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The yolk sac (YS) generates the first blood and immune cells and provides nutritional and metabolic support to the developing embryo. Our current understanding of its functions derives from pivotal studies in model systems and insights from human studies are limited. Single-cell genomics technologies have facilitated the interrogation of human developmental tissues at unprecedented resolution. Atlases of blood and immune cells from multiple organs have been greatly enhanced by focused, time-resolved analyses of specific tissues.

Hematopoietic stem cells (HSCs) are a rare hematopoietic cell type that can entirely reconstitute the blood and immune systems following transplantation. Allogeneic HSC transplantation (HSCT) is used clinically as a curative therapy for a range of hematolymphoid diseases, but remains a high-risk therapy due to potential side effects including poor graft function and graft 6 vs-host disease (GvHD). Ex vivo HSC expansion has been suggested as an approach to improve hematopoietic reconstitution from low-cell dose grafts. Here, we demonstrate that we can improve the selectivity of polyvinyl alcohol (PVA)-based mouse HSC cultures through the use of physioxic culture conditions. Single-cell transcriptomic analysis confirmed inhibition of lineage-committed progenitor cells in physioxic cultures. Long-term physioxic expansion also afforded culture-based ex vivo HSC selection from whole bone marrow, spleen, and embryonic tissues. Furthermore, we provide evidence that HSC-selective ex vivo cultures deplete GvHD 13 causing T cells and that this approach can be combined with genotoxic-free antibody-based conditioning HSCT approaches. Our results offer a simple approach to improve PVA-based HSC cultures and the underlying molecular phenotype, as well as highlight the potential translational implications of selective HSC expansion systems for allogeneic HSCT.

Hematopoietic aging is marked by a loss of regenerative capacity and skewed differentiation from hematopoietic stem cells (HSC) leading to impaired blood production. Signals from the bone marrow (BM) niche tailor blood production, but the contribution of the old niche to hematopoietic aging remains unclear. Here, we characterize the inflammatory milieu that drives both niche and hematopoietic remodeling. We find decreased numbers and functionality of osteoprogenitors (OPr) at the endosteum and expansion of central marrow LepR+ mesenchymal stromal cells (MSC-L) associated with deterioration of the sinusoidal vasculature, which together create a degraded and inflamed old BM niche. Niche inflammation, in turn, drives chronic activation of emergency myelopoiesis pathways in old HSCs and multipotent progenitors (MPP), which promotes myeloid differentiation at the expense of lymphoid and erythroid commitment and hinders hematopoietic regeneration. Moreover, we show how production of IL-1 by the damaged endosteum acts in trans to drive the proinflammatory nature of the central marrow with damaging consequences for the old blood system. Remarkably, niche deterioration, HSC dysfunction, and defective regeneration can all be ameliorated by blocking IL-1 signaling. Our results demonstrate that targeting IL-1 as a key mediator of niche inflammation is a tractable strategy to improve blood production during aging

Hematopoietic aging is marked by a loss of regenerative capacity and skewed differentiation from hematopoietic stem cells (HSC) leading to impaired blood production. Signals from the bone marrow (BM) niche tailor blood production, but the contribution of the old niche to hematopoietic aging remains unclear. Here, we characterize the inflammatory milieu that drives both niche and hematopoietic remodeling. We find decreased numbers and functionality of osteoprogenitors (OPr) at the endosteum and expansion of central marrow LepR+ mesenchymal stromal cells (MSC-L) associated with deterioration of the sinusoidal vasculature, which together create a degraded and inflamed old BM niche. Niche inflammation, in turn, drives chronic activation of emergency myelopoiesis pathways in old HSCs and multipotent progenitors (MPP), which promotes myeloid differentiation and hinders hematopoietic regeneration. Moreover, we show how production of IL- 1b by the damaged endosteum acts in trans to drive the proinflammatory nature of the central marrow with damaging consequences for the old blood system. Remarkably, niche deterioration, HSC dysfunction, and defective regeneration can all be ameliorated by blocking IL-1 signaling. Our results demonstrate that targeting IL-1 as a key mediator of niche inflammation is a tractable strategy to improve blood production during aging.

Funder: Royal Australasian College of Physicians, Funder: NIHR, Funder: UKRI, Funder: Chief Scientist Office, Clotting Factor V (FV) is primarily synthesized in the liver and when cleaved by thrombin forms pro-coagulant Factor Va (FVa). Using whole blood RNAseq and scRNAseq of peripheral blood mononuclear cells, we find that FV mRNA is expressed in leukocytes, and identify neutrophils, monocytes, and T regulatory cells as sources of increased FV in hospitalized patients with COVID-19. Proteomic analysis confirms increased FV in circulating neutrophils in severe COVID-19, and immunofluorescence microscopy identifies FV in lung-infiltrating leukocytes in COVID-19 lung disease. Increased leukocyte FV expression in severe disease correlates with T-cell lymphopenia. Both plasma-derived and a cleavage resistant recombinant FV, but not thrombin cleaved FVa, suppress T-cell proliferation in vitro. Anticoagulants that reduce FV conversion to FVa, including heparin, may have the unintended consequence of suppressing the adaptive immune system.

The sympathetic nervous system has been evolutionary selected to respond to stress and activates haematopoietic stem cells via noradrenergic signals. However, the pathways preserving haematopoietic stem cell quiescence and maintenance under proliferative stress remain largely unknown. Here we found that cholinergic signals preserve haematopoietic stem cell quiescence in bone-associated (endosteal) bone marrow niches. Bone marrow cholinergic neural signals increase during stress haematopoiesis and are amplified through cholinergic osteoprogenitors. Lack of cholinergic innervation impairs balanced responses to chemotherapy or irradiation and reduces haematopoietic stem cell quiescence and self-renewal. Cholinergic signals activate α7 nicotinic receptor in bone marrow mesenchymal stromal cells leading to increased CXCL12 expression and haematopoietic stem cell quiescence. Consequently, nicotine exposure increases endosteal haematopoietic stem cell quiescence in vivo and impairs hematopoietic regeneration after haematopoietic stem cell transplantation in mice. In humans, smoking history is associated with delayed normalisation of platelet counts after allogeneic haematopoietic stem cell transplantation. These results suggest that cholinergic signals preserve stem cell quiescence under proliferative stress., The sympathetic nervous system has been shown to respond to stress and activate haematopoietic stem cells. Here they show that cholinergic signals in the bone marrow preserve haematopoietic stem cell quiescence and self-renewal under proliferative stress.

International audience; In 2016, the European Hematology Association (EHA) published the EHA Roadmap for European Hematology Research1 aiming to highlight achievements in the diagnostics and treatment of blood disorders, and to better inform European policy makers and other stakeholders about the urgent clinical and scientific needs and priorities in the field of hematology. Each section was coordinated by 1–2 section editors who were leading international experts in the field. In the 5 years that have followed, advances in the field of hematology have been plentiful. As such, EHA is pleased to present an updated Research Roadmap, now including 11 sections, each of which will be published separately. The updated EHA Research Roadmap identifies the most urgent priorities in hematology research and clinical science, therefore supporting a more informed, focused, and ideally a more funded future for European hematology research. The 11 EHA Research Roadmap sections include Normal Hematopoiesis; Malignant Lymphoid Diseases; Malignant Myeloid Diseases; Anemias and Related Diseases; Platelet Disorders; Blood Coagulation and Hemostatic Disorders; Transfusion Medicine; Infections in Hematology; Hematopoietic Stem Cell Transplantation; CAR-T and Other Cell-based Immune Therapies; and Gene Therapy.

The Human Developmental Cell Atlas (HDCA) initiative, part of the Human Cell Atlas, aims to create a comprehensive reference map of cells during development. This will be critical to understand normal organogenesis, the impact of mutations, environmental factors and infectious agents on human development, its relevance to congenital and childhood disorders, and the cellular basis of ageing, cancer and regenerative medicine. In this perspective, we outline the HDCA initiative and the challenges of mapping and modelling human development using state-of-the-art technologies in order to create a reference atlas across gestation. Like the Human Genome Project, the HDCA project will integrate the output from a growing community of scientists mapping human development into a unified atlas. We describe the early milestones achieved and the use of human stem cell-derived cultures, organoids and animal models to inform the HDCA, especially for prenatal tissues that are hard to acquire. Finally, we provide a roadmap towards a complete atlas of human development.

It is not fully understood why COVID-19 is typically milder in children To examine differences in response to SARS-CoV-2 infection in children and adults, we analysed paediatric and adult COVID-19 patients and healthy controls (total n=93) using single-cell multi-omic profiling of matched nasal, tracheal, bronchial and blood samples. In healthy paediatric airways, we observed cells already in an interferon-activated state, that upon SARS-CoV-2 infection was further induced especially in airway immune cells. We postulate that higher paediatric innate interferon-responses restrict viral replication and disease progression. The systemic response in children was characterised by increases in naive lymphocytes and a depletion of natural killer cells, while in adults cytotoxic T cells and interferon-stimulated subpopulations were significantly increased. We provide evidence that dendritic cells initiate interferon signaling in early infection, and identify novel epithelial cell states that associate with COVID-19 and age. Our matching nasal and blood data showed a strong interferon response in the airways with the induction of systemic interferon-stimulated populations, which were massively reduced in paediatric patients. Together, we provide several mechanisms that explain the milder clinical syndrome observed in children.

Frameshift mutations in CALR (calreticulin) are associated with essential thrombocythaemia (ET), but the stages at and mechanisms by which mutant CALR drives transformation remain incompletely defined. Here, we use single-cell approaches to examine the haematopoietic stem/progenitor cell (HSPC) landscape in a mouse model of mutant CALR-driven ET. We identify a trajectory linking HSCs with megakaryocytes and prospectively identify a novel intermediate population that is overrepresented in the disease state. We also show that mutant CALR drives transformation primarily from the earliest stem cell compartment, with some contribution from megakaryocyte progenitors. Finally, relative to wild-type HSCs, mutant CALR HSCs show increases in JAK-STAT signalling, the unfolded protein response, cell cycle, and a previously undescribed upregulation of cholesterol biosynthesis. Overall, we have identified a novel megakaryocyte-biased cell population that is increased in a mouse model of ET and described transcriptomic changes linking CALR mutations to increased HSC proliferation and megakaryopoiesis., Work in the Göttgens lab is supported by the Medical Research Council (MR/M008975/1), Wellcome (206328/Z/17/Z), Blood Cancer UK (18002), and Cancer Research UK (RG83389, jointly with A.R.G). Work in the Green lab is supported by Wellcome (RG74909), WBH Foundation (RG91681), and Cancer Research UK (RG83389, jointly with B.G.).

Technological advances play a key role in furthering our understanding of stem cell biology, and advancing the prospects of regenerative therapies. Highly parallelized methods, developed in the last decade, can profile DNA, RNA or proteins in thousands of cells and even capture data across two or more modalities (multi-omics). This allows unbiased and precise definition of molecular cell states, thus allowing classification of cell types, tracking of differentiation trajectories and discovery of underlying mechanisms. Despite being based on destructive techniques, novel experimental and bioinformatic approaches enable embedding and extraction of temporal information, which is essential for deconvolution of complex data and establishing cause and effect relationships. Here we provide an overview of recent studies pertinent to stem cell biology, followed by an outlook on how further advances in single cell molecular profiling and computational analysis have the potential to shape the future of both basic and translational research., Work in the Gottgens Laboratory is funded by grants from Wellcome Trust; MRC; Bloodwise; Cancer Research UK; National Institutes of Health (NIDDK DK106766); and core support grants by the Cancer Research UK Cambridge Centre and by Wellcome to the Cambridge Institute for Medical Research and Wellcome–Medical Research Council Cambridge Stem Cell Institute.

The murine developing epicardium heterogeneously expresses the transcription factors TCF21 and WT1. Here, we show that this cell heterogeneity is conserved in human epicardium, regulated by BNC1 and associated with cell fate and function. Single cell RNAseq of epicardium derived from human pluripotent stem cells (hPSC-epi) revealed that distinct epicardial sub-populations are defined by high levels of expression for the transcription factors BNC1 or TCF21. WT1+ cells are included in the BNC1+ population, which was confirmed in human foetal hearts. THY1 emerged as a membrane marker of the TCF21 population. We show that THY1+ cells can differentiate into cardiac fibroblast (CF) and smooth muscle cells (SMC), while THY1-cells were predominantly restricted to SMC. Knocking down BNC1 during the establishment of the epicardial populations resulted in a homogeneous, predominantly, TCF21high population. Network inference methods using transcriptomic data from the different cell lineages derived from the hPSC-epi, delivered a core transcriptional network organized around WT1, TCF21 and BNC1. This study is a step towards engineering sub-populations of epicardial cells with selective biological activities and unveils a list of epicardial regulators.