HSPCs display within-family homogeneity in differentiation and proliferation despite population heterogeneity

Tak, Tamar; Prevedello, Giulio; Simon, Gaël; Paillon, Noémie; Benlabiod, Camélia; Marty, Caroline; Plo, Isabelle; Duffy, Ken R.; Perié, Leïla

doi:10.7554/elife.60624

Cited by 9 publications

(10 citation statements)

References 55 publications

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“…After filtering, read counts of each barcode in the different cell lineages were normalized enabling categorization into classes of biased output toward the analyzed lineages using a threshold of 10% of barcode reads (other thresholds in Figure 1—figure supplement 3c and Figure 7—figure supplement 1c ). Statistics on barcoding results were performed using a permutation test as in Tak et al, 2021 . Significance of flow cytometry results was assessed using Student’s t -test.…”

Section: Methodsmentioning

confidence: 99%

Erythropoietin directly remodels the clonal composition of murine hematopoietic multipotent progenitor cells

Eisele

Cosgrove

Magniez

et al. 2022

eLife

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The cytokine erythropoietin (EPO) is a potent inducer of erythrocyte development and one of the most prescribed biopharmaceuticals. The action of EPO on erythroid progenitor cells is well established, but its direct action on hematopoietic stem and progenitor cells (HSPCs) is still debated. Here, using cellular barcoding, we traced the differentiation of hundreds of single murine HSPCs, after ex vivo EPO-exposure and transplantation, in five different hematopoietic cell lineages, and observed the transient occurrence of high-output Myeloid-Erythroid-megaKaryocyte (MEK)-biased and Myeloid-B-cell-Dendritic cell (MBDC)-biased clones. Single-cell RNA sequencing (ScRNAseq) analysis of ex vivo EPO-exposed HSPCs revealed that EPO induced the upregulation of erythroid associated genes in a subset of HSPCs, overlapping with multipotent progenitor (MPP) 1 and MPP2. Transplantation of Barcoded EPO-exposed-MPP2 confirmed their enrichment in Myeloid-Erythroid-biased clones. Collectively, our data show that EPO does act directly on MPP independent of the niche, and modulates fate by remodeling the clonal composition of the MPP pool.

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Section: Methodsmentioning

confidence: 99%

Erythropoietin directly remodels the clonal composition of murine hematopoietic multipotent progenitor cells

Eisele

Cosgrove

Magniez

et al. 2022

eLife

Self Cite

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“…graphically represents the output of the statistical testing performed in the script "2_bar_plot". A formal description of the statistical framework is available 26 . In brief, this framework enables statistical hypothesis testing while assuming that cells from the same family are dependent (an assumption that is itself testable), contrary to classical statistics that would require independence between all observed cells.…”

Section: Figure 5biimentioning

confidence: 99%

Simultaneous Assessment of Kinship, Division Number, and Phenotype <em>via</em> Flow Cytometry for Hematopoietic Stem and Progenitor Cells

Donada

Tak

Prevedello

et al. 2023

JoVE

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Few techniques can assess phenotype and fate for the same cell simultaneously. Most of the current protocols used to characterize phenotype, although able to generate large datasets, necessitate the destruction of the cell of interest, making it impossible to assess its functional fate. Heterogeneous biological differentiating systems like hematopoiesis are therefore difficult to describe. Building on cell division tracking dyes, we further developed a protocol to simultaneously determine kinship, division number, and differentiation status for many single hematopoietic progenitors. This protocol allows the assessment of the ex vivo differentiation potential of murine and human hematopoietic progenitors, isolated from various biological sources. Moreover, as it is based on flow cytometry and a limited number of reagents, it can quickly generate a large amount of data, at the single-cell level, in a relatively inexpensive manner. We also provide the analytical pipeline for single-cell analysis, combined with a robust statistical framework. As this protocol allows the linking of cell division and differentiation at the single-cell level, it can be used to quantitatively assess symmetric and asymmetric fate commitment, the balance between self-renewal and differentiation, and the number of divisions for a given commitment fate. Altogether, this protocol can be used in experimental designs aiming to unravel the biological differences between hematopoietic progenitors, from a single-cell perspective.

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“…HSC transplantation, barcoding and fate mapping experiments showed that only a few HSCs can produce all blood cells, while the majority of HSC differentiation is restricted or imbalanced to few lineages (2)(3)(4)(5)(6)(7). Furthermore, HSCs are different in their proliferative capacity which influence self-renewal kinetics (8,9), with some HSCs been able to generate specific blood cells without undergoing cell division (10).…”

Section: Introductionmentioning

confidence: 99%

Hematopoietic stem and progenitor cell heterogeneity is inherited from the embryonic hemogenic endothelium

Ghersi

Baldissera

Hintzen

et al. 2022

Preprint

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Multipotent hematopoietic stem/progenitor cells (HSPCs) generate all mature blood cells in the erythroid, lymphoid, and myeloid lineages. HSPCs are initially produced in the embryo, via transdifferentiation of hemogenic endothelial cells (hemECs) in the aorta-gonad mesonephros (AGM). HSPCs in the AGM are functionally heterogenous in differentiation and proliferative output, but how these intrinsic differences are acquired remains unanswered. This knowledge could inform approaches to overcome the dysregulation of HSPC heterogeneity associated with poor outcomes of autologous transplants. Here we discovered that loss of microRNA (miR)-128 (miR-128Δ/Δ) in zebrafish leads to an expansion of hemECs forming replicative HSPCs in the AGM, and a skew towards the erythroid and lymphoid lineages in larval and adult stages. Furthermore, we found that inhibiting miR-128 during the differentiation of human pluripotent stem cells into hemECs, but not during the endothelial-to-hematopoietic transition, recapitulated the lineage skewing. In vivo, expression of wild-type miR-128 in endothelium restored the blood lineage distribution in miR-128Δ/Δ zebrafish. We found that miR-128 represses the expression of the Wnt inhibitor csnk1a1 and the Notch ligand jag1b, and thus promotes Wnt and Notch signaling in hemECs. De-repression of cskn1a1 resulted in hemECs generating replicative and erythroid-biased HSPCs, whereas de-repression of jag1b resulted in hemECs forming lymphoid-biased HSPCs in the AGM and relative mature blood cells in adult. We propose that HSPC heterogeneity is established in hemogenic endothelium prior to transdifferentiation and is programmed in part by Wnt and Notch signaling modulation.

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HSPCs display within-family homogeneity in differentiation and proliferation despite population heterogeneity

Cited by 9 publications

References 55 publications

Erythropoietin directly remodels the clonal composition of murine hematopoietic multipotent progenitor cells

Erythropoietin directly remodels the clonal composition of murine hematopoietic multipotent progenitor cells

Simultaneous Assessment of Kinship, Division Number, and Phenotype <em>via</em> Flow Cytometry for Hematopoietic Stem and Progenitor Cells

Hematopoietic stem and progenitor cell heterogeneity is inherited from the embryonic hemogenic endothelium

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