(685e) Oxygen Tension Regulates hMSC Expansion and Formation of Microenvironment

Human mesenchymal stem cells (hMSCs) isolated from bone marrow reside in a low oxygen tension environment in vivo. In vitro results from recent studies also showed that MSCs exhibited greater colony forming ability, proliferated faster and longer, and maintained their undifferentiated characteristics better under low oxygen conditions. Hypoxia also influences the secretion of extracellular matrix (ECM) proteins, which formed part of the specialized niche that facilitated expansion of marrow-derived mesenchymal progenitor cells and prevented their differentiation into osteoblasts. As oxygen tension and ECM microenvironment are important components of the hMSC microenvironment, the objective of this study is to investigate the effects of oxygen tension on hMSC ECM secretion and its sequent influences on hMSC morphology, proliferation, and multi-lineage differentiation. We firstly investigated the effect of oxygen tensions on hMSC morphology and proliferation using flow cytometery and image analysis. hMSC morphology has been shown to correlate with proliferation potential, and smaller and more spindle-shaped cells were found under 2% O2. Secondly, hMSCs were cultured on the hMSC-derived ECM produced under normoxia (i.e., 20% oxygen) or hypoxia (i.e., 2% oxygen) conditions to investigate whether hMSC-derived ECM promotes retention of the stem cell characteristic. Cell proliferation will be measured by DNA, CFU-F, and expression of cell cycle markers, whereas Western Blot and immunostaining assays will be used to quantify ECM expression. Cell differentiation potential will be determined using qPCR and functional assays. Together, the results will help determine the role of oxygen tension as a signaling molecule as well as a metabolic substrate.