Download Acute rosiglitazone treatment is cardioprotective against - AJP

Am J Physiol Heart Circ Physiol 301: H895–H902, 2011.
First published June 10, 2011; doi:10.1152/ajpheart.00137.2011.
Acute rosiglitazone treatment is cardioprotective against
ischemia-reperfusion injury by modulating AMPK, Akt, and JNK
signaling in nondiabetic mice
Alex Morrison, Xiaoyan Yan, Chao Tong, and Ji Li
Department of Pharmacology and Toxicology, School of Medicine and Biomedical Sciences, University at Buffalo-State
University of New York, Buffalo, New York
Submitted 8 February 2011; accepted in final form 6 June 2011
thiazolidinedione; peroxisome proliferator-activated receptor-␥; myocardial infarction
remains the number one leading
cause of death for patients with and without diabetes (7).
Current interventions rely on the rapid recanalization of an
occluded coronary artery with percutaneous intervention,
thrombolytics, or anticoagulants. However, these treatments
are accompanied by an unfavorable consequence known as
reperfusion injury. As this is recognized as a major clinical
problem, potential therapies pertaining to cardiac metabolism, inflammation, oxidative stress, and apoptosis have yet
ISCHEMIC HEART DISEASE
Address for reprint requests and other correspondence: J. Li, Dept. of
Pharmacology and Toxicology, School of Medicine and Biomedical Sciences,
Univ. at Buffalo-State Univ. of New York, Buffalo, NY 14214 (e-mail:
[email protected]).
http://www.ajpheart.org
to be implemented (43). Therefore, there is an increasing
need for therapeutic strategies that limit myocardial ischemia-reperfusion (I/R) injury (37).
Rosiglitazone (RGZ), a peroxisome proliferator-activated
receptor (PPAR)-␥ agonist indicated for the treatment of
type 2 diabetes, has been shown to possess cardioprotective
properties in vivo during I/R (1, 23, 36) and in patients after
percutaneous coronary intervention (40). However, the use
of this drug remains controversial as recent meta-analyses
and clinical trials have indicated that long-term use of RGZ
in type 2 diabetic patients is associated with an increased
risk of heart failure and myocardial infarction (54).
Despite the negative effects that have been associated
with RGZ, recent evidence has suggested this thiazolidinedione (TZD) can modulate targets aimed at mitigating I/R
injury. Previous studies have shown that RGZ and other
TZDs can activate AMP-activated protein kinase (AMPK)
(4, 9, 18, 48), a master metabolic regulator that has recently
been demonstrated to be a cardioprotective stress-activated
protein kinase during I/R (22, 24, 29, 32). In addition, RGZ
has been shown to activate Akt, a prosurvival and antiapoptotic protein, during I/R in the heart and in cardiomyocytes
(16, 49, 51). Moreover, RGZ and PPAR-␥ activation have
been demonstrated to possess anti-inflammatory properties
in patients (25), cardiomyocytes (34), and animal models of
I/R injury by decreasing monocyte chemoattractant protein-1 and TNF-␣ levels, NF-␬B activity, and leukocyte
invasion (14, 50).
Interestingly, although the negative effects of RGZ have
been seen with chronic use in type 2 diabetic patients, there
is evidence that RGZ has beneficial cardiovascular effects in
nondiabetic patients (3, 30, 42). Such studies have reported
that RGZ increases plasma adiponectin concentrations, an
adipocytokine that has been shown to have insulin-sensitizing effects as well as protective effects against myocardial
I/R injury (32, 36, 44). It has also been demonstrated that
RGZ treatment possesses cardioprotective effects in nondiabetic animal models of I/R (23, 36, 50). Intriguingly, other
studies (2, 50) have reported that RGZ appears to have
cardioprotective effects when administered acutely to nondiabetic animals. Furthermore, acute RGZ treatment of
nondiabetic patients can result in rapid vasodilation, indicating the potential significance of its acute effects (39).
In this study, we aimed to determine whether acute RGZ
treatment is cardioprotective in nondiabetic mice against I/R
injury and to investigate whether the mechanism is via a
PPAR-␥-dependent process. We attempt to provide evi-
0363-6135/11 Copyright © 2011 the American Physiological Society
H895
Downloaded from http://ajpheart.physiology.org/ by 10.220.33.2 on May 6, 2017
Morrison A, Yan X, Tong C, Li J. Acute rosiglitazone treatment
is cardioprotective against ischemia-reperfusion injury by modulating
AMPK, Akt, and JNK signaling in nondiabetic mice. Am J Physiol
Heart Circ Physiol 301: H895–H902, 2011. First published June 10,
2011; doi:10.1152/ajpheart.00137.2011.—Rosiglitazone (RGZ), a
peroxisome proliferator-activated receptor (PPAR)-␥ agonist, has
been demonstrated to possess cardioprotective properties during ischemia-reperfusion. However, this notion remains controversial as recent
evidence has suggested an increased risk in cardiac events associated
with long-term use of RGZ in patients with type 2 diabetes. In this
study, we tested the hypothesis that acute RGZ treatment is beneficial
during I/R by modulating cardioprotective signaling pathways in a
nondiabetic mouse model. RGZ (1 ␮g/g) was injected intravenously
via the tail vein 5 min before reperfusion. Myocardial infarction was
significantly reduced in mice treated with RGZ compared with vehicle
controls (8.7% ⫾ 1.1% vs. 20.2% ⫾ 2.5%, P ⬍ 0.05). Moreover,
isolated hearts were subjected to 20 min of global, no-flow ischemia
in an ex vivo heart perfusion system. Postischemic recovery was
significantly improved with RGZ treatment administered at the onset
of reperfusion compared with vehicle (P ⬍ 0.001). Immunoblot
analysis data revealed that the levels of both phospho-AMP-activated
protein kinase (Thr172) and phospho-Akt (Ser473) were significantly
upregulated when RGZ was administered 5 min before reperfusion
compared with vehicle. On the other hand, inflammatory signaling
[phospho-JNK (Thr183/Tyr185)] was significantly downregulated as a
result of RGZ treatment compared with vehicle (P ⬍ 0.05). Intriguingly, pretreatment with the selective PPAR-␥ inhibitor GW-9662 (1
␮g/g iv) 10 min before reperfusion significantly attenuated these
beneficial effects of RGZ on the ischemic heart. Taken together, acute
treatment with RGZ can reduce ischemic injury in a nondiabetic
mouse heart via modulation of AMP-activated protein kinase, Akt,
and JNK signaling pathways, which is dependent on PPAR-␥ activation.
H896
CARDIOPROTECTIVE EFFECTS OF ACUTE ROSIGLITAZONE TREATMENT
dence that RGZ can be used as a cardioprotective drug for
acute ischemic events in patients without diabetes.
MATERIALS AND METHODS
Fig. 1. Rosiglitazone (RGZ) reduces myocardial infarct size after ischemia-reperfusion (I/
R). In vivo hearts were subjected to 20 min of
ischemia followed by 4 h reperfusion. RGZ (1
␮g/g) or vehicle was administered via the tail
vein 5 min before reperfusion. The extent of
myocardial necrosis was assessed as described
in MATERIALS AND METHODS. A: representative
sections of the extent of myocardial infarction.
B: ratio of the area at risk (AAR) to the total
myocardial area (left) and ratio of the infarcted
area to the AAR (right). Values are means ⫾
SE for 5 independent experiments. *P ⬍ 0.05
vs. vehicle.
AJP-Heart Circ Physiol • VOL
301 • SEPTEMBER 2011 •
www.ajpheart.org
Downloaded from http://ajpheart.physiology.org/ by 10.220.33.2 on May 6, 2017
Experimental animals. Male FVB/NJ mice (4 – 6 mo of age) were
used in all experiments. All animal procedures carried out in this study
were approved by the Institutional Animal Care and Use Committee
of University at Buffalo-State University of New York.
Experimental myocardial infarction. Mice were anesthetized with
pentobarbital sodium (100 mg/kg ip) and placed on a ventilator
(Harvard Rodent Ventilator, Harvard Apparatus, Holliston, MA), and
core temperature was maintained at 37°C with a heating pad. After a
left lateral thoracotomy, the left anterior descending coronary artery
was occluded for 20 min with an 8-0 nylon suture and polyethylene
tubing to prevent arterial injury and then reperfused for 4 h. The ECG
(AD Instruments, Colorado Springs, CO) showing ST segment elevation and blanching of the left ventricle (LV) confirmed myocardial
ischemia as a result of coronary occlusion. Vehicle (1:3 DMSO-saline)
or RGZ [1 ␮g/g (50), Enzo Life Sciences] was injected via tail vein 5 min
before reperfusion. Compound C [10 ␮g/g (20), Enzo Life Sciences] was
injected intraperitoneally 30 min before coronary occlusion. Hearts were
then excised, perfusion fixed, and stained to delineate the extent of
myocardial necrosis as a percentage of the nonperfused ischemic area at
risk (AAR) (22). Viable tissue in the ischemic region was stained red by
2,3,5-triphenyltetrazolium, and the nonischemic region was stained blue
with Evans blue dye (22). Hearts were fixed, sectioned, photographed
with a Leica microscope, and analyzed with National Institutes of Health
ImageJ software.
Mouse heart perfusion and measurement of cardiac function. Mice
were deeply anesthetized with pentobarbital sodium (100 mg/kg ip),
and hearts were excised and placed in the Langendorff perfusion
mode with Krebs-Henseleit buffer containing 7 mmol/l glucose, 0.4
mmol/l oleate, 1% BSA, and 10 ␮U/ml insulin. Hearts were perfused
at a flow of 4 ml/min, and a fluid-inflated balloon connected to the
LabChart5 system (AD Instruments) was inflated to achieve an
end-diastolic pressure of 5 mmHg, which was kept constant during the
baseline measurement of cardiac function. Global, “no-flow” ischemia
was induced by the termination of flow to the heart for 20 min, and
reperfusion was reinstated for 30 min at 4 ml/min. RGZ (5 ␮M)
treatment or vehicle (DMSO) was administered at the onset of
reperfusion. Cardiac function was measured as the heart rate-LV
developed pressure product (in mmHg·beats·min⫺1) (17, 24).
In vivo regional ischemia. Mice were anesthetized with pentobarbital sodium (100 mg/kg ip) and placed on a ventilator (Harvard
Rodent Ventilator, Harvard Apparatus), and core temperature was
maintained at 37°C with a heating pad. After a left lateral thoracotomy, the left anterior descending coronary artery was occluded for 20
min followed by 10 min of reperfusion. An ECG and blanching of the
CARDIOPROTECTIVE EFFECTS OF ACUTE ROSIGLITAZONE TREATMENT
H897
RESULTS
Acute RGZ treatment decreases myocardial infarction. To
determine the ability of RGZ to protect against myocardial
infarction, nondiabetic mice were subjected to 20 min of
ischemia followed by 4 h of reperfusion. A single dose of RGZ
(1 ␮g/g) or vehicle was injected intravenously via the tail vein
5 min before reperfusion. Myocardial infarction was significantly reduced in mice treated with RGZ compared with
vehicle (8.7 ⫾ 1.1% vs. 20.2 ⫾ 2.5%, respectively, P ⬍ 0.05).
The infarct area was determined as a ratio of the ischemic AAR
(Fig. 1).
RGZ administered at the onset of reperfusion improves
postischemic recovery. To further confirm that RGZ exhibits
protective effects against I/R injury, mouse hearts were subjected to Langendorff heart perfusion. Hearts were first perfused for 20 min to get a stable baseline measurement followed
by 20 min of global, no-flow ischemia followed by 30 min of
reperfusion. Postischemic recovery was significantly improved
with RGZ administered at the onset of reperfusion compared
with vehicle (P ⬍ 0.001), as indicated by the dramatic increase
in the heart rate-LV pressure product (in mmHg·beats·min⫺1;
Fig. 2).
Cardioprotective signaling activated by acute RGZ treatment.
Since RGZ has previously been shown to activate AMPK and
Akt (18, 51), we wanted to investigate whether acute treatment
would activate such cardioprotective pathways. To investigate
these mechanisms in vivo, mouse hearts were subjected to 20
min of ischemia and 10 min of reperfusion. RGZ (1 ␮g/g iv) or
vehicle was injected 5 min before reperfusion (Fig. 3). The
results demonstrated that phosphorylation of AMPK at Thr172
of the ␣-catalytic subunit was significantly upregulated when
RGZ treatment was administered compared with vehicle after
I/R (P ⬍ 0.01; Fig. 3). Confirmation of AMPK activation was
determined by phosphorylation of downstream acetyl-CoA
carboxylase (Fig. 3). Furthermore, another cardioprotective
signaling protein, p-Akt (Ser473), was also significantly upregulated after I/R as a result of RGZ treatment compared with
vehicle (P ⬍ 0.05; Fig. 4). To address whether these effects
were dependent or independent on PPAR-␥ activation, mice
were pretreated with GW-9662, a selective PPAR-␥ receptor
antagonist (19), 10 min before reperfusion (Figs. 3–5). When
AJP-Heart Circ Physiol • VOL
Fig. 2. RGZ ameliorates postischemic cardiac dysfunction. The heart rate and
left ventricular pressure during baseline perfusion and postischemic reperfusion with or without administration of RGZ were assessed as described in
MATERIALS AND METHODS. RGZ (5 ␮M) or vehicle treatment (DMSO) was
initiated at the onset of reperfusion. Cardiac function is presented as the heart
rate-left ventricular developed pressure product (in mmHg·beats·min⫺1). Values are means ⫾ SE; n ⫽ 4 – 6 hearts/group. *P ⬍ 0.001 vs. vehicle during
reperfusion.
RGZ was administered after GW-9662, AMPK and Akt activation were significantly blunted after I/R compared with
RGZ alone (P ⬍ 0.05, Fig. 3, and P ⬍ 0.05, Fig. 4,
respectively). There appeared to be no effect of RGZ or
vehicle during sham operation. However, there was a slight
increase in p-AMPK when RGZ was administered during
sham operation [P ⫽ not significant (NS); Fig. 3].
It has also been demonstrated that RGZ promotes cardioprotection through the activation of endothelial nitric oxide
(NO) synthase (eNOS) (10). Therefore, we sought to determine
whether acute RGZ treatment at this dose would be able to
enhance the phosphorylation of eNOS at Ser1177. RGZ administered at the onset of reperfusion appeared to have no significant effect, although there was a modest increase in p-eNOS
seen when RGZ was given during the sham condition (P ⫽ NS;
Fig. 5).
Acute RGZ treatment inhibits JNK activation. We next
wanted to determine whether acute RGZ treatment would be
able to attenuate JNK activation due to its role in inflammation
and apoptosis as a result of I/R in the heart (15, 27). The results
revealed that p-JNK (Thr183/Tyr185) was significantly downregulated when RGZ was administered during ischemia 5 min
before reperfusion compared with vehicle alone (P ⬍ 0.05;
Fig. 6). Furthermore, when GW-9662 was administered before
RGZ, the ability of RGZ to inhibit JNK phosphorylation was
abolished (P ⬍ 0.01; Fig. 6).
Inhibition of AMPK limits the ability of RGZ to reduce
myocardial infarction. To examine whether AMPK activation
is necessary for RGZ-induced cardioprotection against myocardial infarction, we treated mice with compound C [10 ␮g/g
ip (20)], an AMPK inhibitor (52), 30 min before myocardial
ischemia. Interestingly, pretreatment with compound C followed by RGZ administration 5 min before the onset of
reperfusion largely inhibited the reduction in infarct size seen
301 • SEPTEMBER 2011 •
www.ajpheart.org
Downloaded from http://ajpheart.physiology.org/ by 10.220.33.2 on May 6, 2017
LV confirmed ischemic repolarization changes (ST segment elevation) during coronary occlusion (AD Instruments). RGZ (1 ␮g/g) or
vehicle (1:3 DMSO-saline) was injected via the tail vein 5 min before
reperfusion. GW-9662 [1 ␮g/g (53), Enzo Life Sciences] was injected
via the tail vein 10 min before reperfusion. All hearts were then
rapidly excised, and the ischemic region of the LV was freeze
clamped in liquid nitrogen for biochemical analysis.
Immunoblot analysis. Immunoblots were performed as previously
described (21). Heart homogenates were resolved by SDS-PAGE, and
proteins were transferred onto polyvinylidene difluoride membranes.
For reprobing, membranes were stripped with 50 mmol/l Tris·HCl,
2% SDS, and 0.1 mol/l ␤-mercaptoethanol (pH 6.8). Rabbit polyclonal antibodies against phosphoryalted (p-)AMPK, total AMPK,
p-Akt, p-ACC, p-eNOS, total eNOS, p-JNK, and total JNK were
purchased from Cell Signaling. Rabbit polyclonal antibodies against
total Akt were obtained from Santa Cruz Biotechnology. Anti-rabbit
secondary antibodies were purchased from Cell Signaling.
Statistical analysis. Values are means ⫾ SE. Data were analyzed
by a two-tailed, unpaired Student’s t-test. P values of ⬍0.05 were
considered statistically significant.
H898
CARDIOPROTECTIVE EFFECTS OF ACUTE ROSIGLITAZONE TREATMENT
Fig. 3. RGZ stimulates AMP-activated protein kinase (AMPK) activation
during in vivo I/R. RGZ treatment during regional ischemia followed by
reperfusion stimulated the phosphorylation (p) of both AMPK and downstream
acetyl CoA carboxylase (ACC), as shown by the immunoblots. Pretreatment
with the peroxisome proliferator-activated receptor (PPAR)-␥ inhibitor GW9662 blocked the induction of both AMPK and ACC phosphorylation by RGZ.
Values are means ⫾ SE; n ⫽ 3. *P ⬍ 0.05 vs. I/R ⫹ vehicle; †P ⬍ 0.05 vs.
I/R ⫹ RGZ.
compared with RGZ treatment alone (Fig. 7), suggesting that
AMPK activation is significantly related to RGZ-induced cardioprotection.
DISCUSSION
Compared with the other clinically used antidiabetic TZD
compounds, RGZ is the most potent, which makes it an ideal
candidate for studying its effects against I/R injury. Of the
three PPARs expressed in the heart (PPAR-␣, PPAR-␤/␦,
and PPAR-␥), PPAR-␥ is expressed at the lowest abundance
(33), and our study reveals the potential importance of
activating this receptor during acute ischemic events. In this
study, we demonstrated that a single dose of RGZ administered before the onset of reperfusion is efficient at limiting
myocardial I/R injury and improving postischemic cardiac
function in the nondiabetic mouse heart. The improvement
in postischemic recovery seen with RGZ infusion at the
onset of reperfusion is consistent with a previous report (38)
suggesting that RGZ can strengthen cardiac hemodynamics.
Intriguingly, the dose of 1 ␮g/g (⬃1.4 mM) used to achieve
cardioprotection in mice by RGZ in this study is highly
relevant and is lower than the dose used clinically to reduce
hyperglycemia in type 2 diabetic patients (4 – 8 mg per
70-kg human per day, ⬃2.8 mM).
AJP-Heart Circ Physiol • VOL
Fig. 4. RGZ stimulates Akt phosphorylation during in vivo I/R. RGZ treatment
during regional ischemia followed by reperfusion stimulated Akt phosphorylation
during I/R in the heart. Pretreatment with the PPAR-␥ inhibitor GW-9662 blocked
the induction of Akt phosphorylation by RGZ. Values are means ⫾ SE; n ⫽ 3.
*P ⬍ 0.05 vs. I/R ⫹ vehicle; †P ⬍ 0.05 vs. I/R ⫹ RGZ.
301 • SEPTEMBER 2011 •
www.ajpheart.org
Downloaded from http://ajpheart.physiology.org/ by 10.220.33.2 on May 6, 2017
We have also further elucidated the molecular mechanisms whereby RGZ is able to modulate cardioprotective
signaling pathways that appear to be dependent on PPAR-␥
activation. First, our results suggest that one of the mechanisms contributing to cardioprotection is by activation of
AMPK. This result was further confirmed by pretreatment
with the AMPK inhibitor compound C, which largely attenuated the cardioprotective effects seen with RGZ, implying
that AMPK activation is causatively related to the beneficial
acute actions of RGZ. Our group and others have recently
shown that AMPK is cardioprotective during ischemia by
enhancing glucose uptake and glucose transporter (GLUT)4
translocation (24), decreasing apoptosis, enhancing postischemic recovery, and limiting myocardial infarction (22,
29). Second, we demonstrated that acute RGZ treatment can
activate Akt, which has been previously shown to reduce
ischemic injury (49, 51). Other TZDs, such as pioglitazone,
have also been shown to protect against myocardial infarction by activation of Akt (47). Both of these pathways are
known to stimulate GLUT4 translocation to the sarcolemmal membrane; thus, further studies are needed to determine
the effects of RGZ eliciting a potential synergistic effect on
myocardial glucose metabolism during I/R in the nondiabetic
heart. Taking this into consideration, experiments expressing
dominant negative forms of AMPK and phosphoinositol 3-kinase
in cardiomyocytes that were exposed to oxidative stress with
H2O2 found complete abrogation of GLUT4 translocation to the
CARDIOPROTECTIVE EFFECTS OF ACUTE ROSIGLITAZONE TREATMENT
H899
Fig. 5. Effect of RGZ on endothelial nitric oxide synthase (eNOS) phosphorylation during in vivo I/R. RGZ treatment during regional ischemia followed
by reperfusion did not significantly affect eNOS phosphorylation during I/R in the
heart. Pretreatment with the PPAR-␥ inhibitor GW-9662, however, did modestly
inhibit eNOS phosphorylation by RGZ. Values are means ⫾ SE; n ⫽ 3.
membrane, suggesting the significance of both pathways in response to metabolic stress (12).
Acute RGZ treatment also significantly decreased JNK
phosphorylation, suggesting that RGZ appears to be mitigating the inflammatory response against I/R injury. In
parallel with these results, the inhibition of JNK signaling
has been demonstrated to be protective against I/R by
limiting apoptosis in endothelial cells (46). There is also
evidence suggesting that RGZ can inhibit JNK activation in
adipocytes through a PPAR-␥-dependent mechanism (8). In
addition, AMPK activation by the adipokine C1q/TNFrelated protein-13 has been shown to limit JNK signaling in
response to metabolic stress (45).
Most interestingly, our results show that when AMPK
phosphorylation is upregulated, JNK phosphorylation is
downregulated when RGZ is given before reperfusion.
These findings are in agreement with previous studies as the
relationship between AMPK and JNK signaling in the heart
is beginning to be understood. For instance, a recent study
(31) has indicated that pharmacological AMPK activation
was highly effective at reducing and protecting endothelial
cells against ROS-induced activation of JNK. This is consistent with the results demonstrated by Qi et al. (27)
showing that there is greater JNK activation in AMPKdeficient mouse hearts and that inhibiting the JNK pathway
is cardioprotective during I/R. Furthermore, a recent report
from our group (41) demonstrated that pharmacological
activation of AMPK by activated protein C can attenuate
AJP-Heart Circ Physiol • VOL
Fig. 6. RGZ treatment inhibits the activation of JNK during in vivo I/R. RGZ
treatment attenuated cardiac JNK phosphorylation during I/R in the heart,
whereas pretreatment with the PPAR-␥ inhibitor, GW-9662, attenuated the
inhibition of JNK phosphorylation by RGZ. Values are means ⫾ SE; n ⫽ 3.
*P ⬍ 0.01 vs. sham; †P ⬍ 0.05 vs. I/R ⫹ vehicle; #P ⬍ 0.01 vs. I/R ⫹ RGZ.
301 • SEPTEMBER 2011 •
www.ajpheart.org
Downloaded from http://ajpheart.physiology.org/ by 10.220.33.2 on May 6, 2017
JNK signaling and protect the heart against I/R injury. A
study by Khandoudi et al. (15) has also suggested that
inhibition of the JNK pathway is related to the improved
postischemic hemodynamics seen with RGZ infusion ex
vivo. The inhibition of JNK signaling by RGZ in this case
may also be a contributing factor toward improved cardiac
function.
As mentioned previously, acute RGZ treatment was shown
to rapidly induce vasodilation in a small population of healthy
nondiabetic male patients (39). This led us to hypothesize that
another cardioprotective signaling mechanism due to RGZ
treatment would be through the phosphorylation of eNOS. A
study (10) has shown that RGZ is able to upregulate the
synthesis of NO, and another recent finding (4) suggests that
RGZ’s ability to stimulate NO is through an AMPK-dependent
mechanism. In our study, we examined at p-eNOS (Ser1177)
levels after acute RGZ treatment before reperfusion, and there
appeared to be no significant increase in eNOS phosphorylation. However, there was a slight increase in p-eNOS during
the sham condition that did not reach significance at the dose
of 1 ␮g/g.
It is possible, however, that RGZ may be exerting its
cardioprotective functions through its ability to act as an
antioxidant. This may be particularly true for our results
obtained ex vivo as RGZ has been suggested to improve
H900
CARDIOPROTECTIVE EFFECTS OF ACUTE ROSIGLITAZONE TREATMENT
postischemic cardiac function by limiting ROS production (26)
besides modulating the cardioprotective AMPK signaling pathway (28). A recent study (28) provided evidence that RGZ
protects cardiomyocytes against oxidative stress by upregulating Bcl-2. Moreover, RGZ administration to rats reduced
oxidative stress and increased the activity of superoxide dismutase in the vasculature (26). Findings by Tao et al. (35)
demonstrated that RGZ possesses vasculoprotective properties
by reducing superoxide, nitrotyrosine, and total NO levels.
Interestingly, these studies administered RGZ to nondiabetic
animal models of I/R, and pronounced effects were seen. A
recent report (6) has also suggested that RGZ’s ability to act as
an antioxidant is through its ability to activate AMPK, which,
in turn, prevents hyperactivity of NADPH in endothelial cells.
However, whether acute RGZ treatment in vivo can be antioxidative against I/R injury remains elusive.
Taken together, the results of this study demonstrate that
by using RGZ acutely in the incidence of I/R, myocardial
infarction is decreased and postischemic cardiac function is
improved by modulating AMPK, Akt, and JNK signaling
AJP-Heart Circ Physiol • VOL
mechanisms in the nondiabetic mouse heart. Forms of therapy such as RGZ have recently become attractive in the case
of a nondiabetic patient, as clinical studies (11, 13) have
indicated that developed hyperglycemia as a result of acute
myocardial infarction is a strong predictor of morbidity and
mortality in patients with and without diabetes. In parallel
with our study, Calvert et al. (5) demonstrated that an acute
low-dose of metformin, another AMPK activator, reduces
myocardial infarction in nondiabetic mice. Although the use
of RGZ pertains exclusively to patients with type 2 diabetes,
the use of acute RGZ therapy to nondiabetic patients may be
a potential beneficial treatment for I/R injury.
GRANTS
This work was supported by American Diabetes Association Grant 1-11BS-92 and by American Heart Association Grant SDG 0835169N.
DISCLOSURES
No conflicts of interest, financial or otherwise, are declared by the author(s).
301 • SEPTEMBER 2011 •
www.ajpheart.org
Downloaded from http://ajpheart.physiology.org/ by 10.220.33.2 on May 6, 2017
Fig. 7. AMPK signaling mediates RGZ-induced cardioprotection against myocardial infarction. In vivo hearts were subjected to 20 min of ischemia followed
by 4 h of reperfusion. Compound C (10 ␮g/g ip) or vehicle was administered 30 min before myocardial ischemia. RGZ (1 ␮g/g) or vehicle was administered
via the tail vein 5 min before reperfusion. The extent of myocardial necrosis was assessed as described in MATERIALS AND METHODS. A: representative sections of the
extent of myocardial infarction. B: ratio of the AAR to the total myocardial area (left) and ratio of the infarcted area to the AAR (right). Values are means ⫾ SE for
3–5 independent experiments. *P ⬍ 0.05 vs. vehicle without compound C.
CARDIOPROTECTIVE EFFECTS OF ACUTE ROSIGLITAZONE TREATMENT
REFERENCES
AJP-Heart Circ Physiol • VOL
20.
21.
22.
23.
24.
25.
26.
27.
28.
29.
30.
31.
32.
33.
34.
35.
36.
37.
Shenk JL, Stimmel JB, Therapontos C, Willson TM, Blanchard SG.
Functional consequences of cysteine modification in the ligand binding
sites of peroxisome proliferator activated receptors by GW9662. Biochemistry 41: 6640 –6650, 2002.
Li J, Benashski S, McCullough LD. Post-stroke hypothermia provides
neuroprotection through inhibition of AMP-activated protein kinase. J
Neurotrauma 2011.
Li J, Miller EJ, Ninomiya-Tsuji J, Russell RR 3rd, Young LH.
AMP-activated protein kinase activates p38 mitogen-activated protein
kinase by increasing recruitment of p38 MAPK to TAB1 in the ischemic
heart. Circ Res 97: 872–879, 2005.
Ma H, Wang J, Thomas DP, Tong C, Leng L, Wang W, Merk M,
Zierow S, Bernhagen J, Ren J, Bucala R, Li J. Impaired macrophage
migration inhibitory factor-AMP-activated protein kinase activation and
ischemic recovery in the senescent heart. Circulation 122: 282–292, 2010.
Mersmann J, Tran N, Zacharowski PA, Grotemeyer D, Zacharowski
K. Rosiglitazone is cardioprotective in a murine model of myocardial I/R.
Shock 30: 64 –68, 2008.
Miller EJ, Li J, Leng L, McDonald C, Atsumi T, Bucala R, Young LH.
Macrophage migration inhibitory factor stimulates AMP-activated protein
kinase in the ischaemic heart. Nature 451: 578 –582, 2008.
Mohanty P, Aljada A, Ghanim H, Hofmeyer D, Tripathy D, Syed T,
Al-Haddad W, Dhindsa S, Dandona P. Evidence for a potent antiinflammatory effect of rosiglitazone. J Clin Endocrinol Metab 89: 2728 –
2735, 2004.
Potenza MA, Gagliardi S, De Benedictis L, Zigrino A, Tiravanti E,
Colantuono G, Federici A, Lorusso L, Benagiano V, Quon MJ,
Montagnani M. Treatment of spontaneously hypertensive rats with
rosiglitazone ameliorates cardiovascular pathophysiology via antioxidant
mechanisms in the vasculature. Am J Physiol Endocrinol Metab 297:
E685–E694, 2009.
Qi D, Hu X, Wu X, Merk M, Leng L, Bucala R, Young LH. Cardiac
macrophage migration inhibitory factor inhibits JNK pathway activation
and injury during ischemia/reperfusion. J Clin Invest 119: 3807–3816,
2009.
Ren Y, Sun C, Sun Y, Tan H, Wu Y, Cui B, Wu Z. PPAR gamma
protects cardiomyocytes against oxidative stress and apoptosis via Bcl-2
upregulation. Vascul Pharmacol 51: 169 –174, 2009.
Russell RR 3rd Li J, Coven DL, Pypaert M, Zechner C, Palmeri M,
Giordano FJ, Mu J, Birnbaum MJ, Young LH. AMP-activated protein
kinase mediates ischemic glucose uptake and prevents postischemic cardiac dysfunction, apoptosis, and injury. J Clin Invest 114: 495–503, 2004.
Samaha FF, Szapary PO, Iqbal N, Williams MM, Bloedon LT,
Kochar A, Wolfe ML, Rader DJ. Effects of rosiglitazone on lipids,
adipokines, and inflammatory markers in nondiabetic patients with low
high-density lipoprotein cholesterol and metabolic syndrome. Arterioscler
Thromb Vasc Biol 26: 624 –630, 2006.
Schulz E, Dopheide J, Schuhmacher S, Thomas SR, Chen K, Daiber
A, Wenzel P, Munzel T, Keaney JF Jr. Suppression of the JNK pathway
by induction of a metabolic stress response prevents vascular injury and
dysfunction. Circulation 118: 1347–1357, 2008.
Shibata R, Sato K, Pimentel DR, Takemura Y, Kihara S, Ohashi K,
Funahashi T, Ouchi N, Walsh K. Adiponectin protects against myocardial ischemia-reperfusion injury through AMPK- and COX-2-dependent
mechanisms. Nat Med 11: 1096 –1103, 2005.
Son NH, Park TS, Yamashita H, Yokoyama M, Huggins LA, Okajima
K, Homma S, Szabolcs MJ, Huang LS, Goldberg IJ. Cardiomyocyte
expression of PPARgamma leads to cardiac dysfunction in mice. J Clin
Invest 117: 2791–2801, 2007.
Takano H, Nagai T, Asakawa M, Toyozaki T, Oka T, Komuro I, Saito
T, Masuda Y. Peroxisome proliferator-activated receptor activators inhibit lipopolysaccharide-induced tumor necrosis factor-alpha expression
in neonatal rat cardiac myocytes. Circ Res 87: 596 –602, 2000.
Tao L, Liu HR, Gao E, Teng ZP, Lopez BL, Christopher TA, Ma XL,
Batinic-Haberle I, Willette RN, Ohlstein EH, Yue TL. Antioxidative,
antinitrative, and vasculoprotective effects of a peroxisome proliferatoractivated receptor-gamma agonist in hypercholesterolemia. Circulation
108: 2805–2811, 2003.
Tao L, Wang Y, Gao E, Zhang H, Yuan Y, Lau WB, Chan L, Koch
WJ, Ma XL. Adiponectin: an indispensable molecule in rosiglitazone
cardioprotection following myocardial infarction. Circ Res 106: 409 –417,
2010.
Turer AT, Hill JA. Pathogenesis of myocardial ischemia-reperfusion
injury and rationale for therapy. Am J Cardiol 106: 360 –368, 2010.
301 • SEPTEMBER 2011 •
www.ajpheart.org
Downloaded from http://ajpheart.physiology.org/ by 10.220.33.2 on May 6, 2017
1. Abdelrahman M, Sivarajah A, Thiemermann C. Beneficial effects of
PPAR-gamma ligands in ischemia-reperfusion injury, inflammation and
shock. Cardiovasc Res 65: 772–781, 2005.
2. Abe M, Takiguchi Y, Ichimaru S, Kaji S, Tsuchiya K, Wada K.
Different effect of acute treatment with rosiglitazone on rat myocardial
ischemia/reperfusion injury by administration method. Eur J Pharmacol
589: 215–219, 2008.
3. Aquilante CL, Kosmiski LA, Zineh I, Rome LC, Knutsen SD. Pharmacodynamic effects of rosiglitazone in nondiabetic patients with metabolic syndrome. Pharmacotherapy 30: 236 –247, 2010.
4. Boyle JG, Logan PJ, Ewart MA, Reihill JA, Ritchie SA, Connell JM,
Cleland SJ, Salt IP. Rosiglitazone stimulates nitric oxide synthesis in
human aortic endothelial cells via AMP-activated protein kinase. J Biol
Chem 283: 11210 –11217, 2008.
5. Calvert JW, Gundewar S, Jha S, Greer JJ, Bestermann WH, Tian R,
Lefer DJ. Acute metformin therapy confers cardioprotection against
myocardial infarction via AMPK-eNOS-mediated signaling. Diabetes 57:
696 –705, 2008.
6. Ceolotto G, Gallo A, Papparella I, Franco L, Murphy E, Iori E,
Pagnin E, Fadini GP, Albiero M, Semplicini A, Avogaro A. Rosiglitazone reduces glucose-induced oxidative stress mediated by NAD(P)H
oxidase via AMPK-dependent mechanism. Arterioscler Thromb Vasc Biol
27: 2627–2633, 2007.
7. DeVries JH. Therapies for type 2 diabetes and coronary artery disease. N
Engl J Med 361: 1408 –1409; author reply 1409 –1410, 2009.
8. Diaz-Delfin J, Morales M, Caelles C. Hypoglycemic action of thiazolidinediones/peroxisome proliferator-activated receptor gamma by inhibition of the c-Jun NH2-terminal kinase pathway. Diabetes 56: 1865–1871,
2007.
9. Fryer LG, Parbu-Patel A, Carling D. The Anti-diabetic drugs rosiglitazone and metformin stimulate AMP-activated protein kinase through
distinct signaling pathways. J Biol Chem 277: 25226 –25232, 2002.
10. Gonon AT, Bulhak A, Labruto F, Sjoquist PO, Pernow J. Cardioprotection mediated by rosiglitazone, a peroxisome proliferator-activated
receptor gamma ligand, in relation to nitric oxide. Basic Res Cardiol 102:
80 –89, 2007.
11. Goyal A, Mehta SR, Diaz R, Gerstein HC, Afzal R, Xavier D, Liu L,
Pais P, Yusuf S. Differential clinical outcomes associated with hypoglycemia and hyperglycemia in acute myocardial infarction. Circulation 120:
2429 –2437, 2009.
12. Horie T, Ono K, Nagao K, Nishi H, Kinoshita M, Kawamura T, Wada
H, Shimatsu A, Kita T, Hasegawa K. Oxidative stress induces GLUT4
translocation by activation of PI3-K/Akt and dual AMPK kinase in cardiac
myocytes. J Cell Physiol 215: 733–742, 2008.
13. Ishihara M, Kojima S, Sakamoto T, Kimura K, Kosuge M, Asada Y,
Tei C, Miyazaki S, Sonoda M, Tsuchihashi K, Yamagishi M, Shirai M,
Hiraoka H, Honda T, Ogata Y, Ogawa H. Comparison of blood glucose
values on admission for acute myocardial infarction in patients with versus
without diabetes mellitus. Am J Cardiol 104: 769 –774, 2009.
14. Johns DG, Ao Z, Eybye M, Olzinski A, Costell M, Gruver S, Smith
SA, Douglas SA, Macphee CH. Rosiglitazone protects against ischemia/
reperfusion-induced leukocyte adhesion in the zucker diabetic fatty rat. J
Pharmacol Exp Ther 315: 1020 –1027, 2005.
15. Khandoudi N, Delerive P, Berrebi-Bertrand I, Buckingham RE, Staels
B, Bril A. Rosiglitazone, a peroxisome proliferator-activated receptorgamma, inhibits the Jun NH2-terminal kinase/activating protein 1 pathway
and protects the heart from ischemia/reperfusion injury. Diabetes 51:
1507–1514, 2002.
16. Kilter H, Werner M, Roggia C, Reil JC, Schafers HJ, Kintscher U,
Bohm M. The PPAR-gamma agonist rosiglitazone facilitates Akt rephosphorylation and inhibits apoptosis in cardiomyocytes during hypoxia/
reoxygenation. Diabetes Obes Metab 11: 1060 –1067, 2009.
17. Kim AS, Miller EJ, Wright TM, Li J, Qi D, Atsina K, Zaha V,
Sakamoto K, Young LH. A small molecule AMPK activator protects the
heart against ischemia-reperfusion injury. J Mol Cell Cardiol 51: 24 –32,
2011.
18. LeBrasseur NK, Kelly M, Tsao TS, Farmer SR, Saha AK, Ruderman
NB, Tomas E. Thiazolidinediones can rapidly activate AMP-activated
protein kinase in mammalian tissues. Am J Physiol Endocrinol Metab 291:
E175–E181, 2006.
19. Leesnitzer LM, Parks DJ, Bledsoe RK, Cobb JE, Collins JL, Consler
TG, Davis RG, Hull-Ryde EA, Lenhard JM, Patel L, Plunket KD,
H901
H902
CARDIOPROTECTIVE EFFECTS OF ACUTE ROSIGLITAZONE TREATMENT
AJP-Heart Circ Physiol • VOL
48.
49.
50.
51.
52.
53.
54.
Fujiwara T, Fujiwara H, Minatoguchi S. Antidiabetic drug pioglitazone
protects the heart via activation of PPAR-␥ receptors, PI3-kinase, Akt, and
eNOS pathway in a rabbit model of myocardial infarction. Am J Physiol
Heart Circ Physiol 296: H1558 –H1565, 2009.
Ye JM, Dzamko N, Hoy AJ, Iglesias MA, Kemp B, Kraegen E.
Rosiglitazone treatment enhances acute AMP-activated protein kinasemediated muscle and adipose tissue glucose uptake in high-fat-fed rats.
Diabetes 55: 2797–2804, 2006.
Yue TL, Bao W, Gu JL, Cui J, Tao L, Ma XL, Ohlstein EH, Jucker
BM. Rosiglitazone treatment in Zucker diabetic Fatty rats is associated
with ameliorated cardiac insulin resistance and protection from ischemia/
reperfusion-induced myocardial injury. Diabetes 54: 554 –562, 2005.
Yue TL, Chen J, Bao W, Narayanan PK, Bril A, Jiang W, Lysko PG,
Gu JL, Boyce R, Zimmerman DM, Hart TK, Buckingham RE,
Ohlstein EH. In vivo myocardial protection from ischemia/reperfusion
injury by the peroxisome proliferator-activated receptor-gamma agonist
rosiglitazone. Circulation 104: 2588 –2594, 2001.
Zhang XJ, Xiong ZB, Tang AL, Ma H, Ma YD, Wu JG, Dong YG.
Rosiglitazone-induced myocardial protection against ischaemia-reperfusion injury is mediated via a phosphatidylinositol 3-kinase/Akt-dependent
pathway. Clin Exp Pharmacol Physiol 37: 156 –161, 2010.
Zhou G, Myers R, Li Y, Chen Y, Shen X, Fenyk-Melody J, Wu M,
Ventre J, Doebber T, Fujii N, Musi N, Hirshman MF, Goodyear LJ,
Moller DE. Role of AMP-activated protein kinase in mechanism of
metformin action. J Clin Invest 108: 1167–1174, 2001.
Zingarelli B, Hake PW, Mangeshkar P, O’Connor M, Burroughs TJ,
Piraino G, Denenberg A, Wong HR. Diverse cardioprotective signaling
mechanisms of peroxisome proliferator-activated receptor-gamma ligands,
15-deoxy-Delta12,14-prostaglandin J2 and ciglitazone, in reperfusion injury: role of nuclear factor-kappaB, heat shock factor 1, and Akt. Shock
28: 554 –563, 2007.
Zinn A, Felson S, Fisher E, Schwartzbard A. Reassessing the cardiovascular risks and benefits of thiazolidinediones. Clin Cardiol 31: 397–
403, 2008.
301 • SEPTEMBER 2011 •
www.ajpheart.org
Downloaded from http://ajpheart.physiology.org/ by 10.220.33.2 on May 6, 2017
38. Uchida K, Ogino K, Shimoyama M, Hisatome I, Shigemasa C. Acute
hemodynamic effects of insulin-sensitizing agents in isolated perfused rat
hearts. Eur J Pharmacol 400: 113–119, 2000.
39. Walcher T, Walcher D, Hetzel J, Mielke C, Rau M, Rittig K, Balletshofer B, Schwedhelm E, Hombach V, Boger RH, Koenig W, Marx N.
Rapid effect of single-dose rosiglitazone treatment on endothelial function
in healthy men with normal glucose tolerance: Data from a randomized,
placebo-controlled, double-blind study. Diab Vasc Dis Res 2010.
40. Wang G, Wei J, Guan Y, Jin N, Mao J, Wang X. Peroxisome
proliferator-activated receptor-gamma agonist rosiglitazone reduces clinical inflammatory responses in type 2 diabetes with coronary artery disease
after coronary angioplasty. Metabolism 54: 590 –597, 2005.
41. Wang J, Yang L, Rezaie AR, Li J. Activated protein C protects against
myocardial ischemic/reperfusion injury through AMP-activated protein kinase
signaling. J Thromb Haemost; doi:10.1111/j.1538-7836.2011.04331.x.
42. Wang TD, Chen WJ, Lin JW, Chen MF, Lee YT. Effects of rosiglitazone on endothelial function, C-reactive protein, and components of the
metabolic syndrome in nondiabetic patients with the metabolic syndrome.
Am J Cardiol 93: 362–365, 2004.
43. Wang W, Lopaschuk GD. Metabolic therapy for the treatment of ischemic heart disease: reality and expectations. Exp Rev Cardiovasc Ther 5:
1123–1134, 2007.
44. Wang Y, Lau WB, Gao E, Tao L, Yuan Y, Li R, Wang X, Koch WJ,
Ma XL. Cardiomyocyte-derived adiponectin is biologically active in
protecting against myocardial ischemia-reperfusion injury. Am J Physiol
Endocrinol Metab 298: E663–E670, 2010.
45. Wei Z, Peterson JM, Wong GW. Metabolic regulation by C1q/TNFrelated protein-13 (CTRP13): activation of AMP-activated protein kinase
and suppression of fatty acid-induced JNK signaling. J Biol Chem 2011.
46. Yang D, Guo S, Zhang T, Li H. Hypothermia attenuates ischemia/
reperfusion-induced endothelial cell apoptosis via alterations in apoptotic
pathways and JNK signaling. FEBS Lett 583: 2500 –2506, 2009.
47. Yasuda S, Kobayashi H, Iwasa M, Kawamura I, Sumi S, Narentuoya
B, Yamaki T, Ushikoshi H, Nishigaki K, Nagashima K, Takemura G,