This document summarizes research on mouse models of Alzheimer's disease conducted by the RIKEN Brain Science Institute Proteolytic Neuroscience Laboratory in Japan. It discusses several generations of mouse models that were developed, including those that overexpress mutant amyloid precursor protein (APP) alone or crossed with other transgenic mice, as well as knock-in models incorporating specific APP mutations. The models have provided insights into amyloid-beta deposition and inflammation with aging. Ongoing work aims to further elucidate mechanisms of pathology and evaluate modulating factors like proteases, vaccines, and genes.

1. Greetings: RIKEN BSI Proteolytic Neuroscience Laboratory
BSI Central Building, April 17, 2014.

2. Takaomi C. Saido
Laboratory for Proteolytic Neuroscience
RIKEN Brain Science Institute
JAPAN
Alzforum Webiner
April 22, 2014
2nd
generation mouse models of Alzheimer’s disease:
12 years of work and future perspectives

3. Crossing Tg mice overexpressing
mutant APP with other Tg mice is
a risky paradigm.
Example: APP-Tg X Calpastatin-
Tg/KO.

4. 0
20
40
60
80
100
0 10 20 30 40 50 60
Age (weeks)
Proportionsurviving(%)
WT
CS-KO
APP/CS-Tg
APP-TgAPP-Tg
CS-KO (+/-)
APP-Tg
CS-KO (-/-)
Effect of calpastatin (CS) overexpression &
deficiency on life span of APP-Tg mice
Higuchi et al. FASEB J. (2012

5. 2nd
generation KI mouse model of A amyloidosis:β APPNL-F
β-cleavage site γ-cleavage
site
671
Aβ42Aβ40
R Y H
humanization
Aβ:3-fold
NL
SEVKM DAEFGHDSGFEVRHQKLVFFAEDVGSNKGAIIGLMVGGVV IA TVIVITLVMLK
670
Overproduction of Aβ1-42 Aβ42/Aβ40 ratio: 30-fold
F Beyreuther/Iberian
mutation
Saito et al. (Nat Neurosci, 2014
Overproduction of APPsβ
NL-F
Relevant negative control?

6. negative control for the 2nd
generation KI mouse model: APPN
β-cleavage site γ-cleavage
site
671
Aβ42Aβ40
R Y H
humanization
Aβ:3-fold
NL
SEVKM DAEFGHDSGFEVRHQKLVFFAEDVGSNKGAIIGLMVGGVV IA TVIVITLVMLK
670
Overproduction of Aβ1-40/42
Saito et al. (Nat Neurosci, 2014
Age of onset: 50-60 years
Overproduction of APPsβ
NL
Relevant negative control!

7. 500 µm
Amyloidburden(%)
HippocampusCortex
4G8 15 mo 18 mo12 mo9 mo
21 mo 24 mo
Age-dependent Aβ deposition in NL-F/NL-F
Aβ deposition could be detected from 6 mo of NL-F/NL-F brains

8. APP KICS-KO
CS-KO; APP KI
Modulation of calpain activity in APP-KI mouse brain by
crossbreeding with Calpastatin (CS)-KO mice
New!

9. Survival of APPNL/F
-KI x Calpastatin (CS)-KO
mice
CS deficiency did not affect the survival of APPNL-F
KI
mice at least up to 20 months.

10. 1. Role of inflammation
2. Role of various proteases
3. Effect of Aβ vaccination
4. Effect of ApoE4
5. Effect of the environmental
enrichment
6. Effect of various diets and
supplements
7. Others (Tau, Dynamin, Arc, etc.)
Major issues to be re-addressed

11. 3rd
generation KI mouse model of A amyloidosis: APPβ NL-G-F
β-cleavage site γ-cleavage
site
671
Aβ42Aβ40
R Y H
humanization
NL
SEVKM DAEFGHDSGFEVRHQKLVFFAEDVGSNKGAIIGLMVGGVV IA TVIVITLVMLK
670
F Beyreuther/Iberian
mutation
Saito et al. (Nat Neurosci, 2014
SEVNL DAEFRHDSGYEVHHQKLVFFAEDVGSNKGAIIGLMVGGVV IA TVFVITLVMLK
G (Arctic mutation)
Oligomerization-prone
NL-G-F

12. 500 µm
2 months 4 months 7 months
Aβ42 pathology of NL-G-F KI mice

13. Neuroinflammation in APPNL-G-F/NL-G-F
KI
Greater neuroinflammation was found around the plaques
in APPNL-G-F/NL-G-F
mice than aged APPNL-F/NL-F
KI mice.
APPNL-F/NL-F
(24 mo) APPNL-G-F/NL-G-F
(9 mo)
Aβ(82E1) microglia astrocyte merge

14. Differential utility of NL-F and NL-G-F mice
1. NL-F: Mechanism of Aβ deposition
(up-stream targets).
2. NL-G-F: Mechanism of
inflammation, tauopathy and
neurodegeneration (down-stream
targets).

16. Finally, we should compare our mice with APPNL/NL
mice
crossbred with mutant presenilin 1 (PS1) knockin mice24
,
which also overproduce Aβ42 without overexpressing
APP. These APP/PS1 double knockin mice are
comparable to the APPNL-F/NL-F
mice in this respect. There
are, however, two major difficulties that limit the utility
of the double knockin mice. The first is that they are
doubly homozygous: crossing them with other mutant
mice is practically impossible. To our knowledge, no
researchers have done so as yet. The second is that
presenilin mutations affect biological processes other
than γ-cleavage of APP25
; no perfect negative controls
exist. It should also be noted that the function(s) of PS1
may, at least in part, differ between humans and mice26
.

17. RIKEN BSI
Lab. for Proteolytic Neuroscience
Takashi Saito
Nobuhisa Iwata (Nagasaki U)
Makoto Higuchi (NIR)
Jiro Takano
Satoshi Tsubuki
Per Nilsson
Naomasa Kakiya
Kaori Tsukakoshi
Hayato Isshiki
Ko Sato
Shoko Hashimoto
Contributors and Collaborators
RIKEN BSI
Lab. for Behavioral Genetics
Shigeyoshi ItoharaNational
Institute for Geriatrics and Gerontology
Akihiko Takashima
University of Minnesota
Karen Hsiao-Ashe
Novartis Institute of Medical Research
Matthias Staufenbiel
Jichi Medical University
Shin-ichi Muramatsu
Harvard Medical School
Cynthia Lemere