Fig. 1

SHEDs characterization and effects of inorganic phosphate on cell functions. CD45, CD73, CD90, and CD105 expression were examined using flow cytometry (A). For differentiation ability assay, cells were maintained in an osteogenic or adipogenic induction medium. Mineral deposition and intracellular lipid accumulation were evaluated using Alizarin Red S staining on day 14 (B) and Oil Red O staining on day 20 (C), respectively. Cells cultured in a normal growth medium were used as the control (left panels). Cells were treated with 2.5 or 5 mM inorganic phosphate (P_i) for 1, 3, and 7 days. Cell viability was determined using MTT assay (D). Cells were treated with foscarnet for 30 min prior to P_i exposure and cell viability was examined using the MTT assay on day 7 (E). The mRNA expression of Ki67 was determined at 24 h after P_i exposure using real-time polymerase chain reaction (F and G). On day 3, cells were stained with propidium iodide to determine cell cycle progression by flow cytometry, and the quantitative measurement of proliferative cells was illustrated (H). Cells were stained with annexin V/ propidium iodide and further analyzed using flow cytometry. In order to elucidate cell apoptotic numbers, the percentage of cell death was quantified (I). Cell migration was examined using an in vitro scratch assay (J) and the percentage of area closure was calculated (K). ^*P < 0.05 compared to the control, ^#P < 0.05 compared to the P_i group.