| Tooth autotransplantation is a surgical treatment that moves a tooth from its originallocation to another site to recover its physiological function. It was reported that the toothautotransplantation had a success rate of90%, suggesting that it may be a potentiallybeneficial treatment for tooth loss.There are many factors that influence the prognosis of an autotransplanted tooth, such asextraoral time, adaptability between the recipient bone and root surface, age and others.All of these factors can be attributed to the status of periodontal tissue and its healing.Hence, improving the periodontium status to achieve periodontal ligament healing is a keyfactor to increase the success rate of tooth autotransplantation.Periodontal ligament stem cells (PDLSCs) are known to exist in the periodontium and may play an important role in periodontium healing, which can be used to enhance theperiodontium status and the success rate of tooth autotransplantation. However, thePDLSC source is limited because the cells can only be obtained from a planned operationin advance. Bone marrow mesenchymal stem cells (BMMSCs) can be used as substitutemesenchymal stem cells (MSCs) and are multi-potent adult stem cells.7Nevertheless, bonemarrow harvesting is an invasive procedure. Therefore, it is necessary to explore othersubstitute stem cells for practical application.Umbilical cord mesenchymal stem cells (UCMSCs) may be a good choice for this purpose.Wharton·s jelly contains many fibroblast-like mesenchymal cells that have the capacity forself-renewal and multi-potent differentiation, resembling many BMMSC properties. Inaddition, the stem cells harvested from the umbilical cord are abundant in the cell source,are easy to acquire, present no ethical difficulties and exhibit low-immunogenicity. All ofthese characteristics are suitable for tooth autotransplantation application.In this study, in vitro experiments were used to investigate the stem cell markers andosteogenic properties of hUCMSCs. Based on previous findings, in vivo experiments wereused to evaluate the root/periodontal tissue formation. The findings will provide guidelinesfor increasing the success rate of tooth autotransplantation.Part1The primary culture and identification of the humanumbilical cord mesenchymal stem cellsObjective: To obtain the mesenchymal stem cells needed in this research by primaryculture and identify the characteristic of the cells by flow cytometry. Method: The primaryculture of hUCMSCs was obtained using enzymatic isolation method. The related markers(CD29, CD44, CD146, CD105, SOX2, CD34and CD45) were analysed using a flowcytometer. Result: Plenty of hUCMSCs could be obtained by enzymatic isolation method. Flow cytometric analysis showed that the cells expressed CD44, CD105, CD29, CD146and SOX2but not hematopoietic lineage markers (CD34and CD45). Conclusion: Wecould get the hUCMSCs in steady and quantity. The cells could be used for follow-upstudy.Part2The study of osteogenic differentiation on humanumbilical human umbilical cord mesenchymal stem cellsObjective：To investigate the change of stem cell and osteogenic related markers in humanumbilical cord mesenchymal stem cells (hUCMSCs) during osteogenic differentiation.Method: Alizarin red, Van Kossa staining and immunofluorescence were performed toassess hUCMSC proliferation and differentiation potential in vitro. ALP, mRNA level ofOPN/BSP/CD146/SOX2and protein level of CD146/SOX2/OPN/BSP/Runx2weremeasured at0,7,14,21d. Result: After osteogenic induction for21days in vitro, Alizarinred, Van Kossa stain was positive, and immunofluorescence staining showed that OPN/BSPwas greatly expressed in the cytoplasm. The ALP activity increased, The protein level ofCD146/SOX2decreased during the osteogenic procedure. The mRNA and proteinexpression levels of CD146/SOX2/BSP/OPN increased at day7and kept a high level duringosteogenic differentiation. The protein expression levels of Runx2had peak value at day7and then declined gradually. Conclusion: The osteogenic differentiation of hUCMSCs is acontinuous process which started completely after7days induction. The predifferentiatedhUCMSCs may be superior to the undifferentiated cells in bone defect repair.Part3The study of mimicking the periodontal healing usinghUCMSCs in vivoObject: To mimic the periodontal healing procedure using hUCMSCs in vivo. Method: The cells were treated with osteogenic medium for seven days in the experimental group,and the untreated cells served as a control. The osteogenic complex was implanted into thesubcutaneous space on the left side. As a control, the untreated cells were implanted intothe other side of the same host. Eight weeks later, the implant sections were prepared andstained with H&E, Masson trichrome staining and immunohistochemistry. Result: In theexperimental group, specimens of HE staining showed cementum-like depositssurrounded by fiber-like tissue. The Masson trichrome staining further confirmed thatsubstantial amounts of collagen surrounded the new cementum-like layer.Immunohistochemistry staining showed that the new layer was strongly positive for ONP.In contrast, in the control group, the deposits were scarcely detectable, with only a smallamount of fibrous connective tissue wrapped around (not attached to) the implanted dentin.Conclusion: The hUCMSCs at this stage had certain properties of osteoblasts. Day7cellscould be used for osteogenic repair and may be superior to day0cells for periodontalhealing to increase the successful rate of tooth autotransplantation. |
PDF Full Text Request