Figure 2

Inhibition of alkaline phosphatase activity decreases cell viability and increases apoptosis. Knockdown of alkaline phosphatase was confirmed in ARCaPM cells transduced with scrambled shRNA (shSCR) or ALPL-knockdown shRNA (shALPL2 and shALPL3) by measuring (A) ALPL mRNA using qRT–PCR and (B) alkaline phosphatase activity (normalised with total protein) using a quantitative fluorimetric assay, 72 h after transduction. (C) Viability of the transduced cells was measured using Alamar Blue assay and (D) cell death was measured using YO-PRO-1. (E) Apoptotic nuclei, identified from their fragmented appearance, were counted in ImageJ (imagej.nih.gov). (F) Apoptosis was quantitated using annexin V/PI staining. Viability (G) and (H) cell death were also measured in ARCaPM cells treated with increasing concentrations of levamisole using Alamar Blue assay and YO-PRO-1 staining, respectively. (I) ALPL knockdown in PC3 cells was confirmed using qRT–PCR and (J) the effects of viability were measured using Alamar Blue assay and (K) cell death was measured using YO-PRO-1 staining. (*P<0.05, **P<0.01, ***P<0.001, ****P<0.0001).