Improved Culture-Based Isolation of Differentiating Endothelial Progenitor Cells from Mouse Bone Marrow Mononuclear Cells
Figure 4
FL cells changed the morphology into spindle shape and expressed angiogenesis-related genes under shear stress.
BMMNCs are separated to attached (AT) cells and floating (FL) cells 24 hour after culture with 10%FBS/DMEM medium on temperature-sensitive culture plate followed by further culture with 10%FBS/EBM2 growth factor containing medium for 3 days. The non-separated total adhered cells (TT) were uses as a control. (A) Cells were then cultured under circular flow for 48 hours. Morphological changes were observed under a phase-contrast microscope. Arrow indicates the direction of medium flow. (B) Angiogenesis-related cytokine mRNA expressions in morphologically changed cells under shear stress were analyzed by real-time RT-PCR and triplicated. Cells cultured without shear stress were used as a control (Ctrl). Relative mRNA expressions of each gene normalized to GAPDH are indicated in the graphs. *, P<0.05 vs. AT.