tbuckley/cabot-search huggingface dataset:
Abstract 5634: Molecular Imaging of Bone Marrow Mononuclear Cell Survival and Homing in Murine Peripheral Artery Occlusive Disease
Authors: Koen E Van Der Bogt, Alwine A Hellingman, Maarten A Lijkwan, Ernst J Bos, Margreet R de Vries, Michael P Fischbein, Paul H Quax, Robert C Robbins, Jaap F Hamming, Joseph C Wu
Journal: Circulation (2009)

Introduction . Bone marrow mononuclear cell (BMMN) therapy has emerged as a potential treatment for peripheral artery obstructive disease (PAOD). However, much is unknown about cellular kinetics following transplantation. This study aimed to use molecular imaging to provide insight into cellular behavior following different transplantation methods. Methods . BMMN were isolated from male F6 transgenic mice (FVB background) that express firefly luciferase (Fluc) and green fluorescence protein (GFP). BMMNs were characterized by bioluminescence imaging (BLI). Male FVB mice (n=33) underwent ligation of the superficial femoral artery and were randomized into 4 groups to receive: Single intramuscular (i.m.) injection of 2×10 6 MN; 4 weekly i.m. injections of 5×10 5 MN; PBS control; and 5×10 6 MN intravenously (i.v.). Cellular kinetics were measured by BLI. Paw perfusion was monitored by Laser Doppler Perfusion Imaging (LDPI). Results. Fluc expression correlated with cell number in all groups (r 2 =0.97). In vivo BLI revealed dismal cell survival within 4 weeks following both i.m. transplanted groups. Following i.v. transplantation, BLI showed early homing to the injured area as well as to liver, spleen, and bone marrow. LDPI showed no significant differences in paw perfusion among i.m. groups. Conclusion. This is the first study to assess kinetics of transplanted BMMN in PAOD using in vivo molecular imaging. BMMN survival after both i.m. and i.v. transplantation is short lived and BMMN do not preserve perfusion in this model.
Abstract 5472: Nox4 is a Novel Inducible Source of Intracellular Reactive Oxygen Species in Human Monocyte-Derived Macrophages and Mediates Oxidized LDL-induced Macrophage Death
Authors: Chi Fung Lee, Mu Qiao, Katrin Schroeder, Reto Asmis
Journal: Circulation (2009)

Nox2/gp91 phox is the large subunit of the integral membrane component of the phagocytic NADPH oxidase (Nox) complex and was believed to be the sole Nox isoform present in monocytes and macrophages. Previously, we reported that increased intracellular ROS formation is required for OxLDL-induced cell death in human monocyte-derived macrophage (HMDM), but the source of these intracellular ROS was unknown (Asmis, Circ.Res. 2003). We now identified Nox4, not Nox2, as the source of OxLDL-induced ROS generation. Nox4 mRNA was detected in human blood monocytes and HMDM. Using a novel rabbit monoclonal antibody that detects Nox4 as a single 66 kDa band, we localized Nox4 to the ER, the nucleus and focal adhesion sites of HMDM, suggesting that Nox4 may play a role in macrophage redox signaling. After stimulation of HMDM with OxLDL, but not native LDL, Nox4 mRNA and protein expression were up-regulated in a dose- and time-dependent manner. Nox4 protein expression was maximal after 6 h and coincided with both increased p22 expression and increased intracellular ROS formation, suggesting that the Nox4/p22 complex is responsible for OxLDL-induced ROS formation in HMDM. Inhibition of MEK1 and ERK1/2 phosphorylation blocked both the induction of Nox4 expression and intracellular ROS formation induced by OxLDL, and protected HMDM from OxLDL-mediated cytotoxicity. In contrast, inhibitors of p38 MAPK or JNK affect neither Nox4 induction nor ROS formation, and showed no cytoprotection. Knockdown of Nox4 with adenovirus-delivered siRNA decreased OxLDL-induced ROS formation and protected macrophages from OxLDL-induced cell injury. In summary, we identified Nox4 as a novel, inducible source of intracellular ROS in human monocytes and macrophages. Nox4 is likely to play important roles in redox signaling and macrophage function. However, if overinduced Nox4-derived ROS promote macrophage death, a mechansim which may have important implications for the pathogenesis of atherosclerosis. This research has received full or partial funding support from the American Heart Association, South Central Affiliate (Arkansas, New Mexico, Oklahoma &amp; Texas).
Abstract 5569: Neointimal Hyperplasia After Mechanical Endovascular Injury is Augmented Through Angiotensin II-mediated Regional Vascular Inflammation via Activation of the JNK Pathway in Angiotensin-converting Enzyme 2-deficient Mice
Authors: Makoto Sahara, Masataka Sata, Toshihiro Morita, Yasunobu Hirata, Ryozo Nagai
Journal: Circulation (2009)

Background: Angiotensin-converting enzyme 2 (ACE2) is a negative regulator of the renin-angiotensin system, although its role on vascular inflammation and atherosclerosis remains unclear. Methods and Results: Male ACE2-deficient (ACE2 − /y ) and wild-type (ACE2 + /y ) mice underwent wire-mediated endovascular injury in femoral artery. After 4 weeks, the injured artery in ACE2 − /y mice exhibited marked neointimal hyperplasia compared with that in ACE2 + /y mice. The intima/media ratio in the vascular lesion was 1.97±0.31 in ACE2 + /y mice and 2.78±0.49 in ACE2 − /y mice ( P &lt;0.05). There were no significant differences in blood pressure between both mice. In histology, the accumulation of macrophages in lesions was significantly augmented in ACE2 − /y mice. ACE2 protein expression in vascular cells in wild-type mice was detected by immunohistochemistry and western blotting, and the concentration of angiotensin II (Ang-II) in the injured artery was higher in ACE2 − /y mice than in ACE2 + /y mice. In addition, in the injured artery, quantitative mRNA expressions of vascular inflammatory molecules such as VCAM-1, MCP-1, TNF α , and MMP9 were increased more markedly in ACE2 − /y mice. In contrast, an ARB, olmesartan, attenuated the lesion formation and mRNA expressions of these molecules in the injured artery in ACE2 − /y mice. Next, we analyzed phosphorylation status of ERK and JNK in aortic vascular smooth muscle cells (VSMCs), which were derived from ACE2 + /y or ACE2 − /y mice and were stimulated with Ang-II. Although Ang-II increased ERK and JNK phosphorylation in VSMCs of both mice, the increase was greater in VSMCs of ACE2 − /y mice. Moreover, we evaluated which signaling pathway mediated Ang-II-induced upregulation of VCAM-1 and MMP9 in VSMCs. Although an ERK inhibitor PD98059 had no impact on Ang-II-induced upregulation of VCAM-1 and MMP9 mRNA expressions, a JNK inhibitor SP600125 significantly attenuated them, suggesting the central role of the JNK pathway in transducing the impact of Ang-II on vascular inflammation. Conclusions: ACE2 deletion induces marked neointimal formation in femoral artery after vascular injury through Ang-II-mediated regional vascular inflammation via activation of the JNK pathway. ACE2 may confer protection against vascular diseases.
Abstract 5568: Impact of Angiotensin II Receptor Blockers on Regression of Coronary Atherosclerotic Plaque -OVATION Study
Authors: Hideki Ishii, Masakazu Kobayashi, Satoshi Ichimiya, Masaaki Kanashiro, Nobutake Kurebayashi, Takahito Sone, Hideyuki Tsuboi, Hiroaki Mukawa, Tetsuya Amano, Tadayuki Uetani, Nobuyuki Marui, Tatsuaki Matsubara, Toyoaki Murohara
Journal: Circulation (2009)

OBJECTIVES: Coronary intravascular ultrasound (IVUS) can provide an imformation of coronary atherosclerosis. Recent reports suggest that coronary plaques can be reduced by some medications. On the other hand, angiotensin II receptor blockers (ARBs) improve clinical outcomes in patients with hypertension. The aim of this multi-center study was to investigate whether blood pressure-lowering therapy with ARBs had efficacies regarding regression of coronary atherosclerosis. Methods and Results: Seventy one hypertensive patients with stable angina pectoris who underwent elective PCI were enrolled. They received one of two ABRs (olmesartan 20mg/day or valsartan 80 mg/day) after PCI. Non-target coronary lesions with mild to moderate stenosis were measured by volumetric IVUS at both baseline and 6-month follow-up period in those patients. Systolic blood pressure was significantly decreased by 16.7% after treatment with ARBs (P &lt; 0.01 vs baseline). There were no significant differences in vessel volume (103.9±58.4 mm 3 at baseline and 98.7±57.6 mm 3 at 6-month follow-up) and lumen volume (57.2±33.6 mm 3 at baseline and 57.0±34.2 mm 3 at 6-month follow-up) between baseline and 6-month ARB treatment. However, statistically significant reduction in plaque volume was observed in examined coronary arteries (46.7±31.8 mm 3 at baseline and 41.7±28.4 mm 3 at 6-month follow-up, 10.2% decrease from baseline, P = 0.01). Interestingly, serum adiponectine levels increased from 9.0±4.2 μ g/ml to 10.7±5.4 μ g/ml (an increase of 19.4 %, P &lt; 0.01), and serum MDA-LDL cholesterol levels decreased from 107.8±46.4 IU/l to 95.5±28.0 IU/l (a decrease of 11.4 %, P &lt; 0.01). Conclusions: Blood pressure-lowering therapy with ARBs or 6 months significantly reduced the plaque volume at non-target coronary lesions in patients with stable angina pectoris. Furthermore, a significant increase in serum adiponectine levels and a significant decrease in serum MDA-LDL cholesterol levels were also observed after 6-month ARBs treatment.
Abstract 5518: Abnormal Coronary Flow Reserve is Related to Impairment of Left Ventricular Function During Adenosine Stress Echocardiography in Untreated Hypertensive Patients
Authors: Ignatios Ikonomidis, Stavros Tzortzis, John Lekakis, Helen Triantafyllidi, Costas Papadopoulos, Costas Tsitlakidis, Ioannis Paraskevaidis, Dimitrios T Kremastinos
Journal: Circulation (2009)

Objective: Coronary microcirculation is disturbed in hypertensive patients. We investigated the effects of coronary flow reserve (CFR) on the response of Left Ventricular (LV) systolic and diastolic function as assessed by Tissue Doppler Imaging during adenosine stress echocardiography in never-treated hypertensive patients. Methods: We studied 43 patients (mean age: 53±12 years), with untreated arterial hypertension (clinic blood pressure &gt;140/90 mmHg and &gt;125/80 mmHg in 24hour ambulatory BP monitoring) by adenosine stress echocardiography and 22 healthy controls matched for sex, age and atherosclerotic factors. We measured: time integral (VTI-Vd) of diastolic (Vd) coronary blood flow velocity by transthoracic Doppler echocardiography of the LAD Ratio of VTI-Vd after adenosine infusion (140 μ g/kg/min) to VTI-Vd at baseline to assess CFR E and A doppler, Sm, Em, Am lateral mitral annulus velocities with TDI, Em/Am ratio and E/Em ratio before and during adenosine infusion to assess LV systolic and diastolic function, the percent % changes of the measured indices between baseline and peak adenosine infusion. Results: All patients had normal baseline ejection fraction. Compared to controls, hypertensives had lower CFR (2.3±0.7 vs. 2.9±0.9, p=0.03). At baseline, hypertensives compared to controls, had lower Em/Am ratio (0.9±0.3 vs. 1.3±0.4, p=0.002), Sm (8.7±1.7 vs. 9.7±2.6, p=0.07), Em (10.1±2.8 vs 12.1±3.5, p=0.03) and higher Am (11.1±2.5 vs. 9.5±2.0, p=0.03). At peak adenosine infusion there were an increase in Sm, Em and Am in all patients (p&lt;0.05). Furthermore a decreasing CFR was associated with a decreasing Em (r= 0.46, p=0.05), Em/Am (r=0.49, p=0.048) and increasing E/Em (r=−0.4, p=0.04). A reduced CFR was related with a smaller % increase of Sm (r= −0.40, p=0.049), % increase of Em (r=− 0.5, p=0.033) and higher % increase of E/Em (r=−.42, p=0.04) at peak stress. Patients with CFR&lt;2.5 (n=28), compared to those with CFR≥2.5 had lower % change of Sm (27±20% vs. 51±34% p=0.04), % change of Em (8±3% vs. 46 ±3%, p=0.05) and higher % increase of E/Em (24±3% vs −1±2%, p=0.04). Conclusions: An abnormal response of LV function during adenosine stress echocardiography is related with impaired CFR in untreated hypertensive patients.
Abstract 5480: Laminin Receptor Activation Inhibits Endothelial Tissue Factor Expression
Authors: Erik W Holy, Simon F Stämpfli, Alexander Akhmedov, Giovanni G Camici, Thomas F Lüscher, Felix C Tanner
Journal: Circulation (2009)

Background. Tissue factor (TF) is an important trigger of arterial thrombosis. The extracellular matrix glycoprotein laminin is a major compound of the basement membrane contributing to vessel wall integrity. The green tea catechin epigallocatechin-3-gallate (EGCG) is a ligand of the 67 kDa laminin receptor (67-LR). This study investigates the effect of 67-LR activation by laminin and EGCG on endothelial TF expression. Methods and Results . Immunofluorescence demonstrated that human aortic endothelial cells expressed 67-LR abundantly. Western blot analysis demonstrated that cells grown on laminin expressed less TF in response to TNF- α than those grown on fibronectin (n=6; p &lt;0.001). EGCG inhibited TNF- α and histamine induced TF expression and activity in endothelial cells in a concentration-dependent manner (1–30 μ M) resulting in a 87% reduction of TF expression (n=5; p &lt;0.001). In contrast, expression of tissue factor pathway inhibitor was not affected (n=4; p =NS). Real-time PCR and promoter studies revealed that inhibition of TF expression occurred at the transcriptional level. EGCG (30 μ M) impaired activation of the mitogen-activated protein (MAP) kinase c-Jun terminal NH2 kinase (JNK), but not ERK or p38. The JNK inhibitor SP600125 (1 μ M) reduced TF promoter activity (n±4, p ±0.001) and protein expression (n=4; p &lt;0.001). 67-LR blocking antibodies blunted the inhibitory effect of EGCG on JNK activation and TF expression. Expression of Vascular cell adhesion molecule 1 (VCAM-1) was not regulated by JNK, and its expression was not affected by 67-LR activation. Conclusions . Laminin receptor activation inhibits endothelial TF expression by impairing JNK phosphorylation. This study identifies a new mechanism by which the basement membrane contributes to hemostatic balance of the endothelium in intact vessels and suggests 67-LR as a potential target for the development of novel anti-thrombotic therapies.
Abstract 5631: Isolation of a Resident and Dynamic Oct4+ Stem Cell Population From Adult Skin
Authors: Anne Limbourg, Vladimir Lozanovski, Sabine Schnabel, Christian Templin, L C Napp, Helmut Drexler, Florian P Limbourg
Journal: Circulation (2009)

The transcription factor Oct3/4 is a critical regulator of pluripotency in embryonic stem cells and induced pluripotent stem (iPS) cells and iPS generation requires re-activation of endogenous Oct4 expression. Since Oct4 might be a critical regulator of somatic stem cell pluripotency we have employed a genetic, bona-fide reporter strategy to determine the temporo-spatial expression of Oct4 in adulte mice. RESULTS: Postnatal Oct4 expression was determined in transgenic Oct4::eGFP reporter mice by flow cytometry, for hearts and skin the response to injury was also evaluated (n=9 –28). The fidelity of the Oct4-GFP reporter strain was confirmed by immunofluorescence confocal microscopy and by RT-PCR. No GFP+ cells were detected in bone marrow or spleen, nor in the atria or ventricles of the myocardium under baseline and post-ischemic conditions. Importantly, Oct4+ cells resided in skin, where levels increased postnatally over time until puberty and returned to low levels in the adult (Oct4+/500.000 cells: puberty 400±268; adult 53±23). Furthermore, the Oct4+ cell pool increased robustly after wound injury (289±156, n=9, p&lt;0.05) even in remote skin areas. In situ analysis revealed the existence of GFP+ niches associated with distinct regions of hair follicle. Oct4+ cells after FACS-sorting revealed a phenotype of c-kit lo, sca-1 +, CXCR4 lo, alpha6 integrin lo, beta1 integrin -, and expressed by qRT low levels of Oct4, Sox2, Klf4, and high levels of c-myc, factors involved in iPS induction. The differentiation potential of Oct4+ skin cells was evaluated by transplanting dual reporter cells (GFP+; lacZ+) into nude mice following hindlimb ischemia and analysis of contribution to tissue lineages. 4 weeks after transplantation, LacZ+ cells contributed to muscle cells, adipose tissue and vascular structures. CONCLUSION: Genetic reporter studies reveal no evidence for Oct4+ cells in bone marrow or heart but, unexpectedly, show persistence of resident Oct4+ stem cells in adult skin, which contribute to mesodermal cell lineage after transplantation. These findings suggest adult skin as an important source for regenerative cell therapy.
Abstract 5515: Mitochondrial Membrane Hyperpolarization and Reduced Mitochondrial Mass Characterize the Arteriolar Endothelium and Mononuclear Cells of Humans With Type 2 Diabetes Mellitus- <i>in vivo</i> and <i>in vitro</i> Assessment of Endothelial Function
Authors: Michael E Widlansky, Joseph A Vita, Tinoy J Kizhakekuttu, Jingli Wang, Emily I Arthur, David D Gutterman
Journal: Circulation (2009)

Background: Endothelial dysfunction and inflammation are central to the vascular pathology of Type 2 diabetes (DM). In vitro data suggest excessive mitochondrial reactive oxygen species production contributes to endothelial dysfunction in part due to pathological hyperpolarization of mitochondrial membrane potential (ΔΨ m ). ΔΨ m hyperpolarization in leukocytes is also associated with activation of the inflammatory response. Mitochondrial mass and morphology are also central to mitochondrial function. The relevance of these mechanisms to the human vasculature in DM is unknown. Methods: Blood mononuclear cells were isolated from 30 DM patients and 32 non-diabetics. Subcutaneous arterioles were harvested by gluteal adipose tissue biopsy in a subset (15 DM, 17 controls). ΔΨ m was measured in cells and arterioles using the membrane potential-sensitive fluorescent dye JC-1. Mitochondrial mass was measured using the cardiolipin-specific fluorophore nonyl acridine orange. Endothelium dependent vasodilation was assessed using video microscopy (arterioles) and ultrasonography (brachial artery). Arteriolar vasoreactivity was also determined in the setting of low-dose (100 nM) CCCP to reduce ΔΨ m magnitude. Results: ΔΨ m magnitude of the arteriolar endothelium and mononuclear cells were significantly greater in DM patients [P=0.01 (endothelium), P &lt; 0.0001 (mononuclear cells)]. Mitochondrial mass was significantly lower in DM mononuclear cells (P=0.004), and in DM endothelium following age adjustment (P=0.003). Endothelium dependent vasodilation was impaired in DM in the brachial artery and subcutaneous arterioles (both P&lt;0.001). CCCP treatment of the DM arteriolar endothelium reversed microvascular endothelial dysfunction (P&lt;0.001 vs. untreated). Conclusions: ΔΨ m is hyperpolarized and mitochondrial mass is lower in the arteriolar endothelium and mononuclear cells of DM patients with endothelial dysfunction. Pharmacological normalization of ΔΨ m reverses microvascular endothelial dysfunction in DM. These findings are consistent with experimental studies suggesting a link between mitochondria and vascular disease. Interventions that restore mitochondrial membrane potential and mass in DM may have therapeutic potential.
Abstract 5474: Crucial Role of Nrf3 in Smooth Muscle Cell Differentiation From Stem Cells and Vessel Injury-Induced Neointima Formation
Authors: Anna E Pepe, Qingzhong Xiao, Anna Zampetaki, Yanhua Hu, Qingbo Xu
Journal: Circulation (2009)

Nuclear factor erythroid 2-related factor 3 (Nrf3), a member of the cap’n’collar family of transcription factors that binds to the DNA-antioxidant responsive elements, have been reported to be involved in muscle precursor migration during early embryo development. The present study aimed to investigate the functional role of Nrf3 in stem cell differentiation into smooth muscle cells (SMCs). When embryonic stem cells were cultivated on collagen IV-coated plates, Nrf3 was up-regulated significantly following 1–6 days of cell differentiation. Knockdown of Nrf3 resulted in down-regulation of smooth muscle specific markers including smooth muscle actin, SM22, calponin and smooth muscle myosin heavy chain, while enforced expression of Nrf3 enhanced SMC differentiation in a dose-dependent manner. SMC-specific transcription factor myocardin, but not serum response factor, was significant upregulated by Nrf3 overexpression. Strikingly, the binding of serum response factor and myocardin to the promoter of smooth muscle differentiation genes was dramatic increased by Nrf3 overexpres-sion, and Nrf3 can direct bind to the smooth muscle gene promoters as demonstrated by chromatin immunoprecipitation (CHIP) assay. Moreover, NADPH-derived ROS production during SMC differentiation was further enhanced by Nrf3 overexpression through upregualtion of NADPH oxidase (NOX4) and inhibition of anti-oxidant signalling pathway. Furthermore, overexpression of Nrf3 resulted in increase of plasma phospholipase A2 (Pla2g7) production, while knockdown Nrf3 ablated the Pla2g7 gene expression and protein activity. Additionally, both Nrf3 and Pla2g7 were up-regulated in adult SMC phenotype swiching from proliferating to contractile phenotype upon serum starvation. Importantly, in vivo knockdown of Nrf3 in the vessel wall significant increased wire-injury-induced neointima formation in ApoE-deficient mice. Our findings demonstrated for the first time that Nrf3 has a crucial role in SMC differentiation from stem cells and important protective function in vessel injury-induced neointima formation, indicating that they could be potential new therapeutic targets for intervention of SMC proliferative-related vascular diseases.
Abstract 5567: The <i>Ldlr</i> <sup>−/−</sup> Mouse as a Novel Model of Aortic Calcification: Application of Micro-Computerized Tomography
Authors: Zuhier Awan, Maxime Denis, Annik Prat, David Goltzman, Nabil G Seidah, Jacques Genest
Journal: Circulation (2009)

BACKGROUND : Patients with familial hypercholesterolemia (FH) due to loss-of-function mutations in the low-density lipoprotein receptor (LDLR) gene exhibit an age-dependent and LDLR gene dosage effect on aortic calcification. We have previously documented that subjects with homozygous FH show significant aortic calcification by their teens, an effect delayed by 20 years in heterozygotes and observed much later in control subjects. The degree of calcification is independent of initial and life long burden of low-density lipoprotein cholesterol concentration. This clinical observation suggests a potential role for the LDLR in osteoblast signalling pathways. In order to better study this observation, we established an animal model of aortic calcification and a novel technique; based on micro-computerized tomography (CT) to examine vascular calcifications in mice. METHOD AND RESULT: We examined C57BL/6 wild-type (wt) and Ldlr −/− mice at 12 and 18 months. The Ldlr −/− mice develop spontaneous atherosclerosis in normal chow-fed older mice. Vascular calcification has not been previously examined in this model. Under isoflurane anesthesia, the aorta was perfused in-situ with saline. The aorta was then carefully dissected and fixed ex-vivo . The tissue was embedded in paraffin block and calcium content was determined by digital x-ray imaging, dual X-ray densitometry, and micro-CT scanning of the entire aorta and aortic valve. The paraffin blocks were then sectioned and Alizarin Red stain was used to further quantify calcified tissue. Analysis of 12 and 18 month wt and Ldlr −/− mice confirmed our observations in human, with aortic calcifications identified at areas of shear stress. CONCLUSIONS: The Ldlr −/− mouse model develops aortic calcification even when fed a normal chow diet. Calcifications at areas of shear stress suggest that calcific aortic stenosis and arterial wall calcifications may share many similar pathophysiological features. This animal model of aortic calcifications, and the CT imaging technique used in the present study, may provide a better understanding of the potential links between FH and calcium deposition in the vascular wall. This model has the potential to study aortic valve calcifications as well.