Extended Data Fig. 6: Gene expression analysis in CTSK–mGFP cells isolated from mouse femurs. | Nature

Extended Data Fig. 6: Gene expression analysis in CTSK–mGFP cells isolated from mouse femurs.

From: Discovery of a periosteal stem cell mediating intramembranous bone formation

Extended Data Fig. 6

a, Bulk RNA-seq analysis of FACS-isolated PSC, PP1, PP2 and non-CTSK-mGFP MSCs from 6-day-old mouse femurs. Hierarchical clustering analysis was performed on RNA-seq data. b, Heat map generated from bulk RNA-seq of FACS-sorted cells shows differences in gene expression between PSCs and the progenitor populations, PP1 and PP2 cells. c, Von Kossa staining indicates bone organoid formation by PSCs (left), PP1 (middle) and PP2 cells (right) 5 weeks after transplantation into the kidney capsule. Scale bar, 20 µm. Representative images from 3 independent experiments with 3 mice per group. d, Significantly reduced bone formation (bone volume) in PP1 (*P = 0.04) and PP2 (*P = 0.032) cells compared to PSCs after transplantation. Two-tailed Student’s t-test. Data are mean ± s.d., n = 3 independent experiments. e, Relative expression of Tnn (i), Tnmd (ii), Ifitm5 (iii) and Bglap (iv) among the four clusters (identified by 1–4) that were generated through analysis of 658 CTSK–mGFP+ mesenchymal cells using the Seurat package. Cell clusters (1–4) along the x axis. f, Expression of genes such as Bglap (i), Alpl (ii), Ifitm5 (iii), Tnn (iv), Tnmd (v) and Kera (vi) are shown by pseudocolouring of t-SNE plots. g, Heat map generated from bulk RNA-seq shows differences in gene expression between PSCs and the progenitor populations, PP1 and PP2 cells. hk, Monocle analysis of CEL-Seq2 data. h, Bright-field image of a colony that was generated from single-cell sorting of RAW264.7 cells by FACS. Scale bar, 20 µm. Representative image from 3 independent experiments. i, Graphs indicate the percentage of wells that received sorted cells by FACS (left) and the percentage of doublets detected in those wells (right). Data are mean ± s.d., n = 4. j, Data represent the total amount of mRNA in the two 384-well plates (plate 47 and plate 48) that were sequenced using CEL-Seq2. k, Pie chart, showing that the analysed CTSK–mGFP+ cells were mesenchymal in origin.

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