Differentiation of Mesenchymal Stem Cells Transplanted to a Rabbit Degenerative Disc Model: Potential and Limitations for Stem Cell Therapy in Disc Regeneration

Abstract

STUDY DESIGN: An in vivo study to assess the differentiation status of mesenchymal stem cells (MSCs) transplanted to the nucleus pulposus of degenerative discs in a rabbit model. OBJECTIVES: To evaluate the fate of MSCs transplanted to the nucleus pulposus of degenerative discs in a rabbit and to determine whether they are a suitable alternative for cell transplantation therapy for disc degeneration. SUMMARY OF BACKGROUND DATA: Although MSCs have been proposed as candidate donor cells for transplantation to treat intervertebral disc degeneration, their differentiation after transplantation has not been adequately investigated. METHODS: Autologous MSCs, labeled with green fluorescent protein, were transplanted into mature rabbits. Consecutive counts of transplanted MSCs in the nucleus area were performed for 48 weeks after transplantation. Differentiation of transplanted cells was determined by immunohistochemical analysis. The proteoglycan content of discs was measured quantitatively using a dimethylmethylene blue assay, and mRNA expression of Type I and II collagen, aggrecan and versican was measured semi-quantitatively using reverse transcription polymerase chain reaction. RESULTS: Many cells that were positive for green fluorescent protein were observed in the nucleus pulposus of cell-transplanted rabbit discs 2 weeks after transplantation. Their number increased significantly by 48 weeks. Some GFP-positive cells were positive for cell-associated matrix molecules, such as Type II collagen, keratan sulfate, chondroitin sulfate, aggrecan, and the nucleus pulposus phenotypic markers, hypoxia inducible factor 1 alpha, glutamine transporter 1, and matrix metalloproteinase 2. MSCs did not show significant expression of these molecules before transplantation. Biochemical and gene expression analyses showed significant restoration of total proteoglycan content and matrix-related genes compared with nontransplanted discs. CONCLUSIONS: MSCs transplanted to degenerative discs in rabbits proliferated and differentiated into cells expressing some of the major phenotypic characteristics of nucleus pulposus cells, suggesting that these MSCs may have undergone site-dependent differentiation. Further studies are needed to evaluate their functional role.