RESUMO
Somatic mutations commonly occur in hematopoietic stem cells (HSCs). Some mutant clones outgrow through clonal hematopoiesis (CH) and produce mutated immune progenies shaping host immunity. Individuals with CH are asymptomatic but have an increased risk of developing leukemia, cardiovascular and pulmonary inflammatory diseases, and severe infections. Using genetic engineering of human HSCs (hHSCs) and transplantation in immunodeficient mice, we describe how a commonly mutated gene in CH, TET2, affects human neutrophil development and function. TET2 loss in hHSCs produce a distinct neutrophil heterogeneity in bone marrow and peripheral tissues by increasing the repopulating capacity of neutrophil progenitors and giving rise to low-granule neutrophils. Human neutrophils that inherited TET2 mutations mount exacerbated inflammatory responses and have more condensed chromatin, which correlates with compact neutrophil extracellular trap (NET) production. We expose here physiological abnormalities that may inform future strategies to detect TET2-CH and prevent NET-mediated pathologies associated with CH.
Assuntos
Dioxigenases , Neutrófilos , Humanos , Camundongos , Animais , Proteínas Proto-Oncogênicas , Células-Tronco Hematopoéticas/fisiologia , Medula Óssea , Hematopoese/genética , Mutação , Proteínas de Ligação a DNA/genética , Dioxigenases/genéticaRESUMO
Determining how hematopoietic stem and progenitor cells (HSPCs) can be infected by viruses is necessary to understand and predict how the immune system will drive the host response. We present here a protocol to analyze the capacity of SARS-CoV-2 to infect different subsets of human HSPCs, inlcuding procedures for SARS-CoV-2 production and titration, isolation of human HSPCs from different sources (bone marrow, umbilical cord, or peripheral blood), and quantification of SARS-Cov-2 infection capacity by RT-qPCR and colony forming unit assay. For complete details on the use and execution of this protocol, please refer to Huerga Encabo et al. (2021).
Assuntos
Medula Óssea/virologia , Teste de Ácido Nucleico para COVID-19/métodos , COVID-19/virologia , Ensaio de Unidades Formadoras de Colônias/métodos , Sangue Fetal/virologia , Células-Tronco Hematopoéticas/virologia , SARS-CoV-2/isolamento & purificação , COVID-19/patologia , Células-Tronco Hematopoéticas/patologia , HumanosRESUMO
Virtual reality simulators have the capability to automatically record user performance data in an unbiased, cost effective manner that is also less error prone than manual methods. Centralized data recording simplifies proficiency evaluation even more; however is not commonly available to date for surgical skills trainers. We will detail our approach in implementing a framework for distributed score recording over the Internet using a database for persistent storage.