Fig. 1

Osteogenic/dentinogenic differentiation of SCAP. a, b SCAP were cultured with or without osteogenic/dentinogenic medium (Os/Den) for 4 weeks. Alizarin Red-S staining shows the calcium deposition. Representative images of Alizarin Red-S staining. Bars = 25 mm (a). Calcium accumulation assay. ***P < 0.005. Graph bars show the means ± SEM (b). c, d SCAP were cultured with or without osteogenic/dentinogenic medium (Os/Den) for 1 week. Western blot analysis shows the expression of osteogenic/dentinogenic markers including runt-related nuclear factor 2 (RUNX2), bone gamma-carboxyglutamate protein (BGLAP), and dentin phosphoprotein (DSPP) in SCAP. Representative expression of RUNX2, BGLAP, and DSPP (c). Relative expression of RUNX2, BGLAP, and DSPP to β-actin (ACTB) (d). e–g In vivo calcified tissue formation assay was performed in immunocompromised mice using hydroxyl apatite/tricalcium phosphate (HA) as carriers with or without SCAP. Representative histological images of transplant tissues (TP) by hematoxylin and eosin staining (HE). Bars = 100 μm (e). Representative immunofluorescent images staining with anti-human mitochondria (hMt) antibody. Bars = 50 μm (f). RT-qPCR analysis shows the expression of RUNX2, BGLAP, and DSPP in the transplant tissues. Results shown as the ratios to the expression of the corresponding 18S ribosomal RNA (18S). nd, no detect (g). h–k SCAP were cultured with or without osteogenic/dentinogenic medium (Os/Den) for 1 weeks. Western blot analysis shows the phosphorylation of AKT and mammalian target of rapamycin (mTOR) in SCAP. Representative images of the expression of AKT, phosphorylated AKT (p-AKT), mTOR, and phosphorylated mTOR (p-mTOR) (h, j). Relative phosphorylation of AKT and mTOR (i, k). b, d, g, i, k n = 5 for all groups. *P < 0.05, ***P < 0.005. Graph bars show the means ± SEM. e, f CT, connective tissue; HA, hydroxyapatite/triphosphate