Fig. 4

Exosomes from odontoblasts with active mTORC1 and inactive mTORC1 inhibited DPSC odontoblastic differentiation at the same concentration. A DPSCs were incubated with PKH67-labeled exosomes from nontreated MDPC23 cells, MDPC23 cells with active mTORC1 and MDPC23 cells with inactive mTORC1 for 24 h. The nuclei of DPSCs were stained with DAPI. The results showed that exosomes that were isolated from these MDPC23 cells could be successfully taken up by DPSCs (scale bar = 100 μm). B qRT‒PCR analysis indicated that the expression levels of the odontoblastic differentiation-related genes ALP, COL1 and DMP1 in DPSCs could be significantly inhibited by exosomes derived from MDPC23 cells, regardless of whether mTORC1 was inhibited or activated in the MDPC23 cells (n = 3). C, D DPSCs lysates were analyzed by western blotting. The results indicated that the protein expression levels of ALP, COL1 and DMP1 in DPSCs were dramatically decreased by exosomes that were derived from MDPC23 cells, and there was a nonsignificant difference between exosomes that were isolated from MDPC23 cells with different levels of mTORC1 activity. Full-length blots are presented in Additional file 1: Fig. S5. E–G The variation in DPSC mineralization was examined by ALP activity staining and Alizarin Red S staining. The results revealed that compared with differentiated DPSCs with normal mineralization, the activity of ALP E and the formation of mineralized nodules F, G were inhibited by exosomes that were derived from MDPC23 cells, and the effects of exosomes derived from cells with different levels of mTORC1 activity were similar (scale bar = 400 μm). The values are presented as the means ± SDs. *p < 0.05, **p < 0.01, ***p < 0.001. DPSC Dental pulp stem cells, ALP alkaline phosphatase