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Acute myeloid leukemia (AML) driven by the activation of EVI1 due to chromosome 3q26/MECOM rearrangements is incurable. Because transcription factors such as EVI1 are notoriously hard to target, insight into the mechanism by which EVI1 drives myeloid transformation could provide alternative avenues for therapy. Applying protein folding predictions combined with proteomics technologies, we demonstrate that interaction of EVI1 with CTBP1 and CTBP2 via a single PLDLS motif is indispensable for leukemic transformation. A 4× PLDLS repeat construct outcompetes binding of EVI1 to CTBP1 and CTBP2 and inhibits proliferation of 3q26/MECOM rearranged AML in vitro and in xenotransplant models. This proof-of-concept study opens the possibility to target one of the most incurable forms of AML with specific EVI1-CTBP inhibitors. This has important implications for other tumor types with aberrant expression of EVI1 and for cancers transformed by different CTBP-dependent oncogenic transcription factors.
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Oxidorreductasas de Alcohol , Proteínas de Unión al ADN , Leucemia Mieloide Aguda , Proteína del Locus del Complejo MDS1 y EV11 , Animales , Humanos , Ratones , Oxidorreductasas de Alcohol/metabolismo , Oxidorreductasas de Alcohol/genética , Línea Celular Tumoral , Proliferación Celular , Transformación Celular Neoplásica/genética , Transformación Celular Neoplásica/metabolismo , Proteínas Co-Represoras/metabolismo , Proteínas Co-Represoras/genética , Proteínas de Unión al ADN/metabolismo , Proteínas de Unión al ADN/genética , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Leucemia Mieloide Aguda/patología , Proteína del Locus del Complejo MDS1 y EV11/metabolismo , Proteína del Locus del Complejo MDS1 y EV11/genética , Unión Proteica , Factores de Transcripción/metabolismo , Factores de Transcripción/genéticaRESUMEN
Hematopoietic stem cells (HSCs) enable hematopoietic stem cell transplantation (HCT) through their ability to replenish the entire blood system. Proliferation of HSCs is linked to decreased reconstitution potential, and a precise regulation of actively dividing HSCs is thus essential to ensure long-term functionality. This regulation becomes important in the transplantation setting where HSCs undergo proliferation followed by a gradual transition to quiescence and homeostasis. Although mouse HSCs have been well studied under homeostatic conditions, the mechanisms regulating HSC activation under stress remain unclear. Here, we analyzed the different phases of regeneration after transplantation. We isolated bone marrow from mice at 8 time points after transplantation and examined the reconstitution dynamics and transcriptional profiles of stem and progenitor populations. We found that regenerating HSCs initially produced rapidly expanding progenitors and displayed distinct changes in fatty acid metabolism and glycolysis. Moreover, we observed molecular changes in cell cycle, MYC and mTOR signaling in both HSCs, and progenitor subsets. We used a decay rate model to fit the temporal transcription profiles of regenerating HSCs and identified genes with progressively decreased or increased expression after transplantation. These genes overlapped to a large extent with published gene sets associated with key aspects of HSC function, demonstrating the potential of this data set as a resource for identification of novel HSC regulators. Taken together, our study provides a detailed functional and molecular characterization of HSCs at different phases of regeneration and identifies a gene set associated with the transition from proliferation to quiescence.
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Trasplante de Células Madre Hematopoyéticas , Células Madre Hematopoyéticas , Ratones , Animales , Células Madre Hematopoyéticas/metabolismo , Médula Ósea , Ciclo Celular/genética , Transducción de SeñalRESUMEN
Despite their latent neurogenic potential, most normal parenchymal astrocytes fail to dedifferentiate to neural stem cells in response to injury. In contrast, aberrant lineage plasticity is a hallmark of gliomas, and this suggests that tumor suppressors may constrain astrocyte dedifferentiation. Here, we show that p53, one of the most commonly inactivated tumor suppressors in glioma, is a gatekeeper of astrocyte fate. In the context of stab-wound injury, p53 loss destabilized the identity of astrocytes, priming them to dedifferentiate in later life. This resulted from persistent and age-exacerbated neuroinflammation at the injury site and EGFR activation in periwound astrocytes. Mechanistically, dedifferentiation was driven by the synergistic upregulation of mTOR signaling downstream of p53 loss and EGFR, which reinstates stemness programs via increased translation of neurodevelopmental transcription factors. Thus, our findings suggest that first-hit mutations remove the barriers to injury-induced dedifferentiation by sensitizing somatic cells to inflammatory signals, with implications for tumorigenesis.
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Astrocitos , Células-Madre Neurales , Astrocitos/patología , Proteína p53 Supresora de Tumor/genética , Receptores ErbB/genética , MutaciónRESUMEN
Intratumoral heterogeneity (ITH) has been linked to decreased efficacy of clinical treatments. However, although genomic ITH has been characterized in genetic, transcriptomic and epigenetic alterations are hallmarks of esophageal squamous cell carcinoma (ESCC), the extent to which these are heterogeneous in ESCC has not been explored in a unified framework. Further, the extent to which tumor-infiltrated T lymphocytes are directed against cancer cells, but how the immune infiltration acts as a selective force to shape the clonal evolution of ESCC is unclear. In this study, we perform multi-omic sequencing on 186 samples from 36 primary ESCC patients. Through multi-omics analyses, it is discovered that genomic, epigenomic, and transcriptomic ITH are underpinned by ongoing chromosomal instability. Based on the RNA-seq data, we observe diverse levels of immune infiltrate across different tumor sites from the same tumor. We reveal genetic mechanisms of neoantigen evasion under distinct selection pressure from the diverse immune microenvironment. Overall, our work offers an avenue of dissecting the complex contribution of the multi-omics level to the ITH in ESCC and thereby enhances the development of clinical therapy.
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Neoplasias Esofágicas , Carcinoma de Células Escamosas de Esófago , Humanos , Carcinoma de Células Escamosas de Esófago/genética , Neoplasias Esofágicas/genética , Multiómica , Transcriptoma , Perfilación de la Expresión Génica , Microambiente TumoralRESUMEN
The ETV6-RUNX1 onco-fusion arises in utero, initiating a clinically silent pre-leukemic state associated with the development of pediatric B-acute lymphoblastic leukemia (B-ALL). We characterize the ETV6-RUNX1 regulome by integrating chromatin immunoprecipitation- and RNA-sequencing and show that ETV6-RUNX1 functions primarily through competition for RUNX1 binding sites and transcriptional repression. In pre-leukemia, this results in ETV6-RUNX1 antagonization of cell cycle regulation by RUNX1 as evidenced by mass cytometry analysis of B-lineage cells derived from ETV6-RUNX1 knock-in human pluripotent stem cells. In frank leukemia, knockdown of RUNX1 or its co-factor CBFß results in cell death suggesting sustained requirement for RUNX1 activity which is recapitulated by chemical perturbation using an allosteric CBFß-inhibitor. Strikingly, we show that RUNX1 addiction extends to other genetic subtypes of pediatric B-ALL and also adult disease. Importantly, inhibition of RUNX1 activity spares normal hematopoiesis. Our results suggest that chemical intervention in the RUNX1 program may provide a therapeutic opportunity in ALL.
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Subunidad alfa 2 del Factor de Unión al Sitio Principal , Leucemia-Linfoma Linfoblástico de Células Precursoras , Adulto , Niño , Humanos , Subunidad alfa 2 del Factor de Unión al Sitio Principal/genética , Factores de Unión al Sitio Principal , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/terapia , Linfocitos B , Fusión GénicaRESUMEN
BACKGROUND: Tumor heterogeneity of human colorectal cancer (CRC)-initiating cells (CRCICs) in cancer tissues often represents aggressive features of cancer progression. For high-resolution examination of CRCICs, we performed single-cell whole-exome sequencing (scWES) and bulk cell targeted exome sequencing (TES) of CRCICs to investigate stemness-specific somatic alterations or clonal evolution. METHODS: Single cells of three subpopulations of CRCICs (CD133+CD44+, CD133-CD44+, and CD133+CD44- cells), CRC cells (CRCCs), and control cells from one CRC tissue were sorted for scWES. Then, we set up a mutation panel from scWES data and TES was used to validate mutation distribution and clonal evolution in additional 96 samples (20 patients) those were also sorted into the same three groups of CRCICs and CRCCs. The knock-down experiments were used to analyze stemness-related mutant genes. Neoantigens of these mutant genes and their MHC binding affinity were also analyzed. FINDINGS: Clonal evolution analysis of scWES and TES showed that the CD133+CD44- CRCICs were the likely origin of CRC before evolving into other groups of CRCICs/CRCCs. We revealed that AHNAK2, PLIN4, HLA-B, ALK, CCDC92 and ALMS1 genes were specifically mutated in CRCICs followed by the validation of their functions. Furthermore, four predicted neoantigens of AHNAK2 were identified and validated, which might have applications in immunotherapy for CRC patients. INTERPRETATION: All the integrative analyses above revealed clonal evolution of CRC and new markers for CRCICs and demonstrate the important roles of CRCICs in tumorigenesis and progression of CRCs. FUNDING: A full list of funding bodies that contributed to this study can be found in the Acknowledgements section.
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Neoplasias Colorrectales , Antígeno AC133 , Movimiento Celular , Transformación Celular Neoplásica/metabolismo , Neoplasias Colorrectales/patología , Humanos , Receptores de Hialuranos/genética , Receptores de Hialuranos/metabolismo , Mutación , Células Madre Neoplásicas/metabolismo , Secuenciación del ExomaRESUMEN
Kinase signaling fuels growth of B-cell precursor acute lymphoblastic leukemia (BCP-ALL). Yet its role in leukemia initiation is unclear and has not been shown in primary human hematopoietic cells. We previously described activating mutations in interleukin-7 receptor alpha (IL7RA) in poor-prognosis "ph-like" BCP-ALL. Here we show that expression of activated mutant IL7RA in human CD34+ hematopoietic stem and progenitor cells induces a preleukemic state in transplanted immunodeficient NOD/LtSz-scid IL2Rγnull mice, characterized by persistence of self-renewing Pro-B cells with non-productive V(D)J gene rearrangements. Preleukemic CD34+CD10highCD19+ cells evolve into BCP-ALL with spontaneously acquired Cyclin Dependent Kinase Inhibitor 2 A (CDKN2A) deletions, as commonly observed in primary human BCP-ALL. CRISPR mediated gene silencing of CDKN2A in primary human CD34+ cells transduced with activated IL7RA results in robust development of BCP-ALLs in-vivo. Thus, we demonstrate that constitutive activation of IL7RA can initiate preleukemia in primary human hematopoietic progenitors and cooperates with CDKN2A silencing in progression into BCP-ALL.
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Subunidad alfa del Receptor de Interleucina-7/inmunología , Leucemia-Linfoma Linfoblástico de Células Precursoras B/inmunología , Células Precursoras de Linfocitos B/inmunología , Transducción de Señal/inmunología , Animales , Antígenos CD34/genética , Antígenos CD34/inmunología , Antígenos CD34/metabolismo , Secuencia de Bases , Diferenciación Celular/genética , Diferenciación Celular/inmunología , Inhibidor p16 de la Quinasa Dependiente de Ciclina/genética , Inhibidor p16 de la Quinasa Dependiente de Ciclina/inmunología , Inhibidor p16 de la Quinasa Dependiente de Ciclina/metabolismo , Expresión Génica/inmunología , Humanos , Subunidad alfa del Receptor de Interleucina-7/genética , Subunidad alfa del Receptor de Interleucina-7/metabolismo , Ratones Endogámicos NOD , Ratones Noqueados , Ratones SCID , Leucemia-Linfoma Linfoblástico de Células Precursoras B/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras B/metabolismo , Células Precursoras de Linfocitos B/metabolismo , RNA-Seq/métodos , Receptores de Citocinas/genética , Receptores de Citocinas/inmunología , Receptores de Citocinas/metabolismo , Transducción de Señal/genética , Análisis de la Célula Individual/métodos , Trasplante HeterólogoRESUMEN
Hematopoietic stem cells (HSCs) emerge during development from the vascular wall of the main embryonic arteries. The onset of circulation triggers several processes that provide critical external factors for HSC generation. Nevertheless, it is not fully understood how and when the onset of circulation affects HSC emergence. Here we show that in Ncx1-/- mouse embryos devoid of circulation the HSC lineage develops until the phenotypic pro-HSC stage. However, these cells reside in an abnormal microenvironment, fail to activate the hematopoietic program downstream of Runx1, and are functionally impaired. Single-cell transcriptomics shows that during the endothelial-to-hematopoietic transition, Ncx1-/- cells fail to undergo a glycolysis to oxidative phosphorylation metabolic switch present in wild-type cells. Interestingly, experimental activation of glycolysis results in decreased intraembryonic hematopoiesis. Our results suggest that the onset of circulation triggers metabolic changes that allow HSC generation to proceed.
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Diferenciación Celular , Linaje de la Célula , Endotelio Vascular/patología , Glucólisis , Hematopoyesis , Células Madre Hematopoyéticas/patología , Intercambiador de Sodio-Calcio/fisiología , Animales , Endotelio Vascular/metabolismo , Femenino , Células Madre Hematopoyéticas/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Fosforilación Oxidativa , Análisis de la Célula Individual , TranscriptomaRESUMEN
Defective silencing of retrotransposable elements has been linked to inflammageing, cancer and autoimmune diseases. However, the underlying mechanisms are only partially understood. Here we implicate the histone H3.3 chaperone Daxx, a retrotransposable element repressor inactivated in myeloid leukaemia and other neoplasms, in protection from inflammatory disease. Loss of Daxx alters the chromatin landscape, H3.3 distribution and histone marks of haematopoietic progenitors, leading to engagement of a Pu.1-dependent transcriptional programme for myelopoiesis at the expense of B-cell differentiation. This causes neutrophilia and inflammation, predisposing mice to develop an autoinflammatory skin disease. While these molecular and phenotypic perturbations are in part reverted in animals lacking both Pu.1 and Daxx, haematopoietic progenitors in these mice show unique chromatin and transcriptome alterations, suggesting an interaction between these two pathways. Overall, our findings implicate retrotransposable element silencing in haematopoiesis and suggest a cross-talk between the H3.3 loading machinery and the pioneer transcription factor Pu.1.
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Cromatina/patología , Proteínas Co-Represoras/genética , Trastornos Leucocíticos/congénito , Chaperonas Moleculares/genética , Mielopoyesis/genética , Proteínas Proto-Oncogénicas/metabolismo , Transactivadores/metabolismo , Animales , Enfermedades Autoinmunes/genética , Enfermedades Autoinmunes/patología , Linfocitos B/citología , Línea Celular , Cromatina/genética , Células Madre Hematopoyéticas/citología , Histonas/metabolismo , Humanos , Inflamación/patología , Trastornos Leucocíticos/patología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Retroelementos/genética , Enfermedades de la Piel/genética , Enfermedades de la Piel/inmunología , Enfermedades de la Piel/patologíaRESUMEN
Comparison of intratumor genetic heterogeneity in cancer at diagnosis and relapse suggests that chemotherapy induces bottleneck selection of subclonal genotypes. However, evolutionary events subsequent to chemotherapy could also explain changes in clonal dominance seen at relapse. We, therefore, investigated the mechanisms of selection in childhood B-cell precursor acute lymphoblastic leukemia (BCP-ALL) during induction chemotherapy where maximal cytoreduction occurs. To distinguish stochastic versus deterministic events, individual leukemias were transplanted into multiple xenografts and chemotherapy administered. Analyses of the immediate post-treatment leukemic residuum at single-cell resolution revealed that chemotherapy has little impact on genetic heterogeneity. Rather, it acts on extensive, previously unappreciated, transcriptional and epigenetic heterogeneity in BCP-ALL, dramatically reducing the spectrum of cell states represented, leaving a genetically polyclonal but phenotypically uniform population with hallmark signatures relating to developmental stage, cell cycle and metabolism. Hence, canalization of cell state accounts for a significant component of bottleneck selection during induction chemotherapy.
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Linfoma de Burkitt , Leucemia-Linfoma Linfoblástico de Células Precursoras , Linfoma de Burkitt/tratamiento farmacológico , Ciclo Celular , Humanos , Quimioterapia de Inducción , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamiento farmacológico , RecurrenciaRESUMEN
ETV6-RUNX1 is almost exclusively associated with childhood B-cell acute lymphoblastic leukemia (B-ALL), but the consequences of ETV6-RUNX1 expression on cell lineage decisions during B-cell leukemogenesis are completely unknown. Clinically silent ETV6-RUNX1 preleukemic clones are frequently found in neonatal cord blood, but few carriers develop B-ALL as a result of secondary genetic alterations. The understanding of the mechanisms underlying the first transforming steps could greatly advance the development of non-toxic prophylactic interventions. Using genetic lineage tracing, we examined the capacity of ETV6-RUNX1 to instruct a malignant phenotype in the hematopoietic lineage by cell-specific Cre-mediated activation of ETV6-RUNX1 from the endogenous Etv6 gene locus. Here we show that, while ETV6-RUNX1 has the propensity to trigger both T- and B-lymphoid malignancies, it is the second hit that determines tumor cell identity. To instigate leukemia, both oncogenic hits must place early in the development of hematopoietic/precursor cells, not in already committed B-cells. Depending on the nature of the second hit, the resulting B-ALLs presented distinct entities that were clearly separable based on their gene expression profiles. Our findings give a novel mechanistic insight into the early steps of ETV6-RUNX1+ B-ALL development and might have major implications for the potential development of ETV6-RUNX1+ B-ALL prevention strategies.
RESUMEN
Innate lymphoid cells are central to the regulation of immunity at mucosal barrier sites, with group 2 innate lymphoid cells (ILC2s) being particularly important in type 2 immunity. In this study, we demonstrate that microRNA(miR)-142 plays a critical, cell-intrinsic role in the homeostasis and function of ILC2s. Mice deficient for miR-142 expression demonstrate an ILC2 progenitor-biased development in the bone marrow, and along with peripheral ILC2s at mucosal sites, these cells display a greatly altered phenotype based on surface marker expression. ILC2 proliferative and effector functions are severely dysfunctional following Nippostrongylus brasiliensis infection, revealing a critical role for miR-142 isoforms in ILC2-mediated immune responses. Mechanistically, Socs1 and Gfi1 expression are regulated by miR-142 isoforms in ILC2s, impacting ILC2 phenotypes as well as the proliferative and effector capacity of these cells. The identification of these novel pathways opens potential new avenues to modulate ILC2-dependent immune functions.
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Linfocitos/inmunología , MicroARNs/inmunología , Animales , Células HEK293 , Homeostasis , Humanos , Inmunidad Innata/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , MicroARNs/genéticaRESUMEN
In vitro differentiation of human pluripotent stem cells (hPSCs) offers a genetically tractable system to examine the physiology and pathology of human tissue development and differentiation. We have used this approach to model the earliest stages of human B lineage development and characterize potential target cells for the in utero initiation of childhood B acute lymphoblastic leukemia. Herein, we detail critical aspects of the protocol including reagent validation, controls, and examples of surface markers used for analysis and cell sorting. For complete details on the use and execution of this protocol, please refer to Boiers et al. (2018).
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Linfocitos B/citología , Técnicas de Cocultivo/métodos , Células Madre Pluripotentes/citología , Animales , Línea Celular , Separación Celular , Humanos , Leucemia Linfoide , RatonesRESUMEN
Checkpoint inhibitors (CPIs) augment adaptive immunity. Systematic pan-tumor analyses may reveal the relative importance of tumor-cell-intrinsic and microenvironmental features underpinning CPI sensitization. Here, we collated whole-exome and transcriptomic data for >1,000 CPI-treated patients across seven tumor types, utilizing standardized bioinformatics workflows and clinical outcome criteria to validate multivariable predictors of CPI sensitization. Clonal tumor mutation burden (TMB) was the strongest predictor of CPI response, followed by total TMB and CXCL9 expression. Subclonal TMB, somatic copy alteration burden, and histocompatibility leukocyte antigen (HLA) evolutionary divergence failed to attain pan-cancer significance. Dinucleotide variants were identified as a source of immunogenic epitopes associated with radical amino acid substitutions and enhanced peptide hydrophobicity/immunogenicity. Copy-number analysis revealed two additional determinants of CPI outcome supported by prior functional evidence: 9q34 (TRAF2) loss associated with response and CCND1 amplification associated with resistance. Finally, single-cell RNA sequencing (RNA-seq) of clonal neoantigen-reactive CD8 tumor-infiltrating lymphocytes (TILs), combined with bulk RNA-seq analysis of CPI-responding tumors, identified CCR5 and CXCL13 as T-cell-intrinsic markers of CPI sensitivity.
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Inhibidores de Puntos de Control Inmunológico/farmacología , Neoplasias/inmunología , Linfocitos T/inmunología , Biomarcadores de Tumor/metabolismo , Antígenos CD8/metabolismo , Quimiocina CXCL13/metabolismo , Cromosomas Humanos Par 9/genética , Estudios de Cohortes , Ciclina D1/genética , Variaciones en el Número de Copia de ADN/genética , Exoma/genética , Amplificación de Genes , Humanos , Evasión Inmune/efectos de los fármacos , Análisis Multivariante , Mutación/genética , Neoplasias/patología , Polimorfismo de Nucleótido Simple/genética , Receptores CCR5/metabolismo , Linfocitos T/efectos de los fármacos , Carga Tumoral/genéticaRESUMEN
Aging is associated with reduced fitness and increased myeloid bias of the hematopoietic stem cell (HSC) compartment, causing increased risk of immune compromise, anemia, and malignancy. We show that mitochondrial membrane potential (MMP) can be used to prospectively isolate chronologically old HSCs with transcriptional features and functional attributes characteristic of young HSCs, including a high rate of transcription and balanced lineage-affiliated programs. Strikingly, MMP is a stronger determinant of the quantitative and qualitative transcriptional state of HSCs than chronological age, and transcriptional consequences of manipulation of MMP in HSCs within their native niche suggest a causal relationship. Accordingly, we show that pharmacological enhancement of MMP in old HSCs in vivo increases engraftment potential upon transplantation and reverses myeloid-biased peripheral blood output at steady state. Our results demonstrate that MMP is a source of heterogeneity in old HSCs, and its pharmacological manipulation can alter transcriptional programs with beneficial consequences for function.
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Trasplante de Células Madre Hematopoyéticas , Células Madre HematopoyéticasRESUMEN
OBJECTIVES: To estimate the impact of the COVID-19 pandemic on cancer care services and overall (direct and indirect) excess deaths in people with cancer. METHODS: We employed near real-time weekly data on cancer care to determine the adverse effect of the pandemic on cancer services. We also used these data, together with national death registrations until June 2020 to model deaths, in excess of background (pre-COVID-19) mortality, in people with cancer. Background mortality risks for 24 cancers with and without COVID-19-relevant comorbidities were obtained from population-based primary care cohort (Clinical Practice Research Datalink) on 3 862 012 adults in England. RESULTS: Declines in urgent referrals (median=-70.4%) and chemotherapy attendances (median=-41.5%) to a nadir (lowest point) in the pandemic were observed. By 31 May, these declines have only partially recovered; urgent referrals (median=-44.5%) and chemotherapy attendances (median=-31.2%). There were short-term excess death registrations for cancer (without COVID-19), with peak relative risk (RR) of 1.17 at week ending on 3 April. The peak RR for all-cause deaths was 2.1 from week ending on 17 April. Based on these findings and recent literature, we modelled 40% and 80% of cancer patients being affected by the pandemic in the long-term. At 40% affected, we estimated 1-year total (direct and indirect) excess deaths in people with cancer as between 7165 and 17 910, using RRs of 1.2 and 1.5, respectively, where 78% of excess deaths occured in patients with ≥1 comorbidity. CONCLUSIONS: Dramatic reductions were detected in the demand for, and supply of, cancer services which have not fully recovered with lockdown easing. These may contribute, over a 1-year time horizon, to substantial excess mortality among people with cancer and multimorbidity. It is urgent to understand how the recovery of general practitioner, oncology and other hospital services might best mitigate these long-term excess mortality risks.
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COVID-19/epidemiología , Modelos Estadísticos , Neoplasias/epidemiología , Pandemias , Vigilancia de la Población , SARS-CoV-2 , Adulto , Causas de Muerte/tendencias , Inglaterra/epidemiología , Femenino , Estudios de Seguimiento , Humanos , Masculino , Persona de Mediana Edad , Multimorbilidad/tendencias , Tasa de Supervivencia/tendencias , Factores de TiempoRESUMEN
We postulate that exit from pluripotency involves intermediates that retain pluripotency while simultaneously exhibiting lineage-bias. Using a MIXL1 reporter, we explore mesoderm lineage-bias within the human pluripotent stem cell compartment. We identify a substate, which at the single cell level coexpresses pluripotent and mesodermal gene expression programmes. Functionally these cells initiate stem cell cultures and exhibit mesodermal bias in differentiation assays. By promoting mesodermal identity through manipulation of WNT signalling while preventing exit from pluripotency using lysophosphatidic acid, we 'trap' and maintain cells in a lineage-biased stem cell state through multiple passages. These cells correspond to a normal state on the differentiation trajectory, the plasticity of which is evidenced by their reacquisition of an unbiased state upon removal of differentiation cues. The use of 'cross-antagonistic' signalling to trap pluripotent stem cell intermediates with different lineage-bias may have general applicability in the efficient production of cells for regenerative medicine.
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Reprogramación Celular , Mesodermo/metabolismo , Células Madre Pluripotentes/citología , Animales , Diferenciación Celular/genética , Línea Celular , Linaje de la Célula , Plasticidad de la Célula/genética , Autorrenovación de las Células , Medios de Cultivo , Células Madre Embrionarias/citología , Células Madre Embrionarias/metabolismo , Humanos , Ratones , Células Madre Pluripotentes/metabolismo , Transducción de Señal/genéticaRESUMEN
Retinoids regulate a wide spectrum of cellular functions from the embryo throughout adulthood, including cell differentiation, metabolic regulation, and inflammation. These traits make retinoids very attractive molecules for medical purposes. In light of some of the physicochemical limitations of retinoids, the development of drug delivery systems offers several advantages for clinical translation of retinoid-based therapies, including improved solubilization, prolonged circulation, reduced toxicity, sustained release, and improved efficacy. In this Review, we discuss advances in preclinical and clinical tests regarding retinoid formulations, specifically the ones based in natural retinoids, evaluated in the context of regenerative medicine, brain, cancer, skin, and immune diseases. Advantages and limitations of retinoid formulations, as well as prospects to push the field forward, will be presented.
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Sistemas de Liberación de Medicamentos/métodos , Medicina Regenerativa/métodos , Retinoides/administración & dosificación , Animales , Encefalopatías/tratamiento farmacológico , Diferenciación Celular/efectos de los fármacos , Ensayos Clínicos como Asunto , Composición de Medicamentos , Sistemas de Liberación de Medicamentos/tendencias , Células Madre Embrionarias/citología , Células Madre Embrionarias/efectos de los fármacos , Humanos , Enfermedades del Sistema Inmune/tratamiento farmacológico , Neoplasias/tratamiento farmacológico , Medicina Regenerativa/tendencias , Retinoides/química , Retinoides/uso terapéutico , Transducción de Señal , Enfermedades de la Piel/tratamiento farmacológicoRESUMEN
BACKGROUND: The reasons why some patients with COVID-19 develop pneumonia and others do not are unclear. To better understand this, we used multiparameter flow cytometry to profile circulating leukocytes from non-immunocompromised adult patients with PCR-proven COVID-19 and specifically compared those with mild symptoms with those who had developed pneumonia. METHODS: Using clinically validated antibody panels we studied leukocytes from 29 patients with PCR-proven COVID-19. Ten were hypoxic requiring ventilatory support, eleven were febrile but otherwise well, and eight were convalescing having previously required ventilatory support. Additionally, we analysed patients who did not have COVID-19 but received ventilatory support for other reasons. We examined routine Full Blood Count (FBC) specimens that were surplus to routine diagnostic requirements; normal ranges were established in a historic group of healthy volunteers. FINDINGS: We observed striking and unexpected differences in cells of the innate immune system. Levels of CD11b and CD18, which together comprise Complement Receptor 3 (CR3), were increased in granulocytes and monocytes from hypoxic COVID-19 patients, but not in those with COVID-19 who remained well, or in those without COVID-19 but ventilated for other reasons. Granulocyte and monocyte numbers were unchanged, however Natural Killer (NK) cell numbers were two-fold higher than normal in COVID-19 patients who remained well. INTERPRETATION: CR3 is central to leukocyte activation and subsequent cytokine release in response to infection. It is also a fibrinogen receptor, and its over-expression in granulocytes and monocytes of patients with respiratory failure tables it as a candidate effector of both the thrombotic and inflammatory features of COVID-19 pneumonia, and both a biomarker of impending respiratory failure and potential therapeutic target. NK cells are innate immune cells that retain immunological memory. Rapid expansion of memory NK cells targeting common antigens shared with other Coronaviruses may explain why most patients with COVID-19 do not develop respiratory complications. Understanding the innate immune response to SARS-CoV-may uncover why most infected individuals experience mild symptoms, and inform a preventive approach to COVID-19 pneumonia in the future.