1. School of Stomatology, Shandong University, Jinan, China;
2. Shandong provincial key laboratory of oral tissue regeneration, Jinan, China;
3. Qilu hospital of Shandong University, Jinan, China.
* These two authors contributed equally to this work and should be considered as co-first authors.
Objectives: The present study established a non-contact coculture system in vitro, aiming to investigate the crosstalk between human dental pulp stem cells (hDPSCs) and human umbilical cord mesenchymal stem cells (hUCMSCs) on proliferation activity and osteogenic genes expression through paracrine.
Materials and methods: The stemness of hDPSCs and hUCMSCs were identified by flow cytometric analysis and multipotential differentiation assays. With the help of transwell inserts, the non-contact coculture system in vitro was established between hDPSCs and hUCMSCs. EdU labeling analysis and Western Blot were used to detect the proliferation activity. The mRNA and protein levels of osteogenic genes were evaluated by RT-PCR and Western Blot. The expression of elements in Akt/mTOR signaling pathway were detected by Western Blot.
Results: Both hDPSCs and hUCMSCs were positive to MSCs specific surface markers and had multi-differentiation potential. The proportion of EdU-positive cells increased and the expression of CDK6 and CYCLIN A were up-regulated in cocultured hDPSCs. Both prior coculture and persistent coculture improved mRNA and protein levels of osteogenic genes in hDPSCs. While in cocultured hUCMSCs, no statistical differences were observed on proliferation and osteogenesis. The phosphorylation of Akt and mTOR was up-regulated in cocultured hDPSCs.
Conclusions: The crosstalk between hDPSCs and hUCMSCs in coculture system increased the proliferation activity and enhanced osteogenic genes expression in hDPSCs. Akt/mTOR signaling pathway might take part in the enhancing effects in both cell proliferation and gene expression.
Keywords: human dental pulp stem cells, human umbilical cord mesenchymal stem cells, proliferation, osteogenesis, crosstalk.