tbuckley/cabot-search huggingface dataset:
Abstract 5514: Laminar Shear Stress Upregulates Endothelial KCa2.3 and KCa3.1 Calcium-activated Potassium Channels via Activating Calmodulin-dependent Protein Kinase Kinase and AMP-activated Protein Kinase
Authors: Masanori Ohta, Sandhya Nandedkar, Hanjoong Jo, Hiroto Miura
Journal: Circulation (2009)

Aim Endothelial cell (EC) KCa2.3 and KCa3.1, calcium-activated potassium channels, play a crucial role in NO- and EDHF-mediated regulation of vascular tone by controlling membrane potential and intracellular calcium levels. Since laminar shear stress (LS) is a major determinant of EC function in a calcium-dependent manner, we hypothesized that LS upregulates KCa2.3 and KCa3.1 via activating calcium/calmodulin (CaM) kinase cascade. Methods Human coronary artery ECs were exposed to static, LS (15 dyn/cm 2 ) or oscillatory shear stress (OS; +/−5 dyn/cm 2 × 1Hz) condition for 24 hours in the absence or presence of STO-609 (CaM-dependent kinase kinase (CaMKK) inhibitor, 10 μ g/ml), Compound C (AMP-activated protein kinase (AMPK) inhibitor, 10 μ M), KN-62 (calcium/CaM-dependent kinase (CaMK) inhibitor, 10 μ M), KG-501 (cAMP response element binding protein (CREB) inhibitor, 25 μ M), or tumor necrosis factor- α (TNF- α ; 1 or 10 ng/ml). The mRNA expression and CREB phosphorylation were determined by real-time PCR or Western blotting. Results LS markedly increased KCa2.3 and KCa3.1 expressions (3.3±0.4 and 6.6±1.8 fold increases vs. static, n=4 –10, p&lt;0.001), whereas OS reduced KCa2.3 and unchanged KCa3.1 expressions (0.4±0.04 and 1.1±0.5 fold, n=3, p=ns). STO-609 significantly inhibited LS-induced increase in KCa2.3 and KCa3.1 expressions (1.8±0.1 fold and 1.5±0.4 fold) and markedly decreased LS-induced CREB phosphorylation (LS; 2.3 fold and LS+STO-609; 1.5 fold vs. static). KCa2.3 and KCa3.1 expressions were also reduced by Compound C (2.2±1.6 and 2.0±1.0 fold, respectively), but not by KN-62 (5.9±1.1 and 6.5±1.2 fold, respectively). KG-501 suppressed LS-induced increase in both channels (KCa2.3; 1.5±0.7 and KCa3.1; 1.9±0.7 fold). TNF- α completely inhibited both channel expressions in a dose-dependent manner (1 and 10 ng/ml; KCa2.3; 0.6±0.01 and 0.1±0.1 fold, and KCa3.1; 0.7±0.1 and 0.3±0.03 fold). Conclusions LS upregulates KCa2.3 and KCa3.1 expressions via CREB phosphorylation through activation of CaMKK and AMPK, a downstream kinase of CaMKK, whereas TNF- α suppresses both channel expressions. Thus KCa2.3 and KCa3.1 may play an important role in endothelial adaptation to hemodynamic changes and in the maintenance of EC function.
Abstract 5483: The Inflammatory Protein Complement C3 is a Novel Substrate for Factor XIII and Impairs Fibrin Clot Lysis
Authors: Angela M Carter, Verena Schroeder, Victoria R Polkinghorne, Kerrie A Smith, Riyaz Somani, Kristina F Standeven, Peter J Grant
Journal: Circulation (2009)

Introduction: Inflammation is fundamentally important in the pathogenesis of cardiovascular disease (CVD) and increasing evidence supports a functional role for complement C3. Using a functional proteomics approach we have identified zymogen and activated C3 in perfused solubilized clots, suggesting C3 influences thrombus formation. Hypothesis: In this study we assessed the hypothesis that complement C3 interacts with coagulation and fibrinolytic factors to influence thrombosis. Methods and Results: Analysis of 123 paired plasma and serum samples revealed that 24.8% (SD 7.6) of plasma C3 was incorporated into the clot. To explore the mechanism of incorporation we investigated binding interactions between C3, fibrin and FXIII using ELISA based binding assays and surface plasmon resonance and found that C3 bound to immobilized fibrinogen and FXIII in a concentration dependent manner. Further analysis of the interactions between C3 and FXIII demonstrated that activated FXIII (FXIIIa) incorporated a known FXIIIa substrate, 5-(biotinamido)pentylamine, into immobilized C3 in a concentration dependent manner and FXIIIa also induced the formation of high-molecular weight C3 polymers, indicating that C3 is a FXIII substrate. Furthermore, FXIIIa incorporated plasma C3 into immobilized fibrin and Western blot analysis of clots formed from purified fibrinogen and C3 indicated high molecular weight bands that cross-reacted with specific antibodies against C3 and fibrinogen, suggesting that C3 is cross-linked to fibrin alpha chains. In turbidimetric assays using purified proteins C3 prolonged fibrin clot lysis times in a concentration dependent manner and Western blot analysis of lysed clots indicated that C3 was cleaved by plasmin, suggesting impaired fibrinolysis is due to substrate competition for plasmin. Increased plasma C3 concentration was significantly associated with increased ex vivo plasma clot density and prolonged fibrinolysis times, after adjusting for fibrinogen concentrations, to support our in vitro results. In conclusion, taken together these data indicate that complement C3 influences key processes involved in fibrin formation and fibrinolysis supporting a role for C3 in cardiovascular thrombosis.
Abstract 5482: Unexpected Role for Connexin37 in Circulating FVIII and Coagulation
Authors: Laurent Burnier, Anna Pfenniger, Steven Taffet, Mario Delmar, Pierre Fontana, Anne Angelillo-Scherrer, Brenda R Kwak
Journal: Circulation (2009)

The gap junction protein connexin37 (Cx37) plays an important role in cell-cell communication in the vasculature. Cx37 is expressed in endothelial cells, platelets and megakaryocytes. We have recently shown that Cx37 limits thrombus propensity by permitting intercellular signaling between aggregating platelets. Here, we have performed high throughput phage display to identify potential binding partners for the regulatory intracellular C-terminus of Cx37 (Cx37CT). We retrieved 2 consensus binding motifs for Cx37CT: WHK…[K,R]XP… and FH-K…[K,R]XXP…. Sequence alignment against the NCBI protein database indicated 66% homology of one the selected peptides with FVIII B-domain. We performed cross-linking reactions using BS3 and confirmed that an 11-mer peptide of the FVIII B-domain sequence linked to recombinant Cx37CT. In vitro binding of this peptide to Cx37CT was also confirmed by surface plasmon resonance. The dissociation constant of FVIII B-domain peptides to Cx37CT was ~20 uM. Other peptide sequences, designed upstream or downstream of the FVIII B-domain sequence, showed very low or no affinity for Cx37CT. Finally, in vivo studies revealed that thrombin generation in platelet-poor plasma from Cx37−/− mice (endogenous thrombin potential: 634±11 nM min, mean±SEM) was increased compared to Cx37+/+ mice (427±12, P&lt;0.001). Moreover, partial activated thromboplastin time (aPTT) was shorter in Cx37−/− (39.7±1.5 s) than in Cx37+/+ mice (45.9±1.8, P=0.03), whereas prothrombin time was comparable. The shorter aPTT in Cx37−/− mice correlated with higher circulating FVIII activity (46.0±0.7 vs. 53.5±2.7 s for Cx37+/+, P=0.03). Overall, our data show for the first time a functional interaction between FVIII and Cx37. This interaction may be relevant for the control of FVIII secretion and, thereby, in the regulation of levels of FVIII circulating in blood. In addition, these results may open new perspectives to improve the efficiency of recombinant FVIII manufacturing.
Abstract 5502: Ultrasound-Controlled Therapeutic Gas Delivery Provides Enhanced Neuroprotection Following Hypoxic-Ischemia
Authors: Hyunggun Kim, George L Britton, Christy K Holland, David D McPherson, Shao-Ling Huang
Journal: Circulation (2009)

Background: Neurologic damage following ischemic insult is a major problem for stroke survivors. Xenon (Xe) exerts favorable neuroprotective properties with few side effects. This study describes a unique application of ultrasound for controlled release of Xe from liposomes (Xe-ELIP) for targeted therapeutic gas delivery and neuroprotection. Method: Xe-ELIP were created by a pressurization-freezing method. One-MHz continuous ultrasound wave with pressure amplitudes of 0.16, 0.22 and 0.33 MPa was used to trigger Xe release from Xe-ELIP, and endothelial cell permeability and cell viability were measured. Male Sprague-Dawley rats underwent right middle cerebral artery occlusion for 2 hours (n=28). In the treatment group (n=8), Xe-ELIP were administered through the right common carotid artery. Continuous ultrasound (0.22 MPa) was applied over the common carotid artery during the Xe-ELIP administration for 4 minutes. Infarct size as well as behavioral outcomes were determined 3 days after treatments. Results: Ultrasound application in the presence of ELIP resulted in a temporary (5 minutes) enhancement of cell permeability. The amount of released Xe depended on the ultrasound pressure amplitude. Xe-ELIP in combination with ultrasound exposure (0.22 MPa) enhanced neuroprotection by decreasing infarct volume by 88% vs the non-ultrasound Xe-ELIP group (64%). Behavioral tests mirrored neurological results. Conclusions: One MHz continuous ultrasound can be used to release therapeutic gas from encapsulated liposomes. Administration of Xe-ELIP with ultrasound exposure over the carotid artery after a cerebral ischemic insult provides enhanced neuroprotection. This research has received full or partial funding support from the American Heart Association, Midwest Affiliate (Illinois, Indiana, Iowa, Kansas, Michigan, Minnesota, Missouri, Nebraska, North Dakota, South Dakota &amp; Wisconsin).
Abstract 5602: The Transcription Factor Islet-1: A Novel Gene Target for Future Cardiac Repair?
Authors: Aya Barzelay, Edith Hochhauser, Gad Keren, Jacob George
Journal: Circulation (2009)

The LIM-homeobox transcription factor isl1 plays a crucial role during heart embryogenesis. Embryonic isl1+ precursors give rise to over two-thirds of the heart and to its subsequent lineages: cardiac muscle, smooth muscle and endothelium. Interestingly, a subset of Isl1+ progenitors remains embedded in the postnatal heart. In the current study, we investigated whether isl1 is expressed in adult mesenchymal stem cells (MSCs), and after experimental myocardial infarction (MI). Additionally, we examined whether isl1 overexpression in MSCs could promote their vasculogenic properties, and whether intramyocardial gene transfer of naked DNA encoding isl1 could promote a functional recovery after MI. We used the transgenic mice isl1/cre/Z/EG , in order to detect isl1 expression in MSCs of adult mice, complemented by qRT-PCR and immunostaining for isl1 detection in rat- MSCs. Four weeks after MI induction, isl1 expression was assessed in bone marrow and spleen cells by qRT-PCR and immunostaining. Isl1 was retrovirally transduced to MSCs. Endothelial markers were examined by FACS and tube formation capacity was assessed on matrigel. Furthermore, intramyocardial injection of plasmid encoding isl1 to mice after ligation of the LAD was performed. We report for the first time, the identification of isl1+ progenitors in adult bone marrow. The number of isl1+ progenitors increased after in vitro cell culture, and also in the splenocytes after acute experimental myocardial infarction. Isl1 overexpression in MSCs promoted their differentiation towards endothelium. Finally, isl1 gene transfer to the peri-infarct region after MI induction resulted in partial salvage of ventricular function evident by echocardiography. Thus, the isl1 gene may stand as an attractive target for future cell based therapy for regenerative myocardial dysfunction.
Abstract 5513: Shear Stress Independent Formation of Collaterals in Porcine Coronaries
Authors: Jeroen P van den Wijngaard, Henny Schulten, Pepijn van Horssen, Rene D ter Wee, Maria Siebes, Mark J Post, Jos A Spaan
Journal: Circulation (2009)

Objectives: To test the hypothesis that intracardiac collateral connections do not result from observable (resolution 40 microns) preexisting vessels and shear stress modulation is not required for collateral formation. Methods: An LCX ameroid constrictor was placed in 6 pigs and the LCX exposed (sham) in 2. After six weeks the hearts were processed and sliced at 40 μ m thickness (3000 slices) in a fluorescence imaging cryomicrotome. High resolution images (2000 * 2000 pixels) were captured, yielding a 3D reconstruction (40 μ m3 voxels) of the vasculature. With custom algorithms, collaterals were detected between (Bridge) and within (Mid) LCX, LAD and RCA. Results: In shams, collaterals were absent. In stimulated hearts 192 collaterals, median ô111 μ m (22–1049), were detected, of which 65% endocardial (p= 0.004). Figure 1 shows a typical case (local opacity altered for visibility). Only 64% of all connections were bridging the LCX to LAD or RCA regions. Mid collaterals (n=24), were found in non-LCX regions with no apparent perturbation of shear stress. Mid collaterals (median 76 μ m, 23–1049) were smaller than bridge (137 μ m, 22–990, Mann-Whitney U p=0.047). Conclusion: Since mid collaterals are parallel to normal vasculature, the novel finding of these vessels contra-indicates a dominant role of shear stress in generating and maintaining collaterals. Higher flow rates are to be expected in Bridge versus Mid connections supporting the hypothesis that shear stress plays a role in outward remodeling. Hence, the detection of substantial numbers of collaterals in regions without flow limitation, suggests additional pathways contributing to collateral growth.
Abstract 5583: Association Between Newly Onset Oral Anticoagulant Treatment With Phenprocoumon and Serum Levels of the Vitamin-k Dependent Calcification Inhibitor Matrix Gla Protein
Authors: Ralf Koos, Vincent Brandenburg, Harald Peter Kühl, Leon Schurgers
Journal: Circulation (2009)

Introduction: Matrix Gla protein (MGP) is a vitamin K-dependent protein. Recently, the uncarboxylated (inactive, uc) form of MGP was associated with vascular tissue calcification. The aim of the present study was to investigate the relationship between a newly initiated oral anticoagulant treatment with the Vitamin-K antagonist phenprocoumon and serum levels of uncarboxylated MGP compared to a control group with aspirine therapy. Methods: We performed a prospective study in 43 non-dialyzed patients (mean age 61±10 years, 28 men) with newly initiated oral anticoagulant therapy with phenprocoumon (n=21) or aged-matched subjects with aspirine therapy (n=22). In all patients circulating ucMGP levels (in pM compared to synthetic ucMGP) were measured by a competitive ELISA-based assay (VitaK BV, Maastricht, The Netherlands) at baseline as well as after 1 month and 3 months follow-up. Results: At baseline, there were no significant differences in patient characteristics regarding cardiovascular risk factors and the prevalence of coronary artery disease between both groups. In addition, at baseline no significant difference in ucMGP levels between patients with newly initiated oral anticoagulant therapy (1003±526 pM) and control subjects with aspirine therapy (842±214 pM, p= 0.40) was detected. At 1 month follow-up, patients with oral anticoagulation therapy demonstrated significantly increased ucMGP levels (1904±402 pM) compared to the aspirine group (628±223 pM, p&lt;0.001). At 3 months-follow up, a further increase in ucMGP levels in patients with oral anticoagulant treatment (2316±344 pM) compared to the aspirine group (685±378, p&lt;0.001) as well as to study inclusion (1003±526 pM, p=0.003) was observed. Conclusion: The present study suggests that newly onset anticoagulation therapy with phenprocoumon is associated with increased serum levels of inactive, uncarboxylated MGP indicating calcification inhibitor deficiency compared to a reference population with aspirine therapy. Long-term follow-up data are needed to evaluate if newly onset oral anticoagulant therapy may be linked to an increased progression of coronary and valvular calcification.
Abstract 5516: Cardioprotective Effects of Hydrogen Peroxide and Erythropoietin During Acute Coronary Occlusion in Canine Coronary Native Collateral Microvessels in vivo
Authors: Toyotaka Yada, Hiroaki Shimokawa, Osamu Hiramatsu, Minoru Satoh, Naoki Kashihara, Yoshiro Shinozaki, Hidezo Mori, Aya Takaki, Masami Goto, Hiroshi Nakamoto, Yasuo Ogasawara, Fumihiko Kajiya
Journal: Circulation (2009)

Background: We have previously demonstrated that endothelium-derived hydrogen peroxide (H 2 O 2 ) is an endothelium-derived hyperpolarizing factor (EDHF) in canine coronary autoregulation in vivo. The aim of this study was to examine the role of H 2 O 2 during acute coronary occlusion with recombinant human erythropoietin (EPO) in the collateral vasodilatation of coronary microcirculation in vivo. Methods and Results: Canine subepicardial native collateral coronary small arteries (CSA, ≥100 μ m) and arterioles (CA, &lt;100 μ m) were observed by an intravital microscope under cyclooxygenase blockade. Experiments were performed after LAD occlusion (90 min) under the following 8 conditions (n=5, each); control, low-dose EPO (100 IU/kg, L-EPO), high-dose EPO (1000 IU/kg, H-EPO), H-EPO+catalase (a H 2 O 2 decomposer), H-EPO+NO synthase inhibitor (L-NMMA), H-EPO+L-NMMA+catalase, H-EPO+L-NMMA+tetraethylammonium (TEA, a K Ca channels blocker) and H-EPO+ wortmannin (PI3K inhibitor). Ischemia caused a significant collateral arteriolar vasodilatation of control, which was significantly inhibited by catalase. After H-EPO, the vasodilatation was significantly increased in both-sized arteries (CSA 6±1%, CA 22±4%) compared with control (CSA 1±1%, CA 16±3%) or L-EPO (CSA 2±1%, CA 18±3%) associated with increase in collateral blood flow (microsphere technique). H-EPO+catalase significantly decreased the vasodilatation in CA (12±2%). The vasodilatation was markedly attenuated after H-EPO+L-NMMA+catalase (CSA −4±1%, CA 6±1%), H-EPO+L-NMMA+TEA (CSA −5±1%, CA 6±1%) and H-EPO+ wortmannin (CSA −5±1%, CA 6±1%). In the control group, expression of Akt phosphorylation in the ischemic LAD area was significantly decreased compared with the H-EPO group. In H-EPO+L-NMMA+catalase or H-EPO+wortmannin group, expression of Akt phosphorylation was further decreased compared with H-EPO or L-NMMA+H-EPO group. Plasma 8-hydroxydeoxyguanosine from coronary sinus as a myocardial oxidative stress marker (HPLC-ECD method) significantly decreased in H-EPO compared with control. Conclusions: H 2 O 2 and EPO play an important collaborative role in native collateral vasodilatation in response to acute coronary occlusion in canine coronary microcirculation in vivo.
Abstract 5509: Association of Pericardial Adipose Tissue With Coronary Atherosclerotic Plaque and Inflammatory Blood Biomarkers
Authors: Christopher L Schlett, Fabian Bamberg, Wolfgang Koenig, Thomas J Brady, John T Nagurney, Udo Hoffmann
Journal: Circulation (2009)

Background: Pericardial adipose tissue (PAT) may exert paracrine effects on the pathogenesis of coronary artery disease (CAD). Thus, we assessed the relationship of PAT and different stages of coronary atherosclerosis, and inflammatory biomarkers. Methods: We included 355 subjects (mean age: 52±11 years, 60% male) without known CAD who had acute chest pain but no acute coronary syndrome. All subjects underwent contrast-enhanced 64-slice cardiac CT. PAT was defined by any pixel with a CT attenuation of −190 HU to −30 HU within the pericardial sac. Blood samples were collected at the time of CT scan, frozen, and subsequently analyzed for inflammatory biomarkers (E-Selectin, hs-CRP, IL-18, MCP-1, MMP-2, MMP-9, PAI-1, TNF- α ). We determined the association of PAT to the presence and extent of coronary atherosclerosis, and inflammatory biomarker levels. Results: In multivariable analyses, age, gender male, BMI, and current smoking were associated with higher levels of PAT (all p≤0.01). Moreover, the presence of any ( β : 18.3 cm 3 , 95%-CI: 7.0 –29.6, p=0.002), non-calcified ( β : 31.5 cm 3 , 95%-CI: 11.3–51.8, p=0.002), and mixed plaque ( β : 15.6 cm 3 , 95%-CI: 1.7–29.5, p=0.03) was associated with increased PAT independent of age, gender, and current smoking. This association persisted even after adjustment for BMI, except for non-calcified plaque. Hs-CRP, IL-18 and PAI-1 were significantly higher in subjects with increased PAT, independent of the presence of CAD (see Table ; following symbols within the table indicate a significant difference (p&lt;0.0253) for the subgroup comparison: * ‘low PAT/no plaque’ vs. ‘low PAT/yes plaque’; ° ‘high PAT/no plaque’ vs. ‘high PAT/yes plaque’; † ‘low PAT/no plaque’ vs. ‘high PAT/no plaque’; ‡ ‘low PAT/yes plaque’ vs. ‘high PAT/yes plaque’). Conclusion: PAT is associated with plaque by CT above and beyond the effect of traditional CVD risk factors. These data may support a central role of PAT in the development of coronary atherosclerosis. Level of blood biomarkers within the four subgroups as median plus lower and upper quartile
Abstract 5612: Evidence for Posttranscriptional Control of Individual Members of the miR-17–92 Cluster
Authors: Angelika Bonauer, Ariane Fischer, Andreas M Zeiher, Stefanie Dimmeler
Journal: Circulation (2009)

MicroRNAs (miRs) are small noncoding RNAs that regulate gene expression by binding to the cellular transcript leading to translational repression or degradation of the target mRNA. The miR-17–92 cluster encodes miR-17, miR-18a, miR-19a, miR-20a, miR-19b and miR-92a. Several members of the miR-17–92 cluster regulate angiogenesis and inhibition of miR-92a was recently shown to enhance recovery after ischemia in animal models. Because all members of the miR-17–92 cluster are under the control of the same promoter and are transcribed by one primary transcript, the mature miRs are expected to be regulated similarly. However, when we assessed the expression of the individual miRs in different cell types using a microRNA array, specific miRs were enriched. In HUVEC, miR-19b (51±3), miR-20a (13±1) and miR-92a (11±0.5) were highly expressed, whereas in CD34+ hematopoietic progenitor cells, miR-17 (27±3), miR-19b (88±13), miR-20a (94±19) were expressed at highest levels. MiR-18a expression was similar in both cell types (1.6±0.1 versus 1.8±0.1) indicating that specific members of the cluster are selectively processed or stabilized. To determine the regulation of the individual mature miRs in endothelial cells, HUVEC were incubated under hypoxia. Whereas the primary miR-17–92 cluster was not regulated, the mature forms of miR-19a (257+145%, 72h) and miR-92a (246±72%, 72h) were increased although with a different time course. Likewise, the individual members of the miR-17–92 cluster were significantly up-regulated after induction of hind limb ischemia (miR-92a: 4-fold; maximum at day 2) and miR-19a (6-fold; maximum at day 3), whereas miR-18a was not significantly regulated. In summary, these data provide first evidence for the posttranscriptional regulation of the miR-17–92 cluster, which controls neovascularization. Ongoing studies aim at identifying miR binding proteins in order to further characterize the mechanisms underlying the posttranscriptional regulation of the miR-17–92 cluster.