Peroxynitrite is an important mediator in thermal injury-induced lung damage.
OBJECTIVE
Intestinal ischemia and reperfusion injury was known to cause postinjury multiple organ failure by neutrophil and unclear nonneutrophil factors. Peroxynitrite formed by the rapid reaction between superoxide and nitric oxide, is a toxic substance that contributes to tissue injury in a number of biological systems. In this study, the role of nitric oxide and neutrophils on lung damage after burn was investigated.


METHODS
Prospective, experimental study.


METHODS
Research laboratory at a university hospital.


METHODS
Thermal injury models in the rat.


METHODS
In experiment 1, specific pathogen-free Sprague-Dawley rats underwent 35% total body surface area burn. At 4, 8, 16, and 24 hrs after burn, intestinal mucosa and lung tissue were harvested for myeloperoxidase (MPO) assay, blood was collected for measurement of peroxynitrite-mediated oxidation of dihydrorhodamine 123, and pulmonary microvascular dysfunction was quantified by measuring the extravasation of Evans blue dye. In experiment 2, polymorphonuclear granulocyte antibody (0.12 mL/100 g administered intraperitoneally 16 hrs before burn), S-methylisothiourea (7.5 mg/kg, intraperitoneally, immediately after burn), a specific inducible nitric oxide synthase inhibitor, and sterile saline (15 mL/kg, intraperitoneally, immediately after burn) were given to different groups of thermally injured animals individually. The plasma dihydrorhodamine 123 oxidation level, intestinal and lung MPO activity, lung permeability, and lung histology were evaluated at 8 hrs after burn. The cellular localization of nitrotyrosine, a marker for peroxynitrite reactivity, was also examined by immunostaining. In experiment 3, 3-morpholinosydnonimine (10 mM, intraperitoneally), a peroxynitrite donor, was given to nonburned rats to examine the peroxynitrite effect on lung inducible nitric oxide synthase expression.


RESULTS
The level of MPO activity in intestine and lung, blood dihydrorhodamine 123 oxidation, and lung permeability were increased up to 2-fold, 2.5-fold, 2-fold, and 2-fold of normal, respectively, at 8 hrs after burn. S-methylisothiourea injection significantly decreased (p <.05) 31% of the lung MPO activity, 41% of the blood peroxynitrite level, 54% of the lung permeability, and the lung peroxynitrite production in burned rats. Polymorphonuclear granulocyte antibody pretreatment significantly decreased 60% of the intestinal MPO, 92% of the blood peroxynitrite level, and 56% the lung MPO activity in burned rats, but the lung permeability was only slightly decreased by polymorphonuclear granulocyte antibody pretreatment. Furthermore, 3-morpholinosydnonimine increased the lung inducible nitric oxide synthase messenger RNA levels.


CONCLUSIONS
Thermal injury induces blood dihydrorhodamine 123 oxidation, intestinal and lung neutrophil deposition, lung nitrotyrosine production, and lung damage. Both specific inhibition of inducible nitric oxide synthase and polymorphonuclear granulocyte antibody pretreatment decrease blood dihydrorhodamine 123 oxidation and intestinal and lung neutrophil deposition, but only inducible nitric oxide synthase inhibition with S-methylisothiourea reduces lung peroxynitrite production and thermal injury-induced lung damage. Nitric oxide and the ensuing peroxynitrite production in lung play a more important role than neutrophil in contributing to thermal injury-induced lung damage.
Chen
Lee-Wei
LW
Department of Surgery, Kaohsiung Veterans General Hospital, National Yang-Ming Medical University, Taipei, Taiwan.
Wang
Jyh-Seng
JS
Chen
Hua-Lin
HL
Chen
Jin-Shyr
JS
Hsu
Ching-Mei
CM
D000818:Animals; D001830:Body Surface Area; D002056:Burns / Q000150:complications / Q000503:physiopathology*; D017931:DNA Primers; D005260:Female; D007150:Immunohistochemistry; D007413:Intestinal Mucosa / Q000378:metabolism; D008297:Male; D009504:Neutrophils / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism; D030421:Peroxynitrous Acid / Q000097:blood; D016133:Polymerase Chain Reaction; D011652:Pulmonary Circulation; D011897:Random Allocation; D051381:Rats; D017207:Rats, Sprague-Dawley; D012128:Respiratory Distress Syndrome / Q000378:metabolism*; D012235:Rhodamines / Q000097:blood; D013047:Specific Pathogen-Free Organisms
D017931:DNA Primers; D012235:Rhodamines; C058319:dihydrorhodamine 123; D030421:Peroxynitrous Acid; D009569:Nitric Oxide