Fig. 5
High-fat diet (HFD) feeding disturbs the skeletal cell phenotype and neural-skeletal cell communication within the periosteal microenvironment. 12 weeks after the initiation of the dietary treatment, four left femurs and tibias from ND and HFD mice were dissected. Periosteal cells were isolated, and scRNA-sequencing and analysis were performed. a UMAP of mesenchymal cell subclusters including mesenchymal progenitors, pre-osteoblasts, and osteoblasts, along with known gene markers for each by violin plot, and the cell number ratio among ND and HFD treated groups. Black indicates ND cells. Gray indicates HFD cells. b Heatmap for differentially expressed genes (DRGs) in each mesenchymal cell subcluster. c UMAP showing the pseudotime trajectory of the mesenchymal cell subclusters along with progenitor gene markers (Pdgfrα and Ly6a) as well as osteoblast markers (Bglap and Alpl). Black line represents the trajectory graph. Plots were generated using Monocle 3.0.1.2. d–g’ Linear graph analysis of phenotype changes across pseudotime including gene modules for stemness, proliferation, adipogenesis and osteoblastogenesis among ND (blue line) and HFD (black line) fed mesenchymal cells, and module index scoring of mesenchymal cell subclusters (mesenchymal progenitors, pre-osteoblasts and osteoblasts). Dashed lines indicate delineates early, mid and late pseudotime. h–k’ Linear graph analysis of dysregulated signaling pathways, including MAPK, TGFβ, Wnt and mTor signaling, across pseudotime among ND (blue line) and HFD (black line) fed groups and module index scoring of mesenchymal cell subclusters. Dashed gray lines in module score graphs represent early, mid, and late pseudotime. Graphs represent average values ± 1 SD. Module scoring data was analyzed using the Kolmogorov-Smirnov test. *P < 0.05, **P < 0.01, ***P < 0.001 and ****P < 0.00 01 in comparison to ND control. 1 255 total mesenchymal cells analyzed
