© 2000 American Heart Association, Inc.
Integrative Physiology |
Targeted Disruption of the Mouse Sod I Gene Makes the Hearts Vulnerable to Ischemic Reperfusion Injury
From the Cardiovascular Research Center (T.Y., N.M., R.M.E., D.K.D.), Department of Surgery, University of Connecticut School of Medicine, Farmington, Conn, and Institute of Chemical Toxicology (Y.-S.H.), Wayne State University, Detroit, Mich.
Correspondence to Dipak K. Das, University of Connecticut, School of Medicine, Cardiovascular Research Center, Department of Surgery, Farmington, CT 06030-1110. E-mail ddas{at}neuron.uchc.edu
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Abstract |
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Abstract—The role of Cu/Zn–superoxide dismutase (SOD) in myocardial ischemic reperfusion injury was studied by using a mouse model with targeted disruption of the mouse Sod I gene. Inactivation of the functional mouse Sod I gene in hearts by gene targeting (Sod I+/-) resulted in a 50% reduction of Cu/Zn-SOD mRNA and significant reduction of Cu/Zn-SOD enzyme activity compared with that of wild-type Sod I+/+ mice. Cu/Zn-SOD mRNA could not be detected in Sod I-/- heart. The isolated buffer-perfused hearts from the knockout mice devoid of any functional copy of the Sod I (Sod I-/-) and matched nontransgenic control mice were subjected to 30 minutes of global ischemia followed by 2 hours of reperfusion. For both groups of mice, the postischemic functional recovery for the hearts was lower than the baseline, but the recovery for the Sod I-/- was less compared with the wild-type mice. Thus, the postischemic recovery of the developed force and the maximum first derivative of the developed force were consistently lower for the Sod I-/- mouse hearts compared with wild-type control hearts. The coronary flow was lower compared with the baseline levels for both groups of hearts, but there was no significant difference between the groups. The myocardial infarction determined from the ratio of infarct size/area of risk was higher for the Sod I-/- mice compared with the control mice. The amount of creatine kinase release from the wild-type mouse hearts was less compared with the Sod I-/- mouse hearts. In concert, a reduced amount of oxidative stress was found in the hearts of wild-type mice compared with Sod I-/- mouse hearts. These results documented that Sod I-/- mouse hearts were more susceptible to ischemic reperfusion injury compared with corresponding wild-type mouse hearts, suggesting that the Sod I gene constitutes an important defense element for the hearts.
Key Words: transgenic • knockout • ischemia/reperfusion • free radicals • oxidative stress
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Introduction |
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Reactive oxygen species, including superoxide and hydroxyl radicals, as well as oxidants such as hydrogen peroxide, have been implicated in the pathogenesis of myocardial ischemic reperfusion injury.1 2 3 In biological tissue, superoxide anions are readily dismutated by superoxide dismutase (SOD) into hydrogen peroxide, which can then be converted into hydroxyl radical by a Fenton-type reaction.4 SOD consists of a transition metal at the active center, and, depending on the transition metal, 3 types of SOD are known to exist, as follows: manganese SOD (Mn-SOD), copper/zinc-SOD (Cu/Zn-SOD), and iron-SOD (Fe-SOD). The best-characterized SOD is Mn-SOD, which is primarily located in the soluble matrix of the mitochondria. In contrast, the Cu/Zn-SOD is exclusively located in the cytosol. Whereas the cardioprotective role of Mn-SOD is well established,5 6 very little is known about the role of Cu/Zn-SOD in myocardial protection.
Several recent studies have documented that Cu/Zn-SOD plays a crucial role in cellular protection associated with ischemia and reperfusion. For example, overexpression of Cu/Zn-SOD reduces hippocampal injury after global ischemia in transgenic mice.7 Transgenic mice with increased Cu/Zn-SOD activity were found to be resistant to hepatic leukostasis and capillary no-reflow after gut ischemia/reperfusion.8 Another recent study demonstrated prevention of postischemic injury by overexpressing Cu/Zn-SOD.9 Adverse effects of Cu/Zn have also been reported. Overexpression of the human Cu/Zn-SOD in mice containing an amyotrophic lateral sclerosis mutation developed a disorder in the transgenic mice that closely resembled human amyotrophic lateral sclerosis.10 Another study implicated Cu/Zn-SOD to be neurotoxic.11
This study sought to examine the cardioprotective effects of Cu/Zn-SOD by using mice with targeted disruption of the mouse Sod I gene. The isolated perfused hearts from the knockout mice devoid of any functional copy of the Sod I gene (Sod I-/-), as well as Sod I+/- and matched wild-type mice, were subjected to an ischemia/reperfusion protocol. The results demonstrated impaired postischemic ventricular recovery and increased infarct size for the Sod I-/- mouse hearts compared with those from Sod I+/- and wild-type controls, suggesting that Sod I-/- mouse hearts are more susceptible to ischemic reperfusion injury.
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Materials and Methods |
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Targeted Disruption of Mouse SOD1 Gene
Mouse Sod1 genomic clones were isolated from a 129/SVJ genomic library (Stratagene) by screening with a rat SOD I cDNA probe.12 An
7.2-kb SacI
genomic fragment, which contains the entire mouse SOD I
gene, was isolated from clone 30 and used in construction of the gene
targeting vector. To inactivate the mouse SOD I
gene, the SmaI and HindIII restriction sites
flanking the SmaI-HindIII fragment, which
contains sequences from introns 1 to 4, were converted into
XhoI sites by linker ligation and then inserted into the
XhoI site in plasmid vector pPNT13
(generously provided by Dr Richard Mulligan, Massachusetts Institute of
Technology) (Figure 1
). Similarly, linker ligation was also used
to clone the EcoRI-SalI fragment containing the
3'-flanking sequence of the gene into the BamHI site in the
pPNT vector.
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The Sod1 targeting vector, in which exon 5 was deleted, was
linearized by HindIII digestion and transfected into R1 ES
cells14 (a generous gift from Dr Andras Nagy, Mount
Sinai Hospital at Toronto, Ontario, Canada). Clones
resistant to G418 and ganciclovir (a gift from Syntex Inc, Palo
Alto, CA) were screened by Southern blot analysis using a probe
5' external to the genomic sequence present in the targeting vector
(Figure 1
). Targeted clones were microinjected into C57BL/6
blastocysts following the standard procedure.15 Chimeric
mice with near 100% chimerism were generated using Sod1
knockout clone 5 and showed 100% transmission of the 129/SVJ
chromosomes.
Isolated Mouse Heart Preparation and Measurement of
Contractile Function
Animals received care in compliance with the Principles of
Laboratory Animal Care formulated by the National Society for
Medical Research and the Guide for the Care and Use of Laboratory
Animals prepared by the Institute of Laboratory Animal
Resources and published by the National Institutes of Health (revised
1996).
Mice (Sod I-/-, Sod I+/-, and wild-type) were anesthetized, and the hearts were excised after thoracotomy. The aorta was cannulated, and the heart was perfused with Krebs-Henseleit bicarbonate buffer by the retrograde Langendorff method.16 A 4-0 silk suture on a round-bodied needle was passed through the apex of the heart and attached to the apex, which in turn was attached to a force transducer. The heart rate, heart-developed force (dF), and first derivative of dF (dF/dt) were recorded.16 The hearts of all mice were subjected to 30 minutes of ischemia followed by 2 hours of reperfusion.
Assessment of Cellular Injury
Creatine kinase (CK) release from the heart was estimated in the
perfusate to assess cellular injury. At the end of reperfusion,
a 10% (wt/vol) solution of triphenyl tetrazolium in phosphate buffer
was infused into a side arm of the aortic cannula until the
myocardium stained deep red.17 The hearts were
excised, weighed, and stored at -70°C for subsequent determination
of infarct size as described previously.16 17
Determination of Reactive Oxygen Species in the Heart
To estimate oxygen free radicals, an additional group of
wild-type, Sod I+/-, and Sod
I-/- hearts were perfused for 15 minutes in the
presence of 2 mmol/L salicylate before ischemia as
described previously.18 At the end of the 2 hours of
reperfusion, hearts were frozen at -70°C for subsequent
analysis of hydroxylated benzoic acids by HPLC using an
electrochemical detector.18 Additionally, malonaldehyde
was assayed19 to monitor the development of oxidative
stress during ischemia/reperfusion.
An expanded Materials and Methods section is available online at http://www.circresaha.org.
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Results |
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Generation and Characterization of Sod I Knockout Mice
As shown in Figure 1
, exon 5 of the
mouse Sod1 gene (which encodes the C terminus of the protein
from amino acid residues 120 to 154, which constitute both the
structure and function of the active site channel20 )
and some of the flanking intron sequences were replaced with the
neomycin resistance cassette (neo). This also creates a new
PstI restriction site, resulting in a shorter
PstI genomic fragment from the targeted allele (12.5 kb)
than that from the wild-type allele (16.5 kb). Mice heterozygous
(Sod I+/-) for the targeted allele
were interbred to generate homozygous knockout
(Sod1-/-) mice. Male and female
Sod1-/- mice grew normally and were
apparently healthy under routine animal husbandry.
Inactivation of the functional mouse Sod1 gene in mouse
hearts by gene targeting was initially determined by RNA blot
analysis. An 50% reduction of Cu/Zn-SOD mRNA was found in
the heart of Sod1+/- mice compared with
that of wild-type (Sod I+/+) mice (Figure 2A). Furthermore, no Cu/Zn-SOD mRNA could
be detected in Sod1-/- heart, indicating
that the truncated Cu/Zn-SOD or Cu/Zn-SOD–neo fusion mRNA
is degraded rapidly in the heart. Reduction of heart Cu/Zn-SOD enzyme
activity in Sod1+/- and
Sod1-/- mice was also confirmed by SOD
activity staining on a native polyacrylamide
gel.21 Figure 2B shows that the Cu/Zn-SOD
activities in the hearts of Sod1+/+,
Sod1+/-, and
Sod1-/- mice are proportional to the mRNA
levels in these mice, as shown in Figure 2A. A decrease in
Cu/Zn-SOD activity apparently had no effects on the activity of other
heart antioxidant enzymes, such as manganese SOD (Mn-SOD) (Figure 2B), catalase; glutathione peroxidase; and enzymes involved in
the recycling of oxidized glutathione, including glutathione reductase
and glucose-6-phosphate dehydrogenase (data not shown). A more detailed
characterization of the Cu/Zn-SOD–deficient mice has been reported
elsewhere.22 Male and female
Sod1+/+ and
Sod1-/- mice at 10 to 12 weeks of age
were used for myocardial ischemia/reperfusion study.
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Myocardial Performance
All hearts recovered their beats spontaneously after 30 minutes of
global ischemia followed by reperfusion. However, the heart
rates remained lowered compared with baseline (Figure 3, top). No significant difference
was found in heart rate between groups throughout the experiment.
Coronary flow was significantly reduced after 30 minutes of
reperfusion compared with baseline for all groups of mouse hearts
(Figure 3, bottom). Although coronary flow was
progressively reduced up to 2 hours of reperfusion, there was no
difference between the 3 groups of hearts. The baseline values of dF
(0.63±0.07 g for wild type, 0.61±0.05 g for Sod
I+/-, and 0.57±0.08 g for Sod
I-/- mice) and maximal first derivative of dF
(dF/dt; 265±33 g/s for wild-type, 251±25 g/s for Sod
I+/-, and 233±27 g/s for Sod
I-/- mice) did not vary significantly
among the groups. The dF during reperfusion was lower for the Sod
I-/- knockout mouse hearts than for control
wild-type hearts throughout the reperfusion (Figure 4, top). In wild-type and Sod
I+/- hearts, dF recovered to the level of
baseline after 30 minutes of reperfusion, and this level was maintained
higher than the baseline through the end of the experiment. On the
other hand, dF did not recover beyond the 90% level of baseline for
the Sod I-/- hearts. At all points, dF
showed significantly lower recovery for Sod
I-/- hearts compared with nontransgenic control
and Sod I+/- hearts after 15 minutes of
reperfusion (P<0.05). A similar trend was seen in
dF/dtmax (Figure 4, bottom). A
significantly lower recovery of dF/dtmax occurred
in Sod I-/- hearts after 15 minutes of
reperfusion (P<0.05). These results demonstrate that
Sod I-/- mouse hearts showed
significantly lower myocardial contractile recovery after 30 minutes of
ischemia.
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), and
Sod1-/- (
) mouse hearts.
Results are expressed as mean±SEM (n=12) for each group.
*P<0.05 compared with control. BL indicates
baseline.
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CK Release From Heart
Total CK release from the heart (Figure 5), which reflects cellular injury or
tissue necrosis and membrane permeability, was only 5 U/mL for all
of the groups before ischemia. After ischemia, CK
release was increased in all 3 groups, but the amount of release was
much higher for the Sod I-/- knockout
mouse hearts. For example, at 30 minutes of reperfusion, CK release was
122±8.7 IU/mL for Sod I-/- mice as
compared with 70±6.8v IU/mL (P<0.05) for nontransgenic
wild-type controls. The amount of CK release was progressively
increased as the duration of reperfusion increased. At 60 minutes of
reperfusion, CK release was 151±7.7 IU/mL for Sod
I-/- mouse heart compared with 98±6.4 IU/mL
(P<0.05) for nontransgenic controls. Similarly, after 120
minutes of reperfusion, CK release from the Sod
I-/- mouse hearts amounted to 163±8.1 IU/mL
compared with that from nontransgenic controls, which was 118±7.4
IU/mL. Amount of CK release from the hearts of Sod
I+/- mice was not significantly different from
that from the wild-type mouse hearts at any point.
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Myocardial Infarction
In this study, global ischemia for 30 minutes was adopted;
therefore, the whole ventricle was regarded as the area of risk. The
area that was not stained by triphenyl tetrazolium was measured and
calculated as the infarct area. Infarct size for each heart was defined
as percentage of area at risk. Mean value of infarct size in the
Sod I-/- mouse heart group was
significantly higher than that in the control group (38.8±3.3% versus
22.5±1.8% for the wild-type, P<0.05) (Figure 6). Infarct size of the hearts from
Sod I+/- mouse hearts was similar to that
of wild-type control. Our results indicated that Sod
I-/- mouse heart mouse hearts had significantly
higher myocardial necrosis.
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Increased OH· Formation in the Sod
I-/- Mouse Hearts
The levels of 2,3-dihydroxy benzoic acid (2,3-DHBA) in the
hearts of wild-type, Sod I+/-, and
Sod I-/- mice are shown in Figure 7. The OH· radical produces both
2,3-DHBA and 2,5-DHBA, but 2,5-DHBA may also be produced by the
cytochrome P450 system.17 We therefore monitored
2,3-DHBA formation, which truly reflects the production of
OH· radical. Similar amounts of 2,3-DHBA were found in all 3
groups of hearts at baseline (before ischemia and reperfusion).
The amount of 2,3-DHBA increased at the end of
ischemia/reperfusion (after 2 hours of reperfusion) in all
hearts, indicating an increased OH· formation. A significantly
higher amount of 2,3-DHBA was noticed in the hearts of Sod
I-/- mouse compared with Sod
I+/- mouse hearts and wild-type mouse hearts,
demonstrating that significantly higher amounts of reactive oxygen
species were formed in the Sod I-/- mouse
hearts on ischemia and reperfusion.
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Malonaldehyde (MDA) Formation
MDA formation is considered a presumptive marker for oxidative
stress. MDA was measured as MDA-DNPH derivative by HPLC. MDA formation
was increased progressively and steadily during the
postischemic reperfusion in all 3 group of hearts (Figure 8). However, the amount of MDA
production was significantly higher at all points in the
Sod I-/- mouse hearts compared with those
for wild-type and Sod I+/- mouse hearts,
demonstrating that Sod I-/- mouse hearts
were subjected to an increased amount of oxidative stress during the
postischemic reperfusion.
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An expanded Materials and Methods section is available online at http://www.circresaha.org.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The results of this study documented that Sod I-/- mouse hearts are more susceptible to ischemic reperfusion injury compared with corresponding wild-type and Sod I+/- mouse hearts, suggesting that Sod I gene constitutes an important defense element for the hearts. This gene encodes a Cu/Zn-dependent cytosolic enzyme of dimeric 32-kDa protein that dismutates O2– into H2O2 and O2. Cu/Zn-SOD is considered primarily an antioxidant enzyme that can reduce oxidative stress. Our results also supported antioxidant action of Cu/Zn-SOD, because Sod I-/- knockout mouse hearts devoid of any Cu//Zn-SOD activities contained the highest amount of 2,3-DHBA, a marker for OH· formation and malonaldehyde, which is a presumptive marker for lipid peroxidation and oxidative stress. Postischemic left ventricular functional recovery of Sod 1-/- mouse hearts was the lowest compared with those of wild-type controls and Sod I+/- mouse hearts, further supporting a cardioprotective role of Cu/Zn-SOD.
The present study utilized both Sod
I+/- and Sod I-/-
mouse hearts. As mentioned earlier, Sod
I+/- mouse hearts showed 50% reduction of
Cu/Zn-SOD mRNA compared with that of wild-type Sod
I+/+ mouse hearts. No Cu/Zn-SOD mRNA was detected
in the hearts of Sod I-/- mice. The
activities of other key antioxidant enzymes, including Mn-SOD,
catalase, glutathione peroxidase, glutathione reductase, and
glucose-6-phosphate dehydrogenase, remained unaltered in the
Sod+/- and Sod
I-/- mouse hearts compared with wild-type
controls. The results thus truly reflect the effects of Cu/Zn-SOD gene
knockout in myocardial ischemic reperfusion injury.
SOD is present in both the cytosolic and the mitochondrial fractions of the mammalian heart. A significant number of studies exist in the literature to show the cardioprotective role of mitochondrial SOD or Mn-SOD. For example, overexpression of Mn-SOD has been found to protect myocardial ischemia/reperfusion injury in transgenic mice.5 Increased expression of Mn-SOD in the hearts transfected with Mn-SOD showed significant improvement in tolerance against ischemia/reperfusion injury.23 The activation of Mn-SOD by the free radicals produced during hyperthermia was found to be important for early-phase and late-phase cardioprotection against ischemia/reperfusion injury in rats. In another related study, the same investigators have found that heat shock–induced Mn-SOD enhanced the tolerance of cardiac myocytes to hypoxia-reoxygenation injury.24 Preconditioning of a heart by cyclic episodes of ischemia and reperfusion has been found to induce Mn-SOD, suggesting its importance in the myocardial defense mechanism.25 Mn-SOD could protect mitochondrial complex I against Adriamycin-induced cardiomyopathy in transgenic mice.26 Another recent study also showed the induction of Mn-SOD,27 supporting this previous observation.
In contrast, only a little information is available regarding the
cardioprotective ability of cytosolic Cu/Zn-SOD. Initial studies have
resulted in conflicting reports, showing Cu/Zn-SOD to be both
protective and nonprotective.28 Efficacy of Cu/Zn-SOD in
cardioprotection was believed to be limited by its inability to enter
the cells,29 as well as by its nonspecific deleterious
effects.30 However, our initial study has shown Cu/Zn-SOD
to be cardioprotective against ischemia/reperfusion
injury.31 A recent study has indicated that overexpression
of human Cu/Zn-SOD prevents postischemic
injury.9 The present study supports this previous
report and further documents that Sod
I-/- mouse hearts are subjected to an increased
amount of oxidative stress during ischemia and reperfusion. The
amount of 2,3-DHBA was the highest in the hearts of Sod
I-/- mouse, indicating a significantly higher
amount of OH· radical formation in these hearts compared with
either wild-type or Sod I+/- mouse hearts.
Sod I+/- mouse hearts contained 50% of
the mRNA and enzyme activity compared with wild-type mouse hearts. Our
results indicate that 50% of the total Sod I activity
provided a comparable degree of cardioprotection (as compared with
wild-type hearts) against ischemia/reperfusion injury.
A large number of studies exist in literature demonstrating development
of oxidative stress due to increased formation of reactive oxygen
species and decreased antioxidant reserve in the ischemic
reperfused myocardium.32 33 The reactive
oxygen species include superoxide anions and hydroxyl and peroxyl
radicals, as well as oxidants such as
H2O2.32 Mammalian
hearts are protected from the oxidant challenge by several defense
systems, which include antioxidant enzymes such as SOD, catalase, and
glutathione peroxidase, and antioxidants, which include glutathione,
ascorbic acid, and 
-tocopherol.33 Our
results imply that Cu/Zn-SOD may also be a part of the antioxidant
reserve, because Sod I-/- mouse
hearts were subjected to increased oxidative stress in concert with
increased ischemic/reperfusion injury.
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Acknowledgments |
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This work was supported by NIH Grants HL 33889, HL 56421, HL 34360, HL 22559, HL 56803, and P30 ES06639, as well as by a Grant-in-Aid from the American Heart Association.
Received August 30, 1999; accepted November 22, 1999.
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 J. Suzuki, F. Broeyer, A. Cohen, M. Takebe, J. Burggraaf, and Y. Mizushima Pharmacokinetics of PC-SOD, a Lecithinized Recombinant Superoxide Dismutase, After Single- and Multiple-Dose Administration to Healthy Japanese and Caucasian Volunteers J. Clin. Pharmacol., February 1, 2008; 48(2): 184 - 192. [Abstract] [Full Text] [PDF]
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A. Heinen, M. Aldakkak, D. F. Stowe, S. S. Rhodes, M. L. Riess, S. G. Varadarajan, and A. K. S. Camara Reverse electron flow-induced ROS production is attenuated by activation of mitochondrial Ca2+-sensitive K+ channels Am J Physiol Heart Circ Physiol, September 1, 2007; 293(3): H1400 - H1407. [Abstract] [Full Text] [PDF]
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 S. Srivastava, B. Chandrasekar, Y. Gu, J. Luo, T. Hamid, B. G. Hill, and S. D. Prabhu Downregulation of CuZn-superoxide dismutase contributes to {beta}-adrenergic receptor-mediated oxidative stress in the heart Cardiovasc Res, June 1, 2007; 74(3): 445 - 455. [Abstract] [Full Text] [PDF]
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 W. Hsueh, E. D. Abel, J. L. Breslow, N. Maeda, R. C. Davis, E. A. Fisher, H. Dansky, D. A. McClain, R. McIndoe, M. K. Wassef, et al. Recipes for Creating Animal Models of Diabetic Cardiovascular Disease Circ. Res., May 25, 2007; 100(10): 1415 - 1427. [Abstract] [Full Text] [PDF]
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 J.-H. Zhu and X. G. Lei Double Null of Selenium-Glutathione Peroxidase-1 and Copper, Zinc-Superoxide Dismutase Enhances Resistance of Mouse Primary Hepatocytes to Acetaminophen Toxicity. Experimental Biology and Medicine, May 1, 2006; 231(5): 545 - 552. [Abstract] [Full Text] [PDF]
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 M. Jain, L. Cui, D. A. Brenner, B. Wang, D. E. Handy, J. A. Leopold, J. Loscalzo, C. S. Apstein, and R. Liao Increased Myocardial Dysfunction After Ischemia-Reperfusion in Mice Lacking Glucose-6-Phosphate Dehydrogenase Circulation, February 24, 2004; 109(7): 898 - 903. [Abstract] [Full Text] [PDF]
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F. Eefting, B. Rensing, J. Wigman, W. J. Pannekoek, W. M. Liu, M. J. Cramer, D. J Lips, and P. A Doevendans Role of apoptosis in reperfusion injury Cardiovasc Res, February 15, 2004; 61(3): 414 - 426. [Abstract] [Full Text] [PDF]
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S. P. Jones, M. R. Hoffmeyer, B. R. Sharp, Y.-S. Ho, and D. J. Lefer Role of intracellular antioxidant enzymes after in vivo myocardial ischemia and reperfusion Am J Physiol Heart Circ Physiol, January 1, 2003; 284(1): H277 - H282. [Abstract] [Full Text] [PDF]
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 Y.-S. Ho Transgenic and Knockout Models for Studying the Role of Lung Antioxidant Enzymes in Defense against Hyperoxia Am. J. Respir. Crit. Care Med., December 15, 2002; 166(12): S51 - 56. [Abstract] [Full Text] [PDF]
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G. Imamura, A. A. Bertelli, A. Bertelli, H. Otani, N. Maulik, and D. K. Das Pharmacological preconditioning with resveratrol: an insight with iNOS knockout mice Am J Physiol Heart Circ Physiol, June 1, 2002; 282(6): H1996 - H2003. [Abstract] [Full Text] [PDF]
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 D.J. Hearse Ischaemia, reperfusion and cardioprotection: successes and failures in the journey from molecule to man Eur. Heart J. Suppl., June 1, 2001; 3(suppl_C): C11 - C21. [Abstract] [PDF]
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T. Yoshida, N. Maulik, Y.-S. Ho, J. Alam, and D. K. Das Hmox-1 Constitutes an Adaptive Response to Effect Antioxidant Cardioprotection : A Study With Transgenic Mice Heterozygous for Targeted Disruption of the Heme Oxygenase-1 Gene Circulation, March 27, 2001; 103(12): 1695 - 1701. [Abstract] [Full Text] [PDF]
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G. K. Asimakis, S. Lick, and C. Patterson Postischemic Recovery of Contractile Function is Impaired in SOD2+/- but Not SOD1+/- Mouse Hearts Circulation, February 26, 2002; 105(8): 981 - 986. [Abstract] [Full Text] [PDF]
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