Journal of Applied Physiology Watch the video to learn how APS reaches out to developing nations.
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

J Appl Physiol 97: 1323-1328, 2004. First published June 4, 2004; doi:10.1152/japplphysiol.00380.2004
8750-7587/04 $5.00
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF) Free
Right arrow All Versions of this Article:
97/4/1323    most recent
00380.2004v1
Right arrow Submit a response
Right arrow Alert me when this article is cited
Right arrow Alert me when eLetters are posted
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via Web of Science (8)
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Qi, W.-N.
Right arrow Articles by Urbaniak, J. R.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Qi, W.-N.
Right arrow Articles by Urbaniak, J. R.

Reperfusion injury in skeletal muscle is reduced in inducible nitric oxide synthase knockout mice

Wen-Ning Qi,1 Long-En Chen,1 Li Zhang,1 Jerry P. Eu,2 Anthony V. Seaber,1 and James R. Urbaniak1

1Orthopaedic Microsurgery Laboratories, Department of Surgery, and 2Pulmonary Divisions, Departments of Medicine and Cell Biology, Duke University Medical Center, Durham, North Carolina 27710

Submitted 7 April 2004 ; accepted in final form 26 May 2004


   ABSTRACT
TOP
 ABSTRACT
MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 
Inducible nitric oxide synthase (iNOS) participates in many pathological events, and selective inhibition of iNOS has been shown to reduce ischemia-reperfusion (I/R) injury in different tissues. To further confirm its role in this injury process, I/R injury was observed in denervated cremaster muscles of iNOS-deficient (iNOS–/–) and wild-type mice. After 3-h ischemia and 90-min reperfusion, blood flow in reperfused muscle was 80 ± 8.5% (mean ± SE) of baseline at 10-min reperfusion and completely returned to the preischemia baseline after 20 min in iNOS–/– mice. In contrast, blood flow was 32 ± 7.4% at 10 min and increased to 60 ± 20% of the baseline level at 90 min in wild-type mice (P < 0.001 vs. iNOS–/– mice at all time points). The increased muscle blood flow in iNOS–/– mice was associated with significantly less vasospasm in all three sizes of arterial vessel size categories. The weight ratio to the contralateral muscle not subjected to I/R was greater in wild-type mice (173 ± 11%) than in iNOS–/– mice (117 ± 3%; P < 0.01). Inflammation and neutrophil extravasation were also more severe in wild-type mice. Western blot analysis demonstrated an absence of iNOS protein band in iNOS–/– mice and upregulation of iNOS protein expression in wild-type mice. Our results confirm the importance of iNOS in I/R injury. Upregulated iNOS exacerbates I/R injury and appears to be a therapeutic target in protection of tissues against this type of injury.

vessel diameter; blood flow; mRNA; protein

ISCHEMIA-REPERFUSION (I/R) injury contributes to tissue morbidity in many clinical situations, such as replantation, free tissue transfer, myocardial infarction, stroke, and organ transplantation (29, 42). Ischemia of sufficient duration results in tissue injury, mainly from depletion of oxygen and energy. Subsequent reperfusion leads to a series of pathological responses, resulting in severe injury, not only at the tissue level, but also at the molecular level (4, 16, 47).

Many cytokines and inflammatory mediators participate in I/R injury (1, 38, 42). Among them, nitric oxide (NO) has emerged as a potential mediator of I/R injury in skeletal muscle (6, 33). In mammals, NO is generated from L-arginine by one of the three NO synthase (NOS) isoforms, and all three isoforms have been found in skeletal muscle. Both neuronal NOS (nNOS) and endothelial NOS (eNOS) are constitutively but variably expressed in mature skeletal muscle fibers (22) in which they are predominantly localized to the sarcolemma or mitochondria, respectively (37). eNOS is also constitutively expressed in the vascular endothelia of skeletal muscle. These two isoforms of NOS are activated by a rise in intracellular calcium concentrations via Ca2+/calmodulin-NOS interaction. Constitutively expressed NOS isoforms may be protective in I/R injury of skeletal muscle, as intravenous infusion of a low dose of NO donor S-nitroso-N-acetyl cysteine has been shown to attenuate I/R injury in rat skeletal muscle (6, 27). In addition, a nonspecific NOS inhibitor, LG-monomethyl-L-arginine, worsens the muscle contractile dysfunction in extensor digitorum longus (EDL) muscle of the rat following I/R (20).

In contrast, inducible NOS (iNOS) is generally upregulated at the transcriptional level during chronic and acute pathological states and produces a high level of NO (9, 28, 40). A high level of NO by itself or in combination with other reactive oxygen species (ROS) such as superoxide are deleterious to tissues. iNOS is expressed in normal skeletal muscle fibers and is upregulated after stimulation with inflammatory cytokines. In addition, leukocytes infiltrating the tissues in I/R injury also express iNOS (3). Thus iNOS participates in many pathological events, including I/R injury. Our previous work suggests that iNOS potentiates I/R injury in skeletal muscle based on the observation that dexamethasone, which suppresses iNOS expression (5) among its vast effects, attenuated I/R injury in rat EDL (7). Specifically, our recent work has shown that a highly selective iNOS inhibitor, [3-(aminomethyl)benzyl]acetamidine (1400W), significantly improves microcirculation in reperfused rat cremaster muscle (34, 48) and contractile function in reperfused rat EDL (32). However, our data also show that 1400W has weak inhibition to other NOS forms (32). To provide unequivocal proof of the importance of iNOS in I/R injury and to avoid the lack of specificity of chemical NOS inhibitors (2), this study investigated I/R injury in denervated rat cremaster muscle in mice with iNOS gene disruption (iNOS–/–).


   MATERIALS AND METHODS
TOP
 ABSTRACT
 MATERIALS AND METHODS
RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 
Homozygous male iNOS–/– (n = 16) mice and their wild-type counterparts with same genetic background (n = 16) were obtained from Jackson Laboratory (Bar Harbor, ME). The protocols were approved by the IACUC of Duke University Medical Center. These mice weighed 20–25 g and received a standard diet and water ad libitum. Each animal underwent unilateral surgical procedures to cause complete ischemia to the left cremaster muscles followed by reperfusion.

Surgical preparation of the cremaster muscle.   The mice were anesthetized with an intraperitoneal injection of pentobarbital sodium (Nembutal, 50 mg/kg body wt; Abbott Laboratories, North Chicago, IL). The cremaster muscle was prepared according to our laboratory's previously described technique (27, 34, 48). The muscle was exposed, opened, and then separated from the testis. The pudic-epigastric artery and vein and the genitofemoral nerve were isolated and separated from each other. The cremaster muscle sac was cut circumferentially, rendering the muscle totally isolated but still attached to the body by its neurovascular pedicle. A 3-mm segment of the genitofemoral nerve was removed proximally to the muscle. The isolated muscle was then spread flat onto the surface of a transparent acrylic microscope stage and secured with five peripheral sutures. The exposed surface of the muscle was moistened with warm (34 ± 0.5°C) lactated Ringer solution (Abbott Laboratories), covered with a thin layer of oxygen-impermeable plastic polymer (Saran Wrap) to prevent diffusion of oxygen and other gases from the environment to the muscle, and warmed by using a heat lamp. The muscle preparation was left undisturbed for 30 min before a baseline measurement was made to allow any effect of the muscle isolation procedure to dissipate. Three hours of complete warm ischemia were achieved by clamping the main vascular pedicle of the isolated muscle by using a microvascular clamp (ST-B-1 VB, ASSI, Westbury, NY). After completion of the ischemic period, the clamp was removed, and the muscle underwent 90 min of reperfusion.

Measurement of blood flow.   See Refs. 34 and 48. Overall blood flow of the cremaster muscle was measured with laser Doppler flowmetry (MBF3, Moor Instruments, Devon, UK). A 1-mm-diameter double-fiber probe was positioned by a manipulator (MM 33, Stoelting, Wood Dale, IL) throughout the consecutive measurements in eight animals from each group. Via a small window on the covering plastic polymer, the tip of the probe was placed as close as possible to the surface of the vascular pedicle of the isolated muscle without causing a compressive effect on the vasculature. The position of the laser probe tip was kept constant throughout the experiment. During reperfusion, blood flow was recorded every 10 min for 90 min. Background flux detected on the vascular pedicle during ischemia was subtracted from measurements.

Measurement of vessel diameter.   See Refs. 34 and 48. Using intravital microscopy (Zeiss binocular microscope, model 473028) equipped with a video monitor (Sony monitor, model PVM-1343MD, a Sony Beta ED video cassette recorder, model EDW-30F), we measured vessel diameters from 10 to 70 µm in a selected arterial tree of each muscle (n = 8 in each group) during the reperfusion phase of the experiment. Ten to twenty vessel measurement sites within the vascular tree were selected in each muscle. Sequential measurements were taken at the same sites throughout the experiment. The internal luminal diameter of each selected vessel site was measured from a recorded image by using a FOR/A IV-560 video-measuring gauge. According to the baseline diameters, the observed vessels were divided into three categories: small arterioles (10–20 µm), large arterioles (21–40 µm), and small arteries (41–70 µm). During reperfusion, the vessel diameters were measured every 10 min for 90 min.

Wet weight ratio evaluation.   To determine the extent of I/R-induced tissue edema, the cremaster muscles from both sides were removed at the end of each experiment (n = 16 in each group), and their weights were measured. The differences between these two groups were expressed as a percentage of the weight of the opposite nontreated cremaster muscle.

Histology.   After weighing was completed, the left cremaster from each animal was immersed in 10% formaldehyde and then histologically examined by using a hematoxylin and eosin stain.

Determination of expressions of NOS protein.   See Ref. 34. After we completed measurement of the vessel diameters and blood flow, four muscle samples were harvested and frozen at –80°C immediately. The muscle samples were homogenized in boiling lysis buffer (1% SDS, 1.0 mM sodium orthovanadate, 10 mM Tris, pH 7.4) and microwaved for 10–15 s. The homogenate was centrifuged, and insoluble material was then discarded. Protein content was determined by using Coomassie Plus Protein Assay Reagent (Pierce, Rockford, IL). Proteins (30 µg) were loaded onto NuPAGE 4–12% bis-(2-hydroxyethyl)amino-tris(hydroxymethyl)methane gel (Invitrogen, Carlsbad, CA). After proteins were transferred onto polyvinylidene difluoride membrane, the membrane was blocked by 5% milk Tris-buffered saline-Tween 20 for 1 h, incubated at 4°C overnight with primary antibody (Ab) [monoclonal Ab for nNOS (1:1,500; BD Transduction, San Diego, CA) and eNOS (1:1,000; BD Transduction) and polyclonal Ab for iNOS (1:750, Upstate, Lake Placid, NY)] and then incubated with second Ab [goat anti-mouse IgG + IgM-horseradish peroxidase (1:5,000, Pierce) for nNOS and eNOS and goat anti-rabbit IgG-horseradish peroxidase (1:12,000, Santa Cruz BioTechnology, Santa Cruz, CA) for iNOS] for 1 h at room temperature. Immunoreactivity was detected with a SuperSignal West Pico Chemiluminescent Substrate detection kit (Pierce). Kodak film was used to visualize chemiluminescent signals. A mouse anti-actin monoclonal Ab (1:1,000, Chemicon, Temecula, CA) was used to detect actin as an internal control protein to demonstrate that all wells were loaded with equal amounts of cytoplasmic protein. The density was measured by Scion Image program.

Statistical analysis.   Vessel diameter for each vessel site and blood flow changes for each muscle at each time point were expressed as percentages of the baseline value. All values are expressed as means ± SE. Statistical analysis was performed by a repeated-measures two-way ANOVA. The post hoc analysis of specific values at each time point was performed by one-way ANOVA. A P value < 0.05 was considered significant. For the measurement of muscle wet weight ratio and NOS protein expressions, statistical analysis was performed by a one-way ANOVA. A P value of < 0.05 was considered statistically significant.


   RESULTS
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
DISCUSSION
 GRANTS
 REFERENCES
 
Measurement of blood flow.   At 10 min of reperfusion, the cremaster muscle blood flow was 32 ± 7.4% (mean ± SE) of baseline in the wild-type mice group and 80 ± 8.5% in the iNOS–/– group. Whereas the blood flow in the wild-type group gradually increased to a maximum of 60 ± 20% at 90 min, blood flow in the iNOS–/– group reached baseline level at 20 min and remained at that level throughout the experiment. Blood flow was significantly greater in the iNOS–/– group than in wild-type animals at all time points (P < 0.01 to <0.001) (Fig. 1).



View larger version (21K):
[in this window]
[in a new window]
  		Fig. 1. Blood flow in denervated cremaster muscle of inducible nitric oxide synthase-deficient mice (iNOS–/–) and wild-type mice (n = 8 for each) during 90-min reperfusion following 3 h of ischemia. Values are means ± SE. ** P < 0.01 and *** P < 0.001 compared with wild-type mice.

 
Measurement of vessel diameter.   The average baseline vessel diameters are summarized in Table 1, with no significant difference in each vessel size category between the iNOS–/– and wild-type groups. As shown in Fig. 2, the average vessel diameters in the wild-type group were between 39 and 59% of baseline at 10 min of reperfusion and increased to a maximum of 72 ± 4% in 10- to 20-µm, 71 ± 5% in 21- to 40-µm, and 64 ± 4% in 41- to 70-µm vessels at 90 min. In contrast, the diameters in the iNOS–/– group sharply increased to >70% of baseline in each vessel category at 10 min and reached maximum diameters of 92 ± 2.8, 88 ± 2.7, and 86 ± 2.3% at 90 min, with a significant (P < 0.05 to <0.001) difference at each time point.


View this table:
[in this window]
[in a new window]
  		Table 1. Baseline values of vessel diameters in the iNOS–/– and wild-type groups

 


View larger version (19K):
[in this window]
[in a new window]
  		Fig. 2. Vascular diameter changes in denervated cremaster muscles of iNOS–/– and wild-type mice (n = 8 for each) during 90-min reperfusion following 3 h of ischemia. Values are means ± SE. A: 10- to 20-µm arterioles. B: 21- to 40-µm arterioles. C: 41- to 70-µm small arteries. * P < 0.05, ** P < 0.01, and *** P < 0.001 compared with wild-type mice.

 
Wet weight ratio evaluation.   The weight ratio (percentage of normal) of the muscles was 173 ± 11% in the wild-type group and 117 ± 3% in the iNOS–/– group, with a significant difference (P < 0.01) between the two groups (Fig. 3).



View larger version (9K):
[in this window]
[in a new window]
  		Fig. 3. The wet weight ratio (means ± SE) of denervated cremaster muscles of iNOS–/– and wild-type mice (n = 16 for each) at the end of 90-min reperfusion. ** P < 0.01 compared with wild-type mice.

 
Histological evaluation.   There was normal skeletal muscle cell shape and no thrombosis in the lumen of vessels in both iNOS–/– and wild-type animals. However, there were reduced edema, muscle necrosis, and neutrophil extravasation in the iNOS–/– group compared with the wild-type group (Fig. 4).



View larger version (133K):
[in this window]
[in a new window]
  		Fig. 4. Photomicrograph of cross section of denervated cremaster muscles of wild-type (A) and iNOS–/– (B) mice at the end of 90-min reperfusion. Inflammatory reaction and edema are more severe in wild-type mice (A) than in iNOS–/– mice (B). Hematoxylin and eosin stain was used. Magnification: x400.

 
NOS protein expression.   The band of iNOS protein was not detected in normal and reperfused cremaster muscle in iNOS–/– mice, whereas iNOS protein expression was upregulated from normal in the reperfused muscle in wild mice (Fig. 5). For nNOS and eNOS proteins, the expression was downregulated to ~50–60% of normal in the reperfused cremaster muscle in both iNOS–/– and wild mice. There was no significant difference between the two groups (Fig. 5).



View larger version (49K):
[in this window]
[in a new window]
  		Fig. 5. Expression of neuronal NOS (nNOS), iNOS, and endothelial NOS (eNOS) protein in denervated rat cremaster muscle at the end of 90-min reperfusion. A: comparison of protein expression between the cremaster muscles of iNOS–/– and wild-type mice (n = 4 for each). Values are means ± SE. B: samples of 3 NOS protein bands. N, normal; P, positive control; I/R, ischemia-reperfusion.

 

   DISCUSSION
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
GRANTS
 REFERENCES
 
In a cremaster muscle model of I/R injury, iNOS–/– mice demonstrated significantly less vasospasm, complete restoration of blood flow, reduced muscle fiber necrosis on histological examination, and decreased edema, as indicated by wet weight ratio, compared with wild-type mice. Those alternations were accompanied by an undetectable band of iNOS protein in the muscle in iNOS–/– mice. Therefore, I/R injury is clearly reduced in iNOS–/– mice in which the iNOS gene has been selectively eliminated.

Our results have confirmed that iNOS is an important factor in the amplification of I/R injury in skeletal muscle, which was previously suggested by studies using the relatively selective iNOS inhibitor dexamethasone (7) or highly selective iNOS inhibitor 1400W, which significantly increased blood flow, reduced vasoconstriction and tissue edema, and preserved muscle contractile function (32, 34, 48). Although 1400W is a highly selective inhibitor of iNOS, there is an unavoidable redundancy in functional actions to nNOS and eNOS isoforms. In fact, 1400W has been shown to have weak inhibition for nNOS and eNOS (12), specifically under a condition of dramatic upregulation of nNOS and eNOS (32). Thus the exact role of iNOS in I/R injury requires further direct confirmation. In this regard, the present study has shown a reduced I/R injury in iNOS–/– mice, in which the iNOS gene is eliminated by state-of-the-art gene knockout technology. Because the control mice and iNOS–/– mice were of the same genetic background and have no known genotypic differences other than the absence of the iNOS gene, the deficiency of this enzyme appears responsible for the reduction of I/R injury in the cremaster muscle.

iNOS has been identified as a major source of NO generation in tissues. It has been reported that ~80% of the basal NO in the respiratory tract originates from iNOS and that 99% of NOS activity is attributable to iNOS after ovalbumin challenge (8). In skeletal muscle, most of the NO production during I/R injury may be generated by iNOS (30). Although the exact role of iNOS in I/R injury is controversial (23), high concentrations of NO produced by iNOS can affect many cellular functions simultaneously by changing ion currents (11), inhibiting cellular respiration (44) and various enzyme activities (45), and mediating DNA damage (24). iNOS appears to be responsible for nitrotyrosine formation, which is a nitration product of tyrosine and is a possible biomarker for cytotoxicity of NO in I/R injury (15). iNOS-mediated apoptosis appears to be through nitrosative mechanisms (10). Other reactive nitrogen species generated from iNOS, including peroxynitrite, may also contribute to I/R injury (43). Furthermore, iNOS may cause alterations in the expression or function of proteins involved in the regulation of tissue function (41). Thus activated iNOS leads to excess NO production, subsequently potentiating I/R injury and the resulting dysfunction and necrosis of reperfused tissues.

Our findings are in agreement with those of others who demonstrated that targeted disruption of the iNOS gene could protect tissues against ischemic and I/R injury. After 70-min tourniquet ischemia and 24-h reperfusion, 40% of muscle fiber necrosis was found in the gatrocnemius muscle in wild-type mice, but this severity of necrosis was seen in iNOS–/– mice only after 90-min ischemia (2). Improved ventricular contractile function and reduced apoptosis were demonstrated in iNOS–/– mice after myocardial infarction (35). Similarly, the infarct and the motor deficits produced by occlusion of the middle cerebral artery were smaller in iNOS–/– mice compared with those in wild-type mice (19). Attenuated renal I/R injury (26), increased resistance to mucosal injury and intestinal I/R-induced bacterial translocation (39), increased microvascular reactivity and improved survival in septic shock (17), diminished myocardial dysfunction in endotoxin-induced murine sepsis (41), and improved apoptosis in ischemic proliferative retinopathy (36) have also been reported in iNOS–/– mice compared with wild-type mice. Furthermore, the protective role in iNOS–/– mice exhibits a gene-dose effect in ischemic brain injury (49). Taken together, the data strongly support the conclusion that upregulated iNOS has detrimental effects on I/R injury to tissues.

Our results, however, are different from those of other studies in which flow, function, and infarct size were not significantly different in isolated, reperfused heart between wild-type and iNOS–/– mice (46). Moreover, evidence showed that NO generated from iNOS might contribute to protection of heart function in isolated mouse heart (13). Extensive myocardial injury associated with a higher mortality rate and neutrophil infiltration were found in the iNOS–/– mouse heart after I/R compared with wild-type mice (50). A similar conclusion was reported in liver I/R injury (14). It appears that different experimental conditions can affect the molecular reactivity of NO. These variables include differences in animal species, rate and timing of NO release, type of tissue, surgical skill, extent of tissue damage, and, consequently, the cellular environment of target molecules. It is noted that the denervated cremaster muscle underwent 3 h of ischemia in the present study and 20 or 60 min of ischemia in those myocardial studies.

Knockout mice are useful tools for studying the contribution of individual NOS isoform to specific pathophysiological processes and can provide many insights into the complex roles of NO (18, 31). Targeted gene disruption or transgenic overexpression will likely augment pharmacological approaches to define the roles of particular gene products in normal tissue function and in pathophysiology of disease processes (18). However, disruption of a single gene may have systemically significant effects, such as increased susceptibility to pathogens in iNOS–/– mice, in addition to those being studied (31). In addition, a potential adaptive mechanism between NOS genes should be considered. Although this adaptation was not found in the present study (Fig. 5), recent data from an ex vivo heart study showed a remarkable iNOS induction in I/R myocardium in eNOS–/– mice (21). Furthermore, the role of NO production generated from NOS-independent pathways (25) should also be considered when evaluating I/R injury in NOS–/– mice. Thus future work needs to include direct measurement of NO to determine the exact mechanism by which iNOS exerts its effect in I/R injury.

In summary, the present study shows that, compared with wild-type mice, I/R injury is reduced in iNOS–/– mice during early reperfusion. Combined with earlier results from inhibiting the iNOS isoform, our results conclusively demonstrate that iNOS is deleterious to I/R injury in skeletal muscle and that selective inhibition of iNOS protects tissues against I/R injury. Whereas our results have demonstrated that knockout mice are valuable tools for the study of the mechanisms of pathophysiological processes, one must be cognizant of unknown systemic effects and adaptive mechanisms related to the genetic manipulation of the knockout animals.


   GRANTS
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
REFERENCES
 
This study was supported by National Heart, Lung, and Blood Institute (NHLBI) Grant 5RO1 HL-36046 (to J. R. Urbaniak), a research grant from the Orthopaedic Research and Education Foundation (to L.-E. Chen), and NHLBI Grant HL-04053-05 (to J. P. Eu).


   FOOTNOTES
 

Address for reprint requests and other correspondence: L.-E. Chen, Orthopaedic Research Laboratory, Duke Univ. Medical Center, Box 3093, Durham, NC 27710 (E-mail: chen0006{at}mc.duke.edu).

The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.


   REFERENCES
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 

  1. Armstrong SC. Protein kinase activation and myocardial ischemia/reperfusion injury. Cardiovasc Res 61: 427–436, 2004.[Abstract/Free Full Text]
  2. Barker JE, Knight KR, Romeo R, Hurley JV, Morrison WA, and Stewart AG. Targeted disruption of the nitric oxide synthase 2 gene protects against ischaemia/reperfusion injury to skeletal muscle. J Pathol 194: 109–115, 2001.[CrossRef][Web of Science][Medline]
  3. Benjamim CF, Silva JS, Fortes ZB, Oliveira MA, Ferreira SH, and Cunha FQ. Inhibition of leukocyte rolling by nitric oxide during sepsis leads to reduced migration of active microbicidal neutrophils. Infect Immun 70: 3602–3610, 2002.[Abstract/Free Full Text]
  4. Blaisdell FW. The pathophysiology of skeletal muscle ischemia and the reperfusion syndrome: a review. Cardiovasc Surg 10: 620–630, 2002.[CrossRef][Web of Science][Medline]
  5. Bradford Sanders D, Hunter K, Wu Y, Jablonowski C, Bahl JJ, and Larson DF. Modulation of the inflammatory response in the cardiomyocyte and macrophage. J Extra Corpor Technol 33: 167–174, 2001.[Medline]
  6. Chen LE, Seaber AV, Nasser RM, Stamler JS, and Urbaniak JR. Effects of S-nitroso-N-acetylcysteine on contractile function of reperfused skeletal muscle. Am J Physiol Regul Integr Comp Physiol 274: R822–R829, 1998.[Abstract/Free Full Text]
  7. Chen LE, Silver WP, Seaber AV, Korompilias AV, and Urbaniak JR. Effects of dexamethasone on the contractile function of reperfused skeletal muscle. Microsurgery 17: 313–320, 1996.[CrossRef][Web of Science][Medline]
  8. De Sanctis GT, MacLean JA, Hamada K, Mehta S, Scott JA, Jiao A, Yandava CN, Kobzik L, Wolyniec WW, Fabian AJ, Venugopal CS, Grasemann H, Huang PL, and Drazen JM. Contribution of nitric oxide synthases 1, 2, and 3 to airway hyperresponsiveness and inflammation in a murine model of asthma. J Exp Med 189: 1621–1630, 1999.[Abstract/Free Full Text]
  9. El-Dwairi Q, Comtois A, Guo Y, and Hussain SN. Endotoxin-induced skeletal muscle contractile dysfunction: contribution of nitric oxide synthases. Am J Physiol Cell Physiol 274: C770–C779, 1998.[Abstract/Free Full Text]
  10. Eu JP, Liu L, Zeng M, and Stamler JS. An apoptotic model for nitrosative stress. Biochemistry 39: 1040–1047, 2000.[CrossRef][Medline]
  11. Eu JP, Xu L, Stamler JS, and Meissner G. Regulation of ryanodine receptors by reactive nitrogen species. Biochem Pharmacol 57: 1079–1084, 1999.[CrossRef][Web of Science][Medline]
  12. Garvey EP, Oplinger JA, Furfine ES, Kiff RJ, Laszlo F, Whittle BJ, and Knowles RG. 1400W is a slow, tight binding, and highly selective inhibitor of inducible nitric-oxide synthase in vitro and in vivo. J Biol Chem 272: 4959–4963, 1997.[Abstract/Free Full Text]
  13. Hampton TG, Amende I, Fong J, Laubach VE, Li J, Metais C, and Simons M. Basic FGF reduces stunning via a NOS2-dependent pathway in coronary-perfused mouse hearts. Am J Physiol Heart Circ Physiol 279: H260–H268, 2000.[Abstract/Free Full Text]
  14. Hines IN, Harada H, Bharwani S, Pavlick KP, Hoffman JM, and Grisham MB. Enhanced post-ischemic liver injury in iNOS-deficient mice: a cautionary note. Biochem Biophys Res Commun 284: 972–976, 2001.[CrossRef][Web of Science][Medline]
  15. Hirabayashi H, Takizawa S, Fukuyama N, Nakazawa H, and Shinohara Y. Nitrotyrosine generation via inducible nitric oxide synthase in vascular wall in focal ischemia-reperfusion. Brain Res 852: 319–325, 2000.[CrossRef][Web of Science][Medline]
  16. Hirose J, Yamaga M, Kato T, Ikebe K, and Takagi K. Effects of a hydroxyl radical scavenger, EPC-K1, and neutrophil depletion on reperfusion injury in rat skeletal muscle. Acta Orthop Scand 72: 404–410, 2001.[CrossRef][Web of Science][Medline]
  17. Hollenberg SM, Broussard M, Osman J, and Parrillo JE. Increased microvascular reactivity and improved mortality in septic mice lacking inducible nitric oxide synthase. Circ Res 86: 774–778, 2000.[Abstract/Free Full Text]
  18. Huang PL. Lessons learned from nitric oxide synthase knockout animals. Semin Perinatol 24: 87–90, 2000.[CrossRef][Web of Science][Medline]
  19. Iadecola C, Zhang F, Casey R, Nagayama M, and Ross ME. Delayed reduction of ischemic brain injury and neurological deficits in mice lacking the inducible nitric oxide synthase gene. J Neurosci 17: 9157–9164, 1997.[Abstract/Free Full Text]
  20. Joneschild ES, Chen LE, Seaber AV, Frankel ES, and Urbaniak JR. Effect of a NOS inhibitor, L-NMMA, on the contractile function of reperfused skeletal muscle. J Reconstr Microsurg 15: 55–60, 1999.[Web of Science][Medline]
  21. Kanno S, Lee PC, Zhang Y, Ho C, Griffith BP, Shears LL 2nd, and Billiar TR. Attenuation of myocardial ischemia/reperfusion injury by superinduction of inducible nitric oxide synthase. Circulation 101: 2742–2748, 2000.[Abstract/Free Full Text]
  22. Kobzik L, Stringer B, Balligand JL, Reid MB, and Stamler JS. Endothelial type nitric oxide synthase in skeletal muscle fibers: mitochondrial relationships. Biochem Biophys Res Commun 211: 375–381, 1995.[CrossRef][Web of Science][Medline]
  23. Kroncke KD, Fehsel K, Suschek C, and Kolb-Bachofen V. Inducible nitric oxide synthase-derived nitric oxide in gene regulation, cell death and cell survival. Int Immunopharmacol 1: 1407–1420, 2001.[CrossRef][Web of Science][Medline]
  24. Kwon NS, Stuehr DJ, and Nathan CF. Inhibition of tumor cell ribonucleotide reductase by macrophage-derived nitric oxide. J Exp Med 174: 761–767, 1991.[Abstract/Free Full Text]
  25. Lepore DA, Kozlov AV, Stewart AG, Hurley JV, Morrison WA, and Tomasi A. Nitric oxide synthase-independent generation of nitric oxide in rat skeletal muscle ischemia-reperfusion injury. Nitric Oxide 3: 75–84, 1999.[CrossRef][Web of Science][Medline]
  26. Ling H, Edelstein C, Gengaro P, Meng X, Lucia S, Knotek M, Wangsiripaisan A, Shi Y, and Schrier R. Attenuation of renal ischemia-reperfusion injury in inducible nitric oxide synthase knockout mice. Am J Physiol Renal Physiol 277: F383–F390, 1999.[Abstract/Free Full Text]
  27. Liu K, Chen LE, Seaber AV, and Urbaniak JR. S-nitroso-N-acetylcysteine protects skeletal muscle against reperfusion injury. Microsurgery 18: 299–305, 1998.[CrossRef][Web of Science][Medline]
  28. Lowenstein CJ, Dinerman JL, and Snyder SH. Nitric oxide: a physiologic messenger. Ann Intern Med 120: 227–237, 1994.[Abstract/Free Full Text]
  29. May JW Jr, Chait LA, O'Brien BM, and Hurley JV. The no-reflow phenomenon in experimental free flaps. Plast Reconstr Surg 61: 256–267, 1978.[Web of Science][Medline]
  30. Messina A, Knight KR, Dowsing BJ, Zhang B, Phan LH, Hurley JV, Morrison WA, and Stewart AG. Localization of inducible nitric oxide synthase to mast cells during ischemia/reperfusion injury of skeletal muscle. Lab Invest 80: 423–431, 2000.[Web of Science][Medline]
  31. Ortiz PA and Garvin JL. Cardiovascular and renal control in NOS-deficient mouse models. Am J Physiol Regul Integr Comp Physiol 284: R628–R638, 2003.[Abstract/Free Full Text]
  32. Patel P, Qi WN, Allen DM, Chen LE, Seaber AV, Stamler JS, and Urbaniak JR. Inhibition of iNOS with 1400W improves contractile function and alters NOS gene and protein expression in reperfused skeletal muscle. Microsurgery 24: 324–331, 2004.
  33. Phan LH, Hickey MJ, Niazi ZB, and Stewart AG. Nitric oxide synthase inhibitor, nitro-iminoethyl-L-ornithine, reduces ischemia-reperfusion injury in rabbit skeletal muscle. Microsurgery 15: 703–707, 1994.[Web of Science][Medline]
  34. Qi WN, Zhang L, Chen LE, Seaber AV, and Urbaniak JR. Inhibition of iNOS improves microcirculation in denervated cremaster muscle during early reperfusion. J Hand Surg [Am] 29A: 638–645, 2004.
  35. Sam F, Sawyer DB, Xie Z, Chang DL, Ngoy S, Brenner DA, Siwik DA, Singh K, Apstein CS, and Colucci WS. Mice lacking inducible nitric oxide synthase have improved left ventricular contractile function and reduced apoptotic cell death late after myocardial infarction. Circ Res 89: 351–356, 2001.[Abstract/Free Full Text]
  36. Sennlaub F, Courtois Y, and Goureau O. Inducible nitric oxide synthase mediates retinal apoptosis in ischemic proliferative retinopathy. J Neurosci 22: 3987–3993, 2002.[Abstract/Free Full Text]
  37. Stamler JS and Meissner G. Physiology of nitric oxide in skeletal muscle. Physiol Rev 81: 209–237, 2001.[Abstract/Free Full Text]
  38. Stangl V, Baumann G, Stangl K, and Felix SB. Negative inotropic mediators released from the heart after myocardial ischaemia-reperfusion. Cardiovasc Res 53: 12–30, 2002.[Abstract/Free Full Text]
  39. Suzuki Y, Deitch EA, Mishima S, Lu Q, and Xu D. Inducible nitric oxide synthase gene knockout mice have increased resistance to gut injury and bacterial translocation after an intestinal ischemia-reperfusion injury. Crit Care Med 28: 3692–3696, 2000.[CrossRef][Web of Science][Medline]
  40. Thompson M, Becker L, Bryant D, Williams G, Levin D, Margraf L, and Giroir BP. Expression of the inducible nitric oxide synthase gene in diaphragm and skeletal muscle. J Appl Physiol 81: 2415–2420, 1996.[Abstract/Free Full Text]
  41. Ullrich R, Scherrer-Crosbie M, Bloch KD, Ichinose F, Nakajima H, Picard MH, Zapol WM, and Quezado ZM. Congenital deficiency of nitric oxide synthase 2 protects against endotoxin-induced myocardial dysfunction in mice. Circulation 102: 1440–1446, 2000.[Abstract/Free Full Text]
  42. Urbaniak JR, Seaber AV, and Chen LE. Assessment of ischemia and reperfusion injury. Clin Orthop Jan: 30–36, 1997.
  43. Wang P and Zweier JL. Measurement of nitric oxide and peroxynitrite generation in the postischemic heart. Evidence for peroxynitrite-mediated reperfusion injury. J Biol Chem 271: 29223–29230, 1996.[Abstract/Free Full Text]
  44. Welter R, Yu L, and Yu CA. The effects of nitric oxide on electron transport complexes. Arch Biochem Biophys 331: 9–14, 1996.[CrossRef][Web of Science][Medline]
  45. Wolosker H, Panizzutti R, and Engelender S. Inhibition of creatine kinase by S-nitrosoglutathione. FEBS Lett 392: 274–276, 1996.[CrossRef][Web of Science][Medline]
  46. Xi L, Jarrett NC, Hess ML, and Kukreja RC. Myocardial ischemia/reperfusion injury in the inducible nitric oxide synthase knockout mice. Life Sci 65: 935–945, 1999.[CrossRef][Web of Science][Medline]
  47. Yan ZQ, Bolognesi MP, Steeber DA, Tedder TF, Chen LE, Seaber AV, and Urbaniak JR. Blockade of L-selectin attenuates reperfusion injury in a rat model. J Reconstr Microsurg 16: 227–233, 2000.[CrossRef][Web of Science][Medline]
  48. Zhang L, Looney CG, Qi WN, Chen LE, Seaber AV, Stamler JS, and Urbaniak JR. Reperfusion injury is reduced in skeletal muscle by inhibition of inducible nitric oxide synthase. J Appl Physiol 94: 1473–1478, 2003.[Abstract/Free Full Text]
  49. Zhao X, Haensel C, Araki E, Ross ME, and Iadecola C. Gene-dosing effect and persistence of reduction in ischemic brain injury in mice lacking inducible nitric oxide synthase. Brain Res 872: 215–218, 2000.[CrossRef][Web of Science][Medline]
  50. Zingarelli B, Hake PW, Yang Z, O'Connor M, Denenberg A, and Wong HR. Absence of inducible nitric oxide synthase modulates early reperfusion- induced NF-{kappa}B and AP-1 activation and enhances myocardial damage. FASEB J 16: 327–342, 2002.[Abstract/Free Full Text]




This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF) Free
Right arrow All Versions of this Article:
97/4/1323    most recent
00380.2004v1
Right arrow Submit a response
Right arrow Alert me when this article is cited
Right arrow Alert me when eLetters are posted
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via Web of Science (8)
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Qi, W.-N.
Right arrow Articles by Urbaniak, J. R.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Qi, W.-N.
Right arrow Articles by Urbaniak, J. R.

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Visit Other APS Journals Online
Copyright © 2004 by the American Physiological Society.