Use of L-arginine in the treatment of experimental necrotizing enterocolitis.
Although multifactorial in etiology, prematurity and feeding are two of the most common risk factors associated with necrotizing enterocolitis (NEC). To understand the pathogenesis of NEC, the complex interaction between intestinal contents and clearing mechanisms in the immature human gut must be elucidated. Nitric oxide (NO) is a proposed mediator of nonadrenergic noncholinergic neural inhibition, causing relaxation in the gut. In addition to its role as a neuroeffector substance, studies suggest that endogenous formation of NO maintains intestinal mucosal integrity, protecting the gut from blood-borne toxins and tissue-destructive mediators. Thus, NO has a dual role in both gut smooth muscle relaxation and mucosal protection. Because two of the primary risk factors in the development of NEC are prematurity (as it relates to gut dysmotility) and enteral feeding (as it relates to mucosal damage by intraluminal substrate), the authors chose to investigate the role of NO in the pathogenesis of NEC induced by intraluminal injection of acidified casein solution in neonatal piglets.


METHODS
Having confirmed the consistent induction of NEC both macroscopically and histologically with this model (n = 32), the following were undertaken. Neonatal piglets (< 3 days old) were laparotomized, and intestinal loops were created from the terminal ileum to the proximal colon. The loops were injected with acidified casein solution and separated by saline-injected control loops. When the abdomen was closed, a continuous peripheral intravenous infusion of L-arginine, an NO synthase substrate (600 mg/kg/h [n = 6]), or N-omega-nitro-L-arginine methyl ester (L-NAME), an NO synthase inhibitor (20 mg/kg/h [n = 6]), was begun. Gut segments were harvested 3 hours later and processed for evaluation of the extent of necrosis.


RESULTS
Macroscopically, the L-NAME-treated group showed areas of hemorrhagic necrosis in the NEC-induced loops. The L-arginine-treated group had greatly diminished or virtually absent lesions. H&E-stained sections were graded microscopically, using a scale from 0 to 4, ranging from intact villi (grade 0) to transmural necrosis (grade 4). In the untreated NEC group, intestinal damage in the acidified casein loops was exhibited by areas of necrosis (extending, in some cases, transmurally), submucosal edema, and inflammatory cell infiltrate (average grade, 3.5). In the L-NAME-treated group, the intestinal damage was similar to that of the NEC-induced group (average grade, 3.5), but also presented with marked hemorrhagic congestion. In the L-arginine group, NEC-induced tissue damage was greatly attenuated, with necrosis limited primarily to the villus tips (average grade, 1). Nevertheless, inflammatory cell infiltrate and mild submucosal edema were still present.


CONCLUSIONS
Continuous intravenous infusion with the NO synthase substrate L-arginine markedly attenuates intestinal injury in this neonatal piglet model of NEC. Intravenous administration of the NO synthase inhibitor L-NAME causes hemorrhagic congestion of the gut wall. Based on these findings, the authors propose that treatment with the amino acid L-arginine should be considered as a potential therapeutic modality for NEC.
Di Lorenzo
M
M
Department of Physiology, University of Ottawa, Ontario, Canada.
Bass
J
J
Krantis
A
A
D000594:Amino Acid Oxidoreductases / Q000378:metabolism; D000818:Animals; D001120:Arginine / Q000627:therapeutic use*; D002364:Caseins; D004195:Disease Models, Animal; D004761:Enterocolitis, Pseudomembranous / Q000139:chemically induced / Q000503:physiopathology / Q000628:therapy*; D005769:Gastrointestinal Motility; D007262:Infusions, Intravenous; D007413:Intestinal Mucosa / Q000503:physiopathology; D019001:Nitric Oxide Synthase; D013552:Swine
D002364:Caseins; D001120:Arginine; D019001:Nitric Oxide Synthase; D000594:Amino Acid Oxidoreductases