Cerebrovascular endothelial cells form transient Notch-dependent cystic structures in zebrafish.

We identify a novel endothelial membrane behaviour in transgenic zebrafish. Cerebral blood vessels extrude large transient spherical structures that persist for an average of 23 min before regressing into the parent vessel. We term these structures "kugeln", after the German for sphere. Kugeln are only observed arising from the cerebral vessels and are present as late as 28 days post fertilization. Kugeln do not communicate with the vessel lumen and can form in the absence of blood flow. They contain little or no cytoplasm, but the majority are highly positive for nitric oxide reactivity. Kugeln do not interact with brain lymphatic endothelial cells (BLECs) and can form in their absence, nor do they perform a scavenging role or interact with macrophages. Inhibition of actin polymerization, Myosin II, or Notch signalling reduces kugel formation, while inhibition of VEGF or Wnt dysregulation (either inhibition or activation) increases kugel formation. Kugeln represent a novel Notch-dependent NO-containing endothelial organelle restricted to the cerebral vessels, of currently unknown function.

Adenosine versus intravenous calcium channel antagonists for supraventricular tachycardia.

BACKGROUND: People with supraventricular tachycardia (SVT) frequently are symptomatic and present to the emergency department for treatment. Although vagal manoeuvres may terminate SVT, they often fail, and subsequently adenosine or calcium channel antagonists (CCAs) are administered. Both are known to be effective, but both have a significant side effect profile. This is an update of a Cochrane review previously published in 2006. OBJECTIVES: To review all randomised controlled trials (RCTs) that compare effects of adenosine versus CCAs in terminating SVT. SEARCH METHODS: We identified studies by searching CENTRAL, MEDLINE, Embase, and two trial registers in July 2017. We checked bibliographies of identified studies and applied no language restrictions. SELECTION CRITERIA: We planned to include all RCTs that compare adenosine versus a CCA for patients of any age presenting with SVT. DATA COLLECTION AND ANALYSIS: We used standard methodological procedures as expected by Cochrane. Two review authors independently checked results of searches to identify relevant studies and resolved differences by discussion with a third review author. At least two review authors independently assessed each included study and extracted study data. We entered extracted data into Review Manager 5. Primary outcomes were rate of reversion to sinus rhythm and major adverse effects of adenosine and CCAs. Secondary outcomes were rate of recurrence, time to reversion, and minor adverse outcomes. We measured outcomes by calculating odds ratios (ORs) and assessed the quality of primary outcomes using the GRADE approach through the GRADEproGDT website. MAIN RESULTS: We identified two new studies for inclusion in the review update; the review now includes seven trials with 622 participants who presented to an emergency department with SVT. All included studies were RCTs, but only three described the randomisation process, and none had blinded participants, personnel, or outcome assessors to the intervention given. Moderate-quality evidence shows no differences in the number of people reverting to sinus rhythm who were treated with adenosine or CCA (89.7% vs 92.9%; OR 1.51, 95% confidence interval (CI) 0.85 to 2.68; participants = 622; studies = 7; I2 = 36%). Low-quality evidence suggests no appreciable differences in major adverse event rates between CCAs and adenosine. Researchers reported only one case of hypotension in the CCA group and none in the adenosine group (0.66% vs 0%; OR 3.09, 95% CI 0.12 to 76.71; participants = 306; studies = 3; I2 = 0%). Included trials did not report length of stay in hospital nor patient satisfaction. AUTHORS' CONCLUSIONS: Moderate-quality evidence shows no differences in effects of adenosine and calcium channel antagonists for treatment of SVT on reverting to sinus rhythm, and low-quality evidence suggests no appreciable differences in the incidence of hypotension. A study comparing patient experiences and prospectively studied adverse events would provide evidence on which treatment is preferable for management of SVT.

Stakeholder-led understanding of the implementation of digital technologies within heart disease diagnosis: a qualitative study protocol.

INTRODUCTION: Cardiovascular diseases are highly prevalent among the UK population, and the quality of care is being reduced due to accessibility and resource issues. Increased implementation of digital technologies into the cardiovascular care pathway has enormous potential to lighten the load on the National Health Service (NHS), however, it is not possible to adopt this shift without embedding the perspectives of service users and clinicians. METHODS AND ANALYSIS: A series of qualitative studies will be carried out with the aim of developing a stakeholder-led perspective on the implementation of digital technologies to improve holistic diagnosis of heart disease. This will be a decentralised study with all data collection being carried out online with a nationwide cohort. Four focus groups, each with 5-6 participants, will be carried out with people with lived experience of heart disease, and 10 one-to-one interviews will be carried out with clinicians with experience of diagnosing heart diseases. The data will be analysed using an inductive thematic analysis approach. ETHICS AND DISSEMINATION: This study received ethical approval from the Sciences and Technology Cross Research Council at the University of Sussex (reference ER/FM409/1). Participants will be required to provide informed consent via a Qualtrics survey before being accepted into the online interview or focus group. The findings will be disseminated through conference presentations, peer-reviewed publications and to the study participants.

JAG1-NOTCH4 mechanosensing drives atherosclerosis.

Endothelial cell (EC) sensing of disturbed blood flow triggers atherosclerosis, a disease of arteries that causes heart attack and stroke, through poorly defined mechanisms. The Notch pathway plays a central role in blood vessel growth and homeostasis, but its potential role in sensing of disturbed flow has not been previously studied. Here, we show using porcine and murine arteries and cultured human coronary artery EC that disturbed flow activates the JAG1-NOTCH4 signaling pathway. Light-sheet imaging revealed enrichment of JAG1 and NOTCH4 in EC of atherosclerotic plaques, and EC-specific genetic deletion of Jag1 (Jag1ECKO) demonstrated that Jag1 promotes atherosclerosis at sites of disturbed flow. Mechanistically, single-cell RNA sequencing in Jag1ECKO mice demonstrated that Jag1 suppresses subsets of ECs that proliferate and migrate. We conclude that JAG1-NOTCH4 sensing of disturbed flow enhances atherosclerosis susceptibility by regulating EC heterogeneity and that therapeutic targeting of this pathway may treat atherosclerosis.

Quantifying endothelial cell proliferation in the zebrafish embryo.

Introduction: Endothelial cell (EC) proliferation is a fundamental determinant of vascular development and homeostasis, and contributes to cardiovascular disease by increasing vascular permeability to blood-borne lipoproteins. Rodents have been traditionally used to analyse EC proliferation mechanisms in vascular health and disease; however, alternative models such as the zebrafish embryo allow researchers to conduct small scale screening studies in a physiologically relevant vasculature whilst reducing the use of mammals in biomedical research. In vitro models of EC proliferation are valuable but do not fully recapitulate the complexity of the in vivo situation. Several groups have used zebrafish embryos for vascular biology research because they offer the advantages of an in vivo model in terms of complexity but are also genetically manipulable and optically transparent. Methods: Here we investigated whether zebrafish embryos can provide a suitable model for the study of EC proliferation. We explored the use of antibody, DNA labelling, and time-lapse imaging approaches. Results: Antibody and DNA labelling approaches were of limited use in zebrafish due to the low rate of EC proliferation combined with the relatively narrow window of time in which they can label proliferating nuclei. By contrast, time-lapse imaging of fluorescent proteins localised to endothelial nuclei was a sensitive method to quantify EC proliferation in zebrafish embryos. Discussion: We conclude that time-lapse imaging is suitable for analysis of endothelial cell proliferation in zebrafish, and that this method is capable of capturing more instances of EC proliferation than immunostaining or cell labelling alternatives. This approach is relevant to anyone studying endothelial cell proliferation for screening genes or small molecules involved in EC proliferation. It offers greater biological relevance than existing in vitro models such as HUVECs culture, whilst reducing the overall number of animals used for this type of research.

Zebrafish xenograft models of cancer and metastasis for drug discovery.

INTRODUCTION: Patients with metastatic cancer suffer the highest rate of cancer-related death, but existing animal models of metastasis have disadvantages that limit our ability to understand this process. The zebrafish is increasingly used for cancer modelling, particularly xenografting of human cancer cell lines, and drug discovery, and may provide novel scientific and therapeutic insights. However, this model system remains underexploited. Areas covered: The authors discuss the advantages and disadvantages of the zebrafish xenograft model for the study of cancer, metastasis and drug discovery. They summarise previous work investigating the metastatic cascade, such as tumour-induced angiogenesis, intravasation, extravasation, dissemination and homing, invasion at secondary sites, assessing metastatic potential and evaluation of cancer stem cells in zebrafish. Expert opinion: The practical advantages of zebrafish for basic biological study and drug discovery are indisputable. However, their ability to sufficiently reproduce and predict the behaviour of human cancer and metastasis remains unproven. For this to be resolved, novel mechanisms must to be discovered in zebrafish that are subsequently validated in humans, and for therapeutic interventions that modulate cancer favourably in zebrafish to successfully translate to human clinical studies. In the meantime, more work is required to establish the most informative methods in zebrafish.