Abstract:

BACKGROUND: Culture in vitro and understanding the biological characteristics of tenocytes are the premise and foundation to study the mechanism and improve the internal environment of tendon healing.
OBJECTIVE: To culture adult rabbit tenocytes in vitro with tissue explant, investigate the morphology, growth and proliferation of tenocytes, and to test the expression of collagen Ⅰ and collagen Ⅲ in the cells.
METHODS: After adult New Zealand rabbit flexor tendon was obtained under aseptic condition, the peritenon of tendon was removed. The tendon was divided into small fragments. The fragments were digested with 0.25% trypsin and 0.1% collagenaseⅠfor 10-15 minutes. The fragments were transferred into culture flasks after centrifugation. And 1 mL culture medium was added into the flasks after the fragments attaching to the wall. Culture medium was added when the cells showed an adhesive growth, and the medium was replaced every 3 days. When 80%-90% of the cells were in confluence, they were passaged at a ratio of 1:3.
RESULTS AND CONCLUSION: The tenocytes showed an adhesive growth at 10 days, and appearance was star-shaped or irregular shaped. The number of tenocytes was increased and the cell appearance changed to fibroblast-like as went on. Passaged cells were round-shaped at the beginning of cell seeding, the cells attached to the wall after 4-6 hours showed a spindle-shape, and the cells gradually arranged in groups. The growth curve of passage cells showed that: the latent period was the first 4 days, the logarithm period was at 5-6 days, and the platform period was at 7 days. Type Ⅰ collagen was positive and type III collagen was negative tested by immunofluorescence assay. The results indicated that tenocytes can be successfully isolated and cultured from adult rabbit tendon in vitro with tissue explant.