Further evidence for the role of free radicals in the limb teratogenicity of L-NAME.
BACKGROUND
L-NAME (N(G)-nitro-(L)-arginine methyl ester), a nitric oxide synthase inhibitor, causes severe limb reduction malformations when gravid rats are treated intraperitoneally on gd-17. Hemorrhages, appearing within hours of L-NAME administration, and defects at term can be significantly reduced by co-treatment with PBN (alpha-phenyl-N-t-butylnitrone), a spin trap antioxidant. We have proposed that limb defects result from ischemia-reperfusion injury. We examine the role of xanthine oxidase and ROS formation in the limb effects of L-NAME.


METHODS
Gravidas were treated with L-NAME (50 mg/kg) in the presence or absence of allopurinol, a xanthine oxidase inhibitor. Spatial patterns of limb hemorrhage were determined promptly and at term as was digit length at the latter interval. Xanthine oxidase activities were assayed in control and treated limbs with and without allopurinol co-treatment.


RESULTS
Allopurinol significantly reduced hemorrhage severity in a dose-responsive fashion when fetuses were examined at term. Higher doses of allopurinol significantly preserved digit length. Xanthine oxidase activities in fetal limb were significantly increased by L-NAME treatment whereas co-treatment with allopurinol restored activities to near-control levels.


CONCLUSIONS
These findings support the role of excess reactive oxygen species (ROS) formation in L-NAME-induced limb reduction. We propose that nitric oxide (NO) depletion by L-NAME interferes with vascular integrity, and causes vasoconstriction. Resultant hypoxia stimulates superoxide formation and nitric oxide formation catalyzed by the inducible isoform of nitric oxide synthase. The reduction products of superoxide or the products of its reaction with nitric oxide oxidize or nitrate endothelial components resulting in limb reduction defects.
Birth Defects Laboratory, Division of Genetics and Development, Department of Pediatrics, University of Washington, Seattle, Washington 98195-6320, USA. agf@u.washington.edu
Alan G
AG
Fantel
Richard E
RE
Person
D000493:Allopurinol / Q000494:pharmacology; D000818:Animals; D005609:Free Radicals; D006470:Hemorrhage / Q000139:chemically induced / Q000196:embryology; D017880:Limb Deformities, Congenital / Q000139:chemically induced*; D019331:NG-Nitroarginine Methyl Ester / Q000633:toxicity*; D017942:Nitric Acid / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D013723:Teratogens / Q000633:toxicity*; D014969:Xanthine Oxidase / Q000037:antagonists & inhibitors / Q000378:metabolism
D005609:Free Radicals; D013723:Teratogens; D017942:Nitric Acid; D000493:Allopurinol; D014969:Xanthine Oxidase; D019331:NG-Nitroarginine Methyl Ester