillesen, K M Welch
Journal: Circulation (2009)

Background: The inflammatory biomarker C-reactive protein (CRP) predicts the risk for cardiovascular events. The placebo-controlled Stroke Prevention by Aggressive Reduction in Cholesterol Levels (SPARCL) trial evaluated the effect of atorvastatin 80 mg/d on the risk of stroke and major coronary events (COR) in patients with a recent stroke or transient ischemic attack and no known coronary heart disease (CHD). This nested case-control analysis of SPARCL data investigated whether CRP levels at enrolment (1– 6 months after stroke) predicted recurrent stroke or first major coronary events (COR: cardiac death or MI) in placebo-treated patients in this unique patient population. Methods: Baseline CRP was measured in 1,243 of 2,366 placebo-treated SPARCL patients, including 278 who had a recurrent stroke, and 94 who had a COR. Baseline characteristics of the sub-study and total SPARCL populations were similar. The predictive value of AHA/CDC recommended cut-points for CRP (&lt;1 [reference group], 1–3, &gt;3 mg/L) was evaluated without and with adjustments for age, gender, smoking, diabetes, entry event, time since entry event, geographic region and race. Results: Baseline CRP (median [interquartile range]) was 2.08 [0.96, 4.66] mg/L. Higher baseline CRP predicted COR but not recurrent stroke (Table ). The results were similar using quartiles of CRP. Conclusion : In this unique cohort of subjects with cerebrovascular disease but no known CHD, baseline CRP predicted COR, but not recurrent stroke. These results point to potentially important differences in the role of inflammation for first coronary vs. recurrent cerebrovascular events.
Abstract 5578: Cell Polarity Factor Par3 Binds SPTLC1 Modulating Monocyte/Macrophage Serine Palmitoyltransferase Activity, De Novo Ceramide Synthesis and Chemotaxis
Authors: Norimasa Tamehiro, Zahedi Mujawar, Michael L Fitzgerald
Journal: Circulation (2009)

Elevated sphingolipids associate with increased cardiovascular disease. Conversely, atherosclerosis is reduced in mice by blocking de novo synthesis of sphingolipids catalyzed by serine palmitoyltransferase (SPT). The SPT enzyme is composed of the SPTLC1 and 2 subunits, and we have identified a novel protein-protein interaction between SPTLC1 and the PDZ protein Partitioning Defective Protein 3 (Par3). Mammalian SPTLC1 orthologs have a highly conserved C-terminus that conforms to a type II PDZ protein interaction motif, and by screening PDZ domain protein arrays with a SPTLC1 C-terminal peptide we found it bound the 3 rd PDZ domain of Par3. Overlay and immuno-precipitation assays confirmed this interaction and indicate Par3 associates with the SPTLC1/2 holoenzyme by binding the C-terminal PDZ motif of SPTLC1, and this binding can re-localize SPTLC1 to the periphery of the cell at sites of cell-cell contact and in cell protrusions. The SPTLC1/2-Par3 complex was detected at endogenous expression levels in mouse liver and primary macrophages demonstrating its physiologic existence, and siRNA inhibition of Par3 in human THP-1 monocytes significantly reduced SPT activity and the de novo synthesis of ceramide and sphingomyelin by nearly 40%. Given monocyte recruitment into inflamed vessels promotes atherosclerosis, and since Par3 and sphingolipids have been associated with polarized cell migration, we tested and found that Par3 and SPTLC1 are required for the ability of THP-1 monocytes to migrate towards monocyte chemoattractant protein 1 (MCP-1). Knockdown of Par3 significantly reduced MCP-1 induced chemotaxis of THP-1 monocytes, as did knockdown of SPTLC1, and this Par3 effect depended upon SPT activity. Migration in response to other inflammatory G-protein coupled chemokines including fractalkine and fMLP was also inhibited in these cells and in primary mouse macrophages treated with the anti-atherosclerotic SPT inhibitor myriocin. In conclusion, we have identified a novel SPTLC1-Par3 interaction that promotes monocyte serine palmitoyltransferase activity and chemotaxis towards inflammatory chemokine signals.
Abstract 5601: Podoplanin Expressing Cells Isolated From Bone Marrow and Peripheral Blood Function as Lymphatic Endothelial Progenitor Cells
Authors: Ji Yoon Lee, Changwon Park, Eugine Lee, Hyongbum Kim, Young-sup Yoon
Journal: Circulation (2009)

Background: Previously, we demonstrated that podoplanin expressing (pod+) cells can be culture-isolated from bone marrow (BM) mononuclear cells (MNCs) and that these cells have a potential to differentiate into lymphatic endothelial cells (LECs). In this study, we investigated whether pod+ cells exist in BM and peripheral blood (PB), and they can function as lymphatic endothelial progenitor cells (LEPCs). Methods and Results. First we performed FACS analysis and found that pod+ cells are present in BM and PB less than 0.5% in C57/BL6 mice. Thus, next to investigate whether the number of pod+ cells would be increased in the BM or PB under conditions which promote lymphatic vessel growth, mice were injected with B16-F1 melanoma cells into the back skins. FACS analysis conducted 7 days later showed that the number of pod+ cells in BM and PB were about 15 fold increased in mice with melanoma growth (normal vs tumor, BM: 0.2±0.5% vs. 3.2±1.0%, PB: 0.4±0.3% vs. 6.7±2.0%, P &lt; 0.01). FACS analysis further revealed that the BM pod+ cells derived from the tumor-bearing mice exhibited high levels of VEGFR-3+ and c-KIT+, but not LYVE-1+ cells, but PB pod+ cells derived from the same mice displayed VEGFR-3+ and LYVE-1+, but not c-KIT+ cells. These data suggest that more undifferentiated LEPC phenotypes exist in BM and undergo differentiation during mobilization into PB. Next, to determine the lymphvasculogenic potential of the isolated BM pod+ cells in vivo, we injected the pod+ cells (Dil-labeled or GFP mouse derived) isolated from BM of tumor-bearing mice into skin and ear wound models. Immunohistochemistry showed that the injected cells were incorporated into lymphatic vessels and expressed LEC-specific markers, suggesting LEC differentiation. We also found that these pod+ cells express CD45 in BM but lost CD45 expression after incorporation into lymphatic vessels, suggesting that pod+ cells are hematopoietic in origin but lost hematopoietic properties during differentiation into LEC. In Conclusion. This study for the first time demonstrated that pod+ cells are present in BM and PB, are augmented under lymphatic vascular growth conditions, and contributed to lymphvasculogenesis, suggesting a role of pod+ cells as LEPCs.
Abstract 5560: Association of 47 Candidate Proteomic Markers of Vascular Disease With the Ankle Brachial Index: The Mayo Clinic Proteomic Markers of Arteriosclerosis Study
Authors: Charles X Kim, George G Klee, Thomas H Mosley, Iftikhar J Kullo
Journal: Circulation (2009)

Objectives: We investigated the association of 47 candidate proteomic markers in pathways of inflammation, thrombosis, lipoprotein metabolism, vascular remodeling, and vasoregulation, with the ankle-brachial index (ABI) and peripheral arterial disease (PAD) in a bi-ethnic cohort of African Americans (AA) and non-Hispanic Whites (NHW). Methods: Multiple assay platforms were used to measure 47 candidate proteomic markers in 1324 AA and 1237 NHW participants belonging to hypertensive sibships. ABI was measured using a standard protocol, with values &gt;1.4 excluded and PAD defined as ABI &lt;0.9. Multivariable regression analyses (with generalized estimating equations) were employed to identify markers independently associated with ABI and PAD after adjusting for age, BMI, smoking, hypertension, diabetes, history of heart attack and stroke, medication use, total and HDL cholesterol, and estimated GFR as well as physical activity, alcohol consumption and education level. Results: Higher circulating levels of CRP, myeloperoxidase, NT-proBNP, osteoprotegerin, D-Dimer and fibrinogen were each associated with lower ABI in both AA and NHW. Additional markers significantly associated with lower ABI were; in AA - serum amyloid A, ICAM, IL-6, TNFR-I, TNFR-II, MCP-1, Lp(a), pro-atrial natriuretic peptide, pro-arginine vasopressin, pro-adrenomedullin, pro-endothelin, osteonectin, Factor VIII; and in NHW- Hsp27, ox-LDL and resistin. Markers associated with presence of PAD were: in AA - CRP, NT-proBNP, Lp(a), pro-arginine vasopressin and osteoprotegerin; in NHW -CRP, NT-proBNP, Hsp27, myeloperoxidase, oxidized-LDL, Factor VIII, and D-Dimer. When the significant markers were included in a final model, the R 2 for ABI increased from 15.9 to 19.4 for AA and from 18.1 to 22.1 for NHW; the c-statistic for PAD increased from 0.791 to 0.833 inAA and from 0.841 to 0.866 in NHW. Conclusions: Novel protein markers significantly add to prediction of ABI and PAD in both AA and NHW and may have clinical utility in diagnosis and prognosis of PAD as well as in identifying new therapies for atherosclerotic vascular disease.
Abstract 5591: KLF2 Restores Neovascularization Capacity of Aged Proangiogenic Cells
Authors: Reinier A Boon, Carmen Urbich, Ariane Fischer, Ruud D Fontijn, Florian H Seeger, Masamichi Koyanagi, Anton J Horrevoets, Stefanie Dimmeler
Journal: Circulation (2009)

Coronary artery disease (CAD) patients have less circulating endothelial progenitor cells (EPCs), which exhibit impaired neovascularization properties. Inverse correlations were also found between EPC function and risk factors like age. Krüppel-like factor 2 (KLF2) is expressed by mature endothelial cells (ECs), is induced by both shear stress and statins and provokes endothelial functional differentiation. Here, we describe that early EPCs, also known as proangiogenic cells (PACs), express KLF2 at a comparable level to mature ECs and that senescence decreases KLF2 levels. To study the effects of aging on KLF2 levels, we compared progenitor cells of 4 weeks and 16 –18 months old C57BL/6 mice. In addition to the 2.5-fold reduction of circulating Sca1+/c-Kit+/Lin- progenitor cells and the 40% reduction of Sca1+/Flk1+ endothelial committed progenitor cells, the spleen-derived PACs isolated from aged mice showed a 60% lower level of KLF2 when compared to PACs isolated from young mice. Moreover, PACs isolated from peripheral blood of aged CAD patients were shown to contain 3-fold less KLF2 than PACs isolated from healthy young controls. Lentiviral overexpression of KLF2 increased human PAC numbers by 60% during in vitro culture. Endothelial lineage-specific KLF2 overexpression in aged bone marrow-derived mononuclear cells strongly augments neovascularization in vivo in a murine hind-limb ischemia model. These results imply that KLF2 is an attractive novel target to rejuvenate PACs before autologous administration to CAD patients.
Abstract 5565: Smoking-Induced Long-lasting Modifications Through a Monoamine Oxidase Epigenetic Regulation
Authors: Francine RENDU, Muriel Del Pino, Gilles Chironi, Jacques Callebert, Katell Peoc’h, Alain Simon, Jean-Marie Launay
Journal: Circulation (2009)

Objectives: Smoking is a major cardiovascular risk factor. We postulated that serotonin, released from activated platelets, could be involved in smoking-induced vascular modifications. Patients &amp; Methods: We studied 115 men with no history of cardiovascular disease and no diabetes, distributed as smokers (S), former smokers (FS) who had stopped smoking for a mean of 13 years, and never smokers (NS). There was no difference in age, BMI, blood pressure or lipid status between the 3 groups. Serotonin, monoaminoxidase (MAO-B) activities and amounts were measured in platelets, and 5-hydroxyindolacetic acid (5-HIAA) - the serotonin/MAO catabolite - in plasma samples. Genomic sequencing of the MAO gene promoter was performed on peripheral blood monocytic cells (PBMC). Results: Both platelet serotonin and plasma 5-HIAA levels were correlated with the 10-year cardiovascular Framingham relative risk (P&lt;0.01), but these correlations became non significant after adjustment for smoking status, underlining that the determining risk factor among those taken into account in the Framingham risk calculation was smoking. Unexpectedly, the platelet serotonin content was similar in S and NS but lower in FS with a parallel higher plasma level of 5-HIAA in FS. This was unforeseen since MAO-B activity was inhibited during smoking (P&lt;0.00001). It was, however, consistent with a higher enzyme amount found in S and FS than in NS (P&lt;0.001). It thus appears that MAO inhibition during smoking was compensated by a higher synthesis. To investigate the persistent increase in MAO-B a study of the methylation of its gene promoter in a supplementary cohort of similar subjects. We found that the methylation frequency of the MAOB gene promoter was markedly lower (P&lt;0.0001) for S and FS vs. NS due to cigarette smoke-induced increase of nucleic acid demethylase activity. Conclusions: This is the first report that smoking induces an epigenetic modification lasting over ten years after quitting. A better understanding of the epigenome may help to further elucidate the development of smoking-induced vascular complications. The results could have a larger impact than cardiovascular damage, considering that MAO-dependent serotonin catabolism is also involved in addiction and lung cancer.
Abstract 5621: A Phase 2 Feasibility Trial of the Oral Direct Thrombin Inhibitor AZD0837 in the Prevention of Stroke and Systemic Embolic Events in Patients With Atrial Fibrillation, Who Are Unable or Unwilling to Take Warfarin Therapy
Authors: Gregory Y Lip, Lars H Rasmussen, S B Olsson, Sofia Zetterstrand, Christina Stahre, Anders Bylock, Maria Aunes-Jansson, Karin Wåhlander
Journal: Circulation (2009)

Many patients with atrial fibrillation (AF) cannot be treated with vitamin K antagonists (VKA) such as warfarin for various reasons, and will therefore not receive effective thromboprophylaxis. Development of a novel oral anticoagulant with an improved safety profile while maintaining the efficacy of VKA would represent a significant advance in the management of AF. The primary objective of the present Phase 2 trial ( NCT00623779 ) was to assess the feasibility of conducting a study in patients with AF unable or unwilling to take warfarin, by evaluation of drop-out rate and compliance with treatment and study procedures. Secondary objectives included assessment of safety/tolerability (including bleeding), and pharmacodynamic properties of AZD0837. Patients were randomised to receive AZD0837 extended-release tablets 150 mg (n=41) or 300 mg (n=42) once daily, or standard therapy (no treatment, ASA 75–325 mg or clopidogrel 75 mg once daily) (n=45). The study was blinded to AZD0837 dose. Treatment duration was between 4 weeks and 6 months (median 6 weeks). Reasons for patients not being treated with warfarin were: permanent cessation or refusal by the patient (62%), physicians assessment that VKA was inappropriate (21%), inability to keep INR between 2.0 –3.0 over a 3-month period (24%), and warfarin allergy (2%). 40% of patients were warfarin-naïve. Compliance with study treatment (mean±SD) was 96.9±16.5% for AZD0837 150 mg and 99.8±11.4% for 300 mg. Compliance with study visits was 93.3±15.0% for AZD0837 150 mg, 95.6±10.4% for 300 mg and 97.5±6.8% for standard therapy. The number of drop-outs were 4, 6 and 3, whilst minor or clinically significant minor bleeds were reported in 0, 5 and 2 with AZD0837 150 mg, 300 mg and standard therapy, respectively. No major bleeding events were reported. Both doses of AZD0837 showed relevant anticoagulation effects in terms of reduced levels of fibrin D-Dimer, and prolongation of ECT and APTT. This Phase 2 trial indicates that a larger study in this group of AF patients in need of effective thromboprophylaxis may be feasible, as judged by compliance and drop-out rates. The oral direct thrombin inhibitor AZD0837 has a favorable safety profile including a low incidence of bleeding events, and seems to offer effective anticoagulation.