Download - PoTingHo-2014 IBC Summer Internship Program
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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
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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.
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3rd month
Genotyping Treatments Sacrifice
10th month
Treatments started
Experimental Timeline
My Internship
Period
Morris Water Maze
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4
11thGenotyping Treatments Sacrifice
Experimental Timeline
Morris Water MazeTreatments started
3rd month 10th month
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5 Chinese Herbal Extracts Treatment
Extract A
Extract B
APP
WT
• Dosage: 250 mg/kg a day by gavage
Mat
eria
l
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11thGenotyping Treatments Sacrifice
Experimental Timeline
Treatments started
3rd month
Morris Water Maze
10th month
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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
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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.
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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
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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.
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11thGenotyping Treatments Sacrifice
Experimental Timeline
3rd month
Treatments started
10th month
Morris Water Maze
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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
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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
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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
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Questions?
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Zhang, C. (2009) Harvard Medical School.
(APP Intracellular Domain)
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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
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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
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E
N
S
W
Morris Water Maze
Q2 Q1
Q3 Q4
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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
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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