Transition of signal requirement in hematopoietic stem cell development from hemogenic endothelial cells.

Hematopoietic stem cells (HSCs) develop from hemogenic endothelial cells (HECs) in vivo during mouse embryogenesis. When cultured in vitro, cells from the embryo phenotypically defined as pre-HSC-I and pre-HSC-II have the potential to differentiate into HSCs. However, minimal factors required for HSC induction from HECs have not yet been determined. In this study, we demonstrated that stem cell factor (SCF) and thrombopoietin (TPO) induced engrafting HSCs from embryonic day (E) 11.5 pre-HSC-I in a serum-free and feeder-free culture condition. In contrast, E10.5 pre-HSC-I and HECs required an endothelial cell layer in addition to SCF and TPO to differentiate into HSCs. A single-cell RNA sequencing analysis of E10.5 to 11.5 dorsal aortae with surrounding tissues and fetal livers detected TPO expression confined in hepatoblasts, while SCF was expressed in various tissues, including endothelial cells and hepatoblasts. Our results suggest a transition of signal requirement during HSC development from HECs. The differentiation of E10.5 HECs to E11.5 pre-HSC-I in the aorta-gonad-mesonephros region depends on SCF and endothelial cell-derived factors. Subsequently, SCF and TPO drive the differentiation of E11.5 pre-HSC-I to pre-HSC-II/HSCs in the fetal liver. The culture system established in this study provides a beneficial tool for exploring the molecular mechanisms underlying the development of HSCs from HECs.

Tuning apicobasal polarity and junctional recycling in the hemogenic endothelium orchestrates the morphodynamic complexity of emerging pre-hematopoietic stem cells.

Hematopoietic stem cells emerge in the embryo from an aortic-derived tissue called the hemogenic endothelium (HE). The HE appears to give birth to cells of different nature and fate but the molecular principles underlying this complexity are largely unknown. Here we show, in the zebrafish embryo, that two cell types emerge from the aortic floor with radically different morphodynamics. With the support of live imaging, we bring evidence suggesting that the mechanics underlying the two emergence types rely, or not, on apicobasal polarity establishment. While the first type is characterized by reinforcement of apicobasal polarity and maintenance of the apical/luminal membrane until release, the second type emerges via a dynamic process reminiscent of trans-endothelial migration. Interfering with Runx1 function suggests that the balance between the two emergence types depends on tuning apicobasal polarity at the level of the HE. In support of this and unexpectedly, we show that Pard3ba - one of the four Pard3 proteins expressed in the zebrafish - is sensitive to interference with Runx1 activity, in aortic endothelial cells. This supports the idea of a signaling cross talk controlling cell polarity and its associated features, between aortic and hemogenic cells. In addition, using new transgenic fish lines that express Junctional Adhesion Molecules and functional interference, we bring evidence for the essential role of ArhGEF11/PDZ-RhoGEF in controlling the HE-endothelial cell dynamic interface, including cell-cell intercalation, which is ultimately required for emergence completion. Overall, we highlight critical cellular and dynamic events of the endothelial-to-hematopoietic transition that support emergence complexity, with a potential impact on cell fate.

In mammals and other animals with backbones, the cells that will make up blood and immune cells are generated during a very narrow timeframe in embryonic development. These cells, called hematopoietic stem cells and progenitors (or HSPCs for short), emerge from tissue known as hemogenic endothelium that makes up the floor of early blood vessels. For HPSCs to eventually specialise into different types of blood and immune cells, they require diverse migratory and homing properties that, ultimately, will determine the specific type of functions they exert. An important question for scientists studying the development of different blood and immune cell types is when this commitment to functional diversity is established. It could, for example, arise due to cells in the hemogenic endothelium having different origins. Alternatively, the signals that generate hemogenic endothelium cells could be responsible. It is also possible that both explanations are true, and that having different mechanisms involved ensures diversity in populations of HSPCs. To investigate differences between the HSPCs emerging from the hemogenic endothelium, Torcq et al. studied zebrafish embryos that had been modified so that one of the proteins involved in sensing cell polarity ­ where the top and bottom of the cell are located ­ was fluorescent. Live imaging of the embryos showed that two types of cells, with striking differences in morphology, emerge from the hemogenic tissue. In addition, one cell type displays the same polarity as the other vessel cells, whereas the other does not. Torcq et al. also present evidence suggesting that the signals responsible for controlling this cell polarity are provided by surrounding blood vessel cells, supporting the idea of an interplay between the different cell types. The finding that two different cell types emerge from the hemogenic endothelium, reveals a potential new source of diversity in HSPCs. Ultimately, this is expected to contribute to their functional complexity, resulting in both long-term stem cells that retain their full regenerative potential into adulthood and more specialized blood and immune cells.

The Onset of Intussusceptive Angiogenesis in COVID-19 Patients Might Come from the Mobilization of Stem Cell Sub-Populations Expressing the Hemangioblast Marker CD143.

COVID-19 and infectious diseases have been included in strategic development goals (SDG) of United Nations (UN). The SARS-CoV-2 pandemic has unveiled complex pathophysiological mechanisms underpinning COVID-19, notably inducing a systemic acquired vascular hemopathy characterized by endothelial dysfunction and intussusceptive angiogenesis, a rapid vascular remodeling process identified as a hallmark in severe COVID-19 cases affecting pulmonary and cardiac tissues. Stem cell migration have been proposed as significant regulators of this neoangiogenic process. In a monocentric cross-sectional study, through spectral flow cytometry analysis of peripheral blood mononuclear cells, we identified a distinct stem cell subpopulation mobilized in critical COVID-19. Indeed, by an unsupervised analysis generating a UMAP representation we highlighted eleven different clusters in critical and non-critical COVID-19 patients. Only one cluster was significantly associated to critical COVID-19 compared to non-critical patients. This cluster expressed the markers: CD45dim, CD34+, CD117+, CD147+, and CD143+, and were negative for CD133. Higher level of expression of hemangioblast markers CD143 were found in critical COVID-19 patients. This population, indicative of hemangioblast-like cells, suggests a key role in COVID-19-related neoangiogenesis, potentially driving the severe vascular complications observed. Our findings underscore the need for further investigation into the contributions of adult stem cells in COVID-19 pathology, offering new insights into therapeutic targets and interventions.

CD32 captures committed haemogenic endothelial cells during human embryonic development.

During embryonic development, blood cells emerge from specialized endothelial cells, named haemogenic endothelial cells (HECs). As HECs are rare and only transiently found in early developing embryos, it remains difficult to distinguish them from endothelial cells. Here we performed transcriptomic analysis of 28- to 32-day human embryos and observed that the expression of Fc receptor CD32 (FCGR2B) is highly enriched in the endothelial cell population that contains HECs. Functional analyses using human embryonic and human pluripotent stem cell-derived endothelial cells revealed that robust multilineage haematopoietic potential is harboured within CD32+ endothelial cells and showed that 90% of CD32+ endothelial cells are bona fide HECs. Remarkably, these analyses indicated that HECs progress through different states, culminating in FCGR2B expression, at which point cells are irreversibly committed to a haematopoietic fate. These findings provide a precise method for isolating HECs from human embryos and human pluripotent stem cell cultures, thus allowing the efficient generation of haematopoietic cells in vitro.

Autophagy regulates the maturation of hematopoietic precursors in the embryo.

An understanding of the mechanisms regulating embryonic hematopoietic stem cell (HSC) development would facilitate their regeneration. The aorta-gonad-mesonephros region is the site for HSC production from hemogenic endothelial cells (HEC). While several distinct regulators are involved in this process, it is not yet known whether macroautophagy (autophagy) plays a role in hematopoiesis in the pre-liver stage. Here, we show that different states of autophagy exist in hematopoietic precursors and correlate with hematopoietic potential based on the LC3-RFP-EGFP mouse model. Deficiency of autophagy-related gene 5 (Atg5) specifically in endothelial cells disrupts endothelial to hematopoietic transition (EHT), by blocking the autophagic process. Using combined approaches, including single-cell RNA-sequencing (scRNA-seq), we have confirmed that Atg5 deletion interrupts developmental temporal order of EHT to further affect the pre-HSC I maturation, and that autophagy influences hemogenic potential of HEC and the formation of pre-HSC I likely via the nucleolin pathway. These findings demonstrate a role for autophagy in the formation/maturation of hematopoietic precursors.