[Fluorescence emission analysis of caspase-3 activation induced by Xiao-Ai-Ping (XAP) inside living human lung adenocarcinoma cells].

ABSTRACT

The CCK-8 was used to measure the inhibition effect of Xiao-Ai-Ping (XAP), a traditional medicine, on the human lung adenocarcinoma (ASTC-a-1) cells viability. The ASTC-a-1 cells expressing stably with SCAT3, a fluorescence resonance energy transfer (FRET) plasmid based on the green fluorescent protein mutants (GFPs), was verified using confocal fluorescence scanning microscopy imaging, fluorescence emission spectra and FRET acceptor photobleaching techniques. The caspase-3 activation can be monitored by the fluorescence emission spectra of SCAT3 inside living cells. The cells expressing stably with SCAT3 were cultured with XAP for 96 hours, and the fluorescence emission spectra of the SCAT3 inside living cells were measured at the time of 0, 24, 72, and 96 hours, respectively. Experimental results showed that (1)XAP inhibited obviously the proliferation of ASTC-a-1 cells and induced the cell death. The inhibition of XAP on the cells was dose-dependent; (2)the SCAT3 inside living cells was cleaved completely 72 hours after the XAP treatment, implying that a great deal of pro-caspase-3 was activated by XAP; (3)24 hours after XAP treatment, the emission spectra of SCAT3 inside living cells cultured in DMEM without XAP for 48 and 72 hours did not change greatly, implying that XAP did not activate obviously caspase-3 within 24 hours.