Repair of articular cartilage defects with three-dimensional bio-printing and its properties

Abstract

Objective To repair the articular cartilage defects of animal models with cartilage tissue block made by multi-nozzle three-dimensional bio-printer and observe its effect. Methods Bio-ink was made by adding silk fibroin, polyethylene glycol and bone mesenchymal stem cells into extracellular matrix (ECM) solution. Rheological properties of biological ink were evaluated by rheometer, the protein secondary structure of biological ink was identified by Fourier transform infrared spectroscopy, and a tissue engineering scaffold with thickness of 2mm and diameter of 6mm was printed by using a pressure sprinkler loaded with cartilage biological ink. The compression modulus of tissue engineering scaffold was measured by tension machine. The degradation rate of each scaffold was evaluated by dry weight loss method, and the viability and proliferation of cells on the scaffold were evaluated by CCK-8 and live&dead cell staining. The differentiation of cellular cartilage on the scaffold was evaluated by real-time fluorescence quantitative PCR. The scaffold was embedded into the defect area of animal articular cartilage to repair articular cartilage defect according to the principle of autogenous cartilage transplantation. The effect of cartilage repair after 3 months was evaluated by histological staining and biochemical detection. Results We found that all biological inks showed the flow characteristics of shear thinning. The absorption peak of biological ink amide I region containing silk fibroin moved to 1 623 cm-1. With the increase of silk fibroin content, the mechanical strength and degradability of biological ink were improved, and the compression modulus of 10% and 15% printing stand reached 19.96±5.66 kpa and 26.87±10.68 kpa, respectively. All biological inks had no obvious cytotoxicity. Real-time quantitative PCR showed that when the content of silk fibroin reached 10%-15%, the bone marrow mesenchymal stem cells in the tissue mass had stronger ability to differentiate into cartilage. In vivo studies showed that after 3 months, the sGAG/DNA content of 10% and 15% silk fibroin scaffolds reached 0.25±0.01 μg/ng and 0.24±0.02 μg/ng, respectively, and the collagen/DNA content reached 17.71±0.83 ng/ng and 16.69±2.39 ng/ng, respectively. Tissue engineered cartilage printed with high concentration silk fibroin can better repair articular cartilage defects. Conclusion TThe chondrogenic differentiation and extracellular matrix (collagen and glycosaminoglycan) secretion of BMSCs were superior to those of the other two scaffolds when the content of silk fibroin reaches 10%-15%. The changes of chondrogenic differentiation ability and extracellular matrix secretion of stem cells from different scaffolds, as well as the repair effect on articular cartilage defects are caused by the differences of mechanical properties of scaffolds, which can be produced by the changes of silk fibroin concentration. Key words: Tissue engineering; Cartilage, articular; Extracellular matrix; Silk