RESUMO
Adult hematopoietic stem and progenitor cells (HSPCs) respond directly to inflammation and infection, causing both acute and persistent changes to quiescence, mobilization, and differentiation. Here we show that murine fetal HSPCs respond to prenatal inflammation in utero and that the fetal response shapes postnatal hematopoiesis and immune cell function. Heterogeneous fetal HSPCs show divergent responses to maternal immune activation (MIA), including changes in quiescence, expansion, and lineage-biased output. Single-cell transcriptomic analysis of fetal HSPCs in response to MIA reveals specific upregulation of inflammatory gene profiles in discrete, transient hematopoietic stem cell (HSC) populations that propagate expansion of lymphoid-biased progenitors. Beyond fetal development, MIA causes the inappropriate expansion and persistence of fetal lymphoid-biased progenitors postnatally, concomitant with increased cellularity and hyperresponsiveness of fetal-derived innate-like lymphocytes. Our investigation demonstrates how inflammation in utero can direct the output and function of fetal-derived immune cells by reshaping fetal HSC establishment.
Assuntos
Hematopoese , Células-Tronco Hematopoéticas , Gravidez , Feminino , Camundongos , Animais , Hematopoese/fisiologia , Células-Tronco Hematopoéticas/metabolismo , Feto , Inflamação/metabolismo , Desenvolvimento FetalRESUMO
RNA-Seq is now a routinely employed assay to measure gene expression. As the technique matured over the last decade, so have dedicated analytic tools. In this chapter, we first describe the mainstream as well as the most up-to-date protocols and their implications on downstream analysis. We then detail the steps entailing RNA-Seq analysis in three main stages: (i) preprocessing and data preparation, (ii) upstream processing, and (iii) high-level analyses. We review the most recent and relevant tools as one workflow following a stepwise order. The chapter further encompasses in-depth features of these tools. Details of the required code are made available throughout the chapter, as well as of the underlying statistics. We illustrate these steps with analysis of publicly available RNA-Seq data.
Assuntos
Perfilação da Expressão Gênica , Sequenciamento de Nucleotídeos em Larga Escala , Expressão Gênica , Perfilação da Expressão Gênica/métodos , Sequenciamento de Nucleotídeos em Larga Escala/métodos , RNA-Seq , Análise de Sequência de RNA/métodos , SoftwareRESUMO
Hematopoietic stem cells (HSCs) rely on complex regulatory networks to preserve stemness. Due to the scarcity of HSCs, technical challenges have limited our insights into the interplay between metabolites, transcription, and the epigenome. In this study, we generated low-input metabolomics, transcriptomics, chromatin accessibility, and chromatin immunoprecipitation data, revealing distinct metabolic hubs that are enriched in HSCs and their downstream multipotent progenitors. Mechanistically, we uncover a non-classical retinoic acid (RA) signaling axis that regulates HSC function. We show that HSCs rely on Cyp26b1, an enzyme conventionally considered to limit RA effects in the cell. In contrast to the traditional view, we demonstrate that Cyp26b1 is indispensable for production of the active metabolite 4-oxo-RA. Further, RA receptor beta (Rarb) is required for complete transmission of 4-oxo-RA-mediated signaling to maintain stem cells. Our findings emphasize that a single metabolite controls stem cell fate by instructing epigenetic and transcriptional attributes.
Assuntos
Células-Tronco Hematopoéticas , Tretinoína , Diferenciação Celular , Ácido Retinoico 4 Hidroxilase/genética , Transdução de Sinais , Tretinoína/farmacologiaRESUMO
The endothelial to haematopoietic transition (EHT) is the process whereby haemogenic endothelium differentiates into haematopoietic stem and progenitor cells (HSPCs). The intermediary steps of this process are unclear, in particular the identity of endothelial cells that give rise to HSPCs is unknown. Using single-cell transcriptome analysis and antibody screening, we identify CD44 as a marker of EHT enabling us to isolate robustly the different stages of EHT in the aorta-gonad-mesonephros (AGM) region. This allows us to provide a detailed phenotypical and transcriptional profile of CD44-positive arterial endothelial cells from which HSPCs emerge. They are characterized with high expression of genes related to Notch signalling, TGFbeta/BMP antagonists, a downregulation of genes related to glycolysis and the TCA cycle, and a lower rate of cell cycle. Moreover, we demonstrate that by inhibiting the interaction between CD44 and its ligand hyaluronan, we can block EHT, identifying an additional regulator of HSPC development.