Detection of caspase-3 activation in single cells by fluorescence resonance energy transfer during photodynamic therapy induced apoptosis.
In many apoptosis pathways, activation of caspase-3 is considered the final stage. In this study, we studied PDT induced caspase-3 activation with fluorescence resonance energy transfer (FRET) technique. A recombinant caspase-3 substrate, SCAT3, was used as the FRET probe. FRET fluorescence images were collected after PDT or TNF-alpha induced apoptosis. By analyzing the dynamic changes of FRET fluorescence, the results indicate that the caspase-3 activation started immediately after the PDT treatment. In contrast, FRET disruption caused by caspase-3 activation started at 3h after TNF-alpha treatment, due to different signaling pathway. The results have proofed, for the first time, that FRET is a sensitive technique that can be used to investigate PDT-induced activation of caspase-3 in real-time and in single cells. By choosing appropriate recombinant substrates as FRET probes, it is likely that FRET technique will provide a new real-time means to study the mechanism of PDT at single cell level.
Wu
Yunxia
Y
Institute of Laser Life Science, South China Normal University, Guangzhou 510631, China.
Xing
Da
D
Luo
Shiming
S
Tang
Yonghong
Y
Chen
Qun
Q
D000230:Adenocarcinoma / Q000188:drug therapy / Q000201:enzymology / Q000473:pathology; D017209:Apoptosis / Q000187:drug effects*; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism*; D045744:Cell Line, Tumor; D004789:Enzyme Activation; D031541:Fluorescence Resonance Energy Transfer; D006801:Humans; D008175:Lung Neoplasms / Q000188:drug therapy / Q000201:enzymology / Q000473:pathology; D010778:Photochemotherapy; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D014409:Tumor Necrosis Factor-alpha; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases