Construction of tissue-engineered cartilage using silk fibroin nets and chondrocytes

Objective: To evaluate the feasibility of non-woven silk fibroin net as a biocompatible scaffold of the auricular chondrocytes for construction of tissue-engineered cartilage. Methods: Chondrocytes were harvested from auricular cartilage by trypsin and type Ⅱ collagenase digestion, and expanded in culture flasks. The chondrocytes(P1) were seeded into non-woven silk fibroin nets, and incubated in CO2 incubator. The adhesion, proliferation and growth of cells were observed under inverted microscope every day and the microstructure of the constructs was observed by using scanning electron microscope(SEM) at 3, 7 and 10 d after coculture. The constructs were harvested and analyzed by histology at 1, 2 , 4 and 6 weeks after coculture, and the mRNA expression of type Ⅱ collagen was examined using RT-PCR. Results: The scaffold was covered with chondrocytes at 48 h and these cells began to secrete extracellular matrix(ECM) at 72 h after coculture. The surface of scaffold was covered with chondrocytes and ECM under SEM. Stent shallow already had some cartilage tissue and a little lacunae at 2 weeks. The cartilage tissue thickened and lacunae increased, but the certer of stent had a little cells, which were star or spindle type and secreted a little ECM at 4 and 6 weeks. The mRNA expression of type Ⅱ collagen was detected by RT-PCR at all time-points. Conclusions: Cartilage-like tissues can form at non-woven silk fibroin net with the auricular chondrocytes, but culture conditions should be further optimized.