Preparation And Cell Evaluation Of A Novel Meniscus Tissue Engineering Scaffold

If 3%-5%of the total volume of the meniscus structure is damaged,its bearing load would be increased to 3-5 times of the original.With the meniscus injury,the degeneration of articular cartilage,knee joint effusion and other problems will gradually aggravate.Among them,one third of the "white-white" area on the medial side of the meniscus is composed of meniscus cells and their extracellular matrix,and its regeneration ability is extremely limited.There is no good cure for the injury in this area.The purpose of this paper is to construct a new scaffold for meniscus tissue engineering.The specific method is to obtain porcine meniscus extracellular matrix(DMECM)by acellular treatment of porcine meniscus tissue.DMECM is mixed with gelatin/chitosan(G/C)in different proportion.Then the optimized DMECM-G/C scaffold was characterized.Based on the SEM image of DMECM-G/C,the stress-strain curve of the whole scaffold was obtained by experiment as the mechanical reference data,and the single hole finite element model of the scaffold was established.The finite element model was used to calculate the stress of DMECM-G/C single hole.The results show that the bottom of the scaffold is first subjected to a larger force,and the upper part is relatively small.When the pressure is 0-0.25 MPa,the cell is in the elastic range.When the pressure is 0.25 MPa,the cell continues to deform,and then the single hole enters the collapse deformation stage and the compaction stage after the collapse.The equivalent elastic modulus of the structure is 0.1288 MPa.The basic physical properties,elastic modulus,infrared spectrum,electron microscope,compression test and calculation results show that the noval scaffold of 1%DMECM-G/C scaffold shows excellent mechanical properties and is expected to become an ideal scaffold material for meniscus tissue engineering.In this paper,the isolation,culture,identification and proliferation of bone marrow mesenchymal stem cells(BMSCs)from SD rats were performed.The biocompatibility of DMECM-G/C was evaluated by BMSCs.The results show that 1%DMECM-G/C is suitable for BMSCs growth and would be used as meniscus tissue engineering scaffold for study in the future.This paper attempts to explore the effect of hydrostatic pressure on BMSCs cultured in DMECM-G/C.In this paper,a bioreactor with batch controlled high pressure is designed and manufactured.In this bioreactor,cell scaffold complex hydrostatic pressure intervention experiment was carried out.Western blot was used to detect the effect of hydrostatic pressure on the apoptosis of BMSCs in 2D and 3D scaffolds.The results showed that BMSCs were cultured in 3D with 1%DMECM-G/C composite scaffolds,and the expression of Caspase-9 and Bad of BMSCs was significantly decreased under the hydrostatic pressure intervention in the intermittent controllable high-pressure bioreactor compared with 2D plane culture.Although both 3D culture and hydrostatic pressure can inhibit the apoptosis of BMSCs,the inhibition of apoptosis is the most significant under the dual intervention of 3D culture and hydrostatic pressure culture.The conclusion of this paper is that the improved controlled high hydrostatic pressure bioreactor could provide the hydrostatic pressure for meniscus tissue engineering research.Prolonged hydrostatic pressure and 1%DMECM-G/C scaffolds significantly inhibited the apoptosis of BMSCs by activating Akt phosphorylation.The results suggest that 1%DMECM-G/C scaffold could provide a better environment for BMSCs culture in vitro.