PGC-1α as therapeutic target to improve the mitochondrial...
15
PGC-1α as therapeutic target to improve the mitochondrial phenotype in chromosome 21 trisomic cells Nunzia Mollo- DMMBM Napoli
Transcript of PGC-1α as therapeutic target to improve the mitochondrial...
PGC-1α as therapeutic target to improve the mitochondrial phenotype in chromosome 21 trisomic cells
Nunzia Mollo- DMMBM Napoli
NRIP1
Hsa21 gene NRIP1 modulates PGC-1a activity
Scarpulla , 2011
* p<0.05; ** p<0.001 Izzo et al., 2014
NRIP1 inhibits PGC-1a expression in trisomic cells
NRIP1
PGC-1α
PPARγ
Can PGC-1α induction reverse the mitochondrial alterations in trisomic cells?
Metformin and pioglitazone induce the PGC-1α signaling pathway in trisomic cells
* p<0.05; ** p<0.01
0,5 M PG 2 M PG0.0
0.5
1.0
1.5
2.0
2.5
Rela
tive P
GC
-1
mR
NA
exp
ressio
n
0,5 M PG 2 M PG0
1
2
3
4
Rela
tive P
PA
R m
RN
A e
xp
ressio
n
*
*
*
*
0,25 M PG 0,5 M PG 1 M PG 2 M PG0
1
2
3
Rela
tive P
GC
-1
pro
tein
exp
ressio
n
B
C D
0,5 M 2 M0.0
0.5
1.0
1.5
2.0
2.5
PG
C-1
mR
NA
exp
ressio
n
0,5 M 2 M0
1
2
3
4
PP
AR m
RN
A e
xp
ressio
n
*
**
*
0,25 M 0,5 M 1 M 2 M0
1
2
3
PG
C-1
pro
tein
exp
ressio
n
A
B C
PG (72h) 0,25 μM 1 μMCTR 0,5 μM 2 μM
PGC1A
ACTIN
A
0,5 M PG 2 M PG0.0
0.5
1.0
1.5
2.0
2.5
Rela
tive P
GC
-1
mR
NA
exp
ressio
n
0,5 M PG 2 M PG0
1
2
3
4
Rela
tive P
PA
R m
RN
A e
xp
ressio
n
*
*
*
*
0,25 M PG 0,5 M PG 1 M PG 2 M PG0
1
2
3
Rela
tive P
GC
-1
pro
tein
exp
ressio
n
B
C D
0,5 M 2 M0.0
0.5
1.0
1.5
2.0
2.5
PG
C-1
mR
NA
exp
ressio
n
0,5 M 2 M0
1
2
3
4
PP
AR m
RN
A e
xp
ressio
n
*
**
*
0,25 M 0,5 M 1 M 2 M0
1
2
3
PG
C-1
pro
tein
exp
ressio
n
A
B C
PG (72h) 0,25 μM 1 μMCTR 0,5 μM 2 μM
PGC1A
ACTIN
A
0,5 M PG 2 M PG0.0
0.5
1.0
1.5
2.0
2.5
Rel
ativ
e P
GC
-1
mR
NA
exp
ress
ion
0,5 M PG 2 M PG0
1
2
3
4
Rel
ativ
e P
PA
R
mR
NA
exp
ress
ion
*
*
*
*
0,25 M PG 0,5 M PG 1 M PG 2 M PG0
1
2
3
Rel
ativ
e P
GC
-1
pro
tein
exp
ress
ion
B
C D
0,5 M 2 M0.0
0.5
1.0
1.5
2.0
2.5
PG
C-1
mR
NA
exp
ress
ion
0,5 M 2 M0
1
2
3
4
PP
AR
mR
NA
exp
ress
ion
*
**
*
0,25 M 0,5 M 1 M 2 M0
1
2
3
PG
C-1
pro
tein
exp
ress
ion
A
B C
PG (72h) 0,25 μM 1 μMCTR 0,5 μM 2 μM
PGC1A
ACTIN
A
0,5 M PG 2 M PG0.0
0.5
1.0
1.5
2.0
2.5
Rel
ativ
e P
GC
-1
mR
NA
exp
ress
ion
0,5 M PG 2 M PG0
1
2
3
4
Rel
ativ
e P
PA
R
mR
NA
exp
ress
ion
*
*
*
*
0,25 M PG 0,5 M PG 1 M PG 2 M PG0
1
2
3
Rel
ativ
e P
GC
-1
pro
tein
exp
ress
ion
B
C D
0,5 M 2 M0.0
0.5
1.0
1.5
2.0
2.5
PG
C-1
mR
NA
exp
ress
ion
0,5 M 2 M0
1
2
3
4
PP
AR
mR
NA
exp
ress
ion
*
**
*
0,25 M 0,5 M 1 M 2 M0
1
2
3
PG
C-1
pro
tein
exp
ress
ion
A
B C
PG (72h) 0,25 μM 1 μMCTR 0,5 μM 2 μM
PGC1A
ACTIN
A
0,5 M PG 2 M PG0.0
0.5
1.0
1.5
2.0
2.5
Rel
ativ
e P
GC
-1
mR
NA
exp
ress
ion
0,5 M PG 2 M PG0
1
2
3
4
Rel
ativ
e P
PA
R
mR
NA
exp
ress
ion
*
*
*
*
0,25 M PG 0,5 M PG 1 M PG 2 M PG0
1
2
3
Rel
ativ
e P
GC
-1
pro
tein
exp
ress
ion
B
C D
0,5 M 2 M0.0
0.5
1.0
1.5
2.0
2.5
PG
C-1
mR
NA
exp
ress
ion
0,5 M 2 M0
1
2
3
4
PP
AR
mR
NA
exp
ress
ion
*
**
*
0,25 M 0,5 M 1 M 2 M0
1
2
3
PG
C-1
pro
tein
exp
ress
ion
A
B C
PG (72h) 0,25 μM 1 μMCTR 0,5 μM 2 μM
PGC1A
ACTIN
A
0,05 mM MTF 0,5 mM MTF0.0
0.5
1.0
1.5
2.0
Rela
tiveT
FA
M m
RN
A e
xp
ressio
n
0,05 mM MTF 0,5 mM MTF0.0
0.5
1.0
1.5
2.0
Rela
tive N
RF
-1 m
RN
A e
xp
ressio
n
0,05 mM MTF 0,5 mM MTF0.0
0.5
1.0
1.5
2.0
Rela
tive P
GC
1-
mR
NA
exp
ressio
n
***
0,05 mM MTF 0,5 mM MTF0.0
0.5
1.0
1.5
2.0
2.5
Rela
tive P
GC
1-
pro
tein
exp
ressio
n
*
**
*
0,05 mM MTF 0,5 mM MTF0.0
0.5
1.0
1.5
2.0
Rela
tive D
-LO
OP
/AC
TIN
exp
ressio
n
* *
**
A
B
D EF
*
CTR 0,05mM 0,5mM
PGC-1α
GAPDH
- 75
- 37
MTF
CPGC-1α expression
0,5 M PG 2 M PG0.0
0.5
1.0
1.5
2.0
2.5
Rela
tive P
GC
-1
mR
NA
exp
ressio
n
0,5 M PG 2 M PG0
1
2
3
4
Rela
tive P
PA
R m
RN
A e
xp
ressio
n
*
*
*
*
0,25 M PG 0,5 M PG 1 M PG 2 M PG0
1
2
3
Rela
tive P
GC
-1
pro
tein
exp
ressio
n
B
C D
0,5 M 2 M0.0
0.5
1.0
1.5
2.0
2.5
PG
C-1
mR
NA
exp
ressio
n
0,5 M 2 M0
1
2
3
4
PP
AR m
RN
A e
xp
ressio
n
*
**
*
0,25 M 0,5 M 1 M 2 M0
1
2
3
PG
C-1
pro
tein
exp
ressio
n
A
B C
PG (72h) 0,25 μM 1 μMCTR 0,5 μM 2 μM
PGC1A
ACTIN
A
0,05 mM MTF 0,5 mM MTF0.0
0.5
1.0
1.5
2.0
Rela
tiveT
FA
M m
RN
A e
xp
ressio
n
0,05 mM MTF 0,5 mM MTF0.0
0.5
1.0
1.5
2.0
Rela
tive N
RF
-1 m
RN
A e
xp
ressio
n
0,05 mM MTF 0,5 mM MTF0.0
0.5
1.0
1.5
2.0
Rela
tive P
GC
1-
mR
NA
exp
ressio
n
***
0,05 mM MTF 0,5 mM MTF0.0
0.5
1.0
1.5
2.0
2.5
Rela
tive P
GC
1-
pro
tein
exp
ressio
n
*
**
*
0,05 mM MTF 0,5 mM MTF0.0
0.5
1.0
1.5
2.0
Rela
tive D
-LO
OP
/AC
TIN
exp
ressio
n
* *
**
A
B
D EF
*
CTR 0,05mM 0,5mM
PGC-1α
GAPDH
- 75
- 37
MTF
C
PG
D
E
F
Izzo et al., 2017
PGC-1α induction improves the bioenergetic profile of trisomic cells
0 10 20 30 40 50 60 70 80 90 1001101200
20
40
60
80
100
120
140CTR
0,5 M PG
2 M PG
Time (minutes)
OC
R (
pM
ole
s/m
in)
Basal ATP-linked Maximal 0
50
100
150
200
2500,5 M PG
2 M PGO
CR
(%
Co
ntr
ol)
Oligomycin FCCP Ant/Rot
0 10 20 30 40 50 60 70 80 90 1001101200
20
40
60
80
100
120
140CTR
0,05 mM MTF
0,5 mM MTF
Time (minutes)
OC
R (
pM
ole
s/m
in)
Olygomycin FCCP Ant/Rot
A
Basal ATP-linked Maximal0
50
100
150
200
2500,05 mM MTF
0,5 mM MTF
OC
R (
% C
on
tro
l)
*
***
*****
C
****
*
D
B
* p<0.05; ** p<0.01
Oxygen production
Izzo et al., 2017
* *
0.5 M PG 2 M PG0.0
0.5
1.0
1.5
2.0
2.5
Rela
tive lu
min
escen
ce in
ten
sit
y (
a.
u.)
0.05 mM MTF 0.5 mM MTF0.0
0.5
1.0
1.5
2.0
Rela
tive lu
min
escen
ce in
ten
sit
y (
a.
u.)
****
A B
Metformin and pioglitazone increase ATP production and reduce ROS production in trisomic cells
ATP production
* p<0.05; ** p<0.01
0.5 mM MTF0
20
40
60
80
100
120
H2D
CF
-DA
Flu
ore
scen
ce i
nte
nsit
y
(% C
on
tro
l)
0.5 M PG 2 M PG0
20
40
60
80
100
120
H2D
CF
-DA
Flu
ore
scen
ce i
nte
nsit
y
(% C
on
tro
l)
A B
ROS production
0,5 M PG 2 M PG0.0
0.5
1.0
1.5
2.0
2.5
Rela
tive P
GC
-1
mR
NA
exp
ressio
n
0,5 M PG 2 M PG0
1
2
3
4
Rela
tive P
PA
R m
RN
A e
xp
ressio
n
*
*
*
*
0,25 M PG 0,5 M PG 1 M PG 2 M PG0
1
2
3
Rela
tive P
GC
-1
pro
tein
exp
ressio
n
B
C D
0,5 M 2 M0.0
0.5
1.0
1.5
2.0
2.5
PG
C-1
mR
NA
exp
ressio
n
0,5 M 2 M0
1
2
3
4
PP
AR m
RN
A e
xp
ressio
n
*
**
*
0,25 M 0,5 M 1 M 2 M0
1
2
3
PG
C-1
pro
tein
exp
ressio
n
A
B C
PG (72h) 0,25 μM 1 μMCTR 0,5 μM 2 μM
PGC1A
ACTIN
A
0,5 M PG 2 M PG0.0
0.5
1.0
1.5
2.0
2.5
Rela
tive P
GC
-1
mR
NA
exp
ressio
n
0,5 M PG 2 M PG0
1
2
3
4
Rela
tive P
PA
R m
RN
A e
xp
ressio
n
*
*
*
*
0,25 M PG 0,5 M PG 1 M PG 2 M PG0
1
2
3
Rela
tive P
GC
-1
pro
tein
exp
ressio
n
B
C D
0,5 M 2 M0.0
0.5
1.0
1.5
2.0
2.5
PG
C-1
mR
NA
exp
ressio
n
0,5 M 2 M0
1
2
3
4
PP
AR m
RN
A e
xp
ressio
n
*
**
*
0,25 M 0,5 M 1 M 2 M0
1
2
3
PG
C-1
pro
tein
exp
ressio
n
A
B C
PG (72h) 0,25 μM 1 μMCTR 0,5 μM 2 μM
PGC1A
ACTIN
A
0,5 M PG 2 M PG0.0
0.5
1.0
1.5
2.0
2.5
Rela
tive P
GC
-1
mR
NA
exp
ressio
n
0,5 M PG 2 M PG0
1
2
3
4
Rela
tive P
PA
R m
RN
A e
xp
ressio
n
*
*
*
*
0,25 M PG 0,5 M PG 1 M PG 2 M PG0
1
2
3
Rela
tive P
GC
-1
pro
tein
exp
ressio
n
B
C D
0,5 M 2 M0.0
0.5
1.0
1.5
2.0
2.5
PG
C-1
mR
NA
exp
ressio
n
0,5 M 2 M0
1
2
3
4
PP
AR m
RN
A e
xp
ressio
n
*
**
*
0,25 M 0,5 M 1 M 2 M0
1
2
3
PG
C-1
pro
tein
exp
ressio
n
A
B C
PG (72h) 0,25 μM 1 μMCTR 0,5 μM 2 μM
PGC1A
ACTIN
A
Izzo et al., 2017
Meformin and pioglitazone restore mitochondrial network organization in trisomic cells
DS-HFFs CTR
mtGFP
Metformin
Pioglitazone
0,05 mM 0,5 mM
0,5 µM 2 µM
N-HFFs
Izzo et al., 2017
mRNA expression Protein expression
Metformin and pioglitazone modulate mitochondrial fission-fusion machinery genes
* p<0.05; ** p<0.01
OPA1 MFN1 MFN2 DRP10.0
0.5
1.0
1.5
2.00.5 M PG
2 M PG
Rela
tive m
RN
A e
xp
ressio
n
OPA1 MFN1 MFN20
1
2
3
4
50.05 mM MTF
0.5 mM MTF
Rela
tive p
rote
in e
xp
ressio
n
OPA1 MFN1 MFN20.0
0.5
1.0
1.5
2.00.05 mM MTF
0.5 mM MTF
Rela
tive m
RN
A e
xp
ressio
n
OPA1 MFN1 MFN20
1
2
3
4
50.5 M PG
2 M PG
Rela
tive p
rote
in e
xp
ressio
n
B C
D
A
E F
OPA1
GAPDH
MFN1
GAPDH
MFN2
GAPDH
OPA1
GAPDH
MFN1
GAPDH
MFN2
GAPDH
**
**
* **
*
*
* * *
*
* ** *
Izzo et al., 2017
Mitostasis : Maintaining Mitochondria in an Extended Cellular Architecture
N1
DS1
MT-REDLC3HOECHST
N1 DS1
Trisomic cells show enlarged autophagosomes
N1 N2 N3 DS1 DS2 DS3 DS4
LC3 ILC3 II
GAPDH0
0,2
0,4
0,6
0,8
1
N DS
LC3II/LC3I protein expression
Take-home messages
Mitochondrial function and network are severely compromised in trisomy 21cells
Targeting PGC-1α improves the mitochondrial phenotype and modulates thefission/fusion machinery
Preliminary data suggest alterations of the mitochondrial clearance in trisomy21 cells
THANKS TO
DMMBMProf. Lucio NitschDott. ssa Anna ContiAntonella IzzoProf.ssa Simona PaladinoMaria CharalambousLucrezia ZerilloMaria Nitti
IEOS-CNRGaetano CalìFlaviana GentileGiuseppe MatareseClaudio ProcacciniDeriggio Faicchia
UNIVERSITY OF FERRARAPaolo PintonSimone Patergnani
Thanks for your attention. I’ll be glad to answer any questions.
« Troveremo. È impossibile non trovare.È uno sforzo intellettuale molto meno difficile che mandare un uomo sulla luna» (JerômeLejeune)Response to a reporter's question on the possibilityof finding a cure for Down syndrome .
0
50
100
150
200
250
CTR PG 500 uM PG 2 uM PG
N° mitocondri
Massa totale
Volume singolo
0
50
100
150
200
250
CTR MTF 0.05 mM MTF 0.5 mM MTF
N° mitocondri
Massa totale
Volume singolo
Morphometric analysis
