Loss of myocardium

1
Loss of myocardium
is a serious consequence
of acute myocardial
infarction.
Loss of more than a third
of the left ventricle results
in CHF and high mortality.
Acute Myocardial
Infarction
2
1972 Eugene Braunwald proposes we search for an
infarct-reduction therapy Many agents were
evaluated with no effect Free radical
scavengers Beta Blockers Calcium
Antagonists Anti-inflammatory agents 1986 Keith
Reimer and Robert Jennings discover Ischemic
Preconditioning
3
ischemic preconditioning
40 min
50
40
30
Infarction of the Ischemic Zone
20
10
0
Control
Murry et al, Circulation 1986741124-1136
4
PC
30 ischemia
Current theory of IPC
5
PC
30 ischemia
6
Trigger Phase
Adenosine
7
ROS
PKC
? Yellon and Downey
8
The ROS step explains why receptor population
during ischemia without brief reperfusion does
not protect
protection
Index ischemia
Receptors populatedmKATP open
no ROS and no protection
Index ischemia
9
Trigger Phase
Adenosine
10
By using activators and blockers we were able to
make a detailed map of the signal transduction
events between the receptor and the production of
reactive oxygen species (ROS). The sequence is as
follows
11
ACh, Brady, Opioid
PI 4,5 P2
EGFR
Gi-??
p85
PI3K
Akt
PDK-1
?
Src
PDK-2
A metaloprotienase releases heparin binding EFG
eNOS
GC
PKG
Activate PKC and Protect
Mitochondrion
12
ACh, Brady, Opioid
PI 4,5 P2
EGFR
Gi-??
p85
PI3K
Akt
PDK-1
?
Src
PDK-2
eNOS
GC
The EGF receptor is populated, dimerized,
autophosphorylated, and PI3 kinase is activated
PKG
Activate PKC and Protect
Mitochondrion
13
ACh, Brady, Opioid
PI 4,5 P2
EGFR
Gi-??
p85
PI3K
Akt
PDK-1
?
Src
PDK-2
eNOS
3 phosphorylated lipid activates AKT via the PDKs
GC
PKG
Activate PKC and Protect
Mitochondrion
14
ACh, Brady, Opioid
PI 4,5 P2
EGFR
Gi-??
p85
PI3K
Akt
PDK-1
?
Src
PDK-2
eNOS
GC
PKG
Activate PKC and Protect
Mitochondrion
Akt activates PKG via eNOS and nitric oxide
15
ACh, Brady, Opioid
PI 4,5 P2
EGFR
Gi-??
p85
PI3K
Akt
PDK-1
?
Src
PDK-2
eNOS
GC
PKG opens potassium channels in the mitochondria
PKG
Activate PKC and Protect
Mitochondrion
16
ACh, Brady, Opioid
PI 4,5 P2
EGFR
Gi-??
p85
PI3K
Akt
PDK-1
?
Src
PDK-2
eNOS
Potassium entry causes free radical production
GC
PKG
Activate PKC and Protect
Mitochondrion
17
Ischemic phasePKC acts as a memory.
PC
30 ischemia
18
The mitochondrial Permeability Transition Pore
forms at reperfusion
VDAC
ANT
matrix
19

• At reperfusion there are 3 populations of cells
• Those that were killed by ischemia
• Those that had sub-lethal injury and will survive

C. Those that are alive but will die from mPTP
opening
20
What inhibits transition pore opening?
PI3 kinase and ERK
21
Inhibiting either PI3 kinase or Erk at
reperfusion Blocks IPC-induced Protection
60
55
50
45
40
35
30
of Ischemic zone infarcted
25
20
15
10
5
0
Control
Hausenloy et al Circulation 108IV62, 2003
22
What turns on PI3-kinase at reperfusion?
ADENOSINE!
23
Adenosine A2b receptor blockade at reperfusion
inhibits protection of IPC
50
40
30
of Ischemic zone infarcted
20

10
0
IPC SPT
IPC MRS1754
Control
IPC
Solenkova et al. Am J Physiol Heart Circ Physiol.
2005 (in press)
24
We used phosphorylation of Akt as a reporter of
PI3-Kinase activation
PI 4,5 P2
PI 3,4,5 P3
PI3K
Akt
PDK-1
S-308
PDK-2
S-473
25
Adenosine receptor blocker given at reperfusion
blocks PI3 kinase activation
1.4
BL
10
control
P-AKT
IPC
IPCSPT
relative density
Solenkova et al. Am J Physiol Heart Circ Physiol.
(in press)
26
What makes the preconditioned heart sensitive to
its own adenosine?
PKC!
27
Activation of PKC at reperfusion protects like
preconditioning
60
PKCActivator
40
of ischemic zone Infarcted
20
0
Control
Philipp et al. Cardiovasc Res in press
28
2.5 nM NECA, a potent A2b agonist, does not cause
phosphorylation of AKT or ERK in isolated hearts.
After treatment with PMA or ischemic
preconditioning 2.5 nM NECA causes robust
phosphorylation of both AKT and ERK. Data is
normalized to the value after treatment but prior
to NECA.
29
(No Transcript)
30
Preconditioning has not been useful in the
patient with acute myocardial infarction because
it must be instituted prior to ischemia.
reperfuse
precondition
See patient
31
In situ dog hearts
30
25
Control
60 min I
3 h R
20
15
Infarction ( of risk zone)
10
5
0
Control
32
Postconditioning depends on ERK, PI3-K, and
critical timing
4 cycles 30 occl
reperfusion
3 h
4 cycles
4 cycles 1delayed
OPEN-CHEST RABBITS
60
p
p
50
40
Infarction ( of risk zone)
30
ERK dependent
20
PI3-K dependent
Timing is critical
10
0
Control


Yang et al., J Am Coll Cardiol. 441103-10, 2004
33
Postconditioning also uses the same steps as seen
in preconditionings trigger pathway
mKATP
34
MTP opens
pH
ROSPKC
(memory)
PKC
pH
We propose that postconditioning protects because
it maintains acidosis during early reoxygenation
which inhibits transition pore formation. That
gives the heart enough time to precondition
itself.
ROS PKC
(Adenosine effect)
pH
(Low pH effect)
35
MTP opens
pH
ROSPKC
(memory)
PKC
pH
ROS PKC
(Adenosine effect)
pH
(Low pH effect)
36
Postconditioning depends on the A2b receptor
Open-chest rabbits
60
40
Infarction ( of risk zone)
20


0
Control
4 cycles
4 cycles
4 cycles
SPT
DPCPX
4 cycles
MRS



SPT
DPCPX
MRS
37
MTP opens
pH
ROSPKC
(memory)
PKC
pH
ROS PKC
(Adenosine effect)
pH
(Low pH effect)
38
Like preconditioning, postconditioning depends
on ROS signaling
80
In Situ rabbit hearts
70
p
60
50
40
of risk zone infarcted
30

20
10
0
4 cycles
Control
4 cycles
MPG

MPG
39
Brief Acidosis in Early Reperfusion
BAER
40
MTP opens
pH
ROSPKC
(memory)
PKC
pH
ROS PKC
(Adenosine effect)
pH
(Low pH effect)
41
pH of the Krebs buffer was lowered by gassing it
with 20 CO2 . Hearts were reperfused for 1 or
2 with Low pH buffer
42
BAER for only 2 min causes salvage equivalent to
that for postconditioning.
43
Like postconditioning, BAER is ROS dependent.
60

p


p
50
40
Infarction ( of risk zone)
30

20

10
0
6 cycles
Control
BAER
BAER
(10")
2'
MPG
44
How can we lower the pH in a patients heart?
The sarcolemma of a cardiomyocyte is impermeable
to H so inorganic acids will be of no help. CO2
diffuses into the heart across the sarcolemma and
then dissociates to carbonic acid.
45
Intracellular pH of the reperfused heart could be
lowered by infusing a short chain carboxylic acid
through the catheter.
46
pH was lowered in rat cardiomyocytes by
superfusing them with buffer gassed with 30 CO2
(solid symbols) or buffer containing 20mM sodium
proprionate (open symbols).
Nomura et al Am J Physiol 283H193 2002
47
Short-chain carboxylic acids may promote
transition pore formation. This shows rat liver
mitochondria in 5 mM succinate and 30 uM calcium.
PPA is propionic acid. Thus propionic acid may
protect or may exacerbate necrosis??
Palmeira et al. Biochem Biophysical Res Com
272431, 2000
48
We also tried having the rabbits breath 20
CO2/80 oxygen for 10 min. The pH of the blood
fell quickly to 7.1 (almost identical to that in
the buffer for the isolated hearts). Other that
strong respiratory movements the CO2 was well
tolerated. Blood pressure and arterial oxygen
content was maintained. Respiration rapidly
returned to normal when they were switched back
to 100 oxygen.