Single cell transcriptome of cultured thymic progenitor and stem cells
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ABSTRACT: Purpose: The goal of this study is to characterise expanding thymic progenitor and stem cells in culture Methods: FACS-isolation protocol to separate epithelial cells from mouse feeder layer cells then loaded into the 10x Genomics Chromium Platform, and sequenced using Illumina HiSeq 4000. Conclusions: Our study dissected in vitro properties of thymic progenitor and stem cells
Project description:Purpose: The goal of this study is to characterise expanding thymic progenitor and stem cells from whole epithelial as well as cortical, medullary thymus sub-compartments. As reference, we have sequenced skin keratinocyte progenitor and stem cells Methods: Cells have been expanded for 4 passages on mouse feeder layer cells then loaded into the 10x Genomics Chromium Platform, and sequenced using Illumina HiSeq 4000. Conclusions: Our study compared thymic progenitor and stem cells from different origin and dissected in vitro properties of thymic and skin progenitor and stem cells
Project description:Purpose: The goal of this study is to characterise expanding epithelial progenitor and stem cells in culture from different organs and compare them to thymic ones Methods: FACS-isolation protocol to separate epithelial cells from mouse feeder layer cells then loaded into the 10x Genomics Chromium Platform, and sequenced using Illumina HiSeq 4000. Conclusions: Our study dissected in vitro properties of airways and skin progenitor and stem cells and showed they differ from thymic ones
Project description:Purpose: The goal of this study is to characterise cortical (cTECs) and medullary (mTECs) epithelial cells of the human thymus. Methods: FACS-isolation protocol to separate 4 different epithelial cells populations from total dissociation of human thymus that is negative for hCD45 and either positive for EPCAM or CD205, then loaded into the 10x Genomics Chromium Platform, and sequenced using Illumina HiSeq 4000. Conclusions: Our study identified and characterised human thymic stem/progenitors cells, as well as identified new specialised cell clusters in each of the two compartments
Project description:Purpose: The goal of this study is to characterise the large variety of stromal cells populations, particularly cortex (cTECs) and medullary (mTECs) epithelial cells. It has been speculated that mTECs and cTECs are composed of functionally distinct subsets with different clonogenic potential. Methods: FACS-isolation protocol to separate 4 different epithelial cells populations from total dissociation of human thymus that is negative for hCD45 then loaded into the 10x Genomics Chromium Platform, and sequenced using Illumina HiSeq 4000. Conclusions: Our study dissected common clusters to human thymic epithelial cells in cortex and medulla containing stem cells markers conserved when expanded in vitro, as well as identified specialised cell clusters in each of the two compartment
Project description:Purpose: The goal of this study is to characterise the follicular dendritic cells (FDCs) upon antigen-immunocomplex immunisation. Methods: FACS sorting of Live/Death- Podoplanin+ CD31- CD45- CXCL13-TdTomato+ after LN disgregation of CXCL13-CreTdTomato mice. Mice has been previously immunised with antigen-Ics at different time-points. Cells were loaded into 10x Genomics Chromium Platform, and sequenced using Illumina HiSeq 4000. Conclusions: Our study detects previously identified stromal cell populations and shows the heterogeneity present on the FDCs. We detect 3 different FDC clusters.
Project description:Purpose: To establish the steady state heterogeneity of murine Dendritic Epidermal T Cells (DETC) and to investigate the function of their constitutive interaction with neighbouring keratinocytes via Skint1 Methods: CD3+ cells were FACS purified from ear epidermal suspensions prepared from FVB mice administered isotype control or anti-Skint1 antibody 7d prior, then loaded into the 10x Genomics Chromium Platform and sequenced using Illumina HiSeq 4000. Conclusions: This analysis identifies four states of DETC in unperturbed epidermis, and reveals that DETC TCR sensing of keratinocytes via Skint1 potentially enables natural immune surveillance and maintains their thymic maturation signature in the periphery
Project description:We report the identification of immature thymic CD4(-),CD8(-) double-negative (DN)1e cells with the NKT cell lineage potential. We also analyzed the gene expression profiles of DN1e thymocytes compared with those of mature thymic NKT cell developmental stages termed NKT stage-1, 2, and -3, which are characterized by differential expression levels of NK1.1 and CD44 antigens in C57BL/6 mouse strain. Next generation sequencing of total transcriptomes using total RNA isolated from FACS sorted ex vivo thymic DN1eP (Lin-/CD44+/CD25-/CD24low/CD5+/CD27+/Ly108-/CXCR3+) fraction, and mature thymic alphaGalCer-loaded CD1d dimer+TCRbeta+ NKT cell developmental stage-1 (CD44-/NK1.1-), stage-2 (CD44+/NK1.1-), and stage-3 (CD44+/NK1.1+) cells.
Project description:The ligand for the c-Kit receptor, KitL, exists as a membrane-associated (mKitL) and a soluble form (sKitL). KitL functions outside c-Kit activation have not been identified. We show that co-culture of c-Kit– and mKitL–expressing NIH3T3 cells results in signaling through mKitL: c-Kit–bound mKitL recruits calcium-modulating cyclophilin ligand (CAML) to selectively activate Akt, leading to CREB phosphorylation, mTOR pathway activation, and increased cell proliferation. Activation of mKitL in thymic vascular endothelial cells (VECs) induces mKitL- and Akt-dependent proliferation, and genetic ablation of mKitL in thymic VECs blocks their c-Kit responsiveness and proliferation during neonatal thymic expansion. Therefore, mKitL–c-Kit form a bi-directional signaling complex that acts in the developing thymus to coordinate thymic VEC and early thymic progenitor (ETP) expansion by simultaneously promoting ETP survival and VEC proliferation. This mechanism may be relevant to both normal tissues and malignant tumors that depend on KitL–c-Kit signaling for their proliferation.
Project description:Purpose: The goal of this study is to understand the progressive patterning of neurogenesis of the developing zebrafish hindbrain. 16hpf, 24hpf and 44hpf zebrafish hindbrains were used for single-cell RNA-sequencing with the aim to uncover hindbrain development. Methods: 40 microdissected hindbrains per each stage were dissociated at loaded into the 10x Genomics Chromium Platform, and sequenced using Illumina HiSeq 4000. Conclusions: Our study constitute a resource of hindbrain gene expression during development. We have identified transcriptional programs involved in: rhombomere segmental identity, dorso-ventral patterning, boundary and centre progenitor cells and temporal regulation of neurogenesis.
Project description:Cells were generated for a single cell atlas of adult human skin to dissect the cellular and molecular organisation underpinning human skin immune barrier function. Surplus skin from breast reconstruction surgery was collected (n=3), then the top 200 μm layer was taken. Epidermis was separated from the dermis, and both layers were separately digested. Single cells were FACS sorted into eight categories, then loaded onto a 10x Genomics Chromium Controller, and sequenced on an Illumina HiSeq 4000.