The Related Research Of The Bone Marrow Mesenchymal Stem Cells

Objective:Bone defect is one of the problems in the orthopaedics. The present treatments have allografts and biomaterial-filling. But they arenot ideal methods. Tissue-engineering has provided a new way to solve the problem and has been one of hotspots recently.Bone Marrow Mesenchymal stem cells(BMSCs) have been focused for own particularity when discovered. Firstly, it is one of primitive bone marrow cells which can differentiate into many kinds of adult animal cells. In a definite condition, it can differentiate into osteoblast, cnondrocyte, tendon cell, adipocyte, bone marrow stroma cell, angio-endothelial cell and nervi-stellate cell, ea tl. And it is easy to be extracted and amplificated. So BMSCs is one of the important tissue engineering seed cells. Secondly, exogenous gene is easy to be inverted and expressed. So BMSCs is the potential target cells which is used as gene therapy. But at present, how to segregate, purify, identify and culture BMSCs in vitro isn't standardized, which doesn't benefit to popularit the BMSCs' culture in vitro.So we have some experiments on how to segregate, purify, identify and culture BMSCs in vitro, at the same time seed me BMSCs into PLGA scoffald manufactured by our own department, find the avidity between BMSCs and biomaterial, show the feasibility of BMSCs as seed cells.Moreover we plan to reform BMSCs by transgene. We shall transfer BMP7 into BMSCs for acquire the new kind of seed cells of bone tissue engineering. For the purpose, we have reconstructed padtrackCMV withBMP7.Simultaneously, at the cell level, from the cell pharmacology, study Tetracycline HCl has effects on proliferation and differentiation of BMSCs. Try to study the factors which affect and intervent bone development, so that we can find the mechanism that clinical medicines affect bone developmentMethod:Segregate BMSCs from five-month fetal's humerus and femur:1.Morphological observation of fetal BMSCs cultured with PLGA scaffold. After Seeded into PLGA scaffold for a week, BMSCs have been followed by observation under scanning electron microscope.2.Tetracycline HCl has effects on proliferation of fBMSCs. Add Tetracycline'HCl into low-glucose DMEM and high-glucose DMEM, then measure the MTT of BMSCs and 293 cells cultured with Tetracycline- HCl after 2d, then find whether a significant difference exists between two cells by a statistical method.3.The reconstruction of adenoviral vector with BMPT.Cut down BMP7 by Enzyme from BMP7-pcDNA3, then link BMP7 into padtrackCMV.Result:1 .After fBMSCs cultured with PLGA for a week, cells array with PLGA's surface character, and grow long-fusiform shape, have many microvillus and slender apophysis, connect between cells.2.The statistical discrepancy exists between fBMSCs and 293 cells cultured with culture medium Added Tetracycline HCl.3.Cut down BMP7 from pcDNA3, then link BMP7 into PadtrackCMV, by enzyme-cutting evaluation, acquire ideal electrophoresis zone.Conduaon:1.Naive fBMSCs grows well with PLGA, it have good abilities ofproliferation and adsorption, so fBMCs is ideal seeding cells for bone tissue engineering.2.PLGA scaffold by mild heating under high pressure loaded with fMSCs may be used well as bone formation.3.Tetracycline HCl has negative effects on proliferation of fBMSCs.4.Adenoviral vector with BMP7 (pAdtackCMV-BMP7) is successful to be reconstructed.