Comprehensive profiling of zebrafish hepatic proximal promoter CpG island methylation and its modification during chemical carcinogenesis

Background\ud DNA methylation is an epigenetic mechanism associated with regulation of gene expression and it is modulated during chemical carcinogenesis. The zebrafish is increasingly employed as a human disease model; however there is a lack of information on DNA methylation in zebrafish and during fish tumorigenesis. \ud \ud Results\ud A novel CpG island tiling array containing 44,000 probes, in combination with immunoprecipitation of methylated DNA, was used to achieve the first comprehensive methylation profiling of normal adult zebrafish liver. DNA methylation alterations were detected in zebrafish liver tumors induced by the environmental carcinogen 7, 12-dimethylbenz(a)anthracene. Genes significantly hypomethylated in tumors were associated particularly with proliferation, glycolysis, transcription, cell cycle, apoptosis, growth and metastasis. Hypermethylated genes included those associated with anti-angiogenesis and cellular adhesion. Of 49 genes that were altered in expression within tumors, and which also had appropriate CpG islands and were co-represented on the tiling array, approximately 45% showed significant changes in both gene expression and methylation. \ud \ud Conclusion\ud The functional pathways containing differentially methylated genes in zebrafish hepatocellular carcinoma have also been reported to be aberrantly methylated during tumorigenesis in humans. These findings increase the confidence in the use of zebrafish as a model for human cancer in addition to providing the first comprehensive mapping of DNA methylation in the normal adult zebrafish liver. \ud \u

GFP Transgenic Medaka (Oryzias latipes) under the Inducible cyp1a Promoter Provide a Sensitive and Convenient Biological Indicator for the Presence of TCDD and Other Persistent Organic Chemicals

10.1371/journal.pone.0064334PLoS ONE85

Localized rbp4 expression in the yolk syncytial layer plays a role in yolk cell extension and early liver development

<p>Abstract</p> <p>Background</p> <p>The number of genes characterized in liver development is steadily increasing, but the origin of liver precursor cells and the molecular control of liver formation remain poorly understood. Existing theories about formation of zebrafish visceral organs emphasize either their budding from the endodermal rod or formation of independent anlage followed by their later fusion, but none of these is completely satisfactory in explaining liver organogenesis in zebrafish.</p> <p>Results</p> <p>Expression of a gene encoding the retinol binding protein 4 (Rbp4) was analyzed in zebrafish. <it>rbp4</it>, which is expressed mainly in the liver in adults, was shown to be expressed in the yolk syncytial layer (YSL) during early embryogenesis. At 12–16 hpf <it>rbp4 </it>expression was restricted to the ventro-lateral YSL and later expanded to cover the posterior YSL. We demonstrated that <it>rbp4 </it>expression was negatively regulated by Nodal and Hedgehog (Hh) signalling and positively controlled by retinoic acid (RA). Knockdown of Rbp4 in the YSL resulted in shortened yolk extension as well as the formation of two liver buds, which could be due to impaired migration of liver progenitor cells. <it>rbp4 </it>appears also to regulate the extracellular matrix protein Fibronectin1 (Fn1) specifically in the ventro-lateral yolk, indicating a role of Fn1 in liver progenitor migration. Since exocrine pancreas, endocrine pancreas, intestine and heart developed normally in Rbp4 morphants, we suggest that <it>rbp4 </it>expression in the YSL is required only for liver development.</p> <p>Conclusion</p> <p>The characteristic expression pattern of <it>rbp4 </it>suggests that the YSL is patterned despite its syncytial nature. YSL-expressed Rbp4 plays a role in formation of both yolk extension and liver bud, the latter may also require migration of liver progenitor cells.</p

STORM: A General Model to Determine the Number and Adaptive Changes of Epithelial Stem Cells in Teleost, Murine and Human Intestinal Tracts

Intestinal stem cells play a pivotal role in the epithelial tissue renewal, homeostasis and cancer development. The lack of a general marker for intestinal stem cells across species has hampered analysis of stem cell number in different species and their adaptive changes upon intestinal lesions or during development of cancer. Here a two-dimensional model, named STORM, has been developed to address this issue. By optimizing epithelium renewal dynamics, the model examines the epithelial stem cell number by taking experimental input information regarding epithelium proliferation and differentiation. As the results suggest, there are 2.0–4.1 epithelial stem cells on each pocket section of zebrafish intestine, 2.0–4.1 stem cells on each crypt section of murine small intestine and 1.8–3.5 stem cells on each crypt section of human duodenum. The model is able to provide quick results for stem cell number and its adaptive changes, which is not easy to measure through experiments. Its general applicability to different species makes it a valuable tool for analysis of intestinal stem cells under various pathological conditions.MIT-Singapore AllianceNational University of Singapore. Dept. of Biological Science

K-ras/PI3K-Akt Signaling Is Essential for Zebrafish Hematopoiesis and Angiogenesis

The RAS small GTPases orchestrate multiple cellular processes. Studies on knock-out mice showed the essential and sufficient role of K-RAS, but not N-RAS and H-RAS in embryonic development. However, many physiological functions of K-RAS in vivo remain unclear. Using wild-type and fli1:GFP transgenic zebrafish, we showed that K-ras-knockdown resulted in specific hematopoietic and angiogenic defects, including the impaired expression of erythroid-specific gene gata1 and ße3-hemoglobin, reduced blood circulation and disorganized blood vessels. Expression of either K-rasC40 that links to phosphoinositide 3-kinase (PI3K) activation, or Akt2 that acts downstream of PI3K, could rescue both hematopoietic and angiogenic defects in the K-ras knockdown. Consistently, the functional rescue by k-ras mRNA was significantly suppressed by wortmannin, a PI3K-specific inhibitor. Our results provide direct evidence that PI3K-Akt plays a crucial role in mediating K-ras signaling during hematopoiesis and angiogenesis in vivo, thus offering new targets and alternative vertebrate model for studying these processes and their related diseases

Myc-induced liver tumors in transgenic zebrafish can regress in tp53 null mutation

10.1371/journal.pone.0117249PLoS ONE101e011724

The role of vasculature and blood circulation in zebrafish swimbladder development

<p>Abstract</p> <p>Background</p> <p>Recently we have performed a detailed analysis of early development of zebrafish swimbladder, a homologous organ of tetrapod lung; however, the events of swimbladder development are still poorly characterized. Many studies have implicated the role of vascular system in development of many organs in vertebrates. As the swimbladder is lined with an intricate network of blood capillaries, it is of interest to investigate the role of the vascular system during early development of swimbladder.</p> <p>Results</p> <p>To investigate the role of endothelial cells (ECs) and blood circulation during development of the swimbladder, phenotypes of swimbladder were analysed at three different stages (~2, 3 and 5 dpf [day postfertilization]) in <it>cloche </it>(<it>clo</it>) mutant and Tnnt2 morphants, in the background of transgenic lines <it>Et(krt4:EGFP)</it>^{<it>sq33-2 </it>}and <it>Et(krt4:EGFP)</it>^{<it>sqet3 </it>}which express EGFP in the swimbladder epithelium and outer mesothelium respectively. Analyses of the three tissue layers of the swimbladder were performed using molecular markers <it>hb9</it>, <it>fgf10a</it>, <it>acta2</it>, and <it>anxa5 </it>to distinguish epithelium, mesenchyme, and outer mesothelium. We showed that the budding stage was independent of ECs and blood flow, while early epithelial growth, mesenchymal organization and its differentiation into smooth muscle, as well as outer mesothelial organization, were dependent on ECs. Blood circulation contributed to later stage of epithelial growth, smooth muscle differentiation, and organization of the outer mesothelium. Inflation of the swimbladder was also affected as a result of absence of ECs and blood flow.</p> <p>Conclusion</p> <p>Our data demonstrated that the vascular system, though not essential in swimbladder budding, plays an important role in the development of the swimbladder starting from the early growth stage, including mesenchyme organization and smooth muscle differentiation, and outer mesothelial organization, which in turn may be essential for the function of the swimbladder as reflected in its eventual inflation.</p

Student-centric Model of Learning Management System Activity and Academic Performance: from Correlation to Causation

In recent years, there is a lot of interest in modeling students' digital traces in Learning Management System (LMS) to understand students' learning behavior patterns including aspects of meta-cognition and self-regulation, with the ultimate goal to turn those insights into actionable information to support students to improve their learning outcomes. In achieving this goal, however, there are two main issues that need to be addressed given the existing literature. Firstly, most of the current work is course-centered (i.e. models are built from data for a specific course) rather than student-centered; secondly, a vast majority of the models are correlational rather than causal. Those issues make it challenging to identify the most promising actionable factors for intervention at the student level where most of the campus-wide academic support is designed for. In this paper, we explored a student-centric analytical framework for LMS activity data that can provide not only correlational but causal insights mined from observational data. We demonstrated this approach using a dataset of 1651 computing major students at a public university in the US during one semester in the Fall of 2019. This dataset includes students' fine-grained LMS interaction logs and administrative data, e.g. demographics and academic performance. In addition, we expand the repository of LMS behavior indicators to include those that can characterize the time-of-the-day of login (e.g. chronotype). Our analysis showed that student login volume, compared with other login behavior indicators, is both strongly correlated and causally linked to student academic performance, especially among students with low academic performance. We envision that those insights will provide convincing evidence for college student support groups to launch student-centered and targeted interventions that are effective and scalable.Comment: 43 pages, 9 figures, 18 tables, Journal of Educational Data Mining (Initial Submission

Evaluation Method for Probability of Blowout after the Failure of Offshore Well Killing

249-259With the development of offshore oil industry, the influx and blowout are inevitable. Well control methods have been well researched, but how to recognize the failure of well control earlier and how to evaluate the probability of blowout for taking steps to avoid are imperfect. Based on the two-phase gas-liquid flow, the characteristic of well killing curve before and after killing are analyzed. Then the method for recognizing the failure of well killing is established by the probabilistic and covariance processing method. Then the blowout due to the failure of well killing is studied and the build-up pressure template is established. According to this, three evaluation methods for blowout probability are established, the shut-off pressure, the standing and casing pressure, formation parameters and underbalanced level varying methods. Final, four hardware systems and one evaluation system are recommended for decreasing or avoiding the risk during the failure of well killing