Quantitative imaging of apoptosis commitment in colorectal tumor cells

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• 作者：Michael Keese ; Martin Offterdinger ; Christian Tischer ; Andreas Girod ; Piet H.M. Lommerse ; Vugar Yagublu ; Richard Magdeburg ; Philippe I.H. Bastiaens
• 关键词：fluorescence correlation spectroscopy (FCS) ; FRET ; caspase-3 ; EGFR ; fluorescence lifetime imaging microscopy (FLIM) ; apoptosis
• 刊名：Differentiation
• 出版年：2007
• 出版时间：November 2007
• 年：2007
• 卷：75
• 期：9
• 页码：809-818
• 全文大小：211 K

文摘

We have studied caspase-3 activation by combined DNA damage induction and EGFR kinase inhibition in order to identify potential EGFR-mediated survival signals conferring resistance to apoptosis in human colorectal tumor cells. The onset of apoptosis was microscopically imaged with a newly developed caspase-3 substrate sensor based on EGFP and tHcred1, enabling us to monitor caspase-3 activation in cells by fluorescence lifetime imaging microscopy or fluorescence correlation spectroscopy. Both optical approaches provide parameters quantitatively reporting the ratio between cleaved and uncleaved sensor, thereby facilitating the comparison of caspase-3 activation between different cells. Using these methods, we show that EGFR kinase inhibitors sensitize colorectal SW-480 tumor cells for 5-fluorouracil-induced apoptosis, indicating that EGFR-mediated survival signaling contributes to apoptosis resistance via its intrinsic kinase activity.