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Single cell transcriptomic of ex vivo-generated lymphoid progenitors from mobilised peripheral blood- or corb blood-derived hematopoietic stem and progenitors cells

ABSTRACT: Single cell transcriptomic of ex vivo-generated lymphoid progenitors from mobilised peripheral blood- or corb blood-derived hematopoietic stem and progenitors cells

PROVIDER: PRJNA1241729 | ENA |

REPOSITORIES: ENA

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Single cell transcriptomic of ex vivo-generated lymphoid progenitors from mobilised peripheral blood- or corb blood-derived hematopoietic stem and progenitors cells

Project description:We previously established a feeder-free cell therapy platform for the ex vivo generation of lymphoid-primed progenitors using immobilized Delta-like ligand 4 (DLL4), and hereafter known as ProTcell. We demonstrated these progenitors can be generated from hematopoietic stem and progenitors cells originating from cord blood (CB) and mobilized peripheral blood (mPB). In vivo studies demonstrated that adoptive transfer of these progenitors accelerates T cell reconstitution following thymic engraftment (Moirangthem RD et al., 2021 ; Simons L et al., 2018 ; Reimann et al., 2012). To further explore the full therapeutic potential of this cell product, we performed a comprehensive characterization using single-cell RNA sequencing. We confirmed ProTcell samples derived from CB and mPB were highly similar,as they clustered together. Our analysis also revealed the presence of distinct cell subsets within the cellular product characterized mainly by their commitment to lymphoid lineages. We highlighted strong similarities with early stages of T cell development, underscoring the physiological relevance of our system.

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scRNAseq of innate lymphoid cells generated in vitro from umbilical cord blood derived CD34+ hematopoietic progenitors.

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Project description:RNA-seq of mouse common lymphoid progenitors, innate lymphoid cell progenitors and NK cell progenitors

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RNA-seq of mouse common lymphoid progenitors, innate lymphoid cell progenitors and NK cell progenitors

Project description:Natural Killer (NK) cells at different developmental stages (common lymphoid progenitor (CLP), innate lymphoid cell progenitors (ILCP), and refined NK progenitor (NKP)) were collected from Vav1+iCre FOXO1,3flox/flox mice (C57BL/6 background). Total RNA was harvested and sequenced with a strand-specific paired-end RNA-seq protocol.

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Project description:A common lymphoid progenitor cell culture system was established, maintaining a multipotent, proliferating state of CLPs. Cells can be cultured and expanded in vitro for several months, maintaining their in vitro and in vivo lymphoid differentiation potential. We were interested, whether cCLP are changing, with regards to their gene expression program, compared to in vivo CLPs, during their expansion in culture. Therefore we compared the gene expression profile of cultured cCLPs and their counterparts in vivo, by NextGenSeq. CLPs were sorted from bone marrow of 6-10 weeks old C57BL/6 mice (n=4). Cells were sorted for Lin-, IL7R+, Flk2+, CD27+, Ly6d- marker expressing CLPs by Flow Cytometry for gene expression analysis. In parallel the same cell population was sorted and cultured under cCLP culture conditions. After two months culture the cells sorted again for Lin-, CD27+, Ly6d- cells by Flow Cytometry for gene expression analysis (n=4). The analysis shows that cCLPs maintain a very similar transcription profile to ex vivo sorted common lymphoid progenitors.

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IRF-8 extinguishes neutrophil production and promotes dendritic cell lineage commitment in both myeloid and lymphoid progenitors (Affymetrix).

Project description:While most blood lineages are assumed to mature through a single cellular and developmental route downstream of hematopoietic stem cells (HSCs), dendritic cells (DCs) can be derived from both myeloid and lymphoid progenitors in vivo. To determine how distinct progenitors can generate similar downstream lineages, we examined the transcriptional changes that accompany loss of in vivo myeloid potential as common myeloid progenitors (CMPs) differentiate into common dendritic cell progenitors (CDPs), and as lymphoid-primed multipotent progenitors (LMPPs) differentiate into all lymphoid progenitors (ALPs). Microarray studies revealed that Interferon regulatory factor 8 (IRF-8) expression increased during each of these transitions. Competitive reconstitutions using Irf8-/- bone marrow demonstrated cell-intrinsic defects in the formation of CDPs and all splenic dendritic cell subsets. Irf8-/- CMPs and, unexpectedly, Irf8-/- ALPs produced more neutrophils in vivo than their wild type counterparts at the expense of DCs. Retroviral expression of IRF-8 in multiple progenitors led to reduced neutrophil production and increased numbers of DCs, even in the granulocyte-macrophage progenitor (GMP), which does not normally possess conventional DC potential. These data suggest that IRF-8 represses a neutrophil module of development and promotes convergent DC development from multiple lymphoid and myeloid progenitors autonomously of cellular context. CMP (Lineage-c-kithiSca-1-CD11c- CD34+ Flk2+CD16/32-CD115- ) or CDP (Lin-c-kitintSca-1-CD34+Flk2+CD16/32-CD115+) were double sorted from the bone marrow of wild type C57BL/6 mice. RNA was extracted from 10,000-30,000 sorted cells using Trizol (Invitrogen) and linear acrylamide (Ambion), amplified using Affymetrix Two-Cycle Amplification and IVT kits (Affymetrix), and hybridized to Affymetrix Mouse Genome 430 2.0 chips.

2012-01-07 | E-GEOD-34892 | biostudies-arrayexpress

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