P3032
Increased Adipose Tissue Mitochondrial Oxidation Protects Mice against Obesity
and Insulin resistance.
Vernochet C, Mourier A, Cinti S, Larson NG, Kahn CR.
Department of Integrative Physiology and Metabolism, Joslin Diabetes Center, Harvard Medical School
Max Planck Institute for Biology of Ageing, Robert-Koch-Str. 21, 50931 Cologne, Germany
Department Experimental and Clinical Medicine-Diagnostic Electron Microscopy Unit University-United Hospitals of Ancona, Italy
Results
Lox
**
*
*
* *
*
*
20
10
F-TfKO
250
500
13 18 23 28 33 38
400
*
300
*
Lox
*
100
0
30
60
90
7.0
6.5
6.0
5.5
5.0
4.5
4.0
3.5
3.0
*
F-TfKO
150
100
min
*
50
*
0
120
100
*
80
60
*
40
20
0
independently. (n=6/genotype).
*
8.5
8.0
VO2
7.0
F-TfKO
Night
Food intake (Left) and Oxygen consumption Right) were determined at 810 weeks of age for Lox and F-TFKO male mice using CLAMS metabolic
cages (103 ml/kg of lean mass).
C-II ( =)
C-I
C-III
H+
Adipose Tissue specific TFAM KO protects
mice from obesity, insulin resistance and
increases mice energy expenditure.
H+
C-IV
ATP synthase
H+
H+
Lox
0
0.5
0
AF WAT
BAT
5 targets (Sc, 1,2,3,4,5)
Coupled
Preadipocyte conf 10T1/2
shSc shTfam
3 independent
C3H10T1/2 cell lines
Uncoupled
FCCP
mtDNA quantification
12
10
eADP + Pi
H+
ATP
FADH2
Figure 4 : Complex I Deficiency/Complex II
Compensation
F-TfKO
Tfam
Rotenone
6
4
2
0
shSc
shTfam
Proper
adipose tissue
mitochondrial function
Substrates/Durgs Xs
Stress Mito Kit
WAT
VDAC
oligoA
8
Conclusion
Various
Isolated
Mitochondria
BAT
Western blot of Tfam in isolated mitochondria from BAT and isolated
white adipocytes (AF WAT) of Lox and f-TfKO mice at 12 weeks of age.
Disturbed
adipose tissue
mitochondrial function
Bioenergetics
Profile State 3
Bioenergetics
Profile State 3
CII
Complex I activity
CII
CI
B)
TCA
Malate/
Succinate/
Pyruvate
Rotenone
Beneficial
Metabolic Effect
Aging and High Fat Diet
TFAM KO
CI
Tfam-Flox (Lox)
0.5
1.0
H+
Matrix
BAT
F-TfKO
1.0
1.5
line.
e-
Figure 2 : TFAM KO efficiency in adipose tissues
Lox
1.5
UCP1
O2
Cyt c
+H+
AF WAT
2.0
Lentivirus shTfam
NADH
A
*
Figure 6 : Tfam KO in vitro on preadipocyte cell
Q
In the present study, we have
generated a unique mouse model to
study the impact of an adipose tissuespecific mitochondrial dysfunction on
mouse physiology by disrupting Tfam
selectively in the brown and white
adipose tissues using a Cre-loxmediated recombination.
2.5
2.0
Lox F-TfKO
Lox F-TfKO
TFAM
picture from http://lab.hirschey.org
3.0
*
Tfam KO increased oxygen consumption
through a higher basal uncoupling state
and Complex I deficiency.
6.5
6.0
*
2.5
Respiratory Control Ratio (RCR) from isolated BAT and white
adipocyte (AF WAT) from Lox and F-TFKO CD fed male mice
at 10-12 weeks of age in presence of complex II substrate
(n=9/genotype).
TFAM KO remodels adipose tissue
mitochondria OXPHOS function.
7.5
IM
13 subunits + 22tRNA + 2 rRNA
3.5
AF WAT
BAT
9.0
Day
ETC (Electron transport chain)
F-TFKO
F-TFKO and Lox BAT (B) and white adipocyte (AF WAT) isolated
mitochondria respiratory chain enzyme activities were assessed
Lox
Mitochondria DNA
120
150
Body weight measurements (Right Panel) and intraperitoneal glucose
tolerance testing (GTT Left Panel) of male Lox and F-TfKO mice on CD
(Chow Diet).
B
*
200
200
weeks
140
BAT
0
8
Lox
600
0
3
F-TFKO
Ratio mitochondrial /
genomic DNA
30
Lox
B
A
RCR GDP (State3/State4)
40
700
in adipose tissues.
RCR (State3/State4)
50
A
Enzyme activity (%)
60
GTT Glucose (mg/dl)
70
Food intake g/day
TFAM
A
Body weight (g)
Obesity and type 2 diabetes are
associated
with
mitochondrial
dysfunction in adipose tissue, but
whether this dysfunction participates
directly in the development of these
disorders remains an open question.
Mitochondrial transcription factor A
(TFAM) is necessary for mtDNA
stability by packaging the DNA into
nucleoid-like structures and also
initiates the mtDNA transcription that
is essential for mtDNA replication and
mitochondrial-encoded
gene
transcription.
Figure 5 : Tfam KO Increasing uncoupling state
Figure 3 : Tfam KO direct consequences on
adipose tissue metabolic rate.
Figure 1 : Adipose tissue specific Tfam KO
consequences on animal physiology.
Enzyme activity (%)
Introduction
PalmytoylCarnitin
FAT TFAM KO
Uncoupled respiration
Energy Expenditure
Flux
Adipose Tissue Mass
Insulin sensitivity
Control
mice
Adipose tissue
specific TFAM KO
mice
BAT F-TFKO
WAT F-TFKO
0.5µm
*(p<0.05) in all panels.
1µm
0.5µm
1µm
Mitochondria morphology of Lox and F-TFKO by electron microscopy
(EM) (picture representative of 5 mice /group)
25
125
62.5
20
100
50.0
15
75
10
5
0
BAT
AF WAT
75.0
*
50
25
0
BAT
AF WAT
FA OCR
pMoles/min/µg
aP2-Cre
F-TfKO
WAT Lox
150
CPLxII OCR
pMoles/min/µg
Lox
BAT Lox
CPLxI OCR
pMoles/min/µg
&
X
*
30
*
37.5
Hepatosteatosis
*
25.0
12.5
0
BAT
AF WAT
State 3 Respiratory rate of F-TFKO and Lox isolated mitochondria from BAT and
white adipocyte (AF WAT) in presence of malate/pyruvate (complex I) (A),
Succinate/Glycerol-3-phosphate/rotenone (complex II) (B), and Palmitoylcarnitine/malate (fatty acid) (C) and ADP from Lox and F-TFKO CD fed male
mice at 10-12 weeks of age (n=9/genotype).
Acknowledgements and References
Vernochet C Cell Metab. 2012 Nov 14. S1550-4131(12)00416-0.
10.1016/j.cmet.2012.10.016. (Epub ahead of print)