Melatonin Rescued Reactive Oxygen Species-Impaired Osteogenesis of Human Bone Marrow Mesenchymal Stem Cells in the Presence of Tumor Necrosis Factor-Alpha
Melatonin decreased TNF-α-induced ROS generation during osteogenesis of BMSCs. BMSCs were cultured and underwent osteogenesis in osteogenesis medium containing vehicle (0.05 mol/L ethanol), TNF-α (10 ng/mL), or both TNF-α (10 ng/mL) and melatonin (100 μmol/L). (a) The adherent cells in a 6-well plate were incubated with DMEM containing CM-H2DCFDA (10 μmol/L) for 30 min at 37°C in darkness. Fluorescence was observed by microscopy in a representative sample on days 3, 7, and 14. Green fluorescence indicates ROS-positive cells. Scale bars: 200 μm. (b, c) The cells were resuspended in DMEM containing CM-H2DCFDA (10 μmol/L) and incubated for 30 min at 37°C in darkness. Measurement of ROS was analyzed with a flow cytometer. The graphs in (b) represent typical results of fluorescence intensity; the results in (c) are representative of three independent experiments. versus the vehicle group, versus the TNF-α group.
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