Unknown,Transcriptomics,Genomics,Proteomics

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Expression data of the endothelial-to-hematopoietic transition in E8.5 concepti

ABSTRACT: During development a specialised subset of endothelial cells, the haemogenic endothelium, undergo an endothelial-to-haematopoietic transition. This process critically involves the transcription factor Runx1. Here we have isolated a specific subpopulation of endothelial cells using a Runx1 enhancer-reporter transgenic mouse line (23GFP). We have compared the gene expression profile of this population to non-23GFP expressing endothelial cells and CD41 expressing haematopoietic progenitor cells to assess whether 23GFP expression marks a biologically distinct subset of endothelium. We used affymetrix microarrays to detail the global programme of gene expression underlying the different populations and identified a distinct expression profile of the 23GFP expressing endothelium. 23GFP transgenic mouse embryos were dissected free of the ectoplacental cone at embryonic day E8.5, pooled, and sorted into 23GFP- and 23GFP+ endothelial (VE-Cad+ Ter119- CD45- CD41-) and CD41+ haematopoietic cells (VE-Cad+ Ter119-CD45-CD41+23GFP+) cells by FACS. Triplicate samples for each cell population were analysed. cDNA synthesis, fragmentation, and hybridisation were performed by the Stanford Protein and Nucleic Acid Facility.

ORGANISM(S): Mus musculus

SUBMITTER: Stephen Taylor

PROVIDER: E-GEOD-52075 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Haematopoietic stem cells (HSCs) are derived early from embryonic precursor cells, such as haemogenic endothelial cells and pre-HSCs. However, the identity of precursor cells remains elusive due to their rareness, transience, and inability to be isolated efficiently. Here we employed potent surface markers to capture the nascent pre-HSCs at 30% purity, as rigorously validated by single-cell-initiated serial transplantation assay. Then we applied single-cell RNA-Seq technique to analyse five populations closely related to HSC formation: endothelial cells, CD45- and CD45+ pre-HSCs in E11 aorta-gonad-mesonephros (AGM) region, and mature HSCs in E12 and E14 foetal liver. In comparison, the pre-HSCs showed unique features in transcriptional machinery, apoptosis, metabolism state, signalling pathway, transcription factor network, and lncRNA expression pattern. Among signalling pathways enriched in pre-HSCs, the mTOR activation was uncovered indispensable for the emergence of HSCs but not haematopoietic progenitors from endothelial cells in vivo. By comparing with proximal populations without HSC potential, the core molecular signature of pre-HSCs was identified. Collectively, our work paves the way for dissection of complex molecular mechanisms regulating the step-wise generation of HSCs in vivo, informing future efforts to engineer HSCs for clinical application. RNA-Seq of 181 single-cell samples from 8 FACS sorted cell types: 1. endothelial cells (samples E11.0_EC_xxxx. CD31+ VE-cadherin+CD41-CD43-CD45-Ter119-); 2. T1 pre-HSCs (samples E11.0_T1_xxxx. CD31+CD45-CD41low c-Kit+CD201high); 3. T1 CD201- cells (samples E11.0_T1CD201neg_xxxx, CD31+CD45-CD41low c-Kit+CD201low/-) ; 4. T2 pre-HSCs (samples E11.0_T2_35xx. CD31+CD45+c-Kit+CD201high), 5. T2 CD41low (samples E11.0_T2_21xx, E11.0_T2_24xx and E11.0_T2_27xx. CD31+CD45+CD41low); 6. E12 HSCs (samples E12.5_FL_xxxx. Lin-Sca-1+Mac-1lowCD201+); 7. E14 HSCs (samples E14.5_FL_xxxx. CD45+CD150+CD48-CD201+); 8. Adult HSCs (samples Adult_HSC_xxxx. CD45+CD150+CD48-CD201+). ECs, T1 pre-HSCs, T1 CD201- cells, T2 pre-HSCs, T2 CD41low cells were sorted from E11 AGM region. Mature HSCs were sorted from E12 or E14 fetal liver and adult bonemarrow.

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Expression data of the endothelial-to-hematopoietic transition in E8.5 concepti

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