Bone Tissue Engineering Program, Institute of Health and Biomedical Innovation, Queensland University of Technology, Brisbane, Australia Adult mesenchymal stem cells (MSCs) are a topic of immense research interest in the field of tissue engineering. Since, depletion of multipotent cells has been implicated in degenerative joint diseases, cell based therapies have been proposed for tissue regeneration, especially for cartilage repair. The aim of the present study is to focus on the possibility of deriving and expanding multipotential MSCs from the heterogeneous bone marrow stromal samples of patients with osteoarthritis (OA) by characterising MSCs at the single cell level. Single cell clonal cultures were established by limiting dilution of marrow stromal cells from three OA patients. A total of 14 clones with a wide variation in their cell doubling time were isolated. The clones were grouped into fast-growing and slow-growing clones. All except one of the fast-growing clones were tripotential. However the slow-growing clones showed limited differentiation potential and morphological changes associated with cellular senescence with extended duration in culture. Flow cytometric analysis did not depict any difference in the expression of the selected putative MSC cell surface markers CD29, CD44, CD90, CD105 and CD166 between fast-growing and slow-growing clones indicating a strong need to investigate for novel cell-surface markers. Further, proteomic analysis to understand the sub-cellular processes responsible for the existence of varying sub-populations identified 11 differentially expressed proteins. These proteins were reported to be associated with cellular organization, signal transduction, energy pathways and stress related proteins. Identification of signaling pathway proteins and cell cycle related proteins, such as calmodulin and caldesmon in the clonal populations, suggest that high-throughput proteomic technologies like two dimensional liquid chromatography (2D LC) coupled with mass spectrometry (MS) may facilitate the discovery of therapeutically useful biomarkers. This study demonstrated the existence of a fast-growing multipotential MSC population from bone marrow samples of patients with OA. Therefore, despite a supposedly smaller stem cell compartment in these patients, we demonstrate here that they can still yield a potentially therapeutically useful source of syngeneic MSCs.
