Temporal patterns of pre- and post-natal target organ damage associated with hypertensive pregnancy: a systematic review.

AIMS: Hypertensive pregnancy is associated with increased risks of developing a range of vascular disorders in later life. Understanding when hypertensive target organ damage first emerges could guide optimal timing of preventive interventions. This review identifies evidence of hypertensive target organ damage across cardiac, vascular, cerebral, and renal systems at different time points from pregnancy to postpartum. METHODS AND RESULTS: Systematic review of Ovid/MEDLINE, EMBASE, and ClinicalTrials.gov up to and including February 2023 including review of reference lists. Identified articles underwent evaluation via a synthesis without meta-analysis using a vote-counting approach based on direction of effect, regardless of statistical significance. Risk of bias was assessed for each outcome domain, and only higher quality studies were used for final analysis. From 7644 articles, 76 studies, including data from 1 742 698 pregnancies, were identified of high quality that reported either blood pressure trajectories or target organ damage during or after a hypertensive pregnancy. Left ventricular hypertrophy, white matter lesions, proteinuria, and retinal microvasculature changes were first evident in women during a hypertensive pregnancy. Cardiac, cerebral, and retinal changes were also reported in studies performed during the early and late post-partum period despite reduction in blood pressure early postpartum. Cognitive dysfunction was first reported late postpartum. CONCLUSION: The majority of target organ damage reported during a hypertensive pregnancy remains evident throughout the early and late post-partum period despite variation in blood pressure. Early peri-partum strategies may be required to prevent or reverse target organ damage in women who have had a hypertensive pregnancy.

This review identifies evidence of damage to the heart, brain, and blood vessels during and after hypertensive disorders of pregnancy and compares the pattern of changes that occur to blood pressure variations. Changes in the heart, brain, and blood vessels are first found in women during a hypertensive pregnancy and are also reported early after pregnancy. The majority of target organ damage reported remains evident long after pregnancy despite variation in blood pressure levels.

Preleukemic single-cell landscapes reveal mutation-specific mechanisms and gene programs predictive of AML patient outcomes.

Acute myeloid leukemia (AML) and myeloid neoplasms develop through acquisition of somatic mutations that confer mutation-specific fitness advantages to hematopoietic stem and progenitor cells. However, our understanding of mutational effects remains limited to the resolution attainable within immunophenotypically and clinically accessible bulk cell populations. To decipher heterogeneous cellular fitness to preleukemic mutational perturbations, we performed single-cell RNA sequencing of eight different mouse models with driver mutations of myeloid malignancies, generating 269,048 single-cell profiles. Our analysis infers mutation-driven perturbations in cell abundance, cellular lineage fate, cellular metabolism, and gene expression at the continuous resolution, pinpointing cell populations with transcriptional alterations associated with differentiation bias. We further develop an 11-gene scoring system (Stem11) on the basis of preleukemic transcriptional signatures that predicts AML patient outcomes. Our results demonstrate that a single-cell-resolution deep characterization of preleukemic biology has the potential to enhance our understanding of AML heterogeneity and inform more effective risk stratification strategies.

Yolk sac cell atlas reveals multiorgan functions during human early development.

The extraembryonic yolk sac (YS) ensures delivery of nutritional support and oxygen to the developing embryo but remains ill-defined in humans. We therefore assembled a comprehensive multiomic reference of the human YS from 3 to 8 postconception weeks by integrating single-cell protein and gene expression data. Beyond its recognized role as a site of hematopoiesis, we highlight roles in metabolism, coagulation, vascular development, and hematopoietic regulation. We reconstructed the emergence and decline of YS hematopoietic stem and progenitor cells from hemogenic endothelium and revealed a YS-specific accelerated route to macrophage production that seeds developing organs. The multiorgan functions of the YS are superseded as intraembryonic organs develop, effecting a multifaceted relay of vital functions as pregnancy proceeds.

Endothelial-specific Gata3 expression is required for hematopoietic stem cell generation.

To generate sufficient numbers of transplantable hematopoietic stem cells (HSCs) in vitro, a detailed understanding of how this process takes place in vivo is essential. The endothelial-to-hematopoietic transition (EHT), which culminates in the production of the first HSCs, is a highly complex process during which key regulators are switched on and off at precise moments, and that is embedded into a myriad of microenvironmental signals from surrounding cells and tissues. We have previously demonstrated an HSC-supportive function for GATA3 within the sympathetic nervous system and the sub-aortic mesenchyme, but show here that it also plays a cell-intrinsic role during the EHT. It is expressed in hemogenic endothelial cells and early HSC precursors, where its expression correlates with a more quiescent state. Importantly, endothelial-specific deletion of Gata3 shows that it is functionally required for these cells to mature into HSCs, placing GATA3 at the core of the EHT regulatory network.

How To Hospital: barriers to developing a patient 'Hospital Survival Guide' to support information transfer during ward-rounds on the patient journey from admission to hospital to discharge.

Clinicians can enable patients to actively participate in their care but communication with patients is often poor and highly variable. The aim of this study was to explore patients' understanding of their current illness while in hospital and using a codesign process to create prototype tools to facilitate better communication during ward rounds.A mixed-methods, multistep design with step 1: Application of a questionnaire addressing domains of care in the acute medical unit; step 2: Development of communication aids that were codesigned with active help of patients, students and a specialist in user centric design to address patient needs and step 3: Evaluation of tools with patients in four Plan-Do-Study-Act cycles.In the initial survey of 30 patients 12 (40%) patients did not know what their diagnosis was and 5 (17%) did not know the results of recent key tests. 20 (67%) patients felt that staff communication and coordination could be improved.An intervention was prototyped with four variations: (1) An A6 ward-round summary sheet completed by doctors during ward rounds. The system worked well but was highly person dependent. (2) An A4 patient-owned diary ('How to Hospital') that contained information about key processes in hospital and space to document conversations from rounds and prompts for questions. 10 patients read the diary and commented favourably but did not complete any pages. (3) 'Diary-cards': a basic set of information cards was given to patients on admission to hospital. (4) Patient specific 'diary-cards' were completed by clinicians-10 forms were piloted during rounds and improved subsequent day information retention of diagnosis to 80%.Our study identified interventions that were feasible but remained person-dependent. The patients' ownership of information in relation to their care might facilitate retention and satisfaction but the optimal format for these interventions for enhancing communication remains unclear.

Interactions between lineage-associated transcription factors govern haematopoietic progenitor states.

Recent advances in molecular profiling provide descriptive datasets of complex differentiation landscapes including the haematopoietic system, but the molecular mechanisms defining progenitor states and lineage choice remain ill-defined. Here, we employed a cellular model of murine multipotent haematopoietic progenitors (Hoxb8-FL) to knock out 39 transcription factors (TFs) followed by RNA-Seq analysis, to functionally define a regulatory network of 16,992 regulator/target gene links. Focussed analysis of the subnetworks regulated by the B-lymphoid TF Ebf1 and T-lymphoid TF Gata3 revealed a surprising role in common activation of an early myeloid programme. Moreover, Gata3-mediated repression of Pax5 emerges as a mechanism to prevent precocious B-lymphoid differentiation, while Hox-mediated activation of Meis1 suppresses myeloid differentiation. To aid interpretation of large transcriptomics datasets, we also report a new method that visualises likely transitions that a progenitor will undergo following regulatory network perturbations. Taken together, this study reveals how molecular network wiring helps to establish a multipotent progenitor state, with experimental approaches and analysis tools applicable to dissecting a broad range of both normal and perturbed cellular differentiation landscapes.

Lysine acetyltransferase Tip60 is required for hematopoietic stem cell maintenance.

Hematopoietic stem cells (HSCs) have the potential to replenish the blood system for the lifetime of the organism. Their 2 defining properties, self-renewal and differentiation, are tightly regulated by the epigenetic machineries. Using conditional gene-knockout models, we demonstrated a critical requirement of lysine acetyltransferase 5 (Kat5, also known as Tip60) for murine HSC maintenance in both the embryonic and adult stages, which depends on its acetyltransferase activity. Genome-wide chromatin and transcriptome profiling in murine hematopoietic stem and progenitor cells revealed that Tip60 colocalizes with c-Myc and that Tip60 deletion suppress the expression of Myc target genes, which are associated with critical biological processes for HSC maintenance, cell cycling, and DNA repair. Notably, acetylated H2A.Z (acH2A.Z) was enriched at the Tip60-bound active chromatin, and Tip60 deletion induced a robust reduction in the acH2A.Z/H2A.Z ratio. These results uncover a critical epigenetic regulatory layer for HSC maintenance, at least in part through Tip60-dependent H2A.Z acetylation to activate Myc target genes.

Single-cell chromatin accessibility maps reveal regulatory programs driving early mouse organogenesis.

During mouse embryonic development, pluripotent cells rapidly divide and diversify, yet the regulatory programs that define the cell repertoire for each organ remain ill-defined. To delineate comprehensive chromatin landscapes during early organogenesis, we mapped chromatin accessibility in 19,453 single nuclei from mouse embryos at 8.25 days post-fertilization. Identification of cell-type-specific regions of open chromatin pinpointed two TAL1-bound endothelial enhancers, which we validated using transgenic mouse assays. Integrated gene expression and transcription factor motif enrichment analyses highlighted cell-type-specific transcriptional regulators. Subsequent in vivo experiments in zebrafish revealed a role for the ETS factor FEV in endothelial identity downstream of ETV2 (Etsrp in zebrafish). Concerted in vivo validation experiments in mouse and zebrafish thus illustrate how single-cell open chromatin maps, representative of a mammalian embryo, provide access to the regulatory blueprint for mammalian organogenesis.

Identification of gene specific cis-regulatory elements during differentiation of mouse embryonic stem cells: An integrative approach using high-throughput datasets.

Gene expression governs cell fate, and is regulated via a complex interplay of transcription factors and molecules that change chromatin structure. Advances in sequencing-based assays have enabled investigation of these processes genome-wide, leading to large datasets that combine information on the dynamics of gene expression, transcription factor binding and chromatin structure as cells differentiate. While numerous studies focus on the effects of these features on broader gene regulation, less work has been done on the mechanisms of gene-specific transcriptional control. In this study, we have focussed on the latter by integrating gene expression data for the in vitro differentiation of murine ES cells to macrophages and cardiomyocytes, with dynamic data on chromatin structure, epigenetics and transcription factor binding. Combining a novel strategy to identify communities of related control elements with a penalized regression approach, we developed individual models to identify the potential control elements predictive of the expression of each gene. Our models were compared to an existing method and evaluated using the existing literature and new experimental data from embryonic stem cell differentiation reporter assays. Our method is able to identify transcriptional control elements in a gene specific manner that reflect known regulatory relationships and to generate useful hypotheses for further testing.

The Transcription Factor ERG Regulates Super-Enhancers Associated With an Endothelial-Specific Gene Expression Program.

RATIONALE: The ETS (E-26 transformation-specific) transcription factor ERG (ETS-related gene) is essential for endothelial homeostasis, driving expression of lineage genes and repressing proinflammatory genes. Loss of ERG expression is associated with diseases including atherosclerosis. ERG's homeostatic function is lineage-specific, because aberrant ERG expression in cancer is oncogenic. The molecular basis for ERG lineage-specific activity is unknown. Transcriptional regulation of lineage specificity is linked to enhancer clusters (super-enhancers). OBJECTIVE: To investigate whether ERG regulates endothelial-specific gene expression via super-enhancers. METHODS AND RESULTS: Chromatin immunoprecipitation with high-throughput sequencing in human umbilical vein endothelial cells showed that ERG binds 93% of super-enhancers ranked according to H3K27ac, a mark of active chromatin. These were associated with endothelial genes such as DLL4 (Delta-like protein 4), CLDN5 (claudin-5), VWF (von Willebrand factor), and CDH5 (VE-cadherin). Comparison between human umbilical vein endothelial cell and prostate cancer TMPRSS2 (transmembrane protease, serine-2):ERG fusion-positive human prostate epithelial cancer cell line (VCaP) cells revealed distinctive lineage-specific transcriptome and super-enhancer profiles. At a subset of endothelial super-enhancers (including DLL4 and CLDN5), loss of ERG results in significant reduction in gene expression which correlates with decreased enrichment of H3K27ac and MED (Mediator complex subunit)-1, and reduced recruitment of acetyltransferase p300. At these super-enhancers, co-occupancy of GATA2 (GATA-binding protein 2) and AP-1 (activator protein 1) is significantly lower compared with super-enhancers that remained constant following ERG inhibition. These data suggest distinct mechanisms of super-enhancer regulation in endothelial cells and highlight the unique role of ERG in controlling a core subset of super-enhancers. Most disease-associated single nucleotide polymorphisms from genome-wide association studies lie within noncoding regions and perturb transcription factor recognition sequences in relevant cell types. Analysis of genome-wide association studies data shows significant enrichment of risk variants for cardiovascular disease and other diseases, at ERG endothelial enhancers and super-enhancers. CONCLUSIONS: The transcription factor ERG promotes endothelial homeostasis via regulation of lineage-specific enhancers and super-enhancers. Enrichment of cardiovascular disease-associated single nucleotide polymorphisms at ERG super-enhancers suggests that ERG-dependent transcription modulates disease risk.

A single-cell hematopoietic landscape resolves 8 lineage trajectories and defects in Kit mutant mice.

Hematopoietic stem and progenitor cells (HSPCs) maintain the adult blood system, and their dysregulation causes a multitude of diseases. However, the differentiation journeys toward specific hematopoietic lineages remain ill defined, and system-wide disease interpretation remains challenging. Here, we have profiled 44 802 mouse bone marrow HSPCs using single-cell RNA sequencing to provide a comprehensive transcriptional landscape with entry points to 8 different blood lineages (lymphoid, megakaryocyte, erythroid, neutrophil, monocyte, eosinophil, mast cell, and basophil progenitors). We identified a common basophil/mast cell bone marrow progenitor and characterized its molecular profile at the single-cell level. Transcriptional profiling of 13 815 HSPCs from the c-Kit mutant (W41/W41) mouse model revealed the absence of a distinct mast cell lineage entry point, together with global shifts in cell type abundance. Proliferative defects were accompanied by reduced Myc expression. Potential compensatory processes included upregulation of the integrated stress response pathway and downregulation of proapoptotic gene expression in erythroid progenitors, thus providing a template of how large-scale single-cell transcriptomic studies can bridge between molecular phenotypes and quantitative population changes.

Critical Modulation of Hematopoietic Lineage Fate by Hepatic Leukemia Factor.

A gradual restriction in lineage potential of multipotent stem/progenitor cells is a hallmark of adult hematopoiesis, but the underlying molecular events governing these processes remain incompletely understood. Here, we identified robust expression of the leukemia-associated transcription factor hepatic leukemia factor (Hlf) in normal multipotent hematopoietic progenitors, which was rapidly downregulated upon differentiation. Interference with its normal downregulation revealed Hlf as a strong negative regulator of lymphoid development, while remaining compatible with myeloid fates. Reciprocally, we observed rapid lymphoid commitment upon reduced Hlf activity. The arising phenotypes resulted from Hlf binding to active enhancers of myeloid-competent cells, transcriptional induction of myeloid, and ablation of lymphoid gene programs, with Hlf induction of nuclear factor I C (Nfic) as a functionally relevant target gene. Thereby, our studies establish Hlf as a key regulator of the earliest lineage-commitment events at the transition from multipotency to lineage-restricted progeny, with implications for both normal and malignant hematopoiesis.

Dynamic Gene Regulatory Networks Drive Hematopoietic Specification and Differentiation.

Metazoan 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.

An experimentally validated network of nine haematopoietic transcription factors reveals mechanisms of cell state stability.

Transcription factor (TF) networks determine cell-type identity by establishing and maintaining lineage-specific expression profiles, yet reconstruction of mammalian regulatory network models has been hampered by a lack of comprehensive functional validation of regulatory interactions. Here, we report comprehensive ChIP-Seq, transgenic and reporter gene experimental data that have allowed us to construct an experimentally validated regulatory network model for haematopoietic stem/progenitor cells (HSPCs). Model simulation coupled with subsequent experimental validation using single cell expression profiling revealed potential mechanisms for cell state stabilisation, and also how a leukaemogenic TF fusion protein perturbs key HSPC regulators. The approach presented here should help to improve our understanding of both normal physiological and disease processes.

Integrated genome-scale analysis of the transcriptional regulatory landscape in a blood stem/progenitor cell model.

Comprehensive study of transcriptional control processes will be required to enhance our understanding of both normal and malignant hematopoiesis. Modern sequencing technologies have revolutionized our ability to generate genome-scale expression and histone modification profiles, transcription factor (TF)-binding maps, and also comprehensive chromatin-looping information. Many of these technologies, however, require large numbers of cells, and therefore cannot be applied to rare hematopoietic stem/progenitor cell (HSPC) populations. The stem cell factor-dependent multipotent progenitor cell line HPC-7 represents a well-recognized cell line model for HSPCs. Here we report genome-wide maps for 17 TFs, 3 histone modifications, DNase I hypersensitive sites, and high-resolution promoter-enhancer interactomes in HPC-7 cells. Integrated analysis of these complementary data sets revealed TF occupancy patterns of genomic regions involved in promoter-anchored loops. Moreover, preferential associations between pairs of TFs bound at either ends of chromatin loops led to the identification of 4 previously unrecognized protein-protein interactions between key blood stem cell regulators. All HPC-7 data sets are freely available both through standard repositories and a user-friendly Web interface. Together with previously generated genome-wide data sets, this study integrates HPC-7 data into a genomic resource on par with ENCODE tier 1 cell lines and, importantly, is the only current model with comprehensive genome-scale data that is relevant to HSPC biology.

Cytokine-induced megakaryocytic differentiation is regulated by genome-wide loss of a uSTAT transcriptional program.

Metazoan development is regulated by transcriptional networks, which must respond to extracellular cues including cytokines. The JAK/STAT pathway is a highly conserved regulatory module, activated by many cytokines, in which tyrosine-phosphorylated STATs (pSTATs) function as transcription factors. However, the mechanisms by which STAT activation modulates lineage-affiliated transcriptional programs are unclear. We demonstrate that in the absence of thrombopoietin (TPO), tyrosine-unphosphorylated STAT5 (uSTAT5) is present in the nucleus where it colocalizes with CTCF and represses a megakaryocytic transcriptional program. TPO-mediated phosphorylation of STAT5 triggers its genome-wide relocation to STAT consensus sites with two distinct transcriptional consequences, loss of a uSTAT5 program that restrains megakaryocytic differentiation and activation of a canonical pSTAT5-driven program which includes regulators of apoptosis and proliferation. Transcriptional repression by uSTAT5 reflects restricted access of the megakaryocytic transcription factor ERG to target genes. These results identify a previously unrecognized mechanism of cytokine-mediated differentiation.

CODEX: a next-generation sequencing experiment database for the haematopoietic and embryonic stem cell communities.

CODEX (http://codex.stemcells.cam.ac.uk/) is a user-friendly database for the direct access and interrogation of publicly available next-generation sequencing (NGS) data, specifically aimed at experimental biologists. In an era of multi-centre genomic dataset generation, CODEX provides a single database where these samples are collected, uniformly processed and vetted. The main drive of CODEX is to provide the wider scientific community with instant access to high-quality NGS data, which, irrespective of the publishing laboratory, is directly comparable. CODEX allows users to immediately visualize or download processed datasets, or compare user-generated data against the database's cumulative knowledge-base. CODEX contains four types of NGS experiments: transcription factor chromatin immunoprecipitation coupled to high-throughput sequencing (ChIP-Seq), histone modification ChIP-Seq, DNase-Seq and RNA-Seq. These are largely encompassed within two specialized repositories, HAEMCODE and ESCODE, which are focused on haematopoiesis and embryonic stem cell samples, respectively. To date, CODEX contains over 1000 samples, including 221 unique TFs and 93 unique cell types. CODEX therefore provides one of the most complete resources of publicly available NGS data for the direct interrogation of transcriptional programmes that regulate cellular identity and fate in the context of mammalian development, homeostasis and disease.