Research of the cartilago regeneration using ultrasonic stimulus and the hyaluronic acid

点击翻译按钮获取中文摘要

Objective. We have shown for the first time that it is possible to enhance the proliferation of cultured chondrocytes and promote the production of type-IX collagen in primary chondrocytes grown on a three-dimensional (3-D) culture system through low-intensity pulsed ultrasound (LIPUS) stimulation. Moreover, one of the underlying mechanisms of intracellular signaling pathways such as MAPK/MAPKK and Integrin/PI3K/Akt involved in mechano-transduction during the proliferation of chondrocytes was identified.Methods. Articular cartilage tissue was obtained from the metatarso-phalangeal joints of freshly slaughtered porcines. Isolated chondrocytes in collagen gel and culture medium composites were added to type-I collagen honeycomb sponges. The final cell density was adjusted to 2×10^6 cells/well/ml. LIPUS stimulation was applied to the chondrocyte cultures for 20 minutes daily. Anti-type-II and anti-type-IX collagen antibodies were used to investigate the phenotype of the chondrocytes and t … More o detect type-II and type-IX collagen production at post-culture week 2. For immuno-staining and Western blot analysis, the following primary antibodies were used: type-II and type-IX collagen, Akt, p-Akt, Cyclin B1 and D1, MAPK, p-MAPK, FAK, p-FAK, Paxillin, p-Paxillin, β-catenin, β-1 Integrin, and α-1, 2 and v Integrins.Results. Chondrocytes proliferated and a collagenous matrix formed on the surface of the collagen sponge. By exposing chondrocytes to LIPUS, thicker layers of matrix containing type-II collagen and proteoglycan accumulated. The rate of proliferation of chondrocytes cultured under LIPUS stimulation was significantly higher than in the control group during the two-week culture period. In Western blot analysis, type-IX collagen, Cyclin B1 and D1, p-FAK, and p-Akt were found to be more strongly expressed in the LIPUS stimulation group than in the control. However, there was no difference between the two groups in MAPK, p-MAPK and type-II collagen expression.Conclusion. LIPUS stimulation promoted the proliferation of cultured chondrocytes and the production of type-IX collagen in a 3-D culture system using collagen sponge as a vehicle to create an environment similar to cartilaginous tissue. Less