Repair Of Articular Cartilage And Subchondral Bone With Human Mesenchymal Stem Cell In Collagen/collagen-hydroxyapatite Scaffold

Articular cartilage and subchondral bone damage is a common orthopedic disease. There are not many treatments that can be used in articular cartilage injury in clinic, yet the effects are not very satisfactory. Studies have shown that bone marrow stromal cells,a kind of pluripotent stem cells,can be induced to differentiate into chondrocytes and osteoblasts under appropriate conditions. This experiment mainly used this feature of bone marrow stromal stem cells to find ways to induce them into chondrocytes and osteoblasts, and also established the induction and appraisal system; Then, on this basis,the bone marrow stromal cells were cultured with the scaffold biomaterials,and the engineering cartilage and subchondral bone restorations were constructed in vitro. The results of this study are as follows:(1) The adherent cells, with long spindle shapes and whirllike distributions, which were successfully isolated from human bone marrow, have also been proved to be human bone marrow stromal cells (hMSCs) by flow cytometry.(2) The isolated hMSCs were cultured with high density in the chondrocytes inducing culture solution,then the micromass with a rough diameter of 0.5 mm were obtained.Through alcian blue staining, type II collagen immunohistochemical staining,glycosaminoglycan quantitative, qRT-PCR and other methods,the results of induction were tested on histochemistry and molecular biology levels, ultimately the inducing and identifying systems of hMSCs differentiating into cartilage were established.(3) The isolated hMSCs were cultured in osteogenic inducing solution by monolayer culture.Cells cultured for 21 days were used to do the experiments of alkaline phosphatase ALP staining, alizarin red staining and type I collagen immunohistochemical staining, quantitative alkaline phosphatase ,qRT-PCR and other methods, the results of induction were tested and verified on histochemistry and molecular biology levels, ultimately the inducing and identifying systems of hMSCs differentiating into osteoblasts were established.(4) The hMSCs were then seeded in the scaffolds of biomaterials. Using the previously established inducing and identifying systems of hMSCs,the scaffolds of the COL and COL-HA were compared in the properties of differentiating into cartilage. The results showed that COL scaffolds were more suitable for cartilage defect repair than the COL-HA scaffolds. (5) The hMSCs were seeded in the scaffolds of biomaterials. Using the previously established inducing and identifying systems of hMSCs,the scaffolds of the COL and COL-HA were compared in the properties of differentiating into osteoblasts. The results showed that COL-HA scaffolds were more suitable for the subchondral bone or bone defects repair than the COL scaffolds.