Panel A shows fluorescence micrographs comparing CellROX staining after ASO and liproxstatin-1 treatments. Panel B shows density plots comparing normalized nuclear-to-cytoplasm CellROX fluorescence ratios across treatment conditions. Panel C shows fluorescence micrographs of oxidized C11 green staining following ASO and liproxstatin-1 treatments. Panel D shows density plots comparing normalized oxidized-to-reduced C11 fluorescence ratios under different treatments. Panel E shows fluorescence micrographs and enlarged views of oxidized and reduced C11 localization.
EPHA2 is required for trafficking of cET-ASOs to leaky endosomes. (A) Fluorescence micrographs of KPC cells treated either with vehicle (untreated) or cET-ASOKras (5 μM) in combination with vehicle or liproxstatin-1 (1 μM) for 4 h. Nuclei (blue) and CellROX green staining are displayed (left panels are merged, right panels are CellROX only). (B) Quantification of CellROX green nuclear translocation. Density plot data are expressed as nuclear/cytoplasmic ratio (log10-transformed pseudo-counts), n = 5 independent experiments, ANOVA Tukey post hoc test. (C) Fluorescence micrographs of KPC cells treated either with vehicle or cET-ASOKras (5 μM) in combination with vehicle (untreated) or liproxstatin-1 (1 μM) for 4 h. Nuclei (blue) and C11-bodipy (red and green denoting reduced and oxidized lipids respectively) staining (left panels are merged, right panels are oxidized C11-bodipy only) are displayed. (D) Quantification of C11-bodipy oxidation. Density plot data are expressed as the oxidized/nonoxidized ratio (log10-transformed pseudo-counts), n = 4 independent experiments, ANOVA Tukey post hoc test. (E) Airyscan micrographs of KPC cells treated with either vehicle or cET-ASOKras (5 μM) and stained with C11-bodipy.