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P1‐298: Characterization of novel apoE isoform‐dependent C‐terminal specific proteolysis of Aβ1‐42 by microglia
Author(s) -
Liu Feng,
Lin Suizhen,
Chai Xiyun,
DeMattos Ronald B.,
Paul Steven M.,
Lilly Eli
Publication year - 2010
Publication title -
alzheimer's and dementia
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 6.713
H-Index - 118
eISSN - 1552-5279
pISSN - 1552-5260
DOI - 10.1016/j.jalz.2010.05.850
Subject(s) - microglia , proteolysis , apolipoprotein e , catabolism , biochemistry , chemistry , microbiology and biotechnology , biology , metabolism , immunology , inflammation , medicine , enzyme , disease
involved in several pathological conditions, like cancer and degenerative disorders. Alzheimer’s disease (AD) is the most common form of dementia involving massive cell death of neurons. However, the cause of AD at the present time is still unknown, although, amyloid-b (Ab) peptide has been suggested to be the triggering factor. Methods: In order to detect localized caspase activation in live cells we designed sensors for caspase-3, -6 and -9 utilizing fluorescence resonance energy transfer (FRET). The FRET-ing sensor molecules, consisting of CFP and YFP separated by a linker containing a specific caspase cleavage motif, were designed to signal caspase cleavage by the loss of FRET. Differentiated SH-SY5Y cells were used as a model system for neurodegeneration. The cells were treated with oligomeric Ab42 or staurosporine as a positive control of apoptosis. The cleavage of the sensors during induced apoptosis was verified by western blot analysis. Time-lapse FRET microscopy was used to monitor caspase activity in different parts of the cells. Results: In our study, when the cells were exposed to staurosporine we were able to detect local activity of caspase-6 initially in the soma of the cells, whereas caspase-6 activity in the neurites was delayed. Furthermore, our study shows that oligomeric Ab42 is able to activate caspase-3, -6 and -9. In contrast to staurosporine, in Ab42 treated cells loss of FRET occurred globally indicating that caspase was activated simultaneously in soma and axons. Conclusions: In conclusion, we show that our caspase-sensors are able to detect local caspase activity in vitro. We also show that exposure to oligomeric Ab42 results in global activation of caspases in differentiated SHSY5Y cells.