Project description:In order to assess the descendants of hypertrophic chondrocytes, we utilized Collagen10-Cre;Rosa26-tdTomato mouse total bone isolated at e16.5 by Collagenase II digestion after mechanical digestion and soft tissue removal. After sequencing and downstream analysis using Seurat, we observed clusters of cells with gene profiles matching classically defined chondrocytes, skeletal stem and progenitor cells (SSPCs), and osteoblasts. Trajectory analysis reveals that the SSPCs lie intermediate to the transition of chondrocyte to osteoblast. We conclude that hypertrophic chondrocytes dedifferentiate to this progenitor stage before further differentiation.
Project description:In order to assess the descendants of hypertrophic chondrocytes, we utilized Collagen10-Cre;Rosa26-tdTomato mouse bone marrow harvested at 2 months of age by centrifugation and light Collagenase II digestion. After sequencing and downstream analysis using Seurat, we observed clusters of cells with gene profiles matching classically defined skeletal stem and progenitor cells as well as CXCL12 abundant reticular (CAR) cells. These cells appear to be upstream of both osteoblasts and adipocytes. We conclude that hypertrophic chondrocytes dedifferentiate to this progentior stage before further differentiation.
Project description:In order to assess the descendants of lateral plate mesoderm within the muscle interstitium, we utilized Prrx1Cre;Rosa26-tdTomato P21 tdTomato+ FACS sorted muscle interstitial cells
Project description:Mesp1-Cre+ cells from E7.5 mouse embryos (from the cross Mesp1-Cre/+ x Rosa26-Gli3R-IRES-YFP/tdTomato) were sorted by FACS where wild type (tdTomato-expressing) and mutant (Gli3R + YFP co-expressing) cells were collected separately from single litters.
Project description:We performed lineage tracing experiments using VE-Cadherin-Cre;LoxP-tdTomato mice. In these mice, endothelial cells (ECs) and their progeny are permanently marked by tdTomato fluorescence. We found that a substantial subset of stromal cells is derived from ECs, as indicated by their tdTomato expression. These findings support the notion that endothelial to mesenchymal transition (EndoMT) contributes to hematopoietic bone marrow niche formation in mice. Here we sought to determine the transcriptomic differences between endothelial-derived (tdTomato-positive) and non-endothelial-derived (tdTomato-negative) bone marrow stromal cells (BMSCs) and osteo/chondrolineage progenitor cells (OLCs). Murine niche populations were obtained from collagenased bone fraction of VE-Cadherin-Cre;LoxP-tdTomato mice at 3 weeks (n=2) or 11 weeks (n=2) of age. BMSCs (CD45-TER119-CD31-CD144-SCA-1+ CD51+ cells) and OLCs (CD45-TER119-CD31-CD144-Sca1-CD51+ cells) were FACS-purified and sequenced.
Project description:The experiment was performed to identify autophagy targets in wildtype and autophagy-deficient forebrain excitatory neurons. Therefore, neurons were isolated from the cortex, hippocampus and striatum of 2-3 weeks old Atg5flox/flox:CamKIIα-Cretg/wt:tdTomato+ (KO) and Atg5wt/wt:CamKIIα-Cretg/wt:tdTomato+ (WT) mice. Neurons in suspension were FACS sorted and excitatory forebrain neurons expressing tdTomato were forwarded to global proteome analysis assessed by LC-MS/MS.