Figure 3.

iOB transcriptomes resemble osteoblast transcriptomes. Genome wide RNA-seq on fibroblasts, calvarial-OBs, and iOBs was performed in triplicate for each group. (A) Osteoblast and bone related gene expression levels. Log₂-fold change of expression level and −log₁₀ (P value) in iOBs versus fibroblasts are shown on the x axis. (B) A plot of gene expression changes in iOBs versus fibroblasts. The log₂-fold change for gene expression level is plotted on the y-axis and the average of the counts normalized by size factor is shown on the x-axis. Each gene is represented with a dot. Differentially expressed genes with an adjusted p (FDR) value below 0.05 are shown in red. Compared to the fibroblast population, 3222 genes were significantly upregulated, and 2427 genes were downregulated in iOB cells. Bglap2 was the most significantly upregulated gene based on the lowest adjusted p value (FDR=1.06E-197). (C) and (D) Gene Ontology Analysis of biological process for significantly upregulated (C) and downregulated (D) genes in iOBs versus fibroblasts. −log₁₀ (P value) of gene enriched terms were shown. (E) Heatmap of sample-to-sample distances using rlog-transformed values of RNA-seq data. For each sample, the distance calculation is based on the contribution of rlog from all the genes. (F) PCA plot using the rlog-transformed values of RNA-seq data. Each color represents a sample type. Each sample has three replicates. (G) Heatmap of relative rlog-transformed values of total genes across samples. Each row represents a gene. Each column represents a cell type. (H) Heatmap of relative rlog-transformed values of differentially expressed genes in iOBs versus fibroblast across samples. Each row represents a gene. Each column represents a cell type. (E-H) Fib: Fibroblast; Cal: Calvarial. (I) Gene set enrichment analysis (GSEA) show osteogenic genes are enriched in iOBs. Normalized enrichment score, NES:1.57, FDR q=0 and p=0.