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EOMES and T-BET are related T-box transcription factors that control natural killer (NK) cell development. Here we demonstrate that EOMES and T-BET regulate largely distinct gene sets during this process. EOMES is dominantly expressed in immature NK cells and drives early lineage specification by inducing hallmark receptors and functions. By contrast, T-BET is dominant in mature NK cells, where it induces responsiveness to IL-12 and represses the cell cycle, likely through transcriptional repressors. Regardless, many genes with distinct functions are co-regulated by the two transcription factors. By generating two gene-modified mice facilitating chromatin immunoprecipitation of endogenous EOMES and T-BET, we show a strong overlap in their DNA binding targets, as well as extensive epigenetic changes during NK cell differentiation. Our data thus suggest that EOMES and T-BET may distinctly govern, via differential expression and co-factors recruitment, NK cell maturation by inserting partially overlapping epigenetic regulations.

The objective of the study was to determine whether the effects of infarct-related artery (IRA) infusion of autologous bone marrow–derived CD34 + cells after ST elevation myocardial infarction (STEMI) are dependent on the dose (quantity and mobility) of the cells infused. Beneficial effects of IRA infusion of mononuclear cells after STEMI have been inconsistent, possibly because of differences in timing, cell type, quantity, and mobility of infused cells. Patients were randomized to bone marrow harvest (n = 16) or control (n = 15). At a median of 8.3 days after coronary stenting for STEMI, CD34 + cells were infused via the IRA at 3 dose levels (5, 10, and 15 × 10 6) in cohorts of 5 patients each. Baseline and follow-up imaging and ex vivo CD34 + cell mobility were performed. Cell harvest and infusion were safe. Quantitative rest hypoperfusion score measured by single-photon emission computed tomography improved at 6 months in the ≥10 million cohorts compared with controls (−256 vs +14, P = .02). There was a trend toward improved ejection fraction at 6 months (+4.5%) in the ≥10 million cohorts compared with no change in the controls and 5 million cohort (+0.7%). Improved perfusion and infarct size reduction correlated with the quantity and mobility of the infused CD34 + cells. The effects of CD34 + cell IRA infusion during the repair phase after STEMI are dose dependent and, at a threshold dose of 10 million CD34 + cells, associated with a significant improvement in perfusion that may limit deterioration in cardiac function (IRA infusion of CD34 + cells in patients with acute myocardial infarction [AMR-01] NCT00313339).

Immune rejection may limit the therapeutic use of induced pluripotent stem cells (iPSCs); here, terminally differentiated mouse iPSCs are shown to generate negligible immune rejection in their host. No immune reaction to iPSCs Induced pluripotent stem cells (iPSCs) derived from a patient's own somatic cells could have great therapeutic potential. The hope is that iPSC-derived differentiated cells would avoid any immunogenic responses. In this study, Masumi Abe and colleagues assess the immunogenicity of skin and bone marrow tissues derived from a large set of isogenic mouse embryonic stem cell and iPSC lines. Their results are consistent with negligible immune rejection by the host. The advantages of using induced pluripotent stem cells (iPSCs) instead of embryonic stem (ES) cells in regenerative medicine centre around circumventing concerns about the ethics of using ES cells and the likelihood of immune rejection of ES-cell-derived tissues 1 , 2 . However, partial reprogramming and genetic instabilities in iPSCs 3 , 4 , 5 , 6 could elicit immune responses in transplant recipients even when iPSC-derived differentiated cells are transplanted. iPSCs are first differentiated into specific types of cells in vitro for subsequent transplantation. Although model transplantation experiments have been conducted using various iPSC-derived differentiated tissues 7 , 8 , 9 , 10 and immune rejections have not been observed, careful investigation of the immunogenicity of iPSC-derived tissue is becoming increasingly critical, especially as this has not been the focus of most studies done so far. A recent study reported immunogenicity of iPSC- but not ES-cell-derived teratomas 11 and implicated several causative genes. Nevertheless, some controversy has arisen regarding these findings 12 . Here we examine the immunogenicity of differentiated skin and bone marrow tissues derived from mouse iPSCs. To ensure optimal comparison of iPSCs and ES cells, we established ten integration-free iPSC and seven ES-cell lines using an inbred mouse strain, C57BL/6. We observed no differences in the rate of success of transplantation when skin and bone marrow cells derived from iPSCs were compared with ES-cell-derived tissues. Moreover, we observed limited or no immune responses, including T-cell infiltration, for tissues derived from either iPSCs or ES cells, and no increase in the expression of the immunogenicity-causing Zg16 and Hormad1 genes in regressing skin and teratoma tissues. Our findings suggest limited immunogenicity of transplanted cells differentiated from iPSCs and ES cells.

Long-lived bone marrow plasma cells (BMPCs) are a persistent and essential source of protective antibodies 1 – 7 . Individuals who have recovered from COVID-19 have a substantially lower risk of reinfection with SARS-CoV-2 8 – 10 . Nonetheless, it has been reported that levels of anti-SARS-CoV-2 serum antibodies decrease rapidly in the first few months after infection, raising concerns that long-lived BMPCs may not be generated and humoral immunity against SARS-CoV-2 may be short-lived 11 – 13 . Here we show that in convalescent individuals who had experienced mild SARS-CoV-2 infections ( n  = 77), levels of serum anti-SARS-CoV-2 spike protein (S) antibodies declined rapidly in the first 4 months after infection and then more gradually over the following 7 months, remaining detectable at least 11 months after infection. Anti-S antibody titres correlated with the frequency of S-specific plasma cells in bone marrow aspirates from 18 individuals who had recovered from COVID-19 at 7 to 8 months after infection. S-specific BMPCs were not detected in aspirates from 11 healthy individuals with no history of SARS-CoV-2 infection. We show that S-binding BMPCs are quiescent, which suggests that they are part of a stable compartment. Consistently, circulating resting memory B cells directed against SARS-CoV-2 S were detected in the convalescent individuals. Overall, our results indicate that mild infection with SARS-CoV-2 induces robust antigen-specific, long-lived humoral immune memory in humans. SARS-CoV-2 infection induces long-lived bone marrow plasma cells that correlate with anti-SARS-CoV-2 spike protein antibody titres in individuals who have recovered from COVID-19.

Single-cell genomics technology has transformed our understanding of complex cellular systems. However, excessive cost and a lack of strategies for the purification of newly identified cell types impede their functional characterization and large-scale profiling. Here, we have generated high-content single-cell proteo-genomic reference maps of human blood and bone marrow that quantitatively link the expression of up to 197 surface markers to cellular identities and biological processes across all main hematopoietic cell types in healthy aging and leukemia. These reference maps enable the automatic design of cost-effective high-throughput cytometry schemes that outperform state-of-the-art approaches, accurately reflect complex topologies of cellular systems and permit the purification of precisely defined cell states. The systematic integration of cytometry and proteo-genomic data enables the functional capacities of precisely mapped cell states to be measured at the single-cell level. Our study serves as an accessible resource and paves the way for a data-driven era in cytometry. Haas, Velten and colleagues use single-cell multiomics of human blood and bone marrow to generate a reference map allowing the quantitative linking of cytometry and proteo-genomic information.

The REGENT-VSEL trial demonstrated a neutral effect of transendocardial injection of autologous bone marrow (BM)-derived CD133+ in regard to myocardial ischemia. The current sub-analysis of the REGENT VSEL trial aims to assess the effect stem cell therapy has on quality of life (QoL) in patients with refractory angina. Thirty-one patients (63.0 ± 6.4 years, 70% male) with recurrent CCS II-IV angina, despite optimal medical therapy, enrolled in the REGENT-VSEL single center, randomized, double-blinded, and placebo-controlled trial. Of the 31 patients, 16 individuals were randomly assigned to the active stem cell group and 15 individuals were randomly assigned to the placebo group on a 1:1 basis. The inducibility of ischemia, (≥ one myocardial segment) was confirmed for each patient using Tc-99m SPECT. QoL was measured using the Seattle Angina Questionnaire. Each patient completed the questionnaire prior to treatment and at the time of their outpatient follow-up visits at 1, 4, 6, and 12 months after cell/placebo treatment. The main finding of the REGENT-VSEL trial sub-analysis was that transendocardial injection of autologous BM-derived CD133+ stem cells in patients with chronic refractory angina did not show significant improvement in QoL in comparison to the control group. Moreover, there was no significant difference between cell therapy and placebo in a number of patients showing improvement of at least 1 Canadian Cardiovascular Society class during the follow-up period. Intra-myocardial delivery of autologous CD133+ stem cells is safe and feasible but does not show a significant improvement in the QoL or angina pectoris symptoms in patients with chronic myocardial ischemia.

The progenitor stage of commitment toward the conventional dendritic cell subsets and the transcriptional networks that control it remain poorly understood. Two articles from Ginhoux and colleagues and Murphy and colleagues offer insight into these processes. Mouse conventional dendritic cells (cDCs) can be classified into two functionally distinct lineages: the CD8α + (CD103 + ) cDC1 lineage, and the CD11b + cDC2 lineage. cDCs arise from a cascade of bone marrow (BM) DC-committed progenitor cells that include the common DC progenitors (CDPs) and pre-DCs, which exit the BM and seed peripheral tissues before differentiating locally into mature cDCs. Where and when commitment to the cDC1 or cDC2 lineage occurs remains poorly understood. Here we found that transcriptional signatures of the cDC1 and cDC2 lineages became evident at the single-cell level from the CDP stage. We also identified Siglec-H and Ly6C as lineage markers that distinguished pre-DC subpopulations committed to the cDC1 lineage (Siglec-H − Ly6C − pre-DCs) or cDC2 lineage (Siglec-H − Ly6C + pre-DCs). Our results indicate that commitment to the cDC1 or cDC2 lineage occurs in the BM and not in the periphery.

Key Points In steady-state conditions, blood monocyte subsets form in a developmental sequence with mouse LY6C hi monocytes giving rise to LY6C low monocytes. LY6C low monocytes act within the vasculature by surveying the vessel wall for injury and LY6C hi monocytes are recruited to sites of inflammation and, after extravasation, differentiate in the tissue into cells with dendritic cell and macrophage activities. Intestinal macrophages are continuously renewed by LY6C hi monocytes and thus differ from most other embryonic-derived tissue macrophages. LY6C hi monocytes are probably recruited in response to the tonic low inflammatory signals that are provided by the commensal gut microbiota. Other tissue macrophages that are derived from monocytes include dermal and heart macrophages. Specific tissue-resident macrophage populations in mice are seeded before birth. At a very early stage, embryonic precursors — such as yolk sac-derived macrophages and fetal liver-derived monocytes — give rise to tissue macrophages that persist and maintain the macrophage pool into adulthood, without being superseded by adult bone marrow-derived or blood monocyte-derived cells. Both yolk sac-derived macrophages and fetal liver-derived monocytes give rise to fetal macrophages. Their relative contribution to adult tissue macrophage populations varies between tissues and remains to be fully elucidated. In adults, tissue macrophages maintain themselves by self-renewal at low levels in the steady state. Importantly, the ability of tissue macrophages to proliferate is enhanced during inflammation. Our understanding of the ontogeny of monocytes and macrophages, as well as their maintenance in the steady state, has recently undergone a renaissance. Here, Ginhoux and Jung discuss the evidence that has changed our view of the relationship between monocytes and tissue macrophages during development and in the steady state. Monocytes and macrophages have crucial and distinct roles in tissue homeostasis and immunity, but they also contribute to a broad spectrum of pathologies and are thus attractive therapeutic targets. Potential intervention strategies that aim to manipulate these cells will require an in-depth understanding of their origins and the mechanisms that ensure their homeostasis. Recent evidence shows that monocytes do not substantially contribute to most tissue macrophage populations in the steady state or during certain types of inflammation. Rather, most tissue macrophage populations in mice are derived from embryonic precursors, are seeded before birth and can maintain themselves in adults by self-renewal. In this Review, we discuss the evidence that has dramatically changed our understanding of monocyte and macrophage development, and the maintenance of these cells in the steady state.

Flow cytometry is an essential tool for dissecting the functional complexity of hematopoiesis. We used single-cell \"mass cytometry\" to examine healthy human bone marrow, measuring 34 parameters simultaneously in single cells (binding of 31 antibodies, viability, DNA content, and relative cell size). The signaling behavior of cell subsets spanning a defined hematopoietic hierarchy was monitored with 18 simultaneous markers of functional signaling states perturbed by a set of ex vivo stimuli and inhibitors. The data set allowed for an algorithmically driven assembly of related cell types defined by surface antigen expression, providing a superimposable map of cell signaling responses in combination with drug inhibition. Visualized in this manner, the analysis revealed previously unappreciated instances of both precise signaling responses that were bounded within conventionally defined cell subsets and more continuous phosphorylation responses that crossed cell population boundaries in unexpected manners yet tracked closely with cellular phenotype. Collectively, such single-cell analyses provide system-wide views of immune signaling in healthy human hematopoiesis, against which drug action and disease can be compared for mechanistic studies and pharmacologic intervention.

Non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH) are prevalent liver conditions that underlie the development of life-threatening cirrhosis, liver failure and liver cancer. Chronic necro-inflammation is a critical factor in development of NASH, yet the cellular and molecular mechanisms of immune dysregulation in this disease are poorly understood. Here, using single-cell transcriptomic analysis, we comprehensively profiled the immune composition of the mouse liver during NASH. We identified a significant pathology-associated increase in hepatic conventional dendritic cells (cDCs) and further defined their source as NASH-induced boost in cycling of cDC progenitors in the bone marrow. Analysis of blood and liver from patients on the NAFLD/NASH spectrum showed that type 1 cDCs (cDC1) were more abundant and activated in disease. Sequencing of physically interacting cDC-T cell pairs from liver-draining lymph nodes revealed that cDCs in NASH promote inflammatory T cell reprogramming, previously associated with NASH worsening. Finally, depletion of cDC1 in XCR1 DTA mice or using anti-XCL1-blocking antibody attenuated liver pathology in NASH mouse models. Overall, our study provides a comprehensive characterization of cDC biology in NASH and identifies XCR1 + cDC1 as an important driver of liver pathology. Single-cell analyses reveal cDC1 as conserved immunological drivers of non-alcoholic steatohepatitis in mice and humans