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Robust hematopoietic stem cell activity is derived from pre-circulation embryos but not yolk sacs

ABSTRACT: Robust hematopoietic stem cell activity is derived from pre-circulation embryos but not yolk sacs

PROVIDER: PRJNA434768 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR6756980.fastq.gz	Fastqsanger.gz
	SRR6756979.fastq.gz	Fastqsanger.gz

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Robust hematopoietic stem cell activity is derived from pre-circulation embryos but not yolk sacs

Project description:The embryonic site of definitive hematopoietic stem cell (dHSC) origination has been debated for decades. Although an intra-embryonic origin is supported, recent data suggest that a large fraction of adult blood derives from the yolk sac (YS). Investigating the origins of hematopoiesis before heartbeat onset (i.e. 5-7 somite pairs (sp)) is precluded by a lack of assays that can distinguish dHSC precursors in early embryos. Here, we report robust, multi-lineage and serially transplantable dHSC activity from cultured 2-7sp murine embryonic explants (Em-Ex). dHSC were undetectable in 2-7sp YS explants (YS-Ex). Our work supports a model in which the embryo, not the YS, is the major source of lifelong hematopoiesis.

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Genome-wide analysis of gene expression of Flk1Cre;ETV2 CKO mouse yolk sacs.

Project description:To identify genes critical for vascular development, we generated mice where ETV2 is inactivated in FLK1+ cells by a loxP-Cre recombination approach. Results provide a detailed insight into the function of ETV2 in emrbyonic vasculare formation. Total RNA obtained from E9.5 yolk sacs from Flk1Cre;ETV2 CKO and control mice.

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Transcription profiling of mouse FVB/N embryonic yolk sacs eythroid and epithelial samples from day E9.5

Project description:Primitive erythropoiesis in the mouse yolk sac is followed by definitive erythropoiesis resulting in adult erythrocytes. In comparison to definitive erythropoiesis little is known about the genes that control the embryonic erythroid program. The purpose of this study was to generate a profile of mouse embryonic yolk sac erythroid cells and identify novel regulatory genes differentially expressed in erythroid compared to non-erythroid (epithelial cells). The identification of these genes will contribute to a greater understanding of how the primitive erythroid program is controlled. This work will have clinical implications for treating sickle cell anemia and Î²-thalassemia. Activating genes in adult erythroid cells that increase embryonic or fetal globin gene expression may be a therapeutic approach to treat individuals with these disorders. Experiment Overall Design: Embryonic day 9.5 (E9.5) yolk sacs were dissected from the embryos of timed-pregnant FVB/N mice. These tissues were frozen in OCT media and 8-micron frozen sections were obtained. Laser capture microdissection (LCM) was used to isolate primitive erythroid precursors and epithelial cells from these E9.5 yolk sac frozen sections using 2 to 4 yolk sacs from 2 different litters per biological replicate. Paired erythroid and epithelial samples were collected from the same microscope slides. Total RNA was isolated from 4 different pairs of erythroid and epithelial samples and hybridized to Affymetrix 430 A 2.0 microarrays.

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Project description:The embryonic site of definitive hematopoietic stem cell (dHSC) origination has been debated for decades. Although an intra-embryonic origin is well supported, the yolk sac (YS) contribution to adult hematopoiesis remains controversial. The same developmental origin makes it difficult to identify specific markers that discern between an intraembryonic versus YS-origin using a lineage trace approach. Additionally, the highly migratory nature of blood cells and the inability of pre-circulatory embryonic cells (i.e., 5-7 somite pairs (sp)) to robustly engraft in transplantation, even after culture, has precluded scientists from properly answering these questions. Here we report robust, multi-lineage and serially transplantable dHSC activity from cultured 2-7sp murine embryonic explants (Em-Ex). dHSC are undetectable in 2-7sp YS explants. Additionally, the engraftment from Em-Ex is confined to an emerging CD31+CD45+c-Kit+CD41- population. In sum, our work supports a model in which the embryo, not the YS, is the major source of lifelong definitive hematopoiesis.

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Next Generation Sequencing Facilitates Quantitative Analysis of Transcriptomes of Yolk Sacs of E16.5 Wild Type and Rsu1-/- mouse embryos

Project description:Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to compare NGS-derived yolk sac transcriptome profiling (RNA-seq) of E16.5 Rsu1-/- mouse embryos to that of the wild-type controls Methods: Yolk sac mRNA profiles of yolk sac isolated from E16.5 wild-type (WT) and ras suppressor 1 (Rsu1−/−) emdbryos were generated by deep sequencing, in triplicate, using Illumina Hiseq 2500 platform. The sequence reads that passed quality filters were were mapped to human reference genome GRCh38 by HISAT2 v2.2.1 with default parameters. Results: Using an optimized data analysis workflow, we mapped about 40 million sequence reads per sample to the mouse genome and identified ___ transcripts in the yolk sacs of E16.5 WT and Rsu1−/− embryos with HISAT2 v2.2.1 workflow . Approximately __ % of the transcripts showed differential expression between the WT and Rsu1−/− yolk sac, with a fold change ≥2.0 and p value <0.05. Altered expression of 12 genes was confirmed with qRT–PCR, demonstrating the high degree of sensitivity of the RNA-seq method. Conclusions: Our study represents the first detailed analysis of E16.5 Rsu1-/- yolk sac transcriptomes, which would expedite genetic network analyses and permit the dissection of complex biologic functions of Rsu1 during late embryogenesis.

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RNA sequencing analysis of embryonic yolk sacs from wildtype and Etv2 mutant embryos at E8.5

Project description:RNA sequencing analysis of embryonic yolk sacs from wildtype and Etv2 mutant embryos at E8.5

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