(11c) Macroscopic Flow Preconditioning Influences Human Mesenchymal Stem Cells Osteogenic Differentiation in Perfusion Bioreactor

Human MSCs have multi-lineage differentiation potential and have attracted considerable attention in bone tissue regeneration. Combining hMSC with 3D scaffolds is an important strategy in bone tissue regeneration. Perfusion bioreactor plays a critical role in regulating 3D construct microenvironment and in providing controlled physiochemical and biomechanical environments. Although media flow is known to influence hMSC growth and construct microenvironment such as ECM secretion and organization, its subsequent impact on the osteogenic differentiation in 3D construct is unknown. We hypothesized that the preconditioning of 3-D hMSC construct by the macroscopic flow (i.e., transverse or parallel) influences constructs microenvironment, and the subsequent osteogenic differentiation upon induction. Two different flow regimens, including the parallel and transverse, modes are produced in the in-housing perfusion bioreactor system. Their effects on hMSCs' proliferation, the ECM microenvironment, and the gene expression are quantitatively analyzed after 7 days at 0.1 ml/min and 0.3 ml/min, respectively. After pre-conditioning, the constructs are induced to undergo osteogenic differentiation for an additional 7-14 days under either parallel or transverse flow to investigate the effects of pre-conditioning on constructs' osteogenic differentiation. The ALP expression and ECM secretions are being analyzed and quantified using ALP assay and Western blot, whereas gene expressions for osteogenic differentiation are being determined by qPCR. Together, the results will provide important insight into the impact of macroscopic flow on construct microenvironment and the effect of the macroscopic flow pre-conditioning on the osteogenic tissue regeneration.