Involvement of the mitochondrial intrinsic pathway in the protective effect of ODN on astroglial cells.

<p>(A) Cultured astrocytes were pre-incubated for 10 min in the absence (a, c) or presence of 0.1 nM ODN (b, d), and then incubated for 1 h with medium alone (a), ODN (b) or with 300 µM H₂O₂ without (c) or with ODN (d). Aggregated (red signal) and monomeric (green signal) fluorescent JC-1 dye revealed active and inactive mitochondria, respectively. Scale bar, 50 µm. (B) Cultured astrocytes were pre-incubated for 10 min in the absence or presence of 0.1 nM ODN, and then incubated for 1 h with medium alone, ODN or with 300 µM H₂O₂ without or with ODN. Bcl-2 and Bax mRNA levels were quantified by RT-PCR. <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0042498#s2" target="_blank">Results</a> are expressed as percentages of control. Each value is the mean (± SEM) of 6 different wells from three independent cultures. ANOVA followed by the Bonferroni's test. * <i>p</i><0.05; NS, not statistically different <i>vs.</i> control. ^#<i>p</i><0.05; ^##<i>p</i><0.01 <i>vs.</i> H₂O₂-treated cells. (C) Cultured astrocytes were pre-incubated for 30 min in the absence or presence of H89 (20 µM), U0126 (20 µM), U73122 (1 µM) or chelerythrine (Chel; 1 µM) and then incubated for 1 h with medium alone, ODN (0.1 nM), dbcAMP (1 mM) or with 300 µM H₂O₂ without or with ODN (0.1 nM) or dbcAMP (1 mM). Caspase-3 activity was quantified by measuring the fluorescence of caspase substrate, Z-DEVD-Rhodamine 110, and the results are expressed as percentages of control. Each value is the mean (± SEM) of at least 12 different wells from three independent cultures. ANOVA followed by the Bonferroni's test. <i>*** p</i><0.001; NS, not statistically different <i>vs.</i> control. ^###<i>p</i><0.001; ns, not statistically different <i>vs.</i> H₂O₂-treated cells.