Cyclic acetal hydroxyapatite composites and endogenous osteogenic gene expression of rat marrow stromal cells

Abstract

In this study, bone marrow stromal cells (BMSCs) were differentiated on cyclic acetal composites containing hydroxyapatite (HA) particles (110 or 550 nm). These composites were evaluated for their role in influencing osteogenic signalling by encapsulated BMSCs. While a number of factors exert influence on osteogenic signalling during the production of an osteogenic matrix, we hypothesize that HA particles may upregulate bone growth factor expression due to enhanced BMSC adhesion. To this end, fluorescence-activated cell sorting (FACS) analysis was performed for the evaluation of BMSC surface marker expression after culture on two-dimensional (2D) cyclic acetal/HA composites. Three-dimensional (3D) composites were then fabricated by incorporating 110 or 550 nm HA particles at 5, 10 and 50 ng/ml concentrations. Bone growth factor molecules (TGFbeta1, FGF-2 and PDGFa), bone biomarker molecules (ALP, OC, OPN and OCN) and extracellular matrix-related molecules (FN, MMP-13, Dmp1 and aggrecan) were selected for evaluation of osteogenic signalling mechanisms when in presence of these composites. FACS results at day 0 demonstrated that BMSCs were a heterogeneous population with a small percentage of cells staining positive for CD29, CD90 and CD51/61, while staining negative for CD34 and CD45. At day 3, a significant enrichment of cells staining strongly for CD29, CD90 and CD51/61 was achieved. Gene expression patterns for bone growth factors and extracellular matrix molecules were found to be largely dependent upon the size of HA particles. Bone marker molecules, except OCN, had unaltered expression patterns in response to the varied size of HA particles. Overall, the results indicate that larger-sized HA particles upregulate PDGF and these groups were also associated with the most significant increase in osteodifferentiation markers, particularly ALP. Our results suggest that endogenous signalling is dependent upon material properties. Furthermore, we propose that studying gene expression patterns induced by the surrounding biomaterials environment is a fundamental step in the creation of engineered tissues.