PoTingHo-2014 IBC Summer Internship Program
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1
2014 IBC Summer Internship Program
Alzheimer’s Disease treatment with Chinese herbal extracts in APP/PSEN1 transgenic mice
Supervisor: Rita P. Y. Chen Ph. D.
Institute of Biological Chemistry, Academia Sinica
Presented by: Po-Ting, HoAug. 29, 2014
2 Mo/HuAPPswe HuPSEN1dE9
Increase total Aβ
Aβ aggregation
plaques
Dementia
Neur
on d
amag
e
Aβ deposit
s
Morris Water Maze
ELISA
Behavior test
Aβ quantification
INTR
OD
UCT
ION
locationIHC/ Thioflavin S stain
Vorhees, C. V. 2006. Nature protocolsZhang, C. (2009) Harvard Medical School.
3
3rd month
Genotyping Treatments Sacrifice
10th month
Treatments started
Experimental Timeline
My Internship
Period
Morris Water Maze
4
11thGenotyping Treatments Sacrifice
Experimental Timeline
Morris Water MazeTreatments started
3rd month 10th month
5 Chinese Herbal Extracts Treatment
Extract A
Extract B
APP
WT
• Dosage: 250 mg/kg a day by gavage
Mat
eria
l
6
11thGenotyping Treatments Sacrifice
Experimental Timeline
Treatments started
3rd month
Morris Water Maze
10th month
7Morris Water Maze
• Behavior testing: to study spatial learning and memory.
N
E
S
W
100 cm
35 cm
Water + milk
Hidden platform
Visual cues
Mat
eria
l
Vorhees, C. V. 2006. Nature protocols.
Q3
8 Spatial learning test Probe test
Day 1 Day 2 Day 3 Day 5Day 4 Day 6
4 trials a day
swim for 90 sec.
guide for 30 sec.
Grab them to the
platform for 30 sec.
Stay on the platform for
30 sec.
Remove platform
1 trial
Swim for 60 sec.
yes
no
no yes Latency time
Duration and cross number of
Q3Met
hods
Vorhees, C. V. 2006. Nature protocols.
9Re
sult
sSpatial Learning trials
D1 D2 D3 D4 D50.0
20.0
40.0
60.0
80.0
100.0897 994 APP WT
Lat
ency
(sec
)Extract A Extract B
N=9 N=9 N=4 N=10
10Re
sult
sProbe trial
897 994 APP WT0.0
2.0
4.0
6.0
8.0
10.0
Q3 (Cross number)
Cro
ss n
umbe
r (b
outs
)
897 994 APP WT0.0
5.0
10.0
15.0
20.0
25.0
30.0
Q3 (Duration, sec)T
ime
(sec
)
Extract AExtract A
Extract BExtract B
*
* P <0.05 compared with APP mice.
11
11thGenotyping Treatments Sacrifice
Experimental Timeline
3rd month
Treatments started
10th month
Morris Water Maze
12 Sacrifice
http://www.zviratka-verunky.estranky.cz/fotoalbum/zvirata/mysi/illustrated_brown_mouse.jpg.-.htmlHatziioannouk T. 2012. Nature Reviews Microbiology 10, 852-867http://www.biologycorner.com/worksheets/rat_head.html
Serum
HeartLungsLiverKidneysspleen
HippocampusCerebral cortexCerebellumBrain StemOlfactory lobes
ELISA
Toxicity test
Met
hods
Brain tissue IHC/ ThS stain
13• The results of probe trial suggested that the treatments with Chinese herbal
extracts A and B might have potential to improve memory deficits.
• Important factors that do matter:
mice should find the platform by memory instead of sights.
a) The proportion of milk and water b) The depth of water
Conc
lusi
onCo
nclu
sion
14
I would like to thank Professor Chen for her expert advice and encouragement throughout this difficult project, as well as Ramus for his brilliance in the lab.
Ackn
owle
dgm
ents
15
Questions?
Zhang, C. (2009) Harvard Medical School.
(APP Intracellular Domain)
B6C3-Tg (APPswe/PSEN1dE9) Double Transgenic Mice
Mo/HuAPPswe
• Swedish mutations amyloid precursor protein
• Elevate the amount of Aβ
• Exon-9-deleted mutant human presenilin 1
• Increase the level of Aβ40 and Aβ42 in the brain
• Both genes are directed to CNS neurons
The Jackson Labortory
Mutant Mouse Regional Resource Centers supported by NIH
Purpose: Make sure there are the presences of transgenes, APP and PSEN, construct in transgenic mice.
Mr. PC 1047 1048 1051 1052 1074
APP377 bps
PSEN608 bps
Genotyping
E
N
S
W
Morris Water Maze
Q2 Q1
Q3 Q4
Treatment NLearning trials
D1 latency (s) D2 latency (s) D3 latency (s) D4 latency (s) D5 latency (s)
Extract A 9 (9M) 76.4 ± 5.3 35.7 ± 9.5 31.7 ± 5.8 49.8 ± 7.8 37.0 ± 9.4
Extract B 9 (9F) 72.9 ± 6.1 48.2 ± 6.6 32.1 ± 4.7 41.8 ± 4.6 34.6 ± 4.5
APP 4 (1M, 3F) 80.8 ± 6.8 55.9 ± 7.7 53.9 ± 7.8 33.7 ± 4.4 59.5 ± 9.3
WT 10 (6M, 4F) 63.0 ± 6.9 41.1 ± 5.8 29.1 ± 5.4 21.2 ± 3.5 29.7 ± 5.0
Results of learning trials
Results of Probe trial
Treatment N
Probe trial: Quadrant 3
Cross number Duration (sec)
Extract A 9 (9M) 5.7 ± 1.4* 20.1 ± 3.7
Extract B 9 (9F) 4.1 ± 0.7 14.8 ± 4.0
APP 4 (1M, 3F) 2.0 ± 1.2 8.7 ± 5.1
WT 10 (6M, 4F) 5.1 ± 1.5 15.1 ± 5.4
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