ABSTRACT: To characterize murine ASPCs we performed bulk RNA-seq (BRB-seq, Alpern et al., 2019) of total, CD142− and CD142+ (Aregs) mouse adipose stem and progenitor cells (ASPCs). ASPCs were collected as Lin− (CD31− CD45− TER119−) CD29+ CD34+ SCA1+ cells of the mouse subcutaneous stromal vascular fraction using FACS. The cells were sequenced directly after sort or after expansion.
Project description:To study the involvement of key Areg (CD142+ ASPC)-specifics factors in the inhibitory capacity of CD142+ ASPCs on adipogenesis, we performed transcriptomic profiling of CD142− ASPCs exposed to the secretome of CD142+ ASPCs carrying knockdowns of the indicated genes. CD142− ASPCs were co-cultured and co-differentiated (within the transwell set-up) with CD142+ ASPCs in which siRNA-based knockdowns of specific genes were performed. ASPCs were collected as Lin− (CD31− CD45− TER119−) CD29+ CD34+ SCA1+ cells of the mouse subcutaneous stromal vascular fraction using FACS.
Project description:To characterize CD142+ ASPCs (Aregs) after exposure to an adipogenic cocktail we performed bulk RNA-seq (using BRB-seq) of total, CD142− and CD142+ mouse adipose stem and progenitor cells (ASPCs), sorted using four different anti-CD142 antibodies. ASPCs were collected as Lin− (CD31− CD45− TER119−) CD29+ CD34+ SCA1+ cells of the mouse subcutaneous stromal vascular fraction using FACS.
Project description:To validate that the robustness of Aregs' (CD142+ ASPCs') molecular identity regardless of the antibody used, we performed transcriptomic profiling of freshly isolated CD142− and CD142+ mouse adipose stem and progenitor cells (ASPCs) sorted using four different anti-CD142 antibodies. ASPCs were collected as Lin− (CD31− CD45− TER119−) CD29+ CD34+ SCA1+ CD142+/− cells of the mouse subcutaneous stromal vascular fraction using FACS.
Project description:To explore the molecular identity of mouse adipose stem and progenitor cells (ASPCs) and more specifically of CD142+ ASPCs (Aregs), we performed bulk RNA-seq on freshly isolated total, CD142− and CD142+ ASPCs of new-born pups (P0), post-natal day 16 mice (P16), 4 week-old (4wo), 7wo and 11wo mice. ASPCs were collected as Lin− (CD31− CD45− TER119−) CD29+CD34+ SCA1+ cells of the mouse subcutaneous stromal vascular fraction using FACS.
Project description:To study the impact of retinoic acid treatment on the transcriptome of CD142− ASPCs, we performed bulk RNA-seq on CD142− ASPCs treated with different RA concentrations ranging between 0.01 to 100 μM, with 100 ng/ml of BSA (negative control), DMSO (negative control) or 100 ng/ml of EGF (positive control).
Project description:Population control for the scRNA-seq based analysis a well-established fraction of mouse subcutaneous adipose-derived stromal vascular fraction (SVF) cells that is generally considered to harbour adipogenic stem and progenitor cells (ASPCs). We collected Lin- (CD31- CD45- TER119-) CD29+ CD34+ SCA1+ cells from the mouse subcutaneous SVF of transgenic mice, in which red fluorescent protein (RFP) is induced in Dlk1-expressing cells upon feeding with tamoxifen. While CD29, CD34, and SCA1 are generally expected to enrich for stem cells, DLK1 has previously been suggested to specifically mark pre-adipocytes.
Project description:We analysed a well-established fraction of mouse subcutaneous adipose-derived SVF cells that is generally considered to harbour adipose stem and progenitor cells (ASPCs). To study the molecular characteristics and the subpopulation structure of ASPCs, we performed three replicate scRNA-seq experiments. We collected Lin- (CD31- CD45- TER119-) CD29+ CD34+ SCA1+ cells from the mouse subcutaneous SVF of transgenic mice, in which red fluorescent protein (RFP) is induced in Dlk1-expressing cells upon feeding with tamoxifen. While CD29, CD34, and SCA1 are generally expected to enrich for stem cells, DLK1 has previously been suggested to specifically mark pre-adipocytes. The submission includes the raw data of all sequenced cells, while we only processed 208 high quality single cells. RFP status is indicated in the processed files.
Project description:To characterize the newly discovered Areg cells, we sampled at least four replicates of mouse adipose stem and progenitor cells (ASPCs), CD142+ ASPCs and CD142- ASPCs at four distinct time-points: (1) immediately post-sort (D0), (2) after attachment (5 hours after plating, 5H), (3) after culturing (24 hours after plating, D1) and (4) after adipogenic differentiation (day 12 after plating, day 8 after induction of differentiation, D12), and investigated their transcriptome through a highly multiplexed 3 polyA-selective mRNA-sequencing assay.
Project description:To validate the anti-adipogenic effect of the newly discovered Areg cells in human, we used FACS to separate the CD142+ from the CD142- population within cultured (passage 1) adipogenic stem and progenitor cells (ASPCs) (CD31- CD45- stromal vascular fraction cells) that were derived from four individuals, and induced adipogenesis. We then performed 3 RNA-seq expression profiling of the induced cells.
Project description:We used the resolving power of single-cell transcriptional profiling to molecularly characterize the mouse adipose stem and progenitor cell-enriched, subcutaneous adipose stromal vascular fraction. We molecularly assessed CD45- CD31- SVF cells using the 10x Genomics Chromium (10x) platform.