Figure 6

Caffeine exacerbates cisplatin-induced cell death in mouse explant cultures and UB/OC-1 cells. (A) Cochlear explants from neonatal (P3–P5) mice treated with cisplatin (20 µM) for 48 h show significant loss of myosin VIIa-stained OHCs (white arrows). The loss was enhanced by pre-treatment with caffeine (100 µM). Representative images are shown. Scale bar represents 25 µm. Data in (B) indicate mean percentage loss of OHCs ± SEM and were derived from (A) (n ≥ 3). (C) UB/OC-1 cells were treated with caffeine (100 μM) for 0.5 h, followed by cisplatin (20 μM) for an additional 24 h. Cell viability as determined by MTS assay showed that caffeine reduced the cell viability of cisplatin treated UB/OC-1 cells. Percent cell viability is presented as mean ± SEM of three independent experiments. (D) RNA was isolated using TRI reagent and expression of TNF-α, iNOS and COX2 were determined by real time RT-PCR and normalized to GAPDH, a house keeping gene. Caffeine treatment enhanced the expression of inflammatory genes in cisplatin-treated UB/OC-1 cells. Data are presented as mean fold change ± SEM (n ≥ 3). Asterisks, *p < 0.05 vs. vehicle, **p < 0.05 vs. cisplatin (one-way ANOVA).