[Preparation and biocompatibility evaluation of novel cartilage acellular matrix sponge]

Tianjun Liu, Bo Tan, Jingcong Luo, Li Deng, Huiqi Xie
Chinese Journal of Reparative and Reconstructive Surgery 2009, 23 (8): 1002-6

OBJECTIVE: To explore the method of preparing spongy and porous scaffold materials with swine articular cartilage acellular matrix and to investigate its applicability for tissue engineered articular cartilage scaffold.

METHODS: Fresh swine articular cartilage was freeze-dried and freeze-ground into microparticles. The microparticles with diameter of less than 90 microm were sieved and treated sequentially with TNE, pepsin and hypotonic solution for decellularization at cryogenic temperatures. Colloidal suspension with a mass/volume ratio of 2% was prepared by dissolving the microparticles into 1.5% HAc, and then was lyophilized for molding and cross-linked by UV radiation to prepare the decellularized cartilage matrix sponge. Physicochemical property detection was performed to identify aperture, porosity and water absorption rate. Histology and scanning electron microscope observations were conducted. The prepared acellular cartilage matrix sponge was implanted into the bilateral area of spine in 24 SD rats subcutaneously (experimental group), and the implantation of Col I sponge served as control group. The rats were killed 1, 2, 4, and 8 weeks after operation to receive histology observation, and the absorption and degeneration conditions of the sponge in vivo were analyzed. BMSCs obtained from femoral marrow of 1-week-old New Zealand white rabbits were cultured. The cells at passage 3 were cultured with acellular cartilage matrix sponge lixivium at 50% (group A), acellular cartilage matrix sponge lixivium at 100% (group B), and DMEM culture medium (group C), respectively. Cell proliferation was detected by MTT method 2, 4, and 6 days after culture.

RESULTS: The prepared acellular cartilage matrix sponge was white and porous. Histology observation suggested that the sponge scaffold consisted primarily of collagen without chondrocyte fragments. Scanning electron microscope demonstrated that the scaffold had porous and honeycomb-shaped structure, the pores were interconnected and even in size. The water absorption rate was 20.29% +/- 25.30%, the aperture was (90.66 +/- 21.26) microm, and the porosity of the scaffold was 90.10% +/- 2.42%. The tissue grew into the scaffold after the subcutaneous implantation of scaffold into the SD rats, angiogenesis was observed, inflammatory reaction was mild compared with the control group, and the scaffold was degraded and absorbed at a certain rate. MTT detection suggested that there were no significant differences among three groups in terms of absorbance (A) value 2 and 4 days after culturing with the lixivium (P > 0.05), but significant differences were evident among three groups 6 days after culturing with the lixivium (P < 0.05).

CONCLUSION: With modified treatment and processing, the cartilage acellular matrix sponge scaffold reserves the main components of cartilage extracellular matrix after thorough decellularization, has appropriate aperture and porosity, and provides even distribution of pores and good biocompatibility without cytotoxicity. It can be used as an ideal scaffold for cartilage tissue engineering.

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