Mechanical tensile stress and SIS membrane stimulate differentiation of rabbit costal chondrocytes into fibrotic cartilage

Intervertebral disc is comprised of two distinct region named anulus fibrosus and nucleus pulposus. Anulus fibrosus(AF) is composed of fibrous cartilage that bears a tension and give flexibility to the vertebrate. For regeneration of intervertebral disc tissue, it must include engineering of these two specialized tissues, but differentiation of in vitro cultured chondrocytes into fibrous cartilage like AF is not well established yet than hyaline cartilage. In this study, we established the differentiating condition into fibrous cartilage using dedifferentiated rabbit costal chondrocytes(CCs) on biomaterials through mechanical and biochemical stimulation. To apply the mechanical stress, dedifferentiated CCs as a cell source were seeded on small intestine submucosa(SIS) membrane as biomaterial. To apply tensile stress into CCs, custom-made bioreactor was used. In mechanical tensile stress applied culture of 20% elongation and 5 Hz frequency, the proliferation rate of CCs on SIS membrane were increased 9.64% and 4.64% after 24 hr and 72 hr. After applying mechanical tensile stress, cell alignment was in parallel with the direction of tension. In reverse transcriptase polymerase chain reaction analysis, expression of type I collagen was up regulated by mechanical stimulation. Based on these results, the differentiation of CCs to fibrotic cartilage like anulus fibrosus can be achieved by applying mechanical stimulation on SIS membrane.