Experimental Study On Full-thickness Cartilage Defects Repaired By The Composite Of Bone Marrow Mesenchymal Stem Cells And The Acellularmatrix Of Articular Cartilage

OBJECTIVE: 1. To investigate the methods of isolation, culturing,purification and indentifying bone marrow-derived mesenchymal stem cells(BMSCs) in vittro.2 To investigate the methods of mading the Acellularmatrix of articular cartilage.3 To repaire full-thickness articular cartilage defects of rabbits by cocultures of autogenic BMSC_S with the Acellularmatrix of articular cartilage ,grade the repaired tissues and evaluate repairing effect of defects in order to provide experimental and theoreticalbasis for clinical application,,METHODS: 1. MSCs derived from Qingzilan rabbits aged 3-4 months weighing 2.5-3.5kg with the methods of adherent culture screening method and density gradient centrifugation were cultured in vitro, the cultured MSCs were investigated with anti-CD44(MSCs specificness labelled antibody) and anti-CD34(hematopoietic cell specificness labelled antibody) by using immunocytochemistry.2.Reforming the methods of Courtman, preparing articular cartilage Acellularmatrix. Evaluated by histology and scanning electron microscope(SEM). 3. Adding TGF (transforming growth factor),DXM (dexamethasone),Vit C (vitamin C) into DMEM(dulbecco's minimum essential medium). Cultured the second filial generation BMSCs suspension and the Acellularmatrix of articular cartilage with DMEM in culture flask. after three days, grafting the compounds to the full-thickness defect zone fossa intercondylaris femoris cartilage. twenty-four healyhy Qingzilan rabbits were divided into three groups randomly. Each group had eight rabbits. The cartilage defects in the intercondylar fossa were filled with the compounds of autogeneous bonemarrow mesenchymal stem cells and the Acellularmatrix of articular cartilage in group A(management A), with only the Acellularmatrix of articular cartilage in group B(management B),with nothing in group C (managementC).Killed the rabbits at 16 weeks after operationg in each group.the reparative tissue sampales were evaluated grossly histologically immunohistochemically and graded according to gross and histological scale. Dataed in SPSS 10.0 software to proceed statistical analysis to find out if the differences between each group had statistical significance.RESULTS: 1. The cultured cells positively reacted to antigen CD44,but negative to antigen CD34 by immuocytochemistry. morphocytology obviously. 2. The articular cartilage Acellularmatrix were asperity,flexibility. The articular cartilage Acellularmatrix were honeycomb appearance and hollowness by SEM(scanning electron microscope). Not founeed reliquus cellular nucleus and cellular organ.3. the defects filled with formed hyaline-like tissue, smooth and glossy,the other defects repaired by fibrous tissue. Gross and histological grading scales were made on 16th week postoperatively. the results showed that the bonemarrow mesenchymal stem cells and the Acellularmatrix of articular cartilage in group A excelled the Acellularmatrix of articular cartilage group B and control group C significantly(P<0.05), differences were statistically significant.CONCLUSIONS: 1. The cells isolated from bone marrow have a morphoginic characteristic of MSCS. They could be used as the sources of seeds cells for cartilage tissue engineering.2. The articular cartilage Acellularmatrix prepared by the reforming methods of Courtman showed hollowness. Not founeed reliquus cellular nucleus and cellular organ. They could be used the scaffold material for cartilage tissue engineering.3. The full-thickness cartilage defects of rabbits were repaired with MSCS and articular cartilage Acellularmatrix complex by cartilage tissue engineering, which is a promising way for treatment cartilage defects.