OBJECTIVETo investigate a method that constructing a tissue-engineered tendon with a continuous and heterogeneous transition region.

METHODSFibroblasts derived from rabbit epithelial tissue were cultured in vitro and collagen gel was prepared. The experimental groups were scaffold only group, fibroblasts+ chondrocytes group (Fb+ CC group), fibroblasts+ osteoblasts group (Fb+ OB group), fibroblasts+ chondrocytes+ osteoblasts group (Fb+ CC+ OB group). Heterogeneous cell populations(fibroblasts, chondrocytes and osteoblasts) with collagen gel were seeded within three predesigned specific regions (fibrogenesis, chondrogenesis, and osteogenesis) of decellularized rabbit achilles tendons to fabricate a stratified scaffold containing three biofunctional regions supporting fibrogenesis, chondrogenesis, and osteogenesis. The tests of morphology, architecture and cytocompatibility of the scaffolds were performed. Gradient tissue-specific matrix formation was analysed within the predesignated regions via histological staining and immunofluorescence assays.

RESULTSThe HE staining and scanning electron microscopy analysis demonstrated that no major cell fragments or nuclear material was evident, and increased intra-fascicular and inter-fascicular spaces were found, the cytocompatibility of the scaffolds showed that the numbers of viable cells on the scaffold surfaces increase steadily, no significant differences were found between the scaffold only containing ordinary culture medium and scaffold containing gel groups. Histological staining and immunofluorescence assays demonstrated that the cartilage-related markers (GAG, COL2A1) were found only in the chondrogenesis region, but bone-related proteins only in the osteogenesis region of bone tunnel, and fibrosis was remarkable for the fibrogenesis region in the joint cavity. The transitional architecture with ligament-fibrocartilage-bone was constructed in the ligament-bone tunnel interface.

CONCLUSIONSA transitional interface (fiber-fiberocartilage-bone) could be replicated in a decellularized tendon through stratified tissue integration in vitro. The cell-tendon complex offers the advantages of a multi-tissue transition involving controlled cellular interactions and matrix heterogeneity.