Introduction: The chondrogenic differentiation of adult mesenchymal stem cells (MSCs) is a promising method for cartilage tissue engineering and repair of cartilage defects. The potential of MSCs to differentiate in chon-drocytes could be proved in many investigations, in matrix-dependent and matrix-free set-ups. A standard system for chondrogenic differentiation of MSCs utilizes alginat beads which allow a high cell density, will not generate a homogenious matrix. Therefore beads may not be optimal for regeneration of cartilage in articulating joints. Furthermore, after chondrogenic differentiation, cells in alginate beads may display signs of hypertrophy, including collagen X, alkaline phosphatase and MMP-13. The aim of our investigation was to explore the chondrogenic differentiation of MSC in a novel collagen-chondroitinsulphat-matrix.
Methods: Adult MSC were harvested from the bone marrow of donators who received a total hip replacement The cells were differentiated in a monolayer culture, on alginat beads and in a novel spongiform collagen-chondroitinsulphat-matrix. For differentiation, the medium was supplemented with dexamethason, ascorbic acid, and TGFβ. The total culture period was 21 days. Afterwards the expression of collagen-I, -II and -X, Interleukin (IL)-1β, IL-6, MMP-1, -3 and -13 was determined by quantitative RT-PCR. Histological analysis of the constructs were performed after 4 weeks of s.c. implantation in immunodeficient SCID-mice.
Results: Human MSC undergo chondrogenic differentiation in the novel collagen-chondroitinsulfat-matrix. In comparison to cells differentiated in alginat beads, a higher expression of collagen II but a comparable expression of collagen I, MMP-1, MMP-3 and IL-1β were observed. Collagen-X, MMP-13 or alkaline phosphatase were not detected in the cells differentiated in the new matrix, but could be found in cells the alginat beads. Furthermore, in comparison to the monolayer cultures, the collagen II expression was 100’000-fold raised, but no difference was found in the expression of collagen I, MMP-1, MMP-3 and IL-1β.
Discussion: The novel collagen-chondroitinsulphat-matrix supports an improved chondrogenic differentiation of MSCs with an elevated expression of collagen-II and very low expression of markers of hypertrophy in comparison to cells in alginate beads or in monolayer cultures. These results are a promising basis for improved tissue engineering of cartilage. The clinical application of these constructs seems to be possible, because the new matrix is approved for autologous chondrocyte transplantation and MSC can be expanded under GMP-compatible conditions.
