Project description:Neutrophils have limited utility as transfusion product due to their short lifespan. Culturing neutrophils from stem cell sources holds potential for fundamental and translational purposes. Here, we cultured CD34+ hematopoietic stem cell-derived neutrophils and compared them with peripheral blood neutrophils in terms of morphology, phenotype, and function. Our culture system resulted in 60% morphologically mature CD15+CD11b+CD16high neutrophils with most effector functions almost indistinguishable from blood neutrophils, confirmed by a high similarity in transcription and protein abundance patterns. While exhibiting strong microbial killing capacity and antibody-dependent cellular cytotoxicity against tumor cells, these cells were deficient in myeloid-derived suppressor cell activity. Upon dissecting the underlying mechanism, this deficiency in immunosuppressive activity correlated with their distinct granular composition in comparison to blood neutrophils. Taken together, our cultured neutrophils closely resemble blood neutrophils, offering a repository for fundamental research and a step towards an effective transfusion product without immunosuppressive activity.
Project description:The variation among induced pluripotent stem cells (iPSCs) in their differentiation capacity to specific lineages is frequently attributed to somatic memory. In this study, we compared hematopoietic differentiation capacity of 35 human iPSC lines derived from four different tissues and four embryonic stem cell lines. The analysis revealed that hematopoietic commitment capacity (PSCs to hematopoietic precursors) is correlated with the expression level of the IGF2 gene independent of the iPSC origins. In contrast, maturation capacity (hematopoietic precursors to mature blood) is affected by iPSC origin; blood-derived iPSCs showed the highest capacity. However, some fibroblast-derived iPSCs showed higher capacity than blood-derived clones. Tracking of DNA methylation changes during reprogramming reveals that maturation capacity is highly associated with aberrant DNA methylation acquired during reprogramming, rather than the types of iPSC origins. These data demonstrated that variations in the hematopoietic differentiation capacity of iPSCs are not attributable to somatic memories of their origins. human pluripotent stem cell-derived hematopoietic precursor cells (n = 33)
Project description:In vertebrates, lifelong supply of all the blood cell types in suitable numbers is maintained by the hematopoietic stem cells (HSCs). During development, these HSCs emerge in the aorta-gonad-mesonephros (AGM) from specialized vascular endothelium through a transdifferentiation process, called as endothelial-to-hematopoietic transition (EHT). During this process, select endothelial cells (CD31+c-kit- or CD31PCKITN) switch to a hematopoietic transcriptional program, undergo morphological changes and become hemogenic (CD31+c-kit+ or CD31PCKITP) and gives rise to hematopoietic cells (CD31-c-kit+ or CD31NCKITP). A complex interplay of key transcription factors and signaling pathways coordinates the whole process. Specific metabolic signature of a cell can precisely define its phenotype. Evidence has emerged that cellular phenotype and function can be driven according to the changes in cellular metabolism. Metabolic programs, which are stage specific, allow stem cells to adapt their function in different microenvironments. In the present study, we performed nano LC-MS/MS based proteomic analysis to understand the molecular program involved in the transdifferentiation of endothelial to hematopoietic cells.
Project description:The variation among induced pluripotent stem cells (iPSCs) in their differentiation capacity to specific lineages is frequently attributed to somatic memory. In this study, we compared hematopoietic differentiation capacity of 35 human iPSC lines derived from four different tissues and four embryonic stem cell lines. The analysis revealed that hematopoietic commitment capacity (PSCs to hematopoietic precursors) is correlated with the expression level of the IGF2 gene independent of the iPSC origins. In contrast, maturation capacity (hematopoietic precursors to mature blood) is affected by iPSC origin; blood-derived iPSCs showed the highest capacity. However, some fibroblast-derived iPSCs showed higher capacity than blood-derived clones. Tracking of DNA methylation changes during reprogramming reveals that maturation capacity is highly associated with aberrant DNA methylation acquired during reprogramming, rather than the types of iPSC origins. These data demonstrated that variations in the hematopoietic differentiation capacity of iPSCs are not attributable to somatic memories of their origins. Bisulfite converted genomic DNA lysates from human pluripotent stem cell-derived hematopoietic precursor cells (CD34+CD38-CD43+ lineage marker-) were hybridized to Illumina HumanMethylation450 BeadChip.
Project description:The variation among induced pluripotent stem cells (iPSCs) in their differentiation capacity to specific lineages is frequently attributed to somatic memory. In this study, we compared hematopoietic differentiation capacity of 35 human iPSC lines derived from four different tissues and four embryonic stem cell lines. The analysis revealed that hematopoietic commitment capacity (PSCs to hematopoietic precursors) is correlated with the expression level of the IGF2 gene independent of the iPSC origins. In contrast, maturation capacity (hematopoietic precursors to mature blood) is affected by iPSC origin; blood-derived iPSCs showed the highest capacity. However, some fibroblast-derived iPSCs showed higher capacity than blood-derived clones. Tracking of DNA methylation changes during reprogramming reveals that maturation capacity is highly associated with aberrant DNA methylation acquired during reprogramming, rather than the types of iPSC origins. These data demonstrated that variations in the hematopoietic differentiation capacity of iPSCs are not attributable to somatic memories of their origins. Bisulfite converted genomic DNA lysates from human pluripotent stem cell-derived hematopoietic precursor cells (CD34+CD38-CD43+ lineage marker-) were hybridized to Illumina HumanMethylation450 BeadChip.