Osteogenic differentiation of bone marrow stromal cells: Implications to bone tissue engineering strategies

Today there exist a significant need for the development of novel bone replacement technologies for the repair of both site-specific skeletal defects and osseous deformities associated with degenerative bone diseases such as osteogenesis imperfecta and osteoporosis. Tissue engineering strategies utilizing bone marrow stromal cells (BMSCs) represent an attractive approach to address this need by creating viable bone implants and novel cell therapies for future options in skeletal reconstructions. The present work details efforts to advance current technologies in bone tissue engineering by increasing the understanding of basic BMSC biology through the elucidation of factors which influence their ability to undergo osteogenic differentiation and subsequently mediate bone formation in vitro and in vivo.; A novel BMSC ex vivo expansion strategy was developed in order to decrease both the loss of proliferative capacity and osteogenic differentiation potential routinely encountered during cultivation on conventional tissue culture polystyrene. It was demonstrated BMSCs ex vivo expanded on a denatured collagen matrix preserved certain aging-related functions normally lost during culture on tissue culture polystyrene such as their proliferative capacity and their ability to induce a major stress-protective protein, heat shock protein 70 (HSP70) in response to cellular stresses. In addition, ex vivo expansion of BMSCs on the denatured collagen matrix also preserved their ability to undergo osteogenic differentiation in vitro . BMSCs cultured to passage 11 and subjected to osteogenic stimulants retained between 69--95% of the levels of several osteogenic markers similar to those expressed by passage 1 cells.; Novel tissue engineered constructs consisting of bone derived-biomaterials seeded with BMSCs were generated and characterized for their ability to support both in vitro osteogenic differentiation and bone formation in vivo. In addition, it was established that the extent of scaffold demineralization, BMSC donor, and cell interactions with matrix-incorporated soluble and insoluble osteogenic factors all played substantial roles in mediating BMSC osteogenic responses within these bone-derived biomaterials both in vitro and in vivo.; A novel bioreactor system was also developed in order to apply physiologically-relevant four point bending strains to BMSCs seeded within a 3-D partially demineralized bone matrix in order investigate the role of this stimuli in modulating BMSC osteogenic differentiation and mineralized matrix production in vitro . Mechanical stimulation in conjunction with specific dexamethasone concentrations was capable of elevating the expression of osteoblast-related markers including alkaline phosphatase activity, osteopontin and alkaline phosphatase mRNA transcript levels, and mineralized matrix production following 16 days of bioreactor cultivation. The results presented within this thesis provide both basic insights into factors which play significant roles in mediating osteogenic differentiation of BMSCs as well as define new approaches and guidelines for the development of future clinical bone tissue engineering strategies.