Mesenchymal stem cells from patients to assay bone graft substitutes

Bio-engineered scaffolds used in orthopedic clinical applications induce different tissue responses after implantation. In this study, non-stoichiometric Mg²⁺ ions and stoichiometric apatites, which are used in orthopedic surgery as bone substitutes, have been assayed in vitro with human adult mesenchymal stem cells (hMSC) to evaluate cytocompatibility and osteoconductivity. hMSCs from the bone marrow aspirates of orthopedic patients were isolated and analyzed by flow cytometry for the surface markers Stro1, CD29, CD44, CD71, CD73, CD90, CD105 (positive) and CD45, CD235 (negative). The hMSC were analyzed for self-renewal capacity and for differentiation potential. The hMSC, which were grown on different biomaterials, were analyzed for (i) cytotoxicity by AlamarBlue metabolic assay, (ii) osteoconductivity by ELISA for activated focal adhesion kinase, (iii) cytoskeleton organization by fluorescence microscopy, and (iv) cell morphology which was investigated by scan electron microscopy (SEM). Results indicate that isolated cell populations agree with minimal criteria for defining hMSC cultures. Non-stoichiometric Mg²⁺ and stoichiometric apatites, in granular form, represent a more favorable environment for mesenchymal stem cell adhesion and growth compared to the non-stoichiometric Mg²⁺ apatite, in nano-structured paste form. This study indicates that different forms of biomaterials modulate osteoconductivity and cellular growth by differential activation focal adhesion kinase.