Acellular Matrix-induced Autologous Chondrocyte Implantation Into Pretreated Microfracture For Treatment Of Cartilage Defects In The Knee Of The Rabit Model

Objectives This study will be decellularized cartilage, the use of acellular matrix material preparation cartilage scaffold with biological source material, the scaffold in vitro inoculation of autologous bone marrow mesenchymal stem cells to build MSCs-scaffold complex, the establishment of rabbit knee full-thickness cartilage defect model, and application of micro fractures combined with implantation of cells scaffold complex repair of rabbit articular cartilage defects, cartilage formation and repair observed effects. Set the control group, microfracture group, blank scaffold group, microfracture combined with scaffold implantation group, microfracture combined with MSCs-scaffold group. Animals were sacrificed after 3 months by gross observation of the specimens and histological detection. Biomimetic compound as a scaffold for cartilage and provide experimental evidence, as well as cartilage repair in clinical and basic research to explore a new and effective method.Material and Methods1. The fresh human cartilage are crushed to particles when immerced in protease inhibitor, then separated by centrifugal and treated with acellular process, then detected by tissues quantitative analysis and biochemical properties. Prepare a 8mm diameter osteochondral block which contain only about 100um hyaline cartialge, then treated with acellular process and have relative detect.2. Chondral scaffold was structured by extrcellular matrix with freeze-drying and crosslinking, then seeded with rabit bone marrow derived mesenchymal stem cells, had histology, histocompatibility and cell toxicity detect after cultured 7 days in vitro.3. Chondral defects was prepared in weight loading area of rabit, defects were separated into control group, microfracture group, blank scaffold group, microfracture combined with scaffold implantation group, microfracture combined with MSCs-scaffold group., each group have 12 defects, then the animals were sacrificed 3 months after the operation and the specimens have relative testing(HE, toluidine blue, Safranin O and typeâ…¡collagen immunohistochemical staining).Results:Knee taken 3 months after the general examination showed that the control group did not completely repair the knee 3,9 no repair; microfracture were 12 cases of articular cartilage defects 8 fully restored, but after the repair of cartilage was significantly higher than the experimental group thinning, HE staining fibrocartilage, Safranin O, toluidine blue and typeâ…¡collagen immunohistochemical stain were negative. Blank scaffold group 8 to 12 incomplete repair, edge repair is poor, in which a cartilage defect was covered by scar tissue,4 free repair. Microfracture knee combined with scaffold group of 12 fully restored in 7 cases,5 were not completely repaired. Experimental group of 12 knees of 10 fully restored, two incomplete repair. Histology showed hyaline cartilage repair tissue area constituted by the cells around the defect of cartilage and number of irregular arrangement, the surface cells are smaller and slightly more volume, deep slightly larger cell volume, extracellular matrix small, Safranin O and toluidine blue staining positive, the defect surface repair smoothly, clear boundaries with adjacent cartilage, and no lymphocyte infiltration, extracellular matrix of typeâ…¡collagen immunohistochemical staining. Micro fracture and blank scaffold graft group, the defect is covered by fibrous tissue, cells of fibroblast-like spindle cells to repair surface is not flat, showing significant depression organizations, most of the degradation of cartilage scaffolds absorbed the side, and no lymphocyte infiltration, vascular scaffold visible chondral invasion, toluidine blue staining weakly positive, Safranin O and typeâ…¡collagen immunohistochemical staining no positive.Conclusion:1. Mesenchymal stem cells in vitro isolation, purification, amplification, can be used as seed cells for cartilage tissue engineering for full-thickness cartilage defects.2. The method acellular cartilage matrix prepared porous scaffold structure, which can be used for implantation with seeding cells, and the scaffold for the whole biological source, no cytotoxicity, completely absorbed in the body can be used as tissue engineering scaffolds.3. This experiment combined acellular cartilage repair capacity of MSCs with microfracture and repair are the best, improve the efficiency of microfracture repair. Prove that the scaffold through the vaccination of autologous MSCs combined with microfracture technique can be better to repair cartilage defects in rabbits loading area, this method prompts in cartilage injury and repair of tissue engineering applications and has some value and prospects.