Figure 6.

In vitro synergistic effect of OLA and AA and evaluation of the cytotoxic mechanisms. (A) Cytotoxic activity of ascorbic acid (75 and 100 µg/mL) and OLA (15 μM) against PANC-1 cell line after 72 hours of exposure. (B) Graphical representation of the real and theoretical inhibition (sum of both treatments separately) exerted by the combination of this AA and OLA doses in vitro and values of the combinatorial index (CI) obtained after combining both treatments in PANC-1 cell line. (C) Cell migration of PANC-1 treated with OLA, NP-ACP-OLA-AA, and their corresponding controls (DMSO and NP-ACP-AA) for 72 hours. (D) Representative immunofluorescence microscopy images of Ki67 protein expression and after performing a TUNEL assay treating PANC-1 cells with 100 μM of each treatment for 24 hours. Pictures were collected with a 10X objective. Scale bar = 200 nm. (E) Graphical representation of the Ki67 nuclear foci detected in each condition relative to the number of cells. (F) Graphical representation of the quantification of TUNEL-stained nuclei in comparison to the total nuclei. Nuclear staining was performed using Hoechst. Images were collected through a 10X objective. (G) Western blot analysis of phosphorylated γ-H2AX after 24-hour treatment of PANC-1 cells with OLA, NP-ACP-AA, and NP-ACP-OLA-AA NPs. All the data are presented as mean ± S.D (n=3). *p ≤ 0.05, **p ≤ 0.01 and ***p ≤ 0.001.

Abbreviation: ns, not significant.