Figure 5.

Association between ROS and intracellular Ca²⁺ in Dex- and Cro-treated MC3T3-E1 osteoblasts. Cells were pretreated with 100 µM Cro, 2 mM NAC, 20 µM BAP, 100 µM H₂O₂ or 0.5 µM Ion for 1 h prior to treatment with 1 µM Dex for 24 h. (A) Visualization of ROS generation and intracellular Ca²⁺ by fluorescence microscopy (magnification, ×100). Quantitative analysis of (B) ROS production and (C) intracellular Ca²⁺ levels following pretreatment with NAC or BAP, and treatment with Dex, as determined via flow cytometry. (D) Visualization of ROS generation and intracellular Ca²⁺ by fluorescence microscopy (magnification, ×100). Quantitative analysis of (E) ROS production and (F) intracellular Ca²⁺ levels following pretreatment with Cro with or without H₂O₂ and Ion, and treatment with Dex, as determined via flow cytometry. Data are presented as the means ± standard deviation of three independent experiments. ***P<0.001 vs. Ctrl; ^###P<0.001 vs. Dex; ^&&&P<0.001 vs. Dex + Cro. BAP, 1,2-bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid; Cro, crocin; Dex, dexamethasone; Ion, ionomycin; NAC, N-acetyl-L-cysteine; ROS, reactive oxygen species.