High Resolution Imaging of Vascular Function in Zebrafish

Rationale: The role of the endothelium in the pathogenesis of cardiovascular disease is an emerging field of study, necessitating the development of appropriate model systems and methodologies to investigate the multifaceted nature of endothelial dysfunction including disturbed barrier function and impaired vascular reactivity. Objective: We aimed to develop and test an optimized high-speed imaging platform to obtain quantitative real-time measures of blood flow, vessel diameter and endothelial barrier function in order to assess vascular function in live vertebrate models. Methods and Results: We used a combination of cutting-edge optical imaging techniques, including high-speed, camera-based imaging (up to 1000 frames/second), and 3D confocal methods to collect real time metrics of vascular performance and assess the dynamic response to the thromboxane A2 (TXA2) analogue, U-46619 (1 μM), in transgenic zebrafish larvae. Data obtained in 3 and 5 day post-fertilization larvae show that these methods are capable of imaging blood flow in a large (1 mm) segment of the vessel of interest over many cardiac cycles, with sufficient speed and sensitivity such that the trajectories of individual erythrocytes can be resolved in real time. Further, we are able to map changes in the three dimensional sizes of vessels and assess barrier function by visualizing the continuity of the endothelial layer combined with measurements of extravasation of fluorescent microspheres. Conclusions: We propose that this system-based microscopic approach can be used to combine measures of physiologic function with molecular behavior in zebrafish models of human vascular disease. © 2012 Watkins et al

Effects of Mindfulness-Based Interventions on Fatigue in Cancer Survivors: A Systematic Review and Meta-Analysis of Randomized Controlled Trials

This systematic review and meta-analysis was designed to determine the efficacy of mindfulness-based interventions (MBIs) in improving fatigue-related outcomes in adult cancer survivors. Randomized controlled trials (RCTs) were identified from PubMed, MEDLINE, PsycINFO, CINAHL, Web of Science, and EMBASE databases and reference lists of included studies. Separate random-effects meta-analyses were conducted for fatigue and vitality/vigor. Twenty-three studies reporting on 21 RCTs (N=2,239) met inclusion criteria. MBIs significantly reduced fatigue compared to controls at post-intervention (g=0.60, 95% CI [0.36, 0.83]) and first follow-up (g=0.42, 95% CI [0.20, 0.64]). Likewise, MBIs significantly improved vitality/vigor at post-intervention (g=0.39, 95% CI [0.25, 0.52]) and first follow-up (g=0.35, 95% CI [0.03, 0.67]). The evidence grade was low due to risk of bias, substantial heterogeneity, and publication bias among studies. MBIs show promise in improving fatigue and vitality/vigor in cancer survivors. More rigorous trials are needed to address current gaps in the evidence base

Next-generation sequencing (NGS) for assessment of microbial water quality: current progress, challenges, and future opportunities

Water quality is an emergent property of a complex system comprised of interacting microbial populations and introduced microbial and chemical contaminants. Studies leveraging next-generation sequencing (NGS) technologies are providing new insights into the ecology of microbially mediated processes that influence fresh water quality such as algal blooms, contaminant biodegradation, and pathogen dissemination. In addition, sequencing methods targeting small subunit (SSU) rRNA hypervariable regions have allowed identification of signature microbial species that serve as bioindicators for sewage contamination in these environments. Beyond amplicon sequencing, metagenomic and metatranscriptomic analyses of microbial communities in fresh water environments reveal the genetic capabilities and interplay of waterborne microorganisms, shedding light on the mechanisms for production and biodegradation of toxins and other contaminants. This review discusses the challenges and benefits of applying NGS-based methods to water quality research and assessment. We will consider the suitability and biases inherent in the application of NGS as a screening tool for assessment of biological risks and discuss the potential and limitations for direct quantitative interpretation of NGS data. Secondly, we will examine case studies from recent literature where NGS based methods have been applied to topics in water quality assessment, including development of bioindicators for sewage pollution and microbial source tracking, characterizing the distribution of toxin and antibiotic resistance genes in water samples, and investigating mechanisms of biodegradation of harmful pollutants that threaten water quality. Finally, we provide a short review of emerging NGS platforms and their potential applications to the next generation of water quality assessment tools.Singapore-MIT Alliance for Research and Technology. Center for Environmental Sensing and Modelin

The thromboxane analogue, U-46619 decreased vessel diameter.

<p>3D imaging of the dorsal aorta in a 5 dpf <i>Tg(kdrl:GFP)^la116</i> zebrafish larva with dorsal downwards and anterior right (<b>A-D</b>). Changes in aortic flow velocity (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0044018#pone-0044018-g005" target="_blank">Figure 5</a>) in response to U-46619 in the 5 dpf embryos, were accompanied by decreases in vessel diameter (<b>E</b>, n = 5, <i>P</i><0.05). χ denotes statistical differences from baseline (time 0).</p

Time-lapse imaging of endothelial barrier dysfunction in response to the serine protease, thrombin.

<p>Extended depth of focus projection of the trunk vessels from a 5 dpf <i>Tg(kdrl:GFP)^la116</i> zebrafish larva (<b>A</b>). The EGFP-labeled endothelium appears green while the 0.1 µm FluoSpheres are pseudocolored red. Data were collected at baseline and at 10 minute intervals following exposure to thrombin (0.01 units/µL) for a total collection period of 3 hours. Localized increases in permeability, as evidenced by accumulation of microspheres outside the vessel wall (white arrows) were apparent by 1 h (panel <b>2</b>) and increased over time (panels 3–6). The zoomed images (<b>B</b>) from boxed region of panel A1 show a clear break in the endothelial layer at 1 hour (panel 4). A total of 8 larvae were examined with equivalent results. These data were confirmed in a subset of larvae (n = 3) using high magnification (60X, 1.4 NA) point scanning confocal imaging and showed extensive accumulation of Fluospheres outside the vessel wall (<b>C</b>). In contrast, 5 dpf zebrafish larvae (n = 8) exposed to the thromboxane mimetic, U-46619 showed no evidence for disrupted barrier function (<b>D</b>).</p

Quantitation of aortic blood flow velocity in zebrafish embryos using intravital imaging.

<p>Image processing was performed using Imaris software (Bitplane, Saint Paul, MN). Red blood cells shown in a representative single time point (<b>A</b>) were segmented by size, shape, and fluorescence intensity (<b>B</b>) and tracked using an auto-regressive tracking algorithm (<b>C</b>). The frequency of image collection was set such that erythrocyte displacement is less than one cell diameter/frame and therefore individual cells can be tracked with confidence. Only cells that could be tracked for >250 µm were included in the analysis. Quantitation revealed pulsatility of blood flow in both the 3 (<b>D</b>) and 5 (<b>E</b>) dpf embryos, and confirmed our ability to measure flow velocities greater than 2 mm/s in response to the thromboxane analogue U46619 (example shown in 5 dpf embryos, panel <b>E,</b> open circles).</p

Time sequence of red blood cell flow through a segment of dorsal aorta.

<p>This montage shows individual images from a time sequence (collected every 2 ms, with a 2 ms exposure). The images shown are semi-sequential (4 ms intervals). Erythrocytes are readily visible, and the tracking of the cells from right to left can be readily seen. (Scale Bar  = 100 microns).</p