| Mesenchymal stem cells (MSCs) are found in the adult bone marrow and they constitute the bone marrow population capable of regenerating mesenchymal tissues. MSCs may differentiate into mesenchymal tissues such as bone, cartilage and fat, as well as into cells of non-mesenchymal lineages such as neurons and hepatocytes. They are advantageous over other stem cell types because of their ease of availability and lack of ethical issues related to their use. Therefore, they have significant potential for use in tissue repair therapies. Glycosaminoglycans (GAGs) are a group of polysaccharides that naturally occur in the extracellular matrix and they have important roles in cell proliferation and differentiation during embryogenesis through their ability to interact with many growth factors and proteins. In this work, an in vitro culture system that mimics the in vivo extracellular environment was developed using GAGs with the aim of manipulating MSC proliferation and differentiation. Additionally, the potential of MSCs in nerve regeneration was explored based on the hypothesis that MSCs produce neurotrophic factors that stimulate neuronal regeneration. MSCs were isolated from adult rat bone marrow and were cultured on chitosan membranes with covalently immobilized GAGs including heparin (HEP), heparan sulfate (HS), dermatan sulfate (DS), chondroitin 4-sulfate (C4S), chondroitin 6-sulfate (C6S) and hyaluronic acid (HA). It was found that MSC proliferation was dependent on GAG type and density present on the membranes. The growth rate was measured to be highest on C4S-modified surfaces followed by those on HEP, HS, DS, C6S and HA in decreasing order. The analysis of membranes for adsorption of serum adhesion proteins demonstrated a correlation between the relative level of adsorbed fibronectin and MSC growth rate on all GAGs except C4S. It was suggested that influence of C4S on MSC proliferation was governed by a separate mechanism, possibly a direct interaction of CD44 with C4S. It was also demonstrated that MSC proliferation was influenced by the thickness of the chitosan membrane used as the immobilization surface due to the changes in the membrane microstructure as a function of thickness. Finally, MSCs were shown to enhance neuronal regeneration from DRG explants in chitosan regeneration tubes. |
