An In Vitro Study On The Interaction Of Human Umbilical Cord Mesenchymal Stem Cells With Dental Pulp Cells In Co-culture System

BackgroundCaries,endodontics,periodontal diseases are common oral disease causing pulp necrosis.More and more researches show that there is of a close bi-directional relationship between these diseases and people's oral health or illness.Therefore,with the development of biotechnology and biomechanical methods such as cell biology and molecular biology,the research of pulp tissue regeneration engineering has become the focus of tissue study in recent years.Objective:To investigated whether human umbilical cord mesenchymal stem cells(hUCMSCs)could be differentiate into odontoblast-like cells by coculture with dental pulp cells(DPCs)which has induced by BMP2 and explored the optimum ratio of two cells.Methods:1. Isolation,culture and identification of human umblilical cord mesenchymal stem cells.Cells were collected to identify their surface markers and examin the multilineage differentiation capability.2 Isolation,culture and identification of human dental pulp cells.Cells were evaluated the expression of for vimentin,cytokeratin by immunohistochemistry to identify cell sources.3 A co-culture system of human umbilical cord mesenchymal stem cells and human dental pulp cells that induced by BMP-2 was established in vitro.Human umbilical cord mesenchymal stem cells and human dental pulp cells were co-culture at the ratio of 1:1,1:5,5:1.The optimum ratio of the group was selected to the further experiment.The formation of calcium nodule were stained by alizarin red staining at 21 days.The expression of DSPP,ALP,DMP1,OCN,NANOG,VEGF,HGF gene were detected by Real-time quantitative polymerase chain reaction(qPCR)after cultured at 7days and 14 days.Results:1 There were spindle-shaped cells moving out from the umbilical cord block at the 5th day.Passaged cells showed a rapid proliferation and growth in vitro.Flow cytometry showed that CD29 and CD44 expreeeion were positive,but the CD34,CD45 were negative.The result of Alizarin red staining and Oil red O staing indicated that the cell could be differentiated into bone and adipose tissue.2 The cells moving out from the dental pulp block at the 3 days.Immunohistochemical staining revealed persian protein positive expression and keratin negative expression.3 The results showed that the formation of calcified nodules in 1:1 group was significantly higher than in hUCMSCs group,but close to that in DPCs.The result of qPCR showed that the mRNA expression of DSPP,ALP,DMP1,OCN,VEGF,HGF in 1:1 group were significantly higher than in hUCMSCs;The mRNA expression of NANOG in 1:1 group were lower than hUCMSCs group.Conclusion:1 The human umbilical cord mesenchymal stem cells and the dental pulp cells show a strong ability to grow in vitro.2 The hUCMSCs can be induced to differentiate into odontoblastoid-like cells by the DPCs.