ROLE OF SIRT3 AS A DISEASE-MODIFYING AGENT
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Transcript of ROLE OF SIRT3 AS A DISEASE-MODIFYING AGENT
ROLE OF SIRT3 AS A DISEASE-MODIFYING AGENT IN PARKINSON’S DISEASE
BIOD98H3Supervisor: Dr. Joanne NashMuhammad Hassan Khan
Introduction- Parkinson’s Disease• Second leading neurodegenerative disorder in the world• Motor and non-motor symptoms • Selective and irreversible loss of DA neurons and Lewy
Body formation intracytoplasmic inclusions containing abnormal aggregates of α-synuclein and ubiquitin
• Genes causing PD encoded mt proteins Mitochondrial dysfunction likely occurs early in PD pathogenesis
• No known cure
Introduction- SIRT3• Sirtuins are a seven-
membered family of NAD+-dependent deacetylases
• SIRT3 is best understood mitochondrial Deacetylase
• Mitochondria are the main source of ROS and energy production in neurons SIRT3 can play a potentially neuroprotective role by improving mitochondrial function resulting in increased survival of neurons
Introduction- α-synuclein rats• Two most widely used approaches:
1. Toxins e.g. MPTP, 6-OHDA2. Genetic e.g. α-synuclein, PINK-1
• Toxin models cause rapid decrease in DA neurons only replicate later stages of PD fail to develop LB and behavioural abnormalities
• Genetic unable to report a significant loss of DA neurons BUT
• Overexpression of human WT or A53T mutant α-synuclein progressive loss of DA neurons, behavioural deficit and α-synuclein cytoplasmic inclusions
Hypothesis• Over-expression of SIRT3 prior to injection of human
A53T α-synuclein will improve the general health of mitochondria and slow or prevent neurodegeneration in the SNc.
Methods
SIRT3/empty vector was infused into the animals on day -7 followed by α-synuclein/empty vector injection on day 0. Three weeks (21 days) after the infusion of α-synuclein behavioural assessment was performed. Brains were removed, tissue sectioned, and stereology performed. The aforementioned steps were again repeated at 6 weeks
Methods• Animal Surgery and Vector Delivery• Behavioural Analysis• Perfusion/Cryostatting/Imunohistochemistry• Antibodies• Stereology Analysis:
estimate the number of dopamine neurons in SNc. TH-postive cells were stained and counted using stereo-investigator.
• HPLC Analysis• Statistical Analysis
Results• SIRT3 was able to reverse behavioural impairment 6
weeks following administration in AAV1/2 A53T α-synuclein rats:
• A B
3 Week Behaviour
EV/EV EV/A53T SIRT3/A53T0
10
20
30
% a
sym
met
ry
6 Week Behaviour
EV/EV EV/A53T SIRT3/A53T0
10
20
30
40
50***
*
% a
sym
met
ry
Results• Effect of SIRT3 over-expression on striatal dopamine
turnover in AAV1/2 A53T α-synuclein rats:
• A B
3 week DA
EV/EV A53T/EV A53T/SIRT30
25
50
75
100
125
150
DA
as
a fu
nct
ion
of
un
lesi
on
ed s
ide
6 week DA
EV/EV A53T/EV A53T/SIRT30
25
50
75
100
125
150
DA
as
a fu
nct
ion
of
un
lesi
on
ed s
ide
Results• Effect of SIRT3 over-expression on striatal dopamine
turnover in AAV1/2 A53T α-synuclein rats:
• C D
3 Week HVA:DA
EV/EV EV/A53T SIRT3/A53T0
25
50
75
100
125
150 *
HV
A:D
A a
s a
fun
ctio
n o
fu
nle
sio
ned
sid
e
6 Week HVA:DA
EV/EV EV/A53T SIRT3/A53T0
25
50
75
100
125
150
175
200
**
HV
A:D
A a
s a
fun
ctio
n o
fu
nle
sio
ned
sid
e
Results• Effect of SIRT3 over-expression on striatal dopamine
turnover in AAV1/2 A53T α-synuclein rats:
• E F
3 Week DOPAC:DA
EV/EV EV/A53T SIRT3/A53T0
25
50
75
100
125
150 **
DO
PA
C:D
Aas
a f
un
ctio
n o
fu
nle
sio
ned
sid
e
6 Week DOPAC:DA
EV/EV EV/A53T SIRT3/A53T0
25
50
75
100
125
150
175
200
****
DO
PA
C:D
Aas
a f
un
ctio
n o
fu
nle
sio
ned
sid
e
Results• Effect of SIRT3 overexpression on dopamine cell number
in the substantia nigra pars compacta:
• A B
3 week Stereology
EV/EV EV/A53T SIRT3/A53T0
20
40
60
80
DA
cel
ls a
s %
of
nai
ve c
ells
6 week Stereology
EV/EV EV/A53T SIRT3/A53T0
20
40
60
80D
A c
ells
as
% o
f n
aive
cel
ls
Discussion• Liu et al (2015) found that SIRT3 plays a protective role
in MPTP mouse model of PD. • Wier et al (2012) found that SIRT3 mRNA levels were
upregulated with increased deposition of amyloid- β and ROS production in an AD model
• Possible protective pathways exploited by SIRT3 include:
aiding DA secretion, protective effect against ROS production, regulation of basal ATP levels, suppression of mtPTP formation, protection against fragmentation of the mitochondria and prevention against age-linked apoptosis
Discussion• Our data suggests that over-expression of SIRT3 prior to
infusion of human A53T α-synuclein is able to play a protective role in slowing or preventing neurodegeneration in the SNc.
• Additional studies investigating the effects of overexpression of SIRT3 need to be performed in other genetic and toxin models of PD.
• Further studies looking at the mechanism of action of SIRT3 in specific neurodegenerative diseases also need to be looked at.