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1.  Consed: a graphical editor for next-generation sequencing 
Bioinformatics  2013;29(22):2936-2937.
Summary: The rapid growth of DNA sequencing throughput in recent years implies that graphical interfaces for viewing and correcting errors must now handle large numbers of reads, efficiently pinpoint regions of interest and automate as many tasks as possible. We have adapted consed to reflect this. To allow full-feature editing of large datasets while keeping memory requirements low, we developed a viewer, bamScape, that reads billion-read BAM files, identifies and displays problem areas for user review and launches the consed graphical editor on user-selected regions, allowing, in addition to longstanding consed capabilities such as assembly editing, a variety of new features including direct editing of the reference sequence, variant and error detection, display of annotation tracks and the ability to simultaneously process a group of reads. Many batch processing capabilities have been added.
Availability: The consed package is free to academic, government and non-profit users, and licensed to others for a fee by the University of Washington. The current version (26.0) is available for linux, macosx and solaris systems or as C++ source code. It includes a user’s manual (with exercises) and example datasets.
Supplementary information: Supplementary data are available at Bioinformatics online.
PMCID: PMC3810858  PMID: 23995391
2.  P2Y6 Receptor Potentiates Pro-Inflammatory Responses in Macrophages and Exhibits Differential Roles in Atherosclerotic Lesion Development 
PLoS ONE  2014;9(10):e111385.
P2Y6, a purinergic receptor for UDP, is enriched in atherosclerotic lesions and is implicated in pro-inflammatory responses of key vascular cell types and macrophages. Evidence for its involvement in atherogenesis, however, has been lacking. Here we use cell-based studies and three murine models of atherogenesis to evaluate the impact of P2Y6 deficiency on atherosclerosis.
Methodology/Principal Findings
Cell-based studies in 1321N1 astrocytoma cells, which lack functional P2Y6 receptors, showed that exogenous expression of P2Y6 induces a robust, receptor- and agonist-dependent secretion of inflammatory mediators IL-8, IL-6, MCP-1 and GRO1. P2Y6-mediated inflammatory responses were also observed, albeit to a lesser extent, in macrophages endogenously expressing P2Y6 and in acute peritonitis models of inflammation. To evaluate the role of P2Y6 in atherosclerotic lesion development, we used P2Y6-deficient mice in three mouse models of atherosclerosis. A 43% reduction in aortic arch plaque was observed in high fat-fed LDLR knockout mice lacking P2Y6 receptors in bone marrow-derived cells. In contrast, no effect on lesion development was observed in fat-fed whole body P2Y6xLDLR double knockout mice. Interestingly, in a model of enhanced vascular inflammation using angiotensin II, P2Y6 deficiency enhanced formation of aneurysms and exhibited a trend towards increased atherosclerosis in the aorta of LDLR knockout mice.
P2Y6 receptor augments pro-inflammatory responses in macrophages and exhibits a pro-atherogenic role in hematopoietic cells. However, the overall impact of whole body P2Y6 deficiency on atherosclerosis appears to be modest and could reflect additional roles of P2Y6 in vascular disease pathophysiologies, such as aneurysm formation.
PMCID: PMC4216081  PMID: 25360548
3.  Both Loved and Feared: Third Party Punishers Are Viewed as Formidable and Likeable, but These Reputational Benefits May Only Be Open to Dominant Individuals 
PLoS ONE  2014;9(10):e110045.
Third party punishment can be evolutionarily stable if there is heterogeneity in the cost of punishment or if punishers receive a reputational benefit from their actions. A dominant position might allow some individuals to punish at a lower cost than others and by doing so access these reputational benefits. Three vignette-based studies measured participants' judgements of a third party punisher in comparison to those exhibiting other aggressive/dominant behaviours (Study 1), when there was variation in the success of punishment (Study 2), and variation in the status of the punisher and the type of punishment used (Study 3). Third party punishers were judged to be more likeable than (but equally dominant as) those who engaged in other types of dominant behaviour (Study 1), were judged to be equally likeable and dominant whether their intervention succeeded or failed (Study 2), and participants believed that only a dominant punisher could intervene successfully (regardless of whether punishment was violent or non-violent) and that subordinate punishers would face a higher risk of retaliation (Study 3). The results suggest that dominance can dramatically reduce the cost of punishment, and that while individuals can gain a great deal of reputational benefit from engaging in third party punishment, these benefits are only open to dominant individuals. Taking the status of punishers into account may therefore help explain the evolution of third party punishment.
PMCID: PMC4210197  PMID: 25347781
4.  Oral neutrophils are an independent marker of the systemic inflammatory response after cardiac bypass 
Cardiopulmonary bypass (CPB) is an immuno-reactive state where neutrophils are activated and accumulate in different tissues. Edema and tissue necrosis are the most common sequelae observed, predominantly in the lungs, kidneys, and heart, heralding significant risk for postoperative complications. No method exists to noninvasively assess in vivo neutrophil activity. The objective of this study was to determine if neutrophil recruitment to the oral cavity would correlate with specific biomarkers after coronary bypass surgery (CPB).
We conducted a single site prospective observational study including non-consecutive adult patients undergoing elective, on-pump CPB. Blood and either oral cavity rinses or swabs were collected pre- and post-CPB. Absolute neutrophil counts from oral samples and serum biomarkers were measured. The association between neutrophil recruitment to the oral cavity, biomarkers and outcomes after CPB were analyzed.
CPB was associated with statistically significant increases in oral and blood neutrophil counts, as well as an increase in certain biomarkers over preoperative baseline. Peripheral blood neutrophil count were increased at all time points however statistically significant differences in median oral neutrophil counts were observed only at the time point immediately postoperative, and in what seems to be two unique patient populations (p < 0.001; group 1, median: 1.6×105, Interquartile range [IQR], 1.1×105 - 4.8×105, and group 2, median: 1.9×106, IQR, 8.7×105 - 4.0×106).
CPB is associated with a transient increase in oral neutrophils that may correlate with the systemic inflammatory response; oral neutrophils may have the ability to discriminate and identify unique patient populations based on their tissue migration.
PMCID: PMC4209230  PMID: 25349536
Biomarkers; Multiple organ failure; Coronary bypass; Critical illness; Neutrophil activation
5.  A Lipid E-MAP Identifies Ubx2 as a Critical Regulator of Lipid Saturation and Lipid Bilayer Stress 
Molecular cell  2013;51(4):519-530.
Biological membranes are complex, and the mechanisms underlying their homeostasis are incompletely understood. Here, we present a quantitative genetic interaction map (E-MAP) focused on various aspects of lipid biology, including lipid metabolism, sorting, and trafficking. This E-MAP contains ~250,000 negative and positive genetic interaction scores and identifies a molecular crosstalk of protein quality control pathways with lipid bilayer homeostasis. Ubx2p, a component of the endoplasmic-reticulum-associated degradation pathway, surfaces as a key upstream regulator of the essential fatty acid (FA) desaturase Ole1p. Loss of Ubx2p affects the transcriptional control of OLE1, resulting in impaired FA desaturation and a severe shift toward more saturated membrane lipids. Both the induction of the unfolded protein response and aberrant nuclear membrane morphologies observed in cells lacking UBX2 are suppressed by the supplementation of unsaturated FAs. Our results point toward the existence of dedicated bilayer stress responses for membrane homeostasis.
PMCID: PMC3791312  PMID: 23891562
6.  Publication of Trials Funded by the National Heart, Lung, and Blood Institute 
The New England journal of medicine  2013;369(20):1926-1934.
Rapid publication of clinical trials is essential in order for the findings to yield maximal benefits for public health and scientific progress. Factors affecting the speed of publication of the main results of government-funded trials have not been well characterized.
We analyzed 244 extramural randomized clinical trials of cardiovascular interventions that were supported by the National Heart, Lung, and Blood Institute (NHLBI). We selected trials for which data collection had been completed between January 1, 2000, and December 31, 2011. Our primary outcome measure was the time between completion of the trial and publication of the main results in a peer-reviewed journal.
As of March 31, 2012, the main results of 156 trials (64%) had been published (Kaplan–Meier median time to publication, 25 months, with 57% published within 30 months). Trials that focused on clinical events were published more rapidly than those that focused on surrogate measures (median, 9 months vs. 31 months; P<0.001). The only independent predictors of more rapid publication were a focus on clinical events rather than surrogate end points (adjusted publication rate ratio, 2.11; 95% confidence interval, 1.26 to 3.53; P = 0.004) and higher costs of conducting the trial, up to a threshold of approximately $5 million (P<0.001). The 37 trials that focused on clinical events and cost at least $5 million accounted for 67% of the funds spent on clinical trials but received 82% of the citations. After adjustment of the analysis for a focus on clinical events and for cost, trial results that were classified as positive were published more quickly than those classified as negative.
Results of less than two thirds of NHLBI-funded randomized clinical trials of cardiovascular interventions were published within 30 months after completion of the trial. Trials that focused on clinical events were published more quickly than those that focused on surrogate end points. (Funded by the National Heart, Lung, and Blood Institute.)
PMCID: PMC3928673  PMID: 24224625
7.  Phase II Clinical Research Design in Cardiology 
Circulation  2013;127(15):1630-1635.
PMCID: PMC3967793  PMID: 23588961
clinical trials; phase II; research design; stem cell; therapy
8.  Stereotactic Radiosurgery with Neoadjuvant Embolization of Larger Arteriovenous Malformations: An Institutional Experience 
BioMed Research International  2014;2014:306518.
Objective. This study investigates the safety and efficacy of a multimodality approach combining staged endovascular embolizations with subsequent SRS for the management of larger AVMs. Methods. Ninety-five patients with larger AVMs were treated with staged endovascular embolization followed by SRS between 1996 and 2011. Results. The median volume of AVM in this series was 28 cm3 and 47 patients (48%) were Spetzler-Martin grade IV or V. Twenty-seven patients initially presented with hemorrhage. Sixty-one patients underwent multiple embolizations while a single SRS session was performed in 64 patients. The median follow-up after SRS session was 32 months (range 9–136 months). Overall procedural complications occurred in 14 patients. There were 13 minor neurologic complications and 1 major complication (due to embolization) while four patients had posttreatment hemorrhage. Thirty-eight patients (40%) were cured radiographically. The postradiosurgery actuarial rate of obliteration was 45% at 5 years, 56% at 7 years, and 63% at 10 years. In multivariate analysis, larger AVM size, deep venous drainage, and the increasing number of embolization/SRS sessions were negative predictors of obliteration. The number of embolizations correlated positively with the number of stereotactic radiosurgeries (P < 0.005). Conclusions. Multimodality endovascular and radiosurgical approach is an efficacious treatment strategy for large AVM.
PMCID: PMC3919115  PMID: 24579080
9.  hMPV Lineage Nomenclature and Heparin Binding 
Viruses  2013;5(10):2546-2547.
PMCID: PMC3814602  PMID: 24136041
10.  Randomized Clinical Trial Of Immunogenicity And Safety Of A Recombinant H1N1/2009 Pandemic Influenza Vaccine Containing Advax™ Polysaccharide Adjuvant 
Vaccine  2012;30(36):5407-5416.
Timely vaccine supply is critical during influenza pandemics. A recombinant hemagglutinin (rHA)-based vaccine could overcome production hurdles of egg-based vaccines but has never previously been tested in a real-life pandemic setting. The primary aim was to determine the efficacy of a recombinant pandemic vaccine and whether its immunogenicity could be enhanced by a novel polysaccharide adjuvant (Advax™).
281 adults aged 18–70 years were recruited in a randomized, subject and observer blinded, parallel-group study of rHA H1N1/2009 vaccine with or without adjuvant. Immunizations were at 0 and 3 weeks with rHA 3, 11 or 45 ug. Serology and safety was followed for 6 months.
At baseline, only 9.1% of subjects (95% CI = 6.0 – 13.2) had seroprotective H1N1/2009 titers. Seroconversion rates varied by rHA dose, presence of adjuvant, subject age and number of immunizations. Eighty percent (95% CI = 52 – 96) of 18 – 49 year olds who received rHA 45ug with adjuvant were seroprotected at week 3, representing a 11.1-fold increase in antibody titers from baseline. Advax™ adjuvant increased seroprotection rates by 1.9 times after the first, and 2.5 times after the second, immunization when compared to rHA alone. Seroprotection was sustained at 26 weeks and the vaccine was well tolerated with no safety issues.
The study confirmed the ability to design, manufacture, and release a recombinant vaccine within a short time from the start of an actual influenza pandemic. Advax™ adjuvant significantly enhanced rHA immunogenicity.
PMCID: PMC3410954  PMID: 22717330
Vaccine; Influenza; Pandemic; Adjuvant; Recombinant; Antigen; Advax™
11.  Contribution of Fibrosis and the Autonomic Nervous System to Atrial Fibrillation Electrograms in Heart Failure 
Fibrotic and autonomic remodeling in heart failure (HF) increase vulnerability to atrial fibrillation (AF). Since AF electrograms (EGMs) are thought to reflect underlying structural substrate, we sought to: a) determine differences in AF-EGMs in normal versus HF atria and b) assess how fibrosis and nerve-rich fat contribute to AF-EGM characteristics in HF.
Methods and Results
AF was induced in 20 normal dogs by vagal stimulation and in 21 HF dogs (subjected to 3 weeks of rapid ventricular pacing at 240/min). AF-EGMs were analyzed for dominant frequency (DF), organizational index (OI), fractionation intervals (FI) and Shannon's entropy (ShEn). In 8 HF dogs, AF-EGM correlation with underlying fibrosis/fat/nerves was assessed. In HF, compared to normals, a) DF was lower and OI/FI/ShEn were greater. DF/FI were more heterogeneous in HF. %fat was greater, and fibrosis and fat more heterogeneously distributed in the posterior left atrium (PLA) than left atrial appendage (LAA). DF/OI correlated closely with %fibrosis. Heterogeneity of DF/FI correlated with heterogeneity of fibrosis. Autonomic blockade caused a greater change in DF/FI/ShEn in the PLA than LAA, with the decrease in ShEn correlating with %fat.
The amount and distribution of fibrosis in the HF atrium appears to contribute to slowing and increased organization of AF-EGMs, while the nerve-rich fat in the HF-PLA is positively correlated with AF-EGM entropy. By allowing for improved detection of regions of dense fibrosis and high autonomic nerve density in the HF atrium, these findings may help enhance the precision and success of substrate-guided ablation for AF.
PMCID: PMC3607299  PMID: 22722658
atrium; fibrillation; heart failure; nervous system; autonomic; fibrosis
12.  Virtual Electrophysiologic Study in a Three-dimensional Cardiac MRI Model of Porcine Myocardial Infarction 
This study sought to test the hypothesis that “virtual” electrophysiologic studies (EPS) on an anatomic platform generated by 3D MRI reconstruction of the left ventricle (LV) can reproduce the reentrant circuits of induced ventricular tachycardia (VT) in a porcine model of myocardial infarction (MI).
Delayed-enhancement MRI has been used to characterize MI and “gray zones”, which are thought to reflect heterogeneous regions of viable and non-viable myocytes.
MI by coronary artery occlusion was induced in eight pigs. After a recovery period, 3D cardiac MRIs were obtained from each pig in-vivo. Normal areas, gray zones, and infarct cores were classified based on voxel intensity. In the computer model, gray zones were assigned slower conduction and longer action potential durations than those for normal myocardium. Virtual EPS was performed and was compared to results of actual in vivo programmed stimulation and non-contact mapping.
The LV volumes ranged from 97.8 to 166.2 cm3 with 4.9 to 17.5% of voxels classified as infarct zones. Six of the seven pigs that developed VT during actual EPS were also inducible with virtual EPS. Four of the six pigs that had simulated VT had reentrant circuits that approximated the circuits seen with non-contact mapping, while the remaining two had similar circuits but propagating in opposite directions.
This initial study demonstrates the feasibility of applying a mathematical model to MRI reconstructions of the LV to predict VT circuits. Virtual EPS may be helpful to plan catheter ablation strategies or to identify patients who are at risk for future episodes of VT.
PMCID: PMC3407287  PMID: 22633654
Ventricular tachycardia; computer-based model; myocardial infarction; magnetic resonance imaging
13.  Effect of the Use and Timing of Bone Marrow Mononuclear Cell Delivery on Left Ventricular Function After Acute Myocardial Infarction: The TIME Randomized Trial 
While the delivery of cell therapy following ST segment myocardial infarction (STEMI) has been evaluated in previous clinical trials, the influence of the timing of cell delivery on the effect on left ventricular (LV) function has not been analyzed in a trial that randomly designated the time of delivery.
To determine 1) the effect of intracoronary autologous bone marrow mononuclear cell (BMC) delivery following STEMI on recovery of global and regional LV function and 2) if timing of BMC delivery (3 versus 7 days following reperfusion) influences this effect.
Design, Setting, and Patients
Between July 17, 2008 and November 15, 2011, 120 patients were enrolled in a randomized, 2×2 factorial, double-blind, placebo-controlled trial of the National Heart, Lung, and Blood Institute (NHLBI)-sponsored Cardiovascular Cell Therapy Research Network (CCTRN) of patients with LV dysfunction (LV Ejection Fraction (LVEF) ≤45%) following successful primary percutaneous coronary intervention (PCI) of anterior STEMI.
Intracoronary infusion of 150 × 106 BMCs or placebo (randomized 2:1 BMC:placebo) within 12 hours of aspiration and processing administered at Day 3 or Day 7 (randomized 1:1) post-PCI.
Main Outcome Measures
Co-primary endpoints were: 1) Change in global (LVEF) and regional (wall motion) LV function in infarct and border zones at 6 months measured by cardiac magnetic resonance imaging and 2) Change in LV function as affected by timing of treatment on Day 3 versus Day 7. Secondary endpoints included major adverse cardiovascular events as well as changes in LV volumes and infarct size.
Patient mean age was 56.9±10.9 years with 87.5% male. At 6 months, LVEF increased similarly in both BMC (45.2±10.6 to 48.3±13.3 %) and placebo groups (44.5±10.8 to 47.8±13.6 %). No detectable treatment effect on regional LV function was observed in either infarct or border zones. Differences between therapy groups in the change in global LV function over time when treated at Day 3 (−0.9±2.9%, 95% CI 6.6 to 4.9%, p=0.763) or Day 7 (1.1±2.9%, 95% CI −4.7 to 6.9, p=0.702) were not significant, nor were they different from each other. Also, timing of treatment had no detectable effect on recovery of regional LV function. Major adverse events were rare with no difference between groups.
Patients with STEMI, who underwent successful primary PCI and administration of intra-coronary BMCs at either 3 or 7 days following the event, had recovery of global and regional LV function similar to placebo
Trial Registration Number, NCT00684021
PMCID: PMC3652242  PMID: 23129008
14.  Primary salivary type lung tumor: Mucoepidermoid carcinoma 
Primary salivary type lung cancer are extremely rare intrathoracic malignancies. Mucoepidermoid tumor is one of the salivary gland tumor which originates from submucosal glands of tracheobronchial tree. These are very slow growing low grade malignant tumors. Surgery is the mainstay of treatment and rarely requires adjuvant therapy. In this case report, we describe a case of a young male who presented with cough and hemoptysis. On further investigation he was found to have mucoepidermoid tumor originating from the left bronchus.
PMCID: PMC3949550
Mucoepidermoid carcinoma; Luftsichel sign; Pneumonectomy
15.  Molecular Characterization of Commensal Escherichia coli Adapted to Different Compartments of the Porcine Gastrointestinal Tract 
Applied and Environmental Microbiology  2012;78(19):6799-6803.
The role of Escherichia coli as a pathogen has been the focus of considerable study, while much less is known about it as a commensal and how it adapts to and colonizes different environmental niches within the mammalian gut. In this study, we characterize Escherichia coli organisms (n = 146) isolated from different regions of the intestinal tracts of eight pigs (dueodenum, ileum, colon, and feces). The isolates were typed using the method of random amplified polymorphic DNA (RAPD) and screened for the presence of bacteriocin genes and plasmid replicon types. Molecular analysis of variance using the RAPD data showed that E. coli isolates are nonrandomly distributed among different gut regions, and that gut region accounted for 25% (P < 0.001) of the observed variation among strains. Bacteriocin screening revealed that a bacteriocin gene was detected in 45% of the isolates, with 43% carrying colicin genes and 3% carrying microcin genes. Of the bacteriocins observed (H47, E3, E1, E2, E7, Ia/Ib, and B/M), the frequency with which they were detected varied with respect to gut region for the colicins E2, E7, Ia/Ib, and B/M. The plasmid replicon typing gave rise to 25 profiles from the 13 Inc types detected. Inc F types were detected most frequently, followed by Inc HI1 and N types. Of the Inc types detected, 7 were nonrandomly distributed among isolates from the different regions of the gut. The results of this study indicate that not only may the different regions of the gastrointestinal tract harbor different strains of E. coli but also that strains from different regions have different characteristics.
PMCID: PMC3457480  PMID: 22798360
16.  Autonomic Remodeling in the Left Atrium and Pulmonary Veins in Heart Failure – Creation of a Dynamic Substrate for Atrial Fibrillation 
Atrial fibrillation (AF) is commonly associated with congestive heart failure (CHF). The autonomic nervous system is involved in the pathogenesis of both AF and CHF. We examined the role of autonomic remodeling in contributing to AF substrate in CHF.
Methods and Results
Electrophysiological mapping was performed in the pulmonary veins (PVs) and left atrium (LA) in 38 rapid-ventricular paced dogs (CHF group) and 39 controls under the following conditions: vagal stimulation, isoproterenol infusion, β-adrenergic blockade, acetylcholinesterase (AChE) inhibition (physostigmine), parasympathetic blockade, and double autonomic blockade. Explanted atria were examined for nerve density/distribution, muscarinic receptor (MR) and beta-adrenergic receptor (βAR) densities, and AChE activity.
In CHF dogs, there was an increase in nerve bundle size, parasympathetic fibers/bundle, and density of sympathetic fibrils and cardiac ganglia, all preferentially in the posterior LA/PVs. Sympathetic hyperinnervation was accompanied by increases in β1AR density and in sympathetic effect on ERPs and activation direction. β-adrenergic blockade slowed AF dominant frequency. Parasympathetic remodeling was more complex, resulting in increased AChE activity, unchanged MR density, unchanged parasympathetic effect on activation direction, and decreased effect of vagal stimulation on ERP (restored by AChE inhibition). Parasympathetic blockade markedly decreased AF duration.
In this heart failure model autonomic and electrophysiologic remodeling occurs involving the posterior left atrium and pulmonary veins. Despite synaptic compensation, parasympathetic hyperinnervation contributes significantly to AF maintenance. Parasympathetic and/or sympathetic signaling may be possible therapeutic targets for AF in CHF.
PMCID: PMC3607326  PMID: 21421805
atrial fibrillation; autonomic nervous system; heart failure
17.  Effect of Transendocardial Delivery of Autologous Bone Marrow Mononuclear Cells on Functional Capacity, Left Ventricular Function, and Perfusion in Chronic Ischemic Heart Failure: The FOCUS-CCTRN Trial 
Previous studies utilizing autologous bone marrow mononuclear cells (BMCs) in patients with ischemic cardiomyopathy have demonstrated safety and suggested efficacy. The FOCUS protocol was designed to assess efficacy of a larger cell dose in an adequately well-powered phase II study.
To determine if administration of BMCs through transendocardial injections improves myocardial perfusion, reduces left ventricular (LV) end systolic volume, or enhances maximal oxygen consumption in patients with coronary artery disease (CAD), LV dysfunction, and limiting heart failure and/or angina.
Design, Setting, and Patients
This is a 100 million cell, first-in-man randomized, double-blind, placebo-controlled trial was performed by the National Heart, Lung, and Blood Institute-sponsored Cardiovascular Cell Therapy Research Network (CCTRN) in symptomatic patients (NYHA II-III and/or CCS II-IV) receiving maximal medical therapy, with a left ventricular ejection fraction (LVEF)≤45%, perfusion defect by single-photon emission tomography (SPECT), and CAD not amenable to revascularization.
All patients underwent bone marrow aspiration, isolation of BMCs using a standardized automated system performed locally, and transendocardial injection of 100 million BMCs or placebo (2:1 BMC: placebo).
Main Outcome Measures
Three co-primary endpoints assessed at 6 months were changes in (a) LV end systolic volume (LVESV) by echocardiography, (b) maximal oxygen consumption (MVO2), and (c) reversibility on SPECT. Secondary measures included other SPECT measures, magnetic resonance imaging (MRI), echocardiography, clinical improvement, and major adverse cardiac events (MACE). Phenotypic and functional analyses of the cell product were performed by the CCTRN Biorepository lab.
Of 153 consented patients, a total of 92 (82 men; average age, 63 years) were randomized (n= 61 BMC, 31 placebo) at 5 sites between April 29, 2009 and April 18, 2011. Changes in LVESV index, (−0.9 ± 11.3 mL/m2; P = 0.733; 95% CI, −6.1 to 4.3), MVO2 (1.0 ± 2.9; P = 0.169; 95% CI, −0.42 to 2.34), percent reversible defect change, (−1.2 ± 23.3; P = 0.835; 95% CI, −12.50 to 10.12), and incidence of MACEwere not statistically significant. However, in an exploratory analysis the change in LVEF across the entire cohort by therapy group was significant (2.7 ± 5.2%; P = 0.030; 95% CI, 0.27 to 5.07).
This is the largest cell therapy trial of autologous BMCs in patients with ischemic LV dysfunction. In patients with chronic ischemic heart disease, transendocardial injection of BMCs compared to placebo did not improve LVESV, MVO2, or reversibility on SPECT.
PMCID: PMC3600947  PMID: 22447880
Chronic CAD; Ischemic Heart Failure; Chronic Angina; bone marrow mononuclear cells; cardiac performance
18.  Effect of Intracoronary Delivery of Autolologous Bone Marrow Mononuclear Cells Two to Three Weeks Following Acute Myocardial Infarction on Left-Ventricular Function: The LateTIME Randomized Trial 
Clinical trial results suggest that intracoronary delivery of autologous bone marrow mononuclear cells (BMCs) may improve left ventricular (LV) function when administered within the first week following myocardial infarction (MI). However, since a substantial number of patients may not present for early cell delivery, we investigated the efficacy of autologous BMC delivery 2–3 weeks post-MI.
To determine if intracoronary delivery of autologous BMCs improves global and regional LV function when delivered 2–3 weeks following first MI.
Design, Setting, and Patients
LateTIME is a randomized, double-blind, placebo-controlled trial of the National Heart, Lung, and Blood Institute - sponsored Cardiovascular Cell Therapy Research Network (CCTRN) of 87 patients with significant LV dysfunction (LVEF ≤ 45%) following successful primary percutaneous coronary intervention (PCI).
Intracoronary infusion of 150 × 106 autologous BMCs (total nucleated cells) or placebo (2:1 BMC:placebo) was performed within 12 hours of bone marrow aspiration after local automated cell processing.
Main Outcome Measures
The primary endpoints were changes in global (LVEF) and regional (wall motion) LV function in the infarct and border zone from baseline to 6 months as measured by cardiac MRI at a core lab blinded to treatment assignment Secondary endpoints included changes in LV volumes and infarct size.
87 patients were randomized between July 2008 and February 2011: mean age = 57 ± 11 yrs, 83% male. Harvesting, processing, and intracoronary delivery of BMCs in this setting was feasible and safe. The change from baseline to six months in the BMC group, when compared to the placebo group, for LVEF (48.7 to 49.2% vs. 45.3 to 48.8%; Difference = −3.0, 95% CI −7.0 to 0.9), wall motion in the infarct zone (6.2 to 6.5 vs. 4.9 to 5.9 mm; Difference = −0.7, 95% CI −2.8 to 1.3), and wall motion in the border zone (16.0 to 16.6 mm vs. 16.1 to 19.3 mm; Difference = −2.6; 95% CI −6.0 to 0.8) were not statistically significant. There was no significant change in LV volumes and infarct volumes decreased by a similar amount in both groups at 6 months compared to baseline.
Among patients with MI and LV dysfunction following reperfusion with PCI, intracoronary infusion of autologous BMCs compared to intracoronary placebo infusion, 2–3 weeks after PCI did not improve global or regional function at 6 months.
PMCID: PMC3600981  PMID: 22084195
Acute myocardial infarction; bone marrow mononuclear cells; LVEF; cardiac MRI
19.  Targeted nonviral gene-based inhibition of Gαi/o-mediated vagal signaling in the posterior left atrium decreases vagal-induced atrial fibrillation 
Pharmacologic and ablative therapies for atrial fibrillation (AF) have suboptimal efficacy. Newer gene-based approaches that target specific mechanisms underlying AF are likely to be more efficacious in treating AF. Parasympathetic signaling appears to be an important contributor to AF substrate.
The purpose of this study was to develop a nonviral gene-based strategy to selectively inhibit vagal signaling in the left atrium and thereby suppress vagal-induced AF.
In eight dogs, plasmid DNA vectors (minigenes) expressing Gαi C-terminal peptide (Gαictp) was injected in the posterior left atrium either alone or in combination with minigene expressing Gαoctp, followed by electroporation. In five control dogs, minigene expressing scrambled peptide (GαRctp) was injected. Vagal- and carbachol-induced left atrial effective refractory periods (ERPs), AF inducibility, and Gαi/octp expression were assessed 3 days following minigene delivery.
Vagal stimulation- and carbachol-induced effective refractory period shortening and AF inducibility were significantly attenuated in atria receiving a Gαi2ctp-expressing minigene and were nearly eliminated in atria receiving both Gαi2ctp- and Gαo1ctp-expressing minigenes.
Inhibition of both Gi and Go proteins is necessary to abrogate vagal-induced AF in the left atrium and can be achieved via constitutive expression of Gαi/octps expressed by nonviral plasmid vectors delivered to the posterior left atrium.
PMCID: PMC3570566  PMID: 21689540
Atrial fibrillation; Atrial fibrillation inducibility; Autonomic nervous system; Effective refractory period; Muscarinic cholinergic receptor; Pertussis toxin-sensitive G proteins; Vagal signaling
20.  11β-Hydroxysteroid Dehydrogenase Type 1 Gene Knockout Attenuates Atherosclerosis and In Vivo Foam Cell Formation in Hyperlipidemic apoE−/− Mice 
PLoS ONE  2013;8(2):e53192.
Chronic glucocorticoid excess has been linked to increased atherosclerosis and general cardiovascular risk in humans. The enzyme 11β-hydroxysteroid dehydrogenase type 1 (11βHSD1) increases active glucocorticoid levels within tissues by catalyzing the conversion of cortisone to cortisol. Pharmacological inhibition of 11βHSD1 has been shown to reduce atherosclerosis in murine models. However, the cellular and molecular details for this effect have not been elucidated.
Methodology/Principal Findings
To examine the role of 11βHSD1 in atherogenesis, 11βHSD1 knockout mice were created on the pro-atherogenic apoE−/− background. Following 14 weeks of Western diet, aortic cholesterol levels were reduced 50% in 11βHSD1−/−/apoE−/− mice vs. 11βHSD1+/+/apoE−/− mice without changes in plasma cholesterol. Aortic 7-ketocholesterol content was reduced 40% in 11βHSD1−/−/apoE−/− mice vs. control. In the aortic root, plaque size, necrotic core area and macrophage content were reduced ∼30% in 11βHSD1−/−/apoE−/− mice. Bone marrow transplantation from 11βHSD1−/−/apoE−/− mice into apoE−/− recipients reduced plaque area 39–46% in the thoracic aorta. In vivo foam cell formation was evaluated in thioglycollate-elicited peritoneal macrophages from 11βHSD1+/+/apoE−/− and 11βHSD1−/−/apoE−/− mice fed a Western diet for ∼5 weeks. Foam cell cholesterol levels were reduced 48% in 11βHSD1−/−/apoE−/− mice vs. control. Microarray profiling of peritoneal macrophages revealed differential expression of genes involved in inflammation, stress response and energy metabolism. Several toll-like receptors (TLRs) were downregulated in 11βHSD1−/−/apoE−/− mice including TLR 1, 3 and 4. Cytokine release from 11βHSD1−/−/apoE−/−-derived peritoneal foam cells was attenuated following challenge with oxidized LDL.
These findings suggest that 11βHSD1 inhibition may have the potential to limit plaque development at the vessel wall and regulate foam cell formation independent of changes in plasma lipids. The diminished cytokine response to oxidized LDL stimulation is consistent with the reduction in TLR expression and suggests involvement of 11βHSD1 in modulating binding of pro-atherogenic TLR ligands.
PMCID: PMC3562192  PMID: 23383297
21.  The VITamin D and OmegA-3 TriaL (VITAL): Rationale and Design of a Large Randomized Controlled Trial of Vitamin D and Marine Omega-3 Fatty Acid Supplements for the Primary Prevention of Cancer and Cardiovascular Disease 
Contemporary Clinical Trials  2011;33(1):159-171.
Data from laboratory studies, observational research, and/or secondary prevention trials suggest that vitamin D and marine omega-3 fatty acids may reduce risk for cancer or cardiovascular disease (CVD), but primary prevention trials with adequate dosing in general populations (i.e., unselected for disease risk) are lacking. The ongoing VITamin D and OmegA-3 TriaL (VITAL) is a large randomized, double-blind, placebo-controlled, 2×2 factorial trial of vitamin D (in the form of vitamin D3 [cholecalciferol], 2000 IU/day) and marine omega-3 fatty acid (Omacor® fish oil, eicosapentaenoic acid [EPA] + docosahexaenoic acid [DHA], 1 g/day) supplements in the primary prevention of cancer and CVD among a multi-ethnic population of 20,000 U.S. men aged ≥50 and women aged ≥55. The mean treatment period will be 5 years. Baseline blood samples will be collected in at least 16,000 participants, with follow-up blood collection in about 6000 participants. Yearly follow-up questionnaires will assess treatment compliance (plasma biomarker measures will also assess compliance in a random sample of participants), use of non-study drugs or supplements, occurence of endpoints, and cancer and vascular risk factors. Self-reported endpoints will be confirmed by medical record review by physicians blinded to treatment assignment, and deaths will be ascertained through national registries and other sources. Ancillary studies will investigate whether these agents affect risk for diabetes and glucose intolerance; hypertension; cognitive decline; depression; osteoporosis and fracture; physical disability and falls; asthma and other respiratory diseases; infections; rheumatoid arthritis, systemic lupus erythematosus, thyroid diseases, and other autoimmune disorders.
PMCID: PMC3253961  PMID: 21986389
Cancer; cardiovascular disease; cholecalciferol; primary prevention; omega-3 fatty acids; vitamin D; randomized controlled trial
22.  First report of Phlebotomine sand flies (Diptera: Psychodidae) in Kansas and Missouri, and a PCR method to distinguish Lutzomyia shannoni from Lutzomyia vexator 
Journal of medical entomology  2012;49(6):1460-1465.
Sand flies Lutzomyia (Psathyromyia) shannoni (Dyar) and Lu. (Helcocyrtomyia) vexator (Coquillet) were collected for the first time in southwest Missouri and southeast Kansas, expanding the known range of these species in North America. Altogether, 680 sand flies (356 males and 324 females) were collected during trapping from May through October 2011 and identified using morphological characters. Of the total sand flies collected 315 were identified as Lu. shannoni, with 181 individuals (or 26.6% of all sand flies) trapped in Missouri and 134 individuals (or 19.7%) trapped in Kansas. Whereas 358 Lu. vexator were identified from SW MO, only a single specimen was trapped in SE KS. One male Lu. vexator with asymmetric gonostyli was trapped in Missouri. We also developed a PCR protocol to consistently and accurately distinguish Lu. shannoni from Lu. vexator based on presence or absence of a 416bp fragment from the cytochrome oxidase I gene.
PMCID: PMC3533249  PMID: 23270176
Sand flies; Lutzomyia shannoni; Lutzomyia vexator
23.  Diversity in Glycosaminoglycan Binding Amongst hMPV G Protein Lineages 
Viruses  2012;4(12):3785-3803.
We have previously shown that hMPV G protein (B2 lineage) interacts with cellular glycosaminoglycans (GAGs). In this study we examined subtypes A1, A2 and B1 for this interaction. GAG-dependent infectivity of available hMPV strains was demonstrated using GAG-deficient cells and heparin competition. We expressed the G protein ectodomains from all strains and analysed these by heparin affinity chromatography. In contrast to the B2 lineage, neither the A2 or B1 G proteins bound to heparin. Sequence analysis of these strains indicated that although there was some homology with the B2 heparin-binding domains, there were less positively charged residues, providing a likely explanation for the lack of binding. Although sequence analysis did not demonstrate well defined positively charged domains in G protein of the A1 strain, this protein was able to bind heparin, albeit with a lower affinity than G protein of the B2 strain. These results indicate diversity in GAG interactions between G proteins of different lineages and suggest that the GAG-dependency of all strains may be mediated by interaction with an alternative surface protein, most probably the conserved fusion (F) protein. Analysis of both native and recombinant F protein confirmed that F protein binds heparin, supporting this conclusion.
PMCID: PMC3528290  PMID: 23242371
human metapneumovirus (hMPV); Glycosaminoglycan (GAG); G protein; F protein; infectivity
24.  Escherichia coli Lacking RpoS Are Rare in Natural Populations of Non-Pathogens 
G3: Genes|Genomes|Genetics  2012;2(11):1341-1344.
The alternative sigma factor RpoS controls a large regulon that allows E. coli to respond to a variety of stresses. Mutations in rpoS can increase rates of nutrient acquisition at the cost of a decrease in stress resistance. These kinds of mutations evolve rapidly under certain laboratory conditions where nutrient acquisition is especially challenging. The frequency of strains lacking RpoS in natural populations of E. coli is less clear. Such strains have been found at frequencies over 20% in some collections of wild isolates. However, laboratory handling can select for RpoS-null strains and may have affected some of these strain collections. Other studies have included an unknown diversity of strains or only used a phenotypic proxy as a measure of RpoS levels. We directly measured RpoS levels in a collection of E. coli that includes the full diversity of the species and that was handled in a manner to minimize the potential for laboratory evolution. We found that only 2% of strains produce no functional RpoS. Comparison of these strains in multiple labs shows that these rpoS mutations occurred in the laboratory. Earlier studies reporting much higher levels of RpoS polymorphism may reflect the storage history of the strains in laboratories rather than true frequency of such strains in natural populations.
PMCID: PMC3484664  PMID: 23173085
SPANC; population genetics; variation; transcription factor
25.  Estimating Global “Blue Carbon” Emissions from Conversion and Degradation of Vegetated Coastal Ecosystems 
PLoS ONE  2012;7(9):e43542.
Recent attention has focused on the high rates of annual carbon sequestration in vegetated coastal ecosystems—marshes, mangroves, and seagrasses—that may be lost with habitat destruction (‘conversion’). Relatively unappreciated, however, is that conversion of these coastal ecosystems also impacts very large pools of previously-sequestered carbon. Residing mostly in sediments, this ‘blue carbon’ can be released to the atmosphere when these ecosystems are converted or degraded. Here we provide the first global estimates of this impact and evaluate its economic implications. Combining the best available data on global area, land-use conversion rates, and near-surface carbon stocks in each of the three ecosystems, using an uncertainty-propagation approach, we estimate that 0.15–1.02 Pg (billion tons) of carbon dioxide are being released annually, several times higher than previous estimates that account only for lost sequestration. These emissions are equivalent to 3–19% of those from deforestation globally, and result in economic damages of $US 6–42 billion annually. The largest sources of uncertainty in these estimates stems from limited certitude in global area and rates of land-use conversion, but research is also needed on the fates of ecosystem carbon upon conversion. Currently, carbon emissions from the conversion of vegetated coastal ecosystems are not included in emissions accounting or carbon market protocols, but this analysis suggests they may be disproportionally important to both. Although the relevant science supporting these initial estimates will need to be refined in coming years, it is clear that policies encouraging the sustainable management of coastal ecosystems could significantly reduce carbon emissions from the land-use sector, in addition to sustaining the well-recognized ecosystem services of coastal habitats.
PMCID: PMC3433453  PMID: 22962585

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