Effect Of Induction On The Differentiation Of Rat Bone Marrow Mesenchymal Stem Cells Co-cultured With Periodontal Ligament Fibroblasts

【Objective】To investigate the feasibility of inducing the differentiation of rat bone mesenchymal stem cells (BMSCs)under the influence of periodontal ligament fibroblasts(PDLFs)in vitro. To determine the mechanism of the induction:whether the contact is the essential condition .If there are only the factors secreted from the periodontal ligament fibroblast,the induction will occur or not. To find a theoretical basis for the differentiation of BMSCs in vivo,to provide a new way for BMSCs as seed cells for periodontal tissue engineering.【Method】We isolated the BMSCs by whole bone marrow adherent culture. Cells were identified by the cell morphology and growth characteristics,growth curve,and the ability to form bone and fat. Cultured the rat PDLFs by enzymatic digestion method combined with explant. Cells were identified by vimentin and keratin immuno-histochemical staining and mineralization. BMSCs were co-cultured with PDLFs at 1:1 ratio in a dual chamber dish separated by a semipermeable membrane. We also set both negative control and Positive control. Alkaline phosphatase (ALP) activity of cells was detected after 3,5 days in 3 groups. The mRNA expression of PDLFs protein markers(OCN,COLⅢ,BSP and Runx2) were measured at 14 days.【Result】Co-cultured with PDLFs could induce BMSCs to obtain periodontal ligament like characteristics. Co-culturing BMSCs with PDLFs led to a significant increase in BMSCs ALP activity and the mRNA expression of OCN,COLⅢand Runx2 ,and a significant decrease in the mRNA expression of BSP .There are no differences of all markers between co-cultured BMSCs and PDLFs cultured individually , and observed changes between co-cultured BMSCs and BMSCs cultured individually .【Conclusion】BMSCs could acquire characteristics of PDLFs in vitro by the non-contact co-culture system. Contact is not the essential condition of induction. The factors secreted by PDLFs may induce the rat BMSCs to obtain periodontal ligament-like marker expression.