Figure 4

DHA-induced apoptosis was attenuated by Z-VAD.fmk and Caspase-3 activation was determined using FRET receptor photobleaching technique in single living cells. a The effect of DHA and STS (as a positive control) on apoptosis attenuated by Z-VAD.fmk was determined using CCK-8 assay. The ASTC-a-1cells were treated with 20 μg/ml DHA and with or without 10 μM of Z-VAD.fmk for 24 and 48 h, respectively. The result was expressed as mean ± SD of three determinations. n = 3 wells for CCK-8 assay. *P < 0.05, **P < 0.01, compared with control. (b, c, d) Fluorescence images of cells stably expressing SCAT3. The circled parts were the bleaching area. (e, f, g) FRET receptor photobleaching assay determined the activation of caspase-3 during DHA- and STS-induced apoptosis in single cells. e Control. f Positive control (1 μM STS-treated cells for 6 h). g After the treatment of 20 μg/ml DHA for 48 h, fluorescence intensity of CFP channel was nearly invariable during the Venus photobleaching, implying that SCAT3 had been cleaved by the activated caspase-3.b-d: Scale Bar: 10 μm.