Osteogenic differentiation of human small salivary gland fibroblast-like monoclonal cells

Objective Explore the method of culturing and proliferating human small salivary gland fibroblast-like monoclonal cells in vitro,identify the character of the cells and study the potential of osteoblast differentiation.Methods Human small salivary gland cells was primary cultured using tissue adherent method,we collect the fibroblast-like cells and clonal culture them using 96-well plates dilution decking method,and detect cell phenotype by immunofluorescence and RT-PCR.Then we start osteogenic inducing,and identify osteogenic differentiation through osteocalcin,collagen typeⅠimmunocytochemistry staining and alkaline phosphatase(AKP),alizarin red staining.Results We successfully amplify human small salivary gland fibroblast-like monoclonal cells in vitro,these cells express CD13,CD29,CD106 and CD44 proved by immunofluorescence,and express gene CK18,α-SMA,vimentin,CD106,nestin revealed by RT-PCR similar to human bone marrow-derived mesenchymal stem cells.After 8 days of osteogenic differentiation,the positive expression rate of AKP is 100%,after 19 days the expressive number of osteocalcin and collagen typeⅠis large,and there are many calcium nodules formation.Conclusion We can harvest a large number of human small gland fibroblast-like monoclonal cells through the isolation and culture system that we built,their immune phenotype is similar to mesenchymal stem cells,and they have good osteogenic potential in osteogenic induction culture conditions.