2015 Fiscal Year Research-status Report
| Project/Area Number |
15K18369
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| Research Institution | Institute of Physical and Chemical Research |
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Principal Investigator |
NILSSON PER 国立研究開発法人理化学研究所, 脳科学総合研究センター, 研究員 (10568934)
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| Project Period (FY) |
2015-04-01 – 2017-03-31
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| Keywords | Autophagy / Aβ metabolism / Neurodegeneration / Alzheimer’s disease / APP knock-in mouse model |
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| Outline of Annual Research Achievements |
To investigate the role of autophagy in amyloid-beta metabolism of Alzheimer’s disease we generated autophagy-deficient amyloid precursor protein (APP) knock-in (KI) mice. APP-KI mice are the most relevant mouse model for AD recently generated in our laboratory which do not harbor the artifacts associated with transgenic mice overexpressing APP. Interestingly, biochemical and immunohistochemical analysis revealed that autophagy-deficiency induced a significant reduction of extracellular amyloid-beta plaques in the APP-KI mice. The reduction of extracellular amyloid beta plaque occurred concomitantly with an accumulation of amyloid-beta inside the neurons. This indicated that autophagy mediates the secretion of amyloid-beta from the neurons. Previous work has shown that autophagosomes can fuse with multivesicular bodies from which exosomes can be released. We therefore manipulated multivesicular body formation in a cell culture system by performing an si-RNA mediated screen to knockdown proteins involved in the generation of multivesicular bodies. The results show that knockdown of such proteins decreased amyloid-beta secretion indicating that amyloid beta is secreted, at least to some extent, through multivesicular body formation. In addition, the neurodegeneration that we have previously observed in autophagy-deficient APP transgenic was not observed in autophagy-deficient APP-KI mice. This shows that the neurodegeneration could be due to APP overexpression and not directly to amyloid beta. Activation of autophagy could hence be a possible treatment for AD.
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| Current Status of Research Progress |
Current Status of Research Progress
2: Research has progressed on the whole more than it was originally planned.
Reason
We have generated autophagy deficient APP knock-in mice by breeding mice with a conditional knock out of autophagy-related gene 7 (Atg7) and a Cre-recombinase transgenic mice (under the control of CamKII promoter) with APPNL-F mice (bearing the Swedish and the Beyreuther mutations). We have confirmed a significant decreased Aβ plaque load, consistent with previous data obtained with autophagy-deficient APP transgenic mice (Atg7 cKO x APP Tg). In addition, genetic deletion of Atg7 leads to a significant increase in intracellular Aβ accumulation, also consistent with previous data obtained from Atg7 x APP Tg mice, showing that autophagy plays an important role in the metabolism of Aβ including the secretion. A preliminary screen to knockdown proteins known to be involved in autophagy-mediated secretory routes has been performed which indicate the involvement of multivesicular bodies in the secretion of Aβ. To investigate if autophagy deficiency together with Aβ amyloidosis induce neurodegeneration, which was observed previously in the Atg7 cKO x APP Tg mice, the brains of the mice were investigated both by histological means of the brains and by MRI in living mice. However, the increased neurodegeneration observed in Atg7 cKO x APP Tg mice were not observed in the Atg7 cKO x APP-KI mice, which may indicate that the neurodegeneration is an artefact from APP overexpression.
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| Strategy for Future Research Activity |
The applicant of this kakenhi will transfer to Karolinska Institutet, Sweden from June 2016. However, the research project will continue and the Atg cKO x APP-KI mice will be transferred to Karolinska Institutet where the analysis will proceed. In addition, some aged mice will be kept during an overlap period and brain samples will be prepared at RIKEN BSI (for example for the IEM experiments).
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