| Periodontitis is one of the most common oral diseases in human, there are somelimitations in the existing treatments which are unable to completely restore themorphology and function of the periodontal tissue. In recent years, with the rapiddevelopment of tissue engineering for regenerative medicine, the biological regenerationof periodontal tissue as a kind of ideal therapeutic method is gaining extensive attention infuture. The periodontal ligament stem cells (human periodontal ligament stem cellshPDLSCs) are considered to be the cytological basis in stability and restoration ofperiodontal tissue, which could be used in the repairmen of periodontitis tissue defect.However, how to abundantly amplify with periodontal ligament stem cells high quality invitro and control the differentiation of periodontal ligament cells has become a keyproblem need to be solved from laboratory study to the clinical application. As we known, microenvironment plays an important regulatory role in themaintenance and differentiation of stem cells, including the matrix microenvironment andthe growth factor microenvironment. Therefore, we focused on periodontal tissueextracellular matrix, and investigated its organization, structure, composition and theeffects on the periodontal ligament stem cells. Based on a large number of previous studies,we have known that bone marrow extracellular matrix between the bone marrowmesenchymal stem cells have significantly promote the cell proliferation and maintained,while the influence of periodontal tissue microenvironment to the periodontal ligamentstem cells has not yet been determined. To this end, we built a special periodontalextracellular matrix (ECM) model, and analyzed its characteristics about structure,composition and effects on the periodontal ligament stem cells.Contents:(1) To define the internal structure and composition of the two different tissue specificECMs.(2) To preliminarily analyze and compare the protein composition of two differentextracellular matrix.(3) To explore the influence of the two tissue specific ECMs to the periodontalligament stem cell proliferation and apotosis in vitro.Methods:(1) We isolated hPDLCs and hBMCs, then used these two kinds of cells to preparecell-derived tissue-specific extracellular matrix and observed these two differentmatrix structural components by scanning electron microscopy (SEM) andtransmission electron microscopy (TEM)(2) We used the Immunofluorescence staining and liquid-mass spectrometry to detectthe element of the two sources of extracellular matrix, and the proteomics analysiswas carried out.(3) We used the limiting dilution technique to isolate hPDLSCs, and incubated them on the different ECMs. We analyzed the discrepancies of proliferation, apoptosiscapacity between two groups using CCK8assays, flow cytometry analysis of cellcycle and apoptosis detection.Results:(1) In two kinds of tissue specific ECM model, although internal cellular structure wasdamaged, the structure of collagen skeletons remains intact and there are differenceson the surface structure of two sources of extracellular matrix. The collagen fromhBMCs-ECM is thicker than its from hPDLCs-ECM.(2) The extracellular matrix protein is preserved basically. hBMCs–ECM may expressmore higher osteogenesis protein than another.(3) Compared with the control group, the cells cultivated on the ECMs have asignificantly higher proliferation ability, while there is no difference in apoptosis.Conclusion:In this study we successfully built two tissue specific ECM from different cell source:hPDLCs-ECM and hBMCs-ECM, and explored their structures. We found the ECMs notonly could keep the original skeleton structure,but retain the base composition，whilesimulate periodontal microenvironment effectively to impace on cells. And hPDLCs-ECM and hBMCs-ECM can promote the proliferation, make the stemness of hPDLSCs，not reducing the apoptosis. This research, not only established an effective model forperiodontal tissue microenviroment about cell development and differentiation, but alsofurther lays a new research strategy for amplifying high quality hPDLSCs in vitro. |
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