Tuning the differentiation of periosteum-derived cartilage using biochemical and mechanical stimulations

Objective: In this study, we aim at tuning the differentiation of periosteum in an organ culture model towards cartilage, rich in collagen type II, using combinations of biochemical and mechanical stimuli. We hypothesize that addition of TGF-beta will stimulate chondrogenesis, whereas sliding indentation will enhance collagen synthesis. Design: Periosteum was dissected from the tibiotarsus of 15-day-old chick embryos. Explants were embedded in between two agarose layers, and cultured without stimulation (control), with biochemical stimulation (10 ng/ml TGF-beta 1), with mechanical stimulation (sliding indentation), or both biochemical and mechanical stimulations. Sliding indentation was introduced as a method to induce tensile tissue strain. Analysis included quantification of DNA, collagen and GAG content, conventional histology, and immunohistochemistry for collagen type I and II at 1 or 2 weeks of culture. Results: Embedding the periosteal explants in between agarose layers induced cartilage formation, confirmed by synthesis of sGAG and collagen type II. Addition of TGF-beta 1 to the culture medium did not further enhance this chondrogenic response. Applying sliding indentation only to the periosteum in between agarose layers enhanced the production of collagen type I, leading to the formation of fibrous tissue without any evidence of cartilage formation. However, when stimulated by both TGF-beta 1 and sliding indentation, collagen production was still enhanced, but now collagen type II, while sGAG was found to be similar to TGF-beta 1 or unloaded samples. Conclusions: The type of tissue produced by periosteal explants can be tuned by combining mechanical stimulation and soluble factors. TGF-beta 1 stimulated a chondrocyte phenotype and sliding indentation stimulated collagen synthesis. Such a combination may be valuable for improvement of the quality of tissue-engineered cartilage