Project description:Many pathways regulating blood formation have been elucidated, yet how each coordinates with embryonic biophysiology to modulate the spatio-temporal production of hematopoietic stem cells (HSCs) is currently unresolved. Here, we report that glucose metabolism impacts the onset and magnitude of HSC induction in vivo. In zebrafish, transient elevations in physiological glucose levels elicited dose-dependent effects on HSC development, including enhanced runx1 expression and hematopoietic cluster formation in the Aorta-Gonad-Mesonephros (AGM) region; embryonic-to-adult transplantation studies confirmed glucose increased functional HSCs. Glucose uptake was required to mediate the enhancement in HSC development; likewise, metabolic inhibitors diminished nascent HSC production and reversed glucose-mediated effects on HSCs. Increased glucose metabolism preferentially impacted hematopoietic and vascular targets, as determined by gene expression analysis, through mitochondrial-derived reactive oxygen species (ROS)-mediated stimulation of hypoxia inducible factor 1α (hif1α); epistasis assays demonstrated hif1α regulates HSC formation in vivo and mediates the dose-dependent effects of glucose metabolism on the timing and magnitude of HSC production. We propose this fundamental metabolic-sensing mechanism enables the embryo to respond to changes in environmental energy input and adjust hematopoietic output to maintain embryonic growth and ensure viability. We performed microarray analysis to explore the changes in gene expression that occur in repsonse to altered metabolism during the induction of developmental hematopoeisis. We compared RNA from zebrafish raised in a solution of 1% glucose from 12-36hpf vs controls. Two biological replicates for each condition were performed.
Project description:Many pathways regulating blood formation have been elucidated, yet how each coordinates with embryonic biophysiology to modulate the spatio-temporal production of hematopoietic stem cells (HSCs) is currently unresolved. Here, we report that glucose metabolism impacts the onset and magnitude of HSC induction in vivo. In zebrafish, transient elevations in physiological glucose levels elicited dose-dependent effects on HSC development, including enhanced runx1 expression and hematopoietic cluster formation in the Aorta-Gonad-Mesonephros (AGM) region; embryonic-to-adult transplantation studies confirmed glucose increased functional HSCs. Glucose uptake was required to mediate the enhancement in HSC development; likewise, metabolic inhibitors diminished nascent HSC production and reversed glucose-mediated effects on HSCs. Increased glucose metabolism preferentially impacted hematopoietic and vascular targets, as determined by gene expression analysis, through mitochondrial-derived reactive oxygen species (ROS)-mediated stimulation of hypoxia inducible factor 1α (hif1α); epistasis assays demonstrated hif1α regulates HSC formation in vivo and mediates the dose-dependent effects of glucose metabolism on the timing and magnitude of HSC production. We propose this fundamental metabolic-sensing mechanism enables the embryo to respond to changes in environmental energy input and adjust hematopoietic output to maintain embryonic growth and ensure viability.
Project description:Comparison of temporal gene expression profiles from Danio rerio skin. The RNA-seq data comprise 3 age groups at 5, 24 and 30 months. Jena Centre for Systems Biology of Ageing - JenAge (www.jenage.de)
Project description:Comparison of temporal small RNA gene expression profiles from Danio rerio brain. The smallRNA-seq data comprise 5 age groups at 6, 12, 24, 36 and 42 months. Jena Centre for Systems Biology of Ageing - JenAge (www.jenage.de)
Project description:Comparison of temporal small RNA gene expression profiles from Danio rerio skin. The smallRNA-seq data comprise 5 age groups at 6, 12, 24, 36 and 42 months. Jena Centre for Systems Biology of Ageing - JenAge (www.jenage.de)
Project description:Zebrafish are an important model organism with inherent advantages that have the potential to make zebrafish a widely applied model for the study of energy homeostasis and obesity. The small size of zebrafish allows for assays on embryos to be conducted in a 96- or 384-well plate format, Morpholino and CRISPR based technologies promote ease of genetic manipulation, and drug treatment by bath application is viable. Moreover, zebrafish are ideal for forward genetic screens allowing for novel gene discovery. Given the relative novelty of zebrafish as a model for obesity, it is necessary to develop tools that fully exploit these benefits. Herein, we describe a method to measure energy expenditure in thousands of embryonic zebrafish simultaneously. We have developed a whole animal microplate platform in which we use 96-well plates to isolate individual fish and we assess cumulative NADH2 production using the commercially available cell culture viability reagent alamarBlue. In poikilotherms the relationship between NADH2 production and energy expenditure is tightly linked. This energy expenditure assay creates the potential to rapidly screen pharmacological or genetic manipulations that directly alter energy expenditure or alter the response to an applied drug (e.g. insulin sensitizers).