FIGURE 4.

Knockdown of Opa1 aggravates cisplatin-induced apoptosis and mitochondrial dysfunction in HEI-OC1 and Neuro 2a cells (A,B) CCK-8 was used to assess cell viability in HEI-OC1 (A) and Neuro 2a (B) cells treated with the indicated doses of cisplatin for 24 h. **p < 0.01, ***p < 0.001 (one-way ANOVA with Dunnett’s multiple comparisons test). (C) Quantification of Opa1 mRNA expression levels in HEI-OC1 and Neuro 2a cells treated with cisplatin for 24 h. **p < 0.01, ***p < 0.001 (two-way ANOVA). (D) Representative Western blot images of Opa1 expression in HEI-OC1 and Neuro 2a cells treated with cisplatin for 24 h. (E) Representative confocal microscopic images of MitoTracker staining at 24 h after cisplatin treatment of HEI-OC1 cells. Right panels show magnified views of the areas marked in the left panels. (F,G) CCK-8 was used to assess cell viability in HEI-OC1 (F) and Neuro 2a (G) cells at 24 h after siRNA transfection with or without cisplatin treatment. N.S., non-significant, *p < 0.05 (two-way ANOVA). (H) Representative flow cytometric plots were generated from HEI-OC1 cells stained with Annexin V-FITC/PI to detect apoptosis at 24 h after siRNA transfection with or without cisplatin treatment. (I) The percentage of late apoptotic cells in each group. ***p < 0.001 (one-way ANOVA with Dunnett’s multiple comparisons test). (J) Representative flow cytometric plots were generated from Neuro 2a cells stained with Annexin V-FITC/PI to detect apoptosis at 24 h after siRNA transfection with or without cisplatin treatment. (K) The percentage of late apoptotic cells in each group. *p < 0.05, ***p < 0.001 (one-way ANOVA with Dunnett’s multiple comparisons test). (L) Intracellular ROS levels were measured by flow cytometric analysis of HEI-OC1 cells stained with DCFH-DA at 24 h after siRNA transfection with or without cisplatin treatment. (M) Quantitative histograms of the mean fluorescence intensity of DCFH-DA in HEI-OC1 cells. *p < 0.05, **p < 0.01 (two-way ANOVA).