Figure 15.

Voltage Dyes Showed that U87 Cells Treated with NS1643 and a Combination of NS1643 and Pantoprazole for 6 Days Showed a Hyperpolarization and YAP Increases its Translocation to the Cytoplasm in NS1643 or Pantoprazole with TMZ, and Pantoprazole with NS1643 Treatment. Immunofluorescence of cells was conducted and analyzed with CellProfiler for integrated fluorescence intensity. Dye assays were analyzed for mean intensity, except for LysoSensor Green which was analyzed for integrated intensity. (A) Stain of lysosomal pH with LysoSensor Green, low levels indicate alkalization. (B) Dye indicator of membrane voltage, DiBAC4(3), low levels indicate hyperpolarization. (C) Dye indicator of cytoplasmic pH, pHRodo Green, low levels indicate alkalization. (D) Dye indicator of cytoplasmic calcium, Fluo-4AM, high levels indicate an increase in calcium. (E) The ratio of nuclear to cytoplasmic Yes-associated protein (YAP), lower levels indicate translocation to the cytoplasm. Treatments corresponding to the colored bars are outlined in the figure itself. The log of the fold change in intensity was compared between single treatments and combined treatments, except the positive control, with significant values shown in the color of the treatment compared. The initial fluorescence intensities were compared to their corresponding control, with significant values shown under the bars as, ***: p < 0.001, **: p < 0.01, *: p < 0.05 (one-way ANOVA with Tukey post hoc analysis n > 3 technical replicates).