Biocompatibility Evaluation Of β-tricalcium Phosphate-hyaluronate Scaffold Materials

Tissue engineering technology is the use of autologous or allogeneic very few organizations,through sterilization,digestion,separation and culture,proliferation to obtain a sufficient number of cells planted in the scaffold material,with a certain shape and function of the active organs,to replace the body A technique for the removal of organs,including seed cells,scaffolds and growth factors.Bone marrow mesenchymal stem cells(BMSCs)are seed cells with significant application prospects,not only have the ability of self-replication and multi-directional differentiation,in vitro can develop into hard bone,cartilage,fat or other types of cells,but also has a convenient collection,Separation and expansion technology is mature and so on.In this study,BMSCs were used to evaluate the biocompatibility of composite β-tricalcium phosphate-sodium Hy Aluronate scaffolds and to explore their application prospect in bone tissue engineering.In this paper,we first explored the methods of extracting and purifying primary rat bone marrow mesenchymal stem cells.The bone marrow mesenchymal stem cells(BMSCs)were extracted by bone marrow adherence method.The cells were cultured at different time to separate the high activity and purity Bone marrow mesenchymal stem cells were identified and confirmed by monoclonal antibody CD34,CD44,CD45,CD90 and CD105.Bone marrow mesenchymal stem cells were confirmed by MTT assay,indicating that the cell proliferation activity was highest in the first to third days,This stage of the cells should be used for follow-up experiments.Hy A/β-TCP composite scaffolds were prepared by physical method to synthesize HYA andβ-TCP according to the principle that the scaffold material in the implantable body should be harmless and beneficial to cell adhesion and growth.Hy A/β-TCPcomposite scaffolds were prepared by modifying the carboxyl groups in HYA and producing hydrophobic groups to prepare spongy,water-insoluble crosslinked modified Hy A/β-TCP composite scaffolds.Finally,The effect of Hy A/β-TCP composite scaffold on cell morphology was observed by co-culture with bone marrow mesenchymal stem cells.The results showed that the cell morphology on the cross-linked material was almost unchanged,the nucleus had no obvious shrinkage or swelling,Indicating that the material is less cytotoxic.The expression of Bax and Bcl-2-related protein was detected by Western Blot,which was tested by CCK-8 kit,cytotoxicity assay,cell Hoechst / PI staining,cell apoptosis rate and flow cytometry It was found that cross-linked Hy A/β-TCPcomposite scaffolds could inhibit apoptosis more effectively from mitochondrial mechanism,and Hy A has better biocompatibility than non-crosslinked materials.After the interaction between the material and the blood,the anticoagulant properties of the material were measured in four time periods 10min、20min、40min、80min.There was no significant difference between the supernatant level of the test substance and the negative control group,and no hemolysis,Indicating that the anticoagulant performance of the non-crosslinked material group was higher than that of the cross-linked material group.The scaffolds were implanted into the subcutaneous tissue of rats for 3 days,15 days and 60 days.There was no obvious abnormality in the activity of the rats,and no other special cases occurred.With the prolongation of implantation time,the inflammation of the tissue gradually decreased,Gradually generated,local hyperplasia is not obvious,implanted tissue scaffold in 60 days after the degradation of complete.The results of the study show that Hy A/β-TCP scaffold material has good cell,blood and tissue compatibility,and it is expected to be widely used in cartilage tissue engineering after further study.