RESUMO
Constitutional trisomy 21 (T21) is a state of aneuploidy associated with high incidence of childhood acute myeloid leukemia (AML). T21-associated AML is preceded by transient abnormal myelopoiesis (TAM), which is triggered by truncating mutations in GATA1 generating a short GATA1 isoform (GATA1s). T21-associated AML emerges due to secondary mutations in hematopoietic clones bearing GATA1s. Since aneuploidy generally impairs cellular fitness, the paradoxically elevated risk of myeloid malignancy in T21 is not fully understood. We hypothesized that individuals with T21 bear inherent genome instability in hematopoietic lineages that promotes leukemogenic mutations driving the genesis of TAM and AML. We found that individuals with T21 show increased chromosomal copy number variations (CNVs) compared to euploid individuals, suggesting that genome instability could be underlying predisposition to TAM and AML. Acquisition of GATA1s enforces myeloid skewing and maintenance of the hematopoietic progenitor state independently of T21; however, GATA1s in T21 hematopoietic progenitor cells (HPCs) further augments genome instability. Increased dosage of the chromosome 21 (chr21) gene DYRK1A impairs homology-directed DNA repair as a mechanism of elevated mutagenesis. These results posit a model wherein inherent genome instability in T21 drives myeloid malignancy in concert with GATA1s mutations.
Assuntos
Síndrome de Down , Leucemia Mieloide Aguda , Reação Leucemoide , Transtornos Mieloproliferativos , Humanos , Criança , Síndrome de Down/complicações , Variações do Número de Cópias de DNA , Transtornos Mieloproliferativos/genética , Instabilidade Genômica , Leucemia Mieloide Aguda/patologia , Aneuploidia , Trissomia , Fator de Transcrição GATA1/genéticaRESUMO
Abundant macrophage infiltration and altered tumor metabolism are two key hallmarks of glioblastoma. By screening a cluster of metabolic small-molecule compounds, we show that inhibiting glioblastoma cell glycolysis impairs macrophage migration and lactate dehydrogenase (LDH) inhibitor stiripentol (an FDA-approved anti-seizure drug for Dravet Syndrome) emerges as the top hit. Combined profiling and functional studies demonstrate that LDHA-directed ERK pathway activates YAP1/STAT3 transcriptional co-activators in glioblastoma cells to upregulate CCL2 and CCL7, which recruit macrophages into the tumor microenvironment. Reciprocally, infiltrating macrophages produce LDHA-containing extracellular vesicles to promote glioblastoma cell glycolysis, proliferation, and survival. Genetic and pharmacological inhibition of LDHA-mediated tumor-macrophage symbiosis markedly suppresses tumor progression and macrophage infiltration in glioblastoma mouse models. Analysis of tumor and plasma samples of glioblastoma patients confirms that LDHA and its downstream signals are potential biomarkers correlating positively with macrophage density. Thus, LDHA-mediated tumor-macrophage symbiosis provides therapeutic targets for glioblastoma.
RESUMO
Many inherited bone marrow failure syndromes (IBMFSs) present a high risk of transformation to myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML). During transformation of IBMFSs, hematopoietic stem and progenitor cells (HSPCs) with poor fitness gain ectopic, dysregulated self-renewal secondary to somatic mutations via undefined mechanisms. Here, in the context of the prototypical IBMFS Fanconi anemia (FA), we performed multiplexed gene editing of mutational hotspots in MDS-associated genes in human induced pluripotent stem cells (iPSCs) followed by hematopoietic differentiation. We observed aberrant self-renewal and impaired differentiation of HSPCs with enrichment of RUNX1 insertions and deletions (indels), generating a model of IBMFS-associated MDS. We observed that compared to the failure state, FA MDS cells show mutant RUNX1-mediated blunting of the G1/S cell cycle checkpoint that is normally activated in FA in response to DNA damage. RUNX1 indels also lead to activation of innate immune signaling, which stabilizes the homologous recombination (HR) effector BRCA1, and this pathway can be targeted to abrogate viability and restore sensitivity to genotoxins in FA MDS. Together, these studies develop a paradigm for modeling clonal evolution in IBMFSs, provide basic understanding of the pathogenesis of MDS, and uncover a therapeutic target in FA-associated MDS.
Assuntos
Anemia de Fanconi , Células-Tronco Pluripotentes Induzidas , Leucemia Mieloide Aguda , Síndromes Mielodisplásicas , Humanos , Anemia de Fanconi/genética , Anemia de Fanconi/patologia , Anemia de Fanconi/terapia , Síndrome Congênita de Insuficiência da Medula Óssea/complicações , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Células-Tronco Pluripotentes Induzidas/patologia , Síndromes Mielodisplásicas/patologia , Mutação , Leucemia Mieloide Aguda/patologiaRESUMO
Hematopoiesis changes over life to meet the demands of maturation and aging. Here, we find that the definitive hematopoietic stem and progenitor cell (HSPC) compartment is remodeled from gestation into adulthood, a process regulated by the heterochronic Lin28b/let-7 axis. Native fetal and neonatal HSPCs distribute with a pro-lymphoid/erythroid bias with a shift toward myeloid output in adulthood. By mining transcriptomic data comparing juvenile and adult HSPCs and reconstructing coordinately activated gene regulatory networks, we uncover the Polycomb repressor complex 1 (PRC1) component Cbx2 as an effector of Lin28b/let-7's control of hematopoietic maturation. We find that juvenile Cbx2-/- hematopoietic tissues show impairment of B-lymphopoiesis, a precocious adult-like myeloid bias, and that Cbx2/PRC1 regulates developmental timing of expression of key hematopoietic transcription factors. These findings define a mechanism of regulation of HSPC output via chromatin modification as a function of age with potential impact on age-biased pediatric and adult blood disorders.
Assuntos
Hematopoese , Células-Tronco Hematopoéticas , MicroRNAs , Complexo Repressor Polycomb 1 , Proteínas de Ligação a RNA , Adulto , Animais , Criança , Redes Reguladoras de Genes , Hematopoese/genética , Células-Tronco Hematopoéticas/metabolismo , Humanos , Recém-Nascido , Linfopoese , Camundongos , Complexo Repressor Polycomb 1/genética , Complexo Repressor Polycomb 1/metabolismo , Proteínas do Grupo Polycomb/metabolismo , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismoRESUMO
High-risk forms of B-acute lymphoblastic leukemia (B-ALL) remain a therapeutic challenge. Leukemia-initiating cells (LICs) self-renew and spark relapse and therefore have been the subject of intensive investigation; however, the properties of LICs in high-risk B-ALL are not well understood. Here, we use single-cell transcriptomics and quantitative xenotransplantation to understand LICs in MLL-rearranged (MLL-r) B-ALL. Compared with reported LIC frequencies in acute myeloid leukemia (AML), engraftable LICs in MLL-r B-ALL are abundant. Although we find that multipotent, self-renewing LICs are enriched among phenotypically undifferentiated B-ALL cells, LICs with the capacity to replenish the leukemic cellular diversity can emerge from more mature fractions. While inhibiting oxidative phosphorylation blunts blast proliferation, this intervention promotes LIC emergence. Conversely, inhibiting hypoxia and glycolysis impairs MLL-r B-ALL LICs, providing a therapeutic benefit in xenotransplantation systems. These findings provide insight into the aggressive nature of MLL-r B-ALL and provide a rationale for therapeutic targeting of hypoxia and glycolysis.
Assuntos
Leucemia Mieloide Aguda , Leucemia-Linfoma Linfoblástico de Células Precursoras , Glicólise , Humanos , Hipóxia , Leucemia Mieloide Aguda/metabolismo , Leucemia-Linfoma Linfoblástico de Células Precursoras/genéticaRESUMO
Fanconi anemia (FA) is a disorder of DNA repair that manifests as bone marrow (BM) failure. The lack of accurate murine models of FA has refocused efforts toward differentiation of patient-derived induced pluripotent stem cells (IPSCs) to hematopoietic progenitor cells (HPCs). However, an intact FA DNA repair pathway is required for efficient IPSC derivation, hindering these efforts. To overcome this barrier, we used inducible complementation of FANCA-deficient IPSCs, which permitted robust maintenance of IPSCs. Modulation of FANCA during directed differentiation to HPCs enabled the production of FANCA-deficient human HPCs that recapitulated FA genotoxicity and hematopoietic phenotypes relative to isogenic FANCA-expressing HPCs. FANCA-deficient human HPCs underwent accelerated terminal differentiation driven by activation of p53/p21. We identified growth arrest specific 6 (GAS6) as a novel target of activated p53 in FANCA-deficient HPCs and modulate GAS6 signaling to rescue hematopoiesis in FANCA-deficient cells. This study validates our strategy to derive a sustainable, highly faithful human model of FA, uncovers a mechanism of HPC exhaustion in FA, and advances toward future cell therapy in FA.
Assuntos
Anemia de Fanconi , Células-Tronco Pluripotentes Induzidas , Animais , Diferenciação Celular , Anemia de Fanconi/genética , Proteína do Grupo de Complementação A da Anemia de Fanconi/genética , Humanos , Camundongos , Proteína Supressora de Tumor p53/genéticaRESUMO
The LIN28:pre-let-7:TUTase ternary complex regulates pluripotency and oncogenesis by controlling processing of the let-7 family of microRNAs. The complex oligouridylates the 3' ends of pre-let-7 molecules, leading to their degradation via the DIS3L2 exonuclease. Previous studies suggest that components of this complex are potential therapeutic targets in malignancies that aberrantly express LIN28. In this study we developed a functional epitope selection approach to identify nanobody inhibitors of the LIN28:pre-let-7:TUT4 complex. We demonstrate that one of the identified nanobodies, Nb-S2A4, targets the 106-residue LIN28:let-7 interaction (LLI) fragment of TUT4. Nb-S2A4 can effectively inhibit oligouridylation and monouridylation of pre-let-7g in vitro. Expressing Nb-S2A4 allows maturation of the let-7 species in cells expressing LIN28, highlighting the therapeutic potential of targeting the LLI fragment.
Assuntos
Proteínas de Ligação a DNA/imunologia , MicroRNAs/metabolismo , Processamento de Terminações 3' de RNA , Anticorpos de Domínio Único/imunologia , Animais , Sítios de Ligação , Proteínas de Ligação a DNA/química , Células HEK293 , Células HeLa , Humanos , Camundongos , MicroRNAs/genética , Ligação Proteica , Estabilidade de RNA , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Células Sf9 , SpodopteraRESUMO
Leukemia phenotypes vary with age of onset. Delineating mechanisms of age specificity in leukemia could improve disease models and uncover new therapeutic approaches. Here, we used heterochronic transplantation of leukemia driven by MLL/KMT2A translocations to investigate the contribution of the age of the hematopoietic microenvironment to age-specific leukemia phenotypes. When driven by MLL-AF9, leukemia cells in the adult microenvironment sustained a myeloid phenotype, whereas the neonatal microenvironment supported genesis of mixed early B cell/myeloid leukemia. In MLL-ENL leukemia, the neonatal microenvironment potentiated B-lymphoid differentiation compared with the adult. Ccl5 elaborated from adult marrow stroma inhibited B-lymphoid differentiation of leukemia cells, illuminating a mechanism of age-specific lineage commitment. Our study illustrates the contribution of the developmental stage of the hematopoietic microenvironment in defining the age specificity of leukemia.
Assuntos
Hematopoese/fisiologia , Leucemia/patologia , Proteínas de Fusão Oncogênica/genética , Envelhecimento , Animais , Animais Recém-Nascidos , Linfócitos B/patologia , Quimiocina CCL5/genética , Quimiocina CCL5/metabolismo , Feminino , Regulação Leucêmica da Expressão Gênica , Células-Tronco Hematopoéticas/metabolismo , Células-Tronco Hematopoéticas/patologia , Histona-Lisina N-Metiltransferase/genética , Leucemia/genética , Antígenos Comuns de Leucócito/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Proteína de Leucina Linfoide-Mieloide/genética , Células Estromais/patologia , Microambiente TumoralRESUMO
The derivation of induced pluripotent stem cells (iPSCs) over a decade ago sparked widespread enthusiasm for the development of new models of human disease, enhanced platforms for drug discovery and more widespread use of autologous cell-based therapy. Early studies using directed differentiation of iPSCs frequently uncovered cell-level phenotypes in monogenic diseases, but translation to tissue-level and organ-level diseases has required development of more complex, 3D, multicellular systems. Organoids and human-rodent chimaeras more accurately mirror the diverse cellular ecosystems of complex tissues and are being applied to iPSC disease models to recapitulate the pathobiology of a broad spectrum of human maladies, including infectious diseases, genetic disorders and cancer.
Assuntos
Doenças Transmissíveis/terapia , Doenças Genéticas Inatas/terapia , Células-Tronco Pluripotentes Induzidas/citologia , Modelos Biológicos , Neoplasias/terapia , Engenharia Tecidual/métodos , Animais , Diferenciação Celular , Linhagem da Célula/genética , Linhagem da Célula/imunologia , Quimera/genética , Quimera/imunologia , Doenças Transmissíveis/genética , Doenças Transmissíveis/imunologia , Doenças Transmissíveis/patologia , Descoberta de Drogas/métodos , Doenças Genéticas Inatas/genética , Doenças Genéticas Inatas/imunologia , Doenças Genéticas Inatas/patologia , Terapia Genética/métodos , Humanos , Células-Tronco Pluripotentes Induzidas/imunologia , Células-Tronco Pluripotentes Induzidas/transplante , Modelos Animais , Neoplasias/genética , Neoplasias/imunologia , Neoplasias/patologia , Organoides/citologia , Organoides/efeitos dos fármacos , Organoides/imunologia , Transplante de Tecidos/métodos , Transplante HeterólogoRESUMO
Platelets are essential for hemostasis; however, several studies have identified age-dependent differences in platelet function. To better understand the origins of fetal platelet function, we have evaluated the contribution of the fetal-specific RNA binding protein Lin28b in the megakaryocyte/platelet lineage. Because activated fetal platelets have very low levels of P-selectin, we hypothesized that the expression of platelet P-selectin is part of a fetal-specific hematopoietic program conferred by Lin28b. Using the mouse as a model, we find that activated fetal platelets have low levels of P-selectin and do not readily associate with granulocytes in vitro and in vivo, relative to adult controls. Transcriptional analysis revealed high levels of Lin28b and Hmga2 in fetal, but not adult, megakaryocytes. Overexpression of LIN28B in adult mice significantly reduces the expression of P-selectin in platelets, and therefore identifies Lin28b as a negative regulator of P-selectin expression. Transplantation of fetal hematopoietic progenitors resulted in the production of platelets with low levels of P-selectin, suggesting that the developmental regulation of P-selectin is intrinsic and independent of differences between fetal and adult microenvironments. Last, we observe that the upregulation of P-selectin expression occurs postnatally, and the temporal kinetics of this upregulation are recapitulated by transplantation of fetal hematopoietic stem and progenitor cells into adult recipients. Taken together, these studies identify Lin28b as a new intrinsic regulator of fetal platelet function.
Assuntos
Plaquetas/metabolismo , Regulação da Expressão Gênica , Proteínas de Ligação a RNA/genética , Fatores Etários , Animais , Biomarcadores , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/metabolismo , Leucócitos/imunologia , Leucócitos/metabolismo , Megacariócitos/citologia , Megacariócitos/metabolismo , Camundongos , Selectina-P/genética , Selectina-P/metabolismo , Ativação Plaquetária , Agregação Plaquetária/genética , Testes de Função Plaquetária , Proteínas de Ligação a RNA/metabolismoRESUMO
LIN28 is an RNA-binding protein that regulates the maturation of the let-7 family of microRNAs by bipartite interactions with let-7 precursors through its two distinct cold shock and zinc-knuckle domains. Through inhibition of let-7 biogenesis, LIN28 functions as a pluripotency factor, as well as a driver of tumorigenesis. Here, we report a fluorescence polarization assay to identify small-molecule inhibitors for both domains of LIN28 involved in let-7 interactions. Of 101,017 compounds screened, six inhibit LIN28:let-7 binding and impair LIN28-mediated let-7 oligouridylation. Upon further characterization, we demonstrate that the LIN28 inhibitor TPEN destabilizes the zinc-knuckle domain of LIN28, while LI71 binds the cold shock domain to suppress LIN28's activity against let-7 in leukemia cells and embryonic stem cells. Our results demonstrate selective pharmacologic inhibition of individual domains of LIN28 and provide a foundation for therapeutic inhibition of the let-7 biogenesis pathway in LIN28-driven diseases.
Assuntos
MicroRNAs/metabolismo , Processamento Pós-Transcricional do RNA , Proteínas de Ligação a RNA/metabolismo , Bibliotecas de Moléculas Pequenas/farmacologia , Uridina/metabolismo , Sítios de Ligação , Linhagem Celular Tumoral , Células-Tronco Embrionárias/efeitos dos fármacos , Células-Tronco Embrionárias/metabolismo , Polarização de Fluorescência , Ensaios de Triagem em Larga Escala , Humanos , MicroRNAs/genética , Modelos Moleculares , Niacina/química , Bibliotecas de Moléculas Pequenas/químicaRESUMO
A better understanding of the cell-fate transitions that occur in complex cellular ecosystems in normal development and disease could inform cell engineering efforts and lead to improved therapies. However, a major challenge is to simultaneously identify new cell states, and their transitions, to elucidate the gene expression dynamics governing cell-type diversification. Here, we present CellRouter, a multifaceted single-cell analysis platform that identifies complex cell-state transition trajectories by using flow networks to explore the subpopulation structure of multi-dimensional, single-cell omics data. We demonstrate its versatility by applying CellRouter to single-cell RNA sequencing data sets to reconstruct cell-state transition trajectories during hematopoietic stem and progenitor cell (HSPC) differentiation to the erythroid, myeloid and lymphoid lineages, as well as during re-specification of cell identity by cellular reprogramming of monocytes and B-cells to HSPCs. CellRouter opens previously undescribed paths for in-depth characterization of complex cellular ecosystems and establishment of enhanced cell engineering approaches.
Assuntos
Células-Tronco Hematopoéticas/citologia , Análise de Célula Única/métodos , Diferenciação Celular , Linhagem da Célula , Expressão Gênica , Humanos , Análise de Sequência de RNA , Análise de Célula Única/instrumentaçãoRESUMO
BACKGROUND: In osteosarcoma, patient survival has not changed in over 30 years. Multiple phase II trials have been conducted in osteosarcoma using the Response Evaluation Criteria in Solid Tumors (RECIST) as a primary endpoint; however, none of these have revealed new treatment strategies. We investigated RECIST in newly diagnosed patients who received neoadjuvant chemotherapy proven to be beneficial. METHODS: Patients treated from 1986 to 2011 for newly diagnosed osteosarcoma with paired tumor imaging before and after adequate neoadjuvant chemotherapy were included in this retrospective study. Two radiologists performed independent, blinded (to image timing) RECIST measurements of primary tumor and lung metastases at diagnosis and post-neoadjuvant chemotherapy. Association between RECIST and histological necrosis and outcome were assessed. RESULTS: Seventy-four patients met inclusion criteria. Five-year overall survival and progression-free survival (PFS) were 77 ± 7% and 61 ± 8%, respectively. No patients had RECIST partial or complete response in the primary tumor. Sixty-four patients (86%) had stable disease, and 10 (14%) had progressive disease (PD). PD in the primary tumor was associated with significantly worse PFS in localized disease patients (P = 0.02). There was no association between RECIST in the primary tumor and necrosis. There were an insufficient number of patients with lung nodules ≥1 cm at diagnosis to evaluate RECIST in pulmonary metastases. CONCLUSIONS: PD by RECIST predicts poor outcome in localized disease patients. In bone lesions, chemotherapy proven to improve overall survival does not result in radiographic responses as measured by RECIST. Further investigation of RECIST in pulmonary metastatic disease in osteosarcoma is needed.
Assuntos
Neoplasias Ósseas , Neoplasias Pulmonares , Terapia Neoadjuvante , Osteossarcoma , Adolescente , Adulto , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/mortalidade , Neoplasias Ósseas/patologia , Criança , Intervalo Livre de Doença , Feminino , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/mortalidade , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/secundário , Masculino , Metástase Neoplásica , Osteossarcoma/tratamento farmacológico , Osteossarcoma/mortalidade , Osteossarcoma/patologia , Estudos Retrospectivos , Taxa de SobrevidaRESUMO
A variety of tissue lineages can be differentiated from pluripotent stem cells by mimicking embryonic development through stepwise exposure to morphogens, or by conversion of one differentiated cell type into another by enforced expression of master transcription factors. Here, to yield functional human haematopoietic stem cells, we perform morphogen-directed differentiation of human pluripotent stem cells into haemogenic endothelium followed by screening of 26 candidate haematopoietic stem-cell-specifying transcription factors for their capacity to promote multi-lineage haematopoietic engraftment in mouse hosts. We recover seven transcription factors (ERG, HOXA5, HOXA9, HOXA10, LCOR, RUNX1 and SPI1) that are sufficient to convert haemogenic endothelium into haematopoietic stem and progenitor cells that engraft myeloid, B and T cells in primary and secondary mouse recipients. Our combined approach of morphogen-driven differentiation and transcription-factor-mediated cell fate conversion produces haematopoietic stem and progenitor cells from pluripotent stem cells and holds promise for modelling haematopoietic disease in humanized mice and for therapeutic strategies in genetic blood disorders.
Assuntos
Diferenciação Celular , Linhagem da Célula , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/metabolismo , Células-Tronco Pluripotentes/citologia , Fatores de Transcrição/metabolismo , Animais , Reprogramação Celular , Subunidade alfa 2 de Fator de Ligação ao Core/metabolismo , Endotélio/citologia , Feminino , Transplante de Células-Tronco Hematopoéticas , Proteínas Homeobox A10 , Proteínas de Homeodomínio/metabolismo , Humanos , Camundongos , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Repressoras/metabolismo , Transativadores/metabolismo , Regulador Transcricional ERG/metabolismoRESUMO
For appropriate development, tissue and organ system morphogenesis and maturation must occur in synchrony with the overall developmental requirements of the host. Mistiming of such developmental events often results in disease. The hematopoietic system matures from the fetal state, characterized by robust erythrocytic output that supports prenatal growth in the hypoxic intrauterine environment, to the postnatal state wherein granulocytes predominate to provide innate immunity. Regulation of the developmental timing of these myeloerythroid states is not well understood. In this study, we find that expression of the heterochronic factor Lin28b decreases in common myeloid progenitors during hematopoietic maturation to adulthood in mice. This decrease in Lin28b coincides with accumulation of mature let-7 microRNAs, whose biogenesis is regulated by Lin28 proteins. We find that inhibition of let-7 in the adult hematopoietic system recapitulates fetal erythroid-dominant hematopoiesis. Conversely, deletion of Lin28b or ectopic activation of let-7 microRNAs in the fetal state induces a shift toward adult-like myeloid-dominant output. Furthermore, we identify Hmga2 as an effector of this genetic switch. These studies provide the first detailed analysis of the roles of endogenous Lin28b and let-7 in the timing of hematopoietic states during development.
Assuntos
Proteínas de Ligação a DNA/metabolismo , Eritropoese/fisiologia , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Proteína HMGA2/biossíntese , MicroRNAs/metabolismo , Células Progenitoras Mieloides/metabolismo , Animais , Proteínas de Ligação a DNA/genética , Proteína HMGA2/genética , Camundongos , Camundongos Knockout , MicroRNAs/genética , Células Progenitoras Mieloides/citologia , Proteínas de Ligação a RNARESUMO
Cell engineering has brought us tantalizingly close to the goal of deriving patient-specific hematopoietic stem cells (HSCs). While directed differentiation and transcription factor-mediated conversion strategies have generated progenitor cells with multilineage potential, to date, therapy-grade engineered HSCs remain elusive due to insufficient long-term self-renewal and inadequate differentiated progeny functionality. A cross-species approach involving zebrafish and mammalian systems offers complementary methodologies to improve understanding of native HSCs. Here, we discuss the role of conserved developmental timing processes in vertebrate hematopoiesis, highlighting how identification and manipulation of stage-specific factors that specify HSC developmental state must be harnessed to engineer HSCs for therapy.
Assuntos
Engenharia Celular/métodos , Desenvolvimento Embrionário , Células-Tronco Hematopoéticas/citologia , Animais , Hematopoese , Humanos , Modelos Biológicos , Fatores de TempoRESUMO
Cytomegalovirus (CMV) infection is a significant source of morbidity and mortality in allogeneic stem cell transplantation (SCT). We identified a cohort of 91 pediatric SCT patients at risk (defined as either donor and/or recipient seropositivity) for CMV infection at our institution. We retrospectively categorized at-risk SCT recipients as those who (1) were at risk of CMV infection in the post-SCT period, (2) had documented CMV infection before SCT, (3) experienced recurrence of post-SCT CMV viremia, or (4) experienced late post-SCT CMV viremia; categories were not mutually exclusive. We analyzed the impact of SCT-related factors on incidence of CMV infection and outcome, and we described the outcome of each of these cohorts. In univariate analysis, recipient CMV seropositivity, use of umbilical cord blood graft, and acute graft-versus-host disease (GVHD) predicted post-SCT CMV viremia, and the effects of acute GVHD (odds ratio, 4.018; 95% confidence interval, 1.032 to 15.643) and CMV seropositivity (odds ratio, 16.525; 95% confidence interval, 2.041 to 133.803) were confirmed in multivariate analysis. Patients with recurrence of post-SCT CMV viremia had a 50% all-cause mortality rate, compared with 12% in all 91 patients. Patients with pre-SCT CMV infection had a high incidence of post-SCT CMV infection but could successfully undergo SCT with antiviral prophylaxis and pre-emptive CMV treatment. All patients with late CMV infection had prior GVHD. Theses findings identify risk factors for post-SCT CMV infection and provide novel descriptions of childhood SCT recipients with pre-SCT, recurrent, and late CMV infection, which may contribute to risk stratification strategies for CMV at-risk patients in pediatric allogeneic SCT.
Assuntos
Infecções por Citomegalovirus/etiologia , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Adolescente , Adulto , Antivirais/uso terapêutico , Criança , Pré-Escolar , Transplante de Células-Tronco de Sangue do Cordão Umbilical/efeitos adversos , Infecções por Citomegalovirus/prevenção & controle , Feminino , Doença Enxerto-Hospedeiro/prevenção & controle , Humanos , Lactente , Masculino , Pré-Medicação/métodos , Recidiva , Estudos Retrospectivos , Fatores de Risco , Fatores de Tempo , Viremia/etiologia , Viremia/prevenção & controle , Adulto JovemRESUMO
We present an 8-year-old male with metastatic alveolar rhabdomyosarcoma (ARMS) who developed precipitous cardiopulmonary collapse with severe tumor lysis syndrome (TLS) 48 hr after initiation of chemotherapy. Despite no detectable pulmonary metastases, acute hypoxemic respiratory failure developed, requiring extracorporeal membrane oxygenation (ECMO). Although TLS has been reported in disseminated ARMS, this singular case of life-threatening respiratory deterioration developing after initiation of chemotherapy presented unique therapeutic dilemmas. We review the clinical aspects of this case, including possible mechanisms of respiratory failure, and discuss the role of ECMO utilization in pediatric oncology.
Assuntos
Oxigenação por Membrana Extracorpórea , Edema Pulmonar , Rabdomiossarcoma Alveolar/tratamento farmacológico , Síndrome de Lise Tumoral , Doença Aguda , Criança , Humanos , Masculino , Metástase Neoplásica , Edema Pulmonar/etiologia , Edema Pulmonar/patologia , Edema Pulmonar/terapia , Rabdomiossarcoma Alveolar/patologia , Síndrome de Lise Tumoral/etiologia , Síndrome de Lise Tumoral/patologia , Síndrome de Lise Tumoral/terapiaRESUMO
Fluid shear stress promotes the emergence of hematopoietic stem cells (HSCs) in the aorta-gonad-mesonephros (AGM) of the developing mouse embryo. We determined that the AGM is enriched for expression of targets of protein kinase A (PKA)-cAMP response element-binding protein (CREB), a pathway activated by fluid shear stress. By analyzing CREB genomic occupancy from chromatin-immunoprecipitation sequencing (ChIP-seq) data, we identified the bone morphogenetic protein (BMP) pathway as a potential regulator of CREB. By chemical modulation of the PKA-CREB and BMP pathways in isolated AGM VE-cadherin(+) cells from mid-gestation embryos, we demonstrate that PKA-CREB regulates hematopoietic engraftment and clonogenicity of hematopoietic progenitors, and is dependent on secreted BMP ligands through the type I BMP receptor. Finally, we observed blunting of this signaling axis using Ncx1-null embryos, which lack a heartbeat and intravascular flow. Collectively, we have identified a novel PKA-CREB-BMP signaling pathway downstream of shear stress that regulates HSC emergence in the AGM via the endothelial-to-hematopoietic transition.
Assuntos
Proteínas Morfogenéticas Ósseas/metabolismo , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Embrião de Mamíferos/embriologia , Células-Tronco Hematopoéticas/metabolismo , Transdução de Sinais/fisiologia , Animais , Proteínas Morfogenéticas Ósseas/genética , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Proteínas Quinases Dependentes de AMP Cíclico/genética , Embrião de Mamíferos/citologia , Células Endoteliais/citologia , Células Endoteliais/metabolismo , Células-Tronco Hematopoéticas/citologia , Mesonefro/citologia , Mesonefro/embriologia , Camundongos , Camundongos Mutantes , Trocador de Sódio e Cálcio/genética , Trocador de Sódio e Cálcio/metabolismoRESUMO
Notch1-Delta-like 4 (Dll4) signalling controls vascular development by regulating endothelial cell (EC) targets that modulate vessel wall remodelling and arterial-venous specification. The molecular effectors that modulate Notch signalling during vascular development remain largely undefined. Here we demonstrate that the transcriptional repressor, Snail1, acts as a VEGF-induced regulator of Notch1 signalling and Dll4 expression. EC-specific Snail1 loss-of-function conditional knockout mice die in utero with defects in vessel wall remodelling in association with losses in mural cell investment and disruptions in arterial-venous specification. Snail1 loss-of-function conditional knockout embryos further display upregulated Notch1 signalling and Dll4 expression that is partially reversed by inhibiting γ-secretase activity in vivo with Dll4 identified as a direct target of Snail1-mediated transcriptional repression. These results document a Snail1-Dll4/Notch1 axis that controls embryonic vascular development.