Summary
Severe congenital neutropenia (SCN) patients are prone to develop myelodysplastic syndrome (MDS) or acute myeloid leukaemia (AML). Leukaemic progression of SCN is associated with the early ...acquisition of CSF3R mutations in haematopoietic progenitor cells (HPCs), which truncate the colony‐stimulating factor 3 receptor (CSF3R). These mutant clones may arise years before MDS/AML becomes overt. Introduction and activation of CSF3R truncation mutants in normal HPCs causes a clonally dominant myeloproliferative state in mice treated with CSF3. Paradoxically, in SCN patients receiving CSF3 therapy, clonal dominance of CSF3R mutant clones usually occurs only after the acquisition of additional mutations shortly before frank MDS or AML is diagnosed. To seek an explanation for this discrepancy, we introduced a patient‐derived CSF3R‐truncating mutation in ELANE‐SCN and HAX1‐SCN derived and control induced pluripotent stem cells and compared the CSF3 responses of HPCs generated from these lines. In contrast to CSF3R‐mutant control HPCs, CSF3R‐mutant HPCs from SCN patients do not show increased proliferation but display elevated levels of inflammatory signalling. Thus, activation of the truncated CSF3R in SCN‐HPCs does not evoke clonal outgrowth but causes a sustained pro‐inflammatory state, which has ramifications for how these CSF3R mutants contribute to the leukaemic transformation of SCN.
Innate and adaptive immune cells participate in the homeostatic regulation of hematopoietic stem cells (HSCs). Here, we interrogate the contribution of myeloid cells, the most abundant cell type in ...the mammalian bone marrow, in a clinically relevant mouse model of neutropenia. Long-term genetic depletion of neutrophils and eosinophils results in activation of multipotent progenitors but preservation of HSCs. Depletion of myeloid cells abrogates HSC expansion, loss of serial repopulation and lymphoid reconstitution capacity and remodeling of HSC niches, features previously associated with hematopoietic aging. This is associated with mitigation of interferon signaling in both HSCs and their niches via reduction of NK cell number and activation. These data implicate myeloid cells in the functional decline of hematopoiesis, associated with activation of interferon signaling via a putative neutrophil-NK cell axis. Innate immunity may thus come at the cost of system deterioration through enhanced chronic inflammatory signaling to stem cells and their niches.
Shwachman-Diamond syndrome is a congenital bone marrow failure disorder characterized by debilitating neutropenia. The disease is associated with loss-of-function mutations in the SBDS gene, ...implicated in ribosome biogenesis, but the cellular and molecular events driving cell specific phenotypes in ribosomopathies remain poorly defined. Here, we established what is to our knowledge the first mammalian model of neutropenia in Shwachman-Diamond syndrome through targeted downregulation of Sbds in hematopoietic stem and progenitor cells expressing the myeloid transcription factor CCAAT/enhancer binding protein α (Cebpa). Sbds deficiency in the myeloid lineage specifically affected myelocytes and their downstream progeny while, unexpectedly, it was well tolerated by rapidly cycling hematopoietic progenitor cells. Molecular insights provided by massive parallel sequencing supported cellular observations of impaired cell cycle exit and formation of secondary granules associated with the defect of myeloid lineage progression in myelocytes. Mechanistically, Sbds deficiency activated the p53 tumor suppressor pathway and induced apoptosis in these cells. Collectively, the data reveal a previously unanticipated, selective dependency of myelocytes and downstream progeny, but not rapidly cycling progenitors, on this ubiquitous ribosome biogenesis protein, thus providing a cellular basis for the understanding of myeloid lineage biased defects in Shwachman-Diamond syndrome.
Bone marrow fibrosis (BMF) develops in various hematological and non-hematological conditions and is a central pathological feature of myelofibrosis. Effective cell-targeted therapeutics are needed, ...but the cellular origin of BMF remains elusive. Here, we show using genetic fate tracing in two murine models of BMF that Gli1+ mesenchymal stromal cells (MSCs) are recruited from the endosteal and perivascular niche to become fibrosis-driving myofibroblasts in the bone marrow. Genetic ablation of Gli1+ cells abolished BMF and rescued bone marrow failure. Pharmacological targeting of Gli proteins with GANT61 inhibited Gli1+ cell expansion and myofibroblast differentiation and attenuated fibrosis severity. The same pathway is also active in human BMF, and Gli1 expression in BMF significantly correlates with the severity of the disease. In addition, GANT61 treatment reduced the myofibroblastic phenotype of human MSCs isolated from patients with BMF, suggesting that targeting of Gli proteins could be a relevant therapeutic strategy.
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•Bone marrow Gli1+ mesenchymal stromal cells (MSCs) differentiate into myofibroblasts•Targeting Gli1+ cells with GANT61 ameliorates bone marrow fibrosis (BMF)•Cxcl4 induces Gli1+ stromal cell migration and differentiation into myofibroblasts•Gli1+ MSCs also expand in human BMF and are sensitive to GANT61 inhibition
Schneider and colleagues show that Gli1+ bone marrow mesenchymal stromal cells are an important source of fibrotic cells during bone marrow fibrosis and that targeting of Gli proteins with GANT61 holds promise for amelioration of this disease.
Surveillance of patients with Barrett's esophagus (BE) aims at early detection and treatment of neoplastic changes, particularly esophageal adenocarcinoma (EAC). The histological evaluation of biopsy ...samples has its limitations, and biomarkers may improve early identification of BE patients at risk for progression to EAC. The aim of this study was to determine the predictive value of p53, Ki67, and aneuploidy as markers of neoplastic progression in BE.
A total of 27 BE patients with histologically proven progression to high-grade dysplasia (HGD) or EAC (cases) and 27 BE patients without progression (controls) were selected and matched for age, gender, and duration of follow-up. Dysplasia grade was determined in 212 biopsy samples obtained during surveillance endoscopies from cases and in 231 biopsy samples collected from controls. DNA ploidy status was determined by flow cytometry, whereas Ki67 and p53 expression was determined by immunohistochemistry. Hazard ratios (HRs) were calculated by Cox regression adjusted for potentially confounding variables.
A univariate analysis showed that low-grade dysplasia (LGD) increased the risk of developing HGD/EAC compared with no dysplasia (HR 3.6; 95% confidence interval (CI): 1.6 - 8.1). Aneuploidy (HR 3.5; 95% CI: 1.3-9.4), strong Ki67 overexpression (HR 5.2; 95% CI: 1.5-17.6), and moderate p53 overexpression (HR 6.5; 95% CI: 2.5-17.1) were also associated with an increased risk of developing HGD/EAC, independent of the histological result. A multivariable analysis showed that in the presence of LGD, p53 overexpression, and to a lesser extent, Ki67 overexpression remained important risk factors for neoplastic progression, whereas aneuploidy was no longer predictive.
p53 overexpression and, to a lesser extent, Ki67 overexpression could predict neoplastic progression in BE irrespective of the histological result. These markers may be useful for identifying patients at an increased risk of developing EAC, either alone or used as a panel.
Neutrophilic differentiation is dependent on CCAAT enhancer-binding protein α (C/EBPα), a transcription factor expressed in multiple organs including the bone marrow. Using functional genomic ...technologies in combination with clustered regularly-interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein 9 genome editing and in vivo mouse modeling, we show that CEBPA is located in a 170-kb topological-associated domain that contains 14 potential enhancers. Of these, 1 enhancer located +42 kb from CEBPA is active and engages with the CEBPA promoter in myeloid cells only. Germ line deletion of the homologous enhancer in mice in vivo reduces Cebpa levels exclusively in hematopoietic stem cells (HSCs) and myeloid-primed progenitor cells leading to severe defects in the granulocytic lineage, without affecting any other Cebpa-expressing organ studied. The enhancer-deleted progenitor cells lose their myeloid transcription program and are blocked in differentiation. Deletion of the enhancer also causes loss of HSC maintenance. We conclude that a single +42-kb enhancer is essential for CEBPA expression in myeloid cells only.
•The CEBPA locus harbors 14 enhancers of which distinct combinations are active in different CEBPA-expressing tissues.•A +42-kb enhancer is required for myeloid-lineage priming to drive adequate CEBPA expression levels necessary for neutrophilic maturation.
Macrophages derive from multiple sources of hematopoietic progenitors. Most macrophages require colony-stimulating factor 1 receptor (CSF1R), but some macrophages persist in the absence of CSF1R. ...Here, we analyzed
:GFP-expressing macrophages in
-deficient zebrafish and report that embryonic macrophages emerge followed by their developmental arrest. In larvae,
+ cell numbers then increased showing two distinct types in the skin: branched, putative Langerhans cells, and amoeboid cells. In contrast, although numbers also increased in
-mutants, exclusively amoeboid
cells were present, which we showed by genetic lineage tracing to have a non-hematopoietic origin. They expressed macrophage-associated genes, but also showed decreased phagocytic gene expression and increased epithelial-associated gene expression, characteristic of metaphocytes, recently discovered ectoderm-derived cells. We further demonstrated that juvenile
-deficient zebrafish exhibit systemic macrophage depletion. Thus,
deficiency disrupts embryonic to adult macrophage development. Zebrafish deficient for
are viable and permit analyzing the consequences of macrophage loss throughout life.
RUNX1 familial platelet disorder (RUNX1‐FPD) is a hematopoietic disorder caused by germline loss‐of‐function mutations in the RUNX1 gene and characterized by thrombocytopathy, thrombocytopenia, and ...an increased risk of developing hematologic malignancies, mostly of myeloid origin. Disease pathophysiology has remained incompletely understood, in part because of a shortage of in vivo models recapitulating the germline RUNX1 loss of function found in humans, precluding the study of potential contributions of non‐hematopoietic cells to disease pathogenesis. Here, we studied mice harboring a germline hypomorphic mutation of one Runx1 allele with a loss‐of‐function mutation in the other Runx1 allele (Runx1L148A/− mice), which display many hematologic characteristics found in human RUNX1‐FPD patients. Runx1L148A/− mice displayed robust and pronounced thrombocytopenia and myeloid‐biased hematopoiesis, associated with an HSC intrinsic reconstitution defect in lymphopoiesis and expansion of myeloid progenitor cell pools. We demonstrate that specific deletion of Runx1 from bone marrow stromal cells in Prrx1‐cre;Runx1fl/fl mice did not recapitulate these abnormalities, indicating that the hematopoietic abnormalities are intrinsic to the hematopoietic lineage, and arguing against a driving role of the bone marrow microenvironment. In conclusion, we report a RUNX1‐FPD mouse model faithfully recapitulating key characteristics of human disease. Findings do not support a driving role of ancillary, non‐hematopoietic cells in the disruption of hematopoiesis under homeostatic conditions.
Mesenchymal niche cells may drive tissue failure and malignant transformation in the hematopoietic system, but the underlying molecular mechanisms and relevance to human disease remain poorly ...defined. Here, we show that perturbation of mesenchymal cells in a mouse model of the pre-leukemic disorder Shwachman-Diamond syndrome (SDS) induces mitochondrial dysfunction, oxidative stress, and activation of DNA damage responses in hematopoietic stem and progenitor cells. Massive parallel RNA sequencing of highly purified mesenchymal cells in the SDS mouse model and a range of human pre-leukemic syndromes identified p53-S100A8/9-TLR inflammatory signaling as a common driving mechanism of genotoxic stress. Transcriptional activation of this signaling axis in the mesenchymal niche predicted leukemic evolution and progression-free survival in myelodysplastic syndrome (MDS), the principal leukemia predisposition syndrome. Collectively, our findings identify mesenchymal niche-induced genotoxic stress in heterotypic stem and progenitor cells through inflammatory signaling as a targetable determinant of disease outcome in human pre-leukemia.
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•Mesenchymal deletion of Sbds in mice recapitulates bone defects in SDS•Mesenchymal niche cells induce genotoxic stress in HSPCs in this model•p53-S100A8/9-TLR4 signaling, activated in SDS and MDS, drives these phenotypes•Mesenchymal S100A8/9 predicts leukemic evolution and disease outcome in human MDS
Cell-extrinsic factors driving malignant transformation remain understudied. In a mouse model of pre-leukemia, Zambetti and colleagues establish a concept of mesenchymal niche-induced genotoxic stress in hematopoietic stem cells through p53-S100A8/9-TLR4 signaling, with relevance to human leukemia. The findings provide conceptual and mechanistic insights into the link between inflammation and cancer.
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