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. 2020 Nov 10;25(22):5234. doi: 10.3390/molecules25225234

Figure 2.

Figure 2

Susceptibility of NB cells to cisplatin is increased by fendiline hydrochloride. (A) Real-time RT-PCR quantification of ABC transporters’ mRNA level (ABCA1, ABCA12, and SLC7A11) in wild type SHSY5Y cells, either treated or untreated with 0.01 μM fendiline for 24 h. Values are reported as the mean ± SD. * indicates p ≤ 0.05, ** indicates p ≤ 0.01; two-tailed Student’s t test was applied. (B) Effects of 0.01 μM fendiline and cisplatin (0.5, 5, 50, 100 μM) or their combination on SHSY5Y cell viability after 24, 48, and 72 h of treatment, as detected by MTT assay. Values are expressed as the mean ± SD. (*) indicates p < 0.05; two-tailed Student’s t test was performed between the group treated with cisplatin at different concentrations and the group with cisplatin in combination with fendiline. (C) Cell viability measured by MTT assay after 24 and 48 h of treatment with cisplatin and Fendiline. Results were reported as the values of each sample previously normalized to its untreated control. (D) Demonstration of the dependence on the efflux pumps of the effects observed in the treatments. Sulf: sulfalazine. Prob: probucol. * indicates p ≤ 0.05, *** indicates p ≤ 0.001 (E) Kinetics of cytotoxicity responses for cisplatin (0.5, 5, 50, 100 μM) in wild type SHSY5Y cells, daily treated or untreated with 0.01 μM fendiline, monitored by RT-CES system. Cell index was recorded every 30 minutes and results for each concentration were expressed as the average of three replicates. Data were normalized to the time the compound was added. Red line: untreated cells (DMSO); green line: cisplatin; blue line: fendiline/cisplatin combination. (F) Colony forming capability of wild type SHSY5Y cells to form colonies in methylcellulose in the presence of different treatments (0.01 μM fendiline and/or 0.5 μM cisplatin). Two representative pictures of colonies at day 12 after seeding are reported for each experimental condition (0.01 μM fendiline and/or 0.5 μM cisplatin). Magnification 20×. Scale bar: 200 μm. (G) Statistical analysis of two independent experiments are reported as the mean ± SD (n = 5 microscope fields for each treatment). ** indicates p ≤ 0.01; two-tailed Student’s t test was applied.