Parathyroid hormone (PTH) plays an essential role in regulating calcium and bone homeostasis in the adult, but whether PTH is required at all for regulating fetal-placental mineral homeostasis is uncertain. To address this we treated Pth-null mice in utero with 1 nmol PTH (1-84) or saline and examined placental calcium transfer 90 minutes later. It was found that placental calcium transfer increased in Pth-null fetuses treated with PTH as compared to Pth-null fetuses treated with saline. Subsequently, to determine the effect of PTH treatment on placental gene expression, in a separate experiment, 90 minutes after the fetal injections the placentas were removed for subsequent RNA extraction and microarray analysis.
Parathyroid hormone regulates fetal-placental mineral homeostasis.
Sex, Specimen part, Treatment
View SamplesWe report high-throughput profiling of gene expression from whole zebrafish ventricles. We profile mRNA in uninjured ventricles and those undergoing regeneration 14 days after genetic ablation. This study provides a framework for understanding transcriptional changes during adult models of regeneration. Overall design: Examination of gene expression in cardiomyocytes under different states of proliferation.
Resolving Heart Regeneration by Replacement Histone Profiling.
No sample metadata fields
View SamplesBackground: Tissue macrophages contribute to development and protection, both requiring appropriately timed and located source(s) of factors controlling growth, cell differentiation and migration. Goal: To understand the role of microglia (tissue macrophages of the central nervous system), in promoting neurodevelopment and controlling neuroinflammation. Summary of findings: We show that microglia fulfill both these roles. In contrast to adult cells, neonatal microglia show a unique neurogenic phenotype with stem cell-like potential. Neonatal microglia are protective against neuroinflammation, and their transplantation ameliorates experimental autoimmune encephalomyelitis. A CD11c+ microglial subset predominates in primary myelinating areas of the developing brain and expresses genes for neuronal and glial survival, migration and differentiation. CD11c+ microglia are also found in clusters of repopulating microglia after experimental ablation and in neuroinflammation in adult mice, but despite some similarities, they do not recapitulate neurogenic neonatal microglia characteristics. Conclusions: We therefore identify a unique phenotype of neonatal microglia that deliver signals necessary for neurogenesis and myelination and suppress neuroinflammation. Overall design: The overall design was to compare transcriptomes of subsets of microglia isolated from neonatal mice, healthy adults, and adult mice with a neuroinflammatory disease (Experimental autoimmune encephalomyelitis, EAE), and to compare anti-inflammatory function of adult and neonatal microglia. Microglia were isolated by cell-sorting based on surface phenotype, and RNAseq data was analyzed using WGCNA, GO and DAVID approaches. Expression of selected genes and pathways was confirmed by histology and flow cytometry. Functional analysis involved transfer of isolated microglia to the central nervous system of animals with EAE and evaluation of outcome. EAE = Experimental autoimmune encephalomyelitis
A novel microglial subset plays a key role in myelinogenesis in developing brain.
Subject
View SamplesBy contrast with mammals, adult zebrafish have a high capacity to regenerate damaged or lost myocardium through proliferation of spared cardiomyocytes. The epicardial sheet covering the heart is activated by injury and aids muscle regeneration through paracrine effects and as a multipotent cell source, and has received recent attention as a target in cardiac repair strategies. While it is recognized that epicardium is required for muscle regeneration and itself has high regenerative potential, the extent of cellular heterogeneity within epicardial tissue is largely unexplored. In this study, we performed transcriptome analysis on dozens of epicardial lineage cells purified from zebrafish harboring a transgenic reporter for the pan-epicardial gene tcf21. Hierarchical clustering analysis suggested the presence of at least three epicardial cell subsets defined by expression signatures. We validated many new pan-epicardial and epicardial markers by alternative expression assays. Additionally, we explored the function of the scaffolding protein and main component of caveolae, caveolin-1 (cav1), which was present in each epicardial subset. In BAC transgenic zebrafish, cav1 regulatory sequences drove strong expression in ostensibly all epicardial cells and in coronary vascular endothelial cells. Moreover, cav1 mutant zebrafish generated by genome editing showed grossly normal heart development and adult cardiac anatomy, but displayed profound defects in injury-induced cardiomyocyte proliferation and heart regeneration. Our study defines a new platform for the discovery of epicardial lineage markers, genetic tools, and mechanisms of heart regeneration. Overall design: Deep sequencing of isolated single epicardial cells
Single epicardial cell transcriptome sequencing identifies Caveolin 1 as an essential factor in zebrafish heart regeneration.
Age, Specimen part, Cell line, Subject
View SamplesA neuronal PI(3,4,5)P3-dependent program of oligodendrocyte precursor recruitment and myelination was identified in mice that conditionally lack PTEN in cerebellar granular cells (PTEN cKO)
A neuronal PI(3,4,5)P<sub>3</sub>-dependent program of oligodendrocyte precursor recruitment and myelination.
Sex, Age, Specimen part
View SamplesWe compared transcriptional profiles of regenerating zebrafish caudal fins following fin amputation with profiles from uninjured zebrafish caudal fins Overall design: Examination of whole fin transcriptional profiles from regenerating fins (2 pools of 10 fins) and uninjured fins (2 pools of 10 fins)
Modulation of tissue repair by regeneration enhancer elements.
No sample metadata fields
View SamplesWe compared transcriptional and chromatin profiles of regenerating zebrafish hearts following genetic ablation with profiles from uninjured zebrafish hearts. Overall design: Examination of whole heart transcriptional profiles from ablated hearts (2 pools of 10 hearts) and uninjured hearts (2 pools of 10 hearts). Examination of differential H3K27Ac marks following genetic ablation of cardiomyocytes (regenerating hearts) and uninjured hearts.
Modulation of tissue repair by regeneration enhancer elements.
No sample metadata fields
View SamplesDevelopmental pathways that orchestrate the fleeting transition of endothelial cells into haematopoietic stem cells remain undefined. Here we demonstrate a tractable approach for fully reprogramming adult mouse endothelial cells to haematopoietic stem cells (rEC-HSCs) through transient expression of the transcription-factor-encoding genes Fosb, Gfi1, Runx1, and Spi1 (collectively denoted hereafter as FGRS) and vascular-niche-derived angiocrine factors. The induction phase (days 0-8) of conversion is initiated by expression of FGRS in mature endothelial cells, which results in endogenous Runx1 expression. During the specification phase (days 8-20), RUNX1+ FGRS-transduced endothelial cells commit to a haematopoietic fate, yielding rEC-HSCs that no longer require FGRS expression. The vascular niche drives a robust self-renewal and expansion phase of rEC-HSCs (days 20-28). rEC-HSCs have a transcriptome and long-term self-renewal capacity similar to those of adult haematopoietic stem cells, and can be used for clonal engraftment and serial primary and secondary multi-lineage reconstitution, including antigen-dependent adaptive immune function. Inhibition of TGF? and CXCR7 or activation of BMP and CXCR4 signalling enhanced generation of rEC-HSCs. Pluripotency-independent conversion of endothelial cells into autologous authentic engraftable haematopoietic stem cells could aid treatment of haematological disorders. Overall design: Expression profiling by high throughput sequencing data; GPL17021 Illumina HiSeq 2500 (Mus musculus)
Conversion of adult endothelium to immunocompetent haematopoietic stem cells.
Specimen part, Subject
View SamplesGene expression changes in 3 human melanoma cell lines were compared to freshly isolated normal primary melanocytes Overall design: Three biological replicates for each melanoma cell line and primary melanocytes were labeled and run Illumina HiSeq2500. The transcriptome of melanocytes was compared to cell line SK-Mel-28, SK-Mel-147 or UACC-62.
Systems analysis identifies melanoma-enriched pro-oncogenic networks controlled by the RNA binding protein CELF1.
Specimen part, Subject
View SamplesSafety sciences and the identification chemical hazard have been seen as one of the most immediate practical applications of human pluripotent stem cell technology. Protocols for the generation of many desirable human cell types have been developed, but optimization of neuronal models for toxicological use has been astonishingly slow, and the wide, clinically- important field of peripheral neurotoxicity is still largely unexplored. Here, a 2-step protocol to generate large lots of identical peripheral human neuronal precursors was characterized and adapted to the measurement of peripheral neurotoxicity. High content imaging allowed an unbiased assessment of cell morphology and viability. The computational quantification of neurite growth as functional parameter highly sensitive to disturbances by toxicants was used as endpoint reflecting specific neurotoxicity. The differentiation of cells towards dorsal root ganglia neurons was tracked in relation to a large background data set based on gene expression microarrays. On this basis, a peripheral neurotoxicity (PeriTox) test was developed as first toxicological assay that harnesses the potential of human pluripotent stem cells to generate cell types/tissues that are not otherwise available for prediction of human systemic organ toxicity. Testing of more than 30 chemicals showed that human neurotoxicants, as well as neurite growth enhancers, were correctly identified. Various classes of chemotherapeutics causing human peripheral neuropathies were identified, while they were missed when tested on human central neurons. The PeriTox-test established here shows the potential of human stem cells for clinically-relevant safety testing of drugs in use and of new emerging candidates.
Stem Cell-Derived Immature Human Dorsal Root Ganglia Neurons to Identify Peripheral Neurotoxicants.
Sex, Specimen part, Cell line
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