SummaryMetazoan development involves the successive activation and silencing of specific gene expression programs and is driven by tissue-specific transcription factors programming the chromatin landscape. To understand how this process executes an entire developmental pathway, we generated global gene expression, chromatin accessibility, histone modification, and transcription factor binding data from purified embryonic stem cell-derived cells representing six sequential stages of hematopoietic specification and differentiation. Our data reveal the nature of regulatory elements driving differential gene expression and inform how transcription factor binding impacts on promoter activity. We present a dynamic core regulatory network model for hematopoietic specification and demonstrate its utility for the design of reprogramming experiments. Functional studies motivated by our genome-wide data uncovered a stage-specific role for TEAD/YAP factors in mammalian hematopoietic specification. Our study presents a powerful resource for studying hematopoiesis and demonstrates how such data advance our understanding of mammalian development.

SummaryRecent reports have shown that somatic cells, under appropriate culture conditions, could be directly reprogrammed to cardiac, hepatic, or neuronal phenotype by lineage-specific transcription factors. In this study, we demonstrate that both embryonic and adult somatic fibroblasts can be efficiently reprogrammed to clonal multilineage hematopoietic progenitors by the ectopic expression of the transcription factors ERG, GATA2, LMO2, RUNX1c, and SCL. These reprogrammed cells were stably expanded on stromal cells and possessed short-term reconstitution ability in vivo. Loss of p53 function facilitated reprogramming to blood, and p53−/− reprogrammed cells efficiently generated erythroid, megakaryocytic, myeloid, and lymphoid lineages. Genome-wide analyses revealed that generation of hematopoietic progenitors was preceded by the appearance of hemogenic endothelial cells expressing endothelial and hematopoietic genes. Altogether, our findings suggest that direct reprogramming could represent a valid alternative approach to the differentiation of embryonic stem cells (ESCs) or induced pluripotent stem cells (iPSCs) for disease modeling and autologous blood cell therapies.

Primitive erythroblasts are the first blood cells generated during embryonic hematopoiesis. Tracking their emergence both in vivo and in vitro has remained challenging due to the lack of specific cell surface markers. To selectively investigate primitive erythropoiesis, we have engineered a new transgenic embryonic stem (ES) cell line, where eGFP expression is driven by the regulatory sequences of the embryonic βH1 hemoglobin gene expressed specifically in primitive erythroid cells. Using this ES cell line, we observed that the first primitive erythroblasts are detected in vitro around day 1.5 of blast colony differentiation, within the cell population positive for the early hematopoietic progenitor marker CD41. Moreover, we establish that these eGFP+ cells emerge from a hemogenic endothelial cell population similarly to their definitive hematopoietic counterparts. We further generated a corresponding βH1-eGFP transgenic mouse model and demonstrated the presence of a primitive erythroid primed hemogenic endothelial cell population in the developing embryo. Taken together, our findings demonstrate that both in vivo and in vitro primitive erythrocytes are generated from hemogenic endothelial cells.

The first hematopoietic cells are generated very early in ontogeny to support the growth of the embryo and to provide the foundation to the adult hematopoietic system. There is a considerable therapeutic interest in understanding how these first blood cells are generated in order to try to reproduce this process in vitro. This would allow generating blood products, or hematopoietic cell populations from embryonic stem (ES) cells, induced pluripotent stem cells or through directed reprogramming. Recent studies have clearly established that the first hematopoietic cells originate from a hemogenic endothelium (HE) through an endothelial to hematopoietic transition (EHT). The molecular mechanisms underlining this transition remain largely unknown with the exception that the transcription factor RUNX1 is critical for this process. In this Extra Views report, we discuss our recent studies demonstrating that the transcriptional repressors GFI1 and GFI1B have a critical role in the EHT. We established that these RUNX1 transcriptional targets are actively implicated in the downregulation of the endothelial program and the loss of endothelial identity during the formation of the first blood cells. In addition, our results suggest that GFI1 expression provides an ideal novel marker to identify, isolate and study the HE cell population.

Hematopoietic cell-based therapies are currently available treatment options for many hematological and nonhematological disorders. However, the scarcity of allogeneic donor-derived cells is a major hurdle in treating these disorders. Embryonic stem cell-based directed differentiation and direct reprogramming of somatic cells provide excellent tools for the potential generation of hematopoietic stem cells usable in the clinic for cellular therapies. In addition to blood stem cell transplantation, mature blood cells such as red blood cells, platelets, and engineered T cells have also been increasingly used to treat several diseases. Besides cellular therapies, induced blood progenitor cells generated from autologous sources (either induced pluripotent stem cells or somatic cells) can be useful for disease modeling of bone marrow failures and acquired blood disorders. However, although great progress has been made toward these goals, we are still far from the use of in vitro-derived blood products in the clinic. We review the current state of knowledge on the directed differentiation of embryonic stem cells and the reprogramming of somatic cells toward the generation of blood stem cells and derivatives. STEM CELLS TRANSLATIONAL MEDICINE 2016;5:1330-1337 SIGNIFICANCEHematopoietic cell-based therapies are currently available treatment options for many hematological and nonhematological disorders. However, the scarcity of allogeneic donor-derived cells is a major hurdle in treating these disorders. The current state of knowledge on the directed differentiation of embryonic stem cells and the reprogramming of somatic cells toward the generation of blood stem cells and derivatives is reviewed.

BackgroundTristetraprolin (TTP) is a key mediator of processes such as inflammation resolution, the inhibition of autoimmunity and in cancer. It carries out this role by the binding and degradation of mRNA transcripts, thereby decreasing their half-life. Transcripts modulated by TTP encode proteins such as cytokines, pro-inflammatory agents and immediate-early response proteins. TTP can also modulate neoplastic phenotypes in many cancers. TTP is induced and functionally regulated by a spectrum of both pro- and anti-inflammatory cytokines, mitogens and drugs in a MAPK-dependent manner. So far the contribution of p38 MAPK to the regulation of TTP expression and function has been best described.ResultsOur results demonstrate the induction of the gene coding TTP (ZFP36) by EGF through the ERK1/2-dependent pathway and implicates the transcription factor ELK-1 in this process. We show that ELK-1 regulates ZFP36 expression by two mechanisms: by binding the ZFP36 promoter directly through ETS-binding site (+ 883 to +905 bp) and by inducing expression of EGR-1, which in turn increases ZFP36 expression through sequences located between -111 and -103 bp.ConclusionsEGF activates TTP expression via ELK-1 and EGR-1 transcription factors.

Well into the second decade of the 21st century, the field of regenerative medicine is bursting with hopes and promises to heal young and old. The bespoken generation of cells is thought to offer unprecedented cures for a vast range of diseases. Haematological disorders have already benefited tremendously from stem cell therapy in the form of bone marrow transplantation. However, lack of compatible donors often means that patients remain on transplantation waiting lists for too long. The in vitro derivation of haematopoietic stem cells offers the possibility to generate tailor-made cells for the treatment of these patients. Promising approaches to generate in vitro-derived blood progenitors include the directed differentiation of pluripotent stem cells and the reprogramming of somatic cells.

Kinins, universal mediators of inflammation, are recognized by two kinds of receptors, B1 and B2, which have been found to be expressed in numerous cell types of several species. However, the knowledge of the regulation of these receptors in leukocytes is still not satisfactory. In the current work, we have demonstrated a constitutive production of B2 receptor mRNA in the human promonocyte U937 cells and its two-fold augmentation after cell differentiation with retinoic acid and phorbol ester. Bradykinin and des-Arg(10)-kallidin induced the expression of both B2 and B1 receptors in cells before and after differentiation. Generally, the undifferentiated cells were more susceptible to bradykinin-dependent induction of kinin receptors (increases by approximately 250% and 200% for B2 and B1 receptors, respectively). The induction, by approx. 200%, of B1 receptor by des-Arg(10)-kallidin was detected on both mRNA and protein levels. In addition, an unexpected strong induction of B2 receptor by this compound was observed in the retinoic acid- and phorbol ester-differentiated cells (by 150% and 200%, respectively) that suggests a possible autoregulation of kinin receptors by own agonists during the inflammatory state. On the other hand, a strong enhancement of the expression of both receptors by interleukin 1beta, especially in the phorbol ester-differentiated cells, indicates the involvement of kinin receptors in the propagation of the inflammatory processes.