Flexible and waterproof micro-sensors to uncover zebrafish circadian rhythms: The next generation of cardiac monitoring for drug screening

Flexible electronics are the next generation of sensors for mobile health and implantation. Zebrafish (Danio rerio) is an emergent strategy for pre-clinical drug development and toxicity testing. To address the confounding effects from sedation of fish and removal from the aquatic habitat for micro-electrocardiogram (µECG) measurements, we developed waterproof and wearable sensors to uncover the circadian variation in heart rate (HR) and heart rate variability (HRV) ( Massin et al., 2000). The parylene-C based ECG sensor consisted of an ultra-soft silicone integrated jacket designed to wrap around the fish during swimming. The Young’s modulus of this silicone jacket matched with the fish surface, and an extended parylene cable connected the underwater chest electrodes with the out-of water electronics. In addition, embedded micro-glass spheres in the silicone effectively reduced the effective density of the jacket to ~1 g cm^(−3). These innovations enabled physiological ECG telemetry in the fish’s natural habitat without the need for sedation. Furthermore, a set of non-linear signal processing techniques filtered out the breathing and electromagnetic artifacts from the recorded signals. We observed a reduction in mean HR and an increase in HRV over 24 h at 10 dpa, accompanied by QT prolongation as well as diurnal variations, followed by normalization in mean HR and QT intervals at 26 days post ventricular amputation (dpa). We revealed Amiodarone-mediated QTc prolongation, HR reduction and HRV increase otherwise masked by sedation. The novel features of the flexible silicon jacket for µECG telemetry unraveled the biological clock and normalization of QT intervals at 26 dpa, providing the first evidence of new physiological phenomena during cardiac injury and repair as well as cardiac drug-mediated aberrant rhythms. Thus, the light weight and waterproof design holds promise to advance the next generation of mobile health and drug discovery

Moving Domain Computational Fluid Dynamics to Interface with an Embryonic Model of Cardiac Morphogenesis

Peristaltic contraction of the embryonic heart tube produces time- and spatial-varying wall shear stress (WSS) and pressure gradients (∇P) across the atrioventricular (AV) canal. Zebrafish (Danio rerio) are a genetically tractable system to investigate cardiac morphogenesis. The use of Tg(fli1a:EGFP)y1 transgenic embryos allowed for delineation and two-dimensional reconstruction of the endocardium. This time-varying wall motion was then prescribed in a two-dimensional moving domain computational fluid dynamics (CFD) model, providing new insights into spatial and temporal variations in WSS and ∇P during cardiac development. The CFD simulations were validated with particle image velocimetry (PIV) across the atrioventricular (AV) canal, revealing an increase in both velocities and heart rates, but a decrease in the duration of atrial systole from early to later stages. At 20-30 hours post fertilization (hpf), simulation results revealed bidirectional WSS across the AV canal in the heart tube in response to peristaltic motion of the wall. At 40-50 hpf, the tube structure undergoes cardiac looping, accompanied by a nearly 3-fold increase in WSS magnitude. At 110-120 hpf, distinct AV valve, atrium, ventricle, and bulbus arteriosus form, accompanied by incremental increases in both WSS magnitude and ∇P, but a decrease in bi-directional flow. Laminar flow develops across the AV canal at 20-30 hpf, and persists at 110-120 hpf. Reynolds numbers at the AV canal increase from 0.07±0.03 at 20-30 hpf to 0.23±0.07 at 110-120 hpf (p< 0.05, n=6), whereas Womersley numbers remain relatively unchanged from 0.11 to 0.13. Our moving domain simulations highlights hemodynamic changes in relation to cardiac morphogenesis; thereby, providing a 2-D quantitative approach to complement imaging analysis. © 2013 Lee et al

Flow-Responsive Vascular Endothelial Growth Factor Receptor-Protein Kinase C Isoform Epsilon Signaling Mediates Glycolytic Metabolites for Vascular Repair

Aims: Hemodynamic shear stress participates in maintaining vascular redox status. Elucidating flow-mediated endothelial metabolites enables us to discover metabolic biomarkers and therapeutic targets. We posited that flow-responsive vascular endothelial growth factor receptor (VEGFR)-protein kinase C isoform epsilon (PKCɛ)-6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 (PFKFB3) signaling modulates glycolytic metabolites for vascular repair. Results: Bidirectional oscillatory flow (oscillatory shear stress [OSS]: 0.1 ± 3 dyne·cm^(−2) at 1 Hz) upregulated VEGFR-dependent PKCɛ expression to a greater degree than did unidirectional pulsatile flow (pulsatile shear stress [PSS]: 23 ± 8 dyne·cm^(−2) at 1 Hz) in human aortic endothelial cells (p < 0.05, n = 3). PSS and OSS further upregulated PKCɛ-dependent PFKFB3 expression for glycolysis (p < 0.05, n = 4). Constitutively active PKCɛ increased, whereas dominant-negative PKCɛ reduced both basal and maximal extracellular acidification rates for glycolytic flux (p < 0.01, n = 4). Metabolomic analysis demonstrated an increase in PKCɛ-dependent glycolytic metabolite, dihydroxyacetone (DHA), but a decrease in gluconeogenic metabolite, aspartic acid (p < 0.05 vs. control, n = 6). In a New Zealand White rabbit model, both PKCɛ and PFKFB3 immunostaining was prominent in the PSS- and OSS-exposed aortic arch and descending aorta. In a transgenic Tg(flk-1:EGFP) zebrafish model, GATA-1a morpholino oligonucleotide injection (to reduce viscosity-dependent shear stress) impaired vascular regeneration after tail amputation (p < 0.01, n = 20), which was restored with PKCɛ messenger RNA (mRNA) rescue (p < 0.05, n = 5). As a corollary, siPKCɛ inhibited tube formation and vascular repair, which were restored by DHA treatment in our Matrigel and zebrafish models. Innovation and Conclusion: Flow-sensitive VEGFR-PKCɛ-PFKFB3 signaling increases the glycolytic metabolite, dihydroxyacetone, to promote vascular repair

Flow-Responsive Vascular Endothelial Growth Factor Receptor-Protein Kinase C Isoform Epsilon Signaling Mediates Glycolytic Metabolites for Vascular Repair

Aims: Hemodynamic shear stress participates in maintaining vascular redox status. Elucidating flow-mediated endothelial metabolites enables us to discover metabolic biomarkers and therapeutic targets. We posited that flow-responsive vascular endothelial growth factor receptor (VEGFR)-protein kinase C isoform epsilon (PKCɛ)-6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 (PFKFB3) signaling modulates glycolytic metabolites for vascular repair. Results: Bidirectional oscillatory flow (oscillatory shear stress [OSS]: 0.1 ± 3 dyne·cm^(−2) at 1 Hz) upregulated VEGFR-dependent PKCɛ expression to a greater degree than did unidirectional pulsatile flow (pulsatile shear stress [PSS]: 23 ± 8 dyne·cm^(−2) at 1 Hz) in human aortic endothelial cells (p < 0.05, n = 3). PSS and OSS further upregulated PKCɛ-dependent PFKFB3 expression for glycolysis (p < 0.05, n = 4). Constitutively active PKCɛ increased, whereas dominant-negative PKCɛ reduced both basal and maximal extracellular acidification rates for glycolytic flux (p < 0.01, n = 4). Metabolomic analysis demonstrated an increase in PKCɛ-dependent glycolytic metabolite, dihydroxyacetone (DHA), but a decrease in gluconeogenic metabolite, aspartic acid (p < 0.05 vs. control, n = 6). In a New Zealand White rabbit model, both PKCɛ and PFKFB3 immunostaining was prominent in the PSS- and OSS-exposed aortic arch and descending aorta. In a transgenic Tg(flk-1:EGFP) zebrafish model, GATA-1a morpholino oligonucleotide injection (to reduce viscosity-dependent shear stress) impaired vascular regeneration after tail amputation (p < 0.01, n = 20), which was restored with PKCɛ messenger RNA (mRNA) rescue (p < 0.05, n = 5). As a corollary, siPKCɛ inhibited tube formation and vascular repair, which were restored by DHA treatment in our Matrigel and zebrafish models. Innovation and Conclusion: Flow-sensitive VEGFR-PKCɛ-PFKFB3 signaling increases the glycolytic metabolite, dihydroxyacetone, to promote vascular repair

Polarity effect in electrovibration for tactile display

Abstract-Electrovibration is the tactile sensation of an alternating potential between the human body and a smooth conducing surface when the skin slides over the surface and where the current is too small to stimulate sensory nerves directly. It has been proposed as a high-density tactile display method, for example to display pictographic information to persons who are blind. Previous models for the electrovibration transduction mechanism are based on a parallel-plate capacitor in which the electrostatic force is insensitive to polarity. We present experimental data showing that electrovibratory perceptual sensitivity to positive pulses is less than that for negative or biphasic pulses and propose that this disparity may be due to the asymmetric electrical properties of human skin. We furthermore propose using negative pulses for insulated tactile displays based on electrovibration because their sensory thresholds were found to be more stable than for waveforms incorporating positive pulses

A framework for the development of a global standardised marine taxon reference image database (SMarTaR-ID) to support image-based analyses

Video and image data are regularly used in the field of benthic ecology to document biodiversity. However, their use is subject to a number of challenges, principally the identification of taxa within the images without associated physical specimens. The challenge of applying traditional taxonomic keys to the identification of fauna from images has led to the development of personal, group, or institution level reference image catalogues of operational taxonomic units (OTUs) or morphospecies. Lack of standardisation among these reference catalogues has led to problems with observer bias and the inability to combine datasets across studies. In addition, lack of a common reference standard is stifling efforts in the application of artificial intelligence to taxon identification. Using the North Atlantic deep sea as a case study, we propose a database structure to facilitate standardisation of morphospecies image catalogues between research groups and support future use in multiple front-end applications. We also propose a framework for coordination of international efforts to develop reference guides for the identification of marine species from images. The proposed structure maps to the Darwin Core standard to allow integration with existing databases. We suggest a management framework where high-level taxonomic groups are curated by a regional team, consisting of both end users and taxonomic experts. We identify a mechanism by which overall quality of data within a common reference guide could be raised over the next decade. Finally, we discuss the role of a common reference standard in advancing marine ecology and supporting sustainable use of this ecosystem

Presbyopia:Effectiveness of correction strategies

Presbyopia is a global problem affecting over a billion people worldwide. The prevalence of unmanaged presbyopia is as high as 50% of those over 50 years of age in developing world populations due to a lack of awareness and accessibility to affordable treatment, and is even as high as 34% in developed countries. Definitions of presbyopia are inconsistent and varied, so we propose a redefinition that states “presbyopia occurs when the physiologically normal age-related reduction in the eye's focusing range reaches a point, when optimally corrected for distance vision, that the clarity of vision at near is insufficient to satisfy an individual's requirements”. Presbyopia is inevitable if one lives long enough, but intrinsic and extrinsic risk factors including cigarette smoking, pregnancy history, hyperopic or astigmatic refractive error, ultraviolet radiation, female sex (although accommodation is similar to males), hotter climates and some medical conditions such as diabetes can accelerate the onset of presbyopic symptoms. Whilst clinicians can ameliorate the symptoms of presbyopia with near vision spectacle correction, bifocal and progressive spectacle lenses, monovision, translating or multifocal contact lenses, monovision, extended depth of focus, multifocal (refractive, diffractive and asymmetric designs) or ‘accommodating’ intraocular lenses, corneal inlays, scleral expansion, laser refractive surgery (corneal monovision, corneal shrinkage, corneal multifocal profiles and lenticular softening), pharmacologic agents, and electro-stimulation of the ciliary muscle, none fully overcome presbyopia in all patients. While the restoration of natural accommodation or an equivalent remains elusive, guidance is gives on presbyopic correction evaluation techniques

Effects of Anacetrapib in Patients with Atherosclerotic Vascular Disease

BACKGROUND: Patients with atherosclerotic vascular disease remain at high risk for cardiovascular events despite effective statin-based treatment of low-density lipoprotein (LDL) cholesterol levels. The inhibition of cholesteryl ester transfer protein (CETP) by anacetrapib reduces LDL cholesterol levels and increases high-density lipoprotein (HDL) cholesterol levels. However, trials of other CETP inhibitors have shown neutral or adverse effects on cardiovascular outcomes. METHODS: We conducted a randomized, double-blind, placebo-controlled trial involving 30,449 adults with atherosclerotic vascular disease who were receiving intensive atorvastatin therapy and who had a mean LDL cholesterol level of 61 mg per deciliter (1.58 mmol per liter), a mean non-HDL cholesterol level of 92 mg per deciliter (2.38 mmol per liter), and a mean HDL cholesterol level of 40 mg per deciliter (1.03 mmol per liter). The patients were assigned to receive either 100 mg of anacetrapib once daily (15,225 patients) or matching placebo (15,224 patients). The primary outcome was the first major coronary event, a composite of coronary death, myocardial infarction, or coronary revascularization. RESULTS: During the median follow-up period of 4.1 years, the primary outcome occurred in significantly fewer patients in the anacetrapib group than in the placebo group (1640 of 15,225 patients [10.8%] vs. 1803 of 15,224 patients [11.8%]; rate ratio, 0.91; 95% confidence interval, 0.85 to 0.97; P=0.004). The relative difference in risk was similar across multiple prespecified subgroups. At the trial midpoint, the mean level of HDL cholesterol was higher by 43 mg per deciliter (1.12 mmol per liter) in the anacetrapib group than in the placebo group (a relative difference of 104%), and the mean level of non-HDL cholesterol was lower by 17 mg per deciliter (0.44 mmol per liter), a relative difference of -18%. There were no significant between-group differences in the risk of death, cancer, or other serious adverse events. CONCLUSIONS: Among patients with atherosclerotic vascular disease who were receiving intensive statin therapy, the use of anacetrapib resulted in a lower incidence of major coronary events than the use of placebo. (Funded by Merck and others; Current Controlled Trials number, ISRCTN48678192 ; ClinicalTrials.gov number, NCT01252953 ; and EudraCT number, 2010-023467-18 .)

Dynamics of Endothelial Metabolomics in Vascular Development and Repair

Hemodynamic shear stress is intimately linked with endothelial metabolic effects, regulating key mechanisms in endothelial function, homeostasis, and repair. In-vivo modulation of shear stress signaling pathways in the zebrafish model allows for identification of mechanisms with potential therapeutic implications. The following studies combine in-vivo zebrafish models with in-vitro shear stress studies to characterize the mechanisms whereby shear stress regulates vascular development and repair. We investigated shear stress-modulated genes via microarray, and identified Angiopoietin-2 (Ang-2), a well known regulator of angiogenesis and vascular development, as a potential target for further mechanistic study. Oscillatory shear stress (OSS) induced Ang-2 mRNA expression in a Wnt-signaling dependent manner. Inhibition of Wnt signaling or Ang-2 expression supressed endothelial cell migration and tube formation, which were rescued by human recombinant Ang-2 treatment. These results were recapitulated in the embryonic zebrafish model using heat-shock inducible transgenic Tg(hsp70l:dkk1-GFP) and Tg(kdrl:GFP) embryos injected with Ang-2 morpholino. Inhibition of Wnt signaling with IWR-1 also impaired vascular repair after tail amputation, which was rescued by injection of zAng-2 mRNA. Taken together, this data demonstrated shear stress activated Ang-2 via canonical Wnt signaling in vascular endothelial cells, and recapitulated shear stress-Wnt-Ang-2 signaling vascular repair in the zebrafish model. Shear stress-modulated gene- and protein-based mechanisms result in significant changes at the metabolomic level. In the second part of my thesis, we examined the role of emerging mechano-sensitive metabolic pathways in vascular repair. Metabolomic analysis revealed both pulsatile shear stress (PSS) and OSS significantly increased endothelial glycolytic metabolites, but decreased gluconeogenic metabolites. Additionally, both OSS and PSS up-regulated the expression of PKCɛ. We therefore tested whether shear stress modulates endothelial metabolomics to promote vascular repair via PKCɛ-mediated glycolytic metabolites. Treatment with pro-glyocolytic metabolites in-vitro rescued tube formation following treatment with siPKCɛ. To recapitulate vascular repair in transgenic Tg(flk1:GFP) zebrafish embryos, we decreased viscosity and fluid shear stress by micro-injection of GATA-1a morpholino oligonucleotide (MO). In the zebrafish tail amputation model, GATA-1a MO impaired and delayed vascular repair. Co-injection of PKCɛ mRNA with GATA-1a MO rescued this phenomenon. Injection of epo mRNA to increase viscosity resulted in enhanced tail repair. Overall, our studies revealed that shear responsive VEGFR- PKCɛ-metabolomic signaling modulates glycolytic metabolites to influence vascular repair.Mitochondria are the metabolic center of the cell, and mitochondrial state is intimately linked to endothelial function. To further investigate the effect of shear stress on endothelial metabolic function, we examined the effect of physiological pulsatile shear stress (PSS) on mitochondrial membrane potential (ΔΨm) and the role of Mn-SOD expression on ΔΨm. PSS induced a dynamic increase in ΔΨm, while silencing Mn-SOD attenuated PSS-mediated ΔΨm increase. Mn-SOD mimetic MnTMPyP increased ΔΨm to the similar extent as induced by PSS. Our findings suggest that PSS modulates mitochondrial function by increasing mitochondrial ΔΨm, in part, via Mn-SOD up-regulation.In addition to using the embryonic zebrafish model to investigate vascular repair, we worked to develop technologies enabling the use of the adult zebrafish as a model for genetic and chemically-induced cardiomyopathies. To address the confounding effects from sedation of fish and removal from the aquatic habitat for micro-electrocardiogram (?ECG) measurements, we developed waterproof and wearable flexible electronic sensors to uncover the circadian variation in heart rate (HR) and heart rate variability (HRV). Innovations including an ultra-soft silicone integrated jacket, matching Young’s modulus to the fish surface, and embedded micro-glass spheres to reduce the effective density enabled physiological ECG telemetry in the fish’s natural habitat without the need for sedation. The novel features of the flexible silicon jacket for ?ECG telemetry unraveled the biological clock and normalization of QT intervals at following ventricular resection at 26 days post ventricular amputation, providing the first evidence of new physiological phenomena during cardiac injury and repair. Additionally, we revealed Amiodarone-mediated QTc prolongation, HR reduction and HRV increase otherwise masked by sedation. This light weight and waterproof design holds promise to advance the next generation of mobile health and drug discovery