Mapping molecular landmarks of human skeletal ontogeny and pluripotent stem cell-derived articular chondrocytes.

Tissue-specific gene expression defines cellular identity and function, but knowledge of early human development is limited, hampering application of cell-based therapies. Here we profiled 5 distinct cell types at a single fetal stage, as well as chondrocytes at 4 stages in vivo and 2 stages during in vitro differentiation. Network analysis delineated five tissue-specific gene modules; these modules and chromatin state analysis defined broad similarities in gene expression during cartilage specification and maturation in vitro and in vivo, including early expression and progressive silencing of muscle- and bone-specific genes.

Differentiation of human embryonic stem cells to HOXA hemogenic vasculature that resembles the aorta-gonad-mesonephros.

The ability to generate hematopoietic stem cells from human pluripotent cells would enable many biomedical applications. We find that hematopoietic CD34 cells in spin embryoid bodies derived from human embryonic stem cells (hESCs) lack HOXA expression compared with repopulation-competent human cord blood CD34 cells, indicating incorrect mesoderm patterning. Using reporter hESC lines to track the endothelial (SOX17) to hematopoietic (RUNX1C) transition that occurs in development, we show that simultaneous modulation of WNT and ACTIVIN signaling yields CD34 hematopoietic cells with HOXA expression that more closely resembles that of cord blood.

MEF2C protects bone marrow B-lymphoid progenitors during stress haematopoiesis.

DNA double strand break (DSB) repair is critical for generation of B-cell receptors, which are pre-requisite for B-cell progenitor survival. However, the transcription factors that promote DSB repair in B cells are not known. Here we show that MEF2C enhances the expression of DNA repair and recombination factors in B-cell progenitors, promoting DSB repair, V(D)J recombination and cell survival.

Critical requirement of VEGF-C in transition to fetal erythropoiesis.

Vascular endothelial growth factor C (VEGF-C) is a major driver of lymphangiogenesis in embryos and adults. Vegfc gene deletion in mouse embryos results in failure of lymphangiogenesis, fluid accumulation in tissues, and lethality. The VEGF-C receptors VEGFR3 and VEGFR2 are required for embryonic blood vessel formation.

Medial HOXA genes demarcate haematopoietic stem cell fate during human development.

Pluripotent stem cells (PSCs) may provide a potential source of haematopoietic stem/progenitor cells (HSPCs) for transplantation; however, unknown molecular barriers prevent the self-renewal of PSC-HSPCs. Using two-step differentiation, human embryonic stem cells (hESCs) differentiated in vitro into multipotent haematopoietic cells that had the CD34(+)CD38(-/lo)CD90(+)CD45(+)GPI-80(+) fetal liver (FL) HSPC immunophenotype, but exhibited poor expansion potential and engraftment ability. Transcriptome analysis of immunophenotypic hESC-HSPCs revealed that, despite their molecular resemblance to FL-HSPCs, medial HOXA genes remained suppressed.

Tracking HSC Origin: From Bench to Placenta.

Reporting in Developmental Cell, Pereira et al. (2016) use in vitro lineage reprogramming insights to inform understanding of hematopoietic stem cell (HSC) development in vivo. They find Prom1(+)Sca1(+)CD34(+)CD45(-) hemogenic precursors, akin to fibroblast-derived hemato-vascular precursors, in mouse placenta and embryo.

GLI2 inhibition abrogates human leukemia stem cell dormancy.

Background: Dormant leukemia stem cells (LSC) promote therapeutic resistance and leukemic progression as a result of unbridled activation of stem cell gene expression programs. Thus, we hypothesized that 1) deregulation of the hedgehog (Hh) stem cell self-renewal and cell cycle regulatory pathway would promote dormant human LSC generation and 2) that PF-04449913, a clinical antagonist of the GLI2 transcriptional activator, smoothened (SMO), would enhance dormant human LSC eradication.

Methods: To test these postulates, whole transcriptome RNA sequencing (RNA-seq), microarray, qRT-PCR, stromal co-culture, confocal fluorescence microscopic, nanoproteomic, serial transplantation and cell cycle analyses were performed on FACS purified normal, chronic phase (CP) chronic myeloid leukemia (CML), blast crisis (BC) phase CML progenitors with or without PF-04449913 treatment.

Scl binds to primed enhancers in mesoderm to regulate hematopoietic and cardiac fate divergence.

Scl/Tal1 confers hemogenic competence and prevents ectopic cardiomyogenesis in embryonic endothelium by unknown mechanisms. We discovered that Scl binds to hematopoietic and cardiac enhancers that become epigenetically primed in multipotent cardiovascular mesoderm, to regulate the divergence of hematopoietic and cardiac lineages. Scl does not act as a pioneer factor but rather exploits a pre-established epigenetic landscape.

GPI-80 defines self-renewal ability in hematopoietic stem cells during human development.

Advances in pluripotent stem cell and reprogramming technologies have given us the hope of generating hematopoietic stem cells (HSCs) in culture. To succeed, greater understanding of the self-renewing HSC during human development is required. We discovered that the glycophosphatidylinositol-anchored surface protein GPI-80 defines a subpopulation of human fetal liver hematopoietic stem/progenitor cells (HSPCs) with self-renewal ability.

The histone methyltransferase activity of MLL1 is dispensable for hematopoiesis and leukemogenesis.

Despite correlations between histone methyltransferase (HMT) activity and gene regulation, direct evidence that HMT activity is responsible for gene activation is sparse. We address the role of the HMT activity for MLL1, a histone H3 lysine 4 (H3K4) methyltransferase critical for maintaining hematopoietic stem cells (HSCs). Here, we show that the SET domain, and thus HMT activity of MLL1, is dispensable for maintaining HSCs and supporting leukemogenesis driven by the MLL-AF9 fusion oncoprotein.

Sex hormone drives blood stem cell reproduction.

Stem cells ensure the maintenance of tissue homeostasis throughout life by tightly regulating their self-renewal and differentiation. In a recent study published in Nature, Nakada et al, 2014 unveil an unexpected endocrine mechanism that regulates hematopoietic stem cell (HSC) self-renewal.

Progesterone receptor in the vascular endothelium triggers physiological uterine permeability preimplantation.

Vascular permeability is frequently associated with inflammation and is triggered by a cohort of secreted permeability factors such as vascular endothelial growth factor (VEGF). Here, we show that the physiological vascular permeability that precedes implantation is directly controlled by progesterone receptor (PR) and is independent of VEGF. Global or endothelial-specific deletion of PR blocks physiological vascular permeability in the uterus, whereas misexpression of PR in the endothelium of other organs results in ectopic vascular leakage.

c-Met-dependent multipotent labyrinth trophoblast progenitors establish placental exchange interface.

The placenta provides the interface for gas and nutrient exchange between the mother and the fetus. Despite its critical function in sustaining pregnancy, the stem/progenitor cell hierarchy and molecular mechanisms responsible for the development of the placental exchange interface are poorly understood. We identified an Epcam(hi) labyrinth trophoblast progenitor (LaTP) in mouse placenta that at a clonal level generates all labyrinth trophoblast subtypes, syncytiotrophoblasts I and II, and sinusoidal trophoblast giant cells.

Selective suppression of endothelial cytokine production by progesterone receptor.

Steroid hormones are well-recognized suppressors of the inflammatory response, however, their cell- and tissue-specific effects in the regulation of inflammation are far less understood, particularly for the sex-related steroids. To determine the contribution of progesterone in the endothelium, we have characterized and validated an in vitro culture system in which human umbilical vein endothelial cells constitutively express human progesterone receptor (PR). Using next generation RNA-sequencing, we identified a selective group of cytokines that are suppressed by progesterone both under physiological conditions and during pathological activation by lipopolysaccharide.

Haemogenic endocardium contributes to transient definitive haematopoiesis.

Haematopoietic cells arise from spatiotemporally restricted domains in the developing embryo. Although studies of non-mammalian animal and in vitro embryonic stem cell models suggest a close relationship among cardiac, endocardial and haematopoietic lineages, it remains unknown whether the mammalian heart tube serves as a haemogenic organ akin to the dorsal aorta. Here we examine the haemogenic activity of the developing endocardium.

Expansion on stromal cells preserves the undifferentiated state of human hematopoietic stem cells despite compromised reconstitution ability.

Lack of HLA-matched hematopoietic stem cells (HSC) limits the number of patients with life-threatening blood disorders that can be treated by HSC transplantation. So far, insufficient understanding of the regulatory mechanisms governing human HSC has precluded the development of effective protocols for culturing HSC for therapeutic use and molecular studies. We defined a culture system using OP9M2 mesenchymal stem cell (MSC) stroma that protects human hematopoietic stem/progenitor cells (HSPC) from differentiation and apoptosis.

Nipah virus envelope-pseudotyped lentiviruses efficiently target ephrinB2-positive stem cell populations in vitro and bypass the liver sink when administered in vivo.

Sophisticated retargeting systems for lentiviral vectors have been developed in recent years. Most seek to suppress the viral envelope's natural tropism while modifying the receptor-binding domain such that its tropism is determined by the specificity of the engineered ligand-binding motif. Here we took advantage of the natural tropism of Nipah virus (NiV), whose attachment envelope glycoprotein has picomolar affinity for ephrinB2, a molecule proposed as a molecular marker of "stemness" (present on embryonic, hematopoietic, and neural stem cells) as well as being implicated in tumorigenesis of specific cancers.

Characterization and therapeutic potential of induced pluripotent stem cell-derived cardiovascular progenitor cells.

Background: Cardiovascular progenitor cells (CPCs) have been identified within the developing mouse heart and differentiating pluripotent stem cells by intracellular transcription factors Nkx2.5 and Islet 1 (Isl1). Study of endogenous and induced pluripotent stem cell (iPSC)-derived CPCs has been limited due to the lack of specific cell surface markers to isolate them and conditions for their in vitro expansion that maintain their multipotency.

Combinatorial assembly of developmental stage-specific enhancers controls gene expression programs during human erythropoiesis.

Gene-distal enhancers are critical for tissue-specific gene expression, but their genomic determinants within a specific lineage at different stages of development are unknown. Here we profile chromatin state maps, transcription factor occupancy, and gene expression profiles during human erythroid development at fetal and adult stages. Comparative analyses of human erythropoiesis identify developmental stage-specific enhancers as primary determinants of stage-specific gene expression programs.

Dynamic distribution of linker histone H1.5 in cellular differentiation.

Linker histones are essential components of chromatin, but the distributions and functions of many during cellular differentiation are not well understood. Here, we show that H1.5 binds to genic and intergenic regions, forming blocks of enrichment, in differentiated human cells from all three embryonic germ layers but not in embryonic stem cells.

Lymphoid priming in human bone marrow begins before expression of CD10 with upregulation of L-selectin.

Expression of the cell-surface antigen CD10 has long been used to define the lymphoid commitment of human cells. Here we report a unique lymphoid-primed population in human bone marrow that was generated from hematopoietic stem cells (HSCs) before onset of the expression of CD10 and commitment to the B cell lineage. We identified this subset by high expression of the homing molecule L-selectin (CD62L).

Scl represses cardiomyogenesis in prospective hemogenic endothelium and endocardium.

Endothelium in embryonic hematopoietic tissues generates hematopoietic stem/progenitor cells; however, it is unknown how its unique potential is specified. We show that transcription factor Scl/Tal1 is essential for both establishing the hematopoietic transcriptional program in hemogenic endothelium and preventing its misspecification to a cardiomyogenic fate. Scl(-/-) embryos activated a cardiac transcriptional program in yolk sac endothelium, leading to the emergence of CD31+Pdgfrα+ cardiogenic precursors that generated spontaneously beating cardiomyocytes.

Trophoblasts regulate the placental hematopoietic niche through PDGF-B signaling.

The placenta is a hematopoietic organ that supports hematopoietic stem/progenitor cell (HSPC) generation and expansion without promoting differentiation. We identified PDGF-B signaling in trophoblasts as a key component of the unique placental hematopoietic microenvironment that protects HSPCs from premature differentiation. Loss of PDGF-B or its receptor, PDGFRβ, induced definitive erythropoiesis in placental labyrinth vasculature.

Defining the nature of human pluripotent stem cell progeny.

While it is clear that human pluripotent stem cells (hPSCs) can differentiate to generate a panoply of various cell types, it is unknown how closely in vitro development mirrors that which occurs in vivo. To determine whether human embryonic stem cells (hESCs) and human-induced pluripotent stem cells (hiPSCs) make equivalent progeny, and whether either makes cells that are analogous to tissue-derived cells, we performed comprehensive transcriptome profiling of purified PSC derivatives and their tissue-derived counterparts. Expression profiling demonstrated that hESCs and hiPSCs make nearly identical progeny for the neural, hepatic, and mesenchymal lineages, and an absence of re-expression from exogenous reprogramming factors in hiPSC progeny.

Return to youth with Sox17.

Maturation of hematopoietic stem cells (HSCs) from fetal to adult state and differentiation to progenitors are thought to follow a one-way street. In this issue of Genes & Development, He and colleagues (pp. 1613-1627) show that overexpression of Sox17 can convert adult multipotential progenitors to self-renewing HSCs that possess fetal properties.