Necrotizing enterocolitis (NEC) is the most frequent and the most lethal disease that affects the gastrointestinal tract of the premature infant. The exact etiology of the disease is undefined. The only consistent identifiable epidemiological precursors for NEC are prematurity and enteral alimentation. We have previously shown upregulation of inducible nitric oxide (NO) synthase (NOS-2) mRNA and protein in intestinal segments from infants with acute NEC. NOS-2 colocalized with enterocyte apoptosis and nitrotyrosine immunoreactivity. NOS-2 was downregulated at the time of intestinal stoma closure when the acute inflammation had subsided. We have developed a reproducible model of gut inflammation in neonatal rats thereby simulating the conditions associated with human NEC. We simply formula-feed hypoxic neonatal rats thereby simulating the conditions associated with human NEC. These pups show NOS-2 mRNA upregulation, nitrosative stress, increased enterocyte apoptosis and decreased enterocyte proliferation in the crypts. Intraepithelial lymphocytes (IEL)from hypoxic formula-fed rats secrete more TNF-alpha and IFN- gamma compared to breast-fed rats. In vitro studies suggest that co- culture of IEL with the rat intestinal epithelial cell line IEC-18, in the presence of IL-1beta induces IFN-gamma, TNF-alpha and NOS-2 production which can be abrogated with antibody to IFN-gamma. Furthermore, peroxynitrite (ONOO) induces apoptosis and inhibits proliferation of IEC-6 cells. The data suggest that intestinal necrosis in NEC may be the result of NO-induced imbalance between tissue injury and repair mechanisms. Mucosal injury resulting from perinatal insults leads to bacterial-epithelial interactions, local release of cytokines such as IFN-gamma and TNF-alpha by IEL and lamina propria (LP) lymphocytes. These mediators induce NOS-2 upregulation with production of NO and ONOO by enterocytes or LP macrophages. NO or ONOO in turn promotes further tissue injury (enterocyte apoptosis) and concurrent inhibition of tissue repair mechanisms (enterocyte proliferation) leading to gut barrier failure and NEC. To test our hypothesis, we propose the following Aims: I) To define the mechanism of NOS-2 upregulation in human and experimental rodent NEC. II) To define the mechanisms by which NOS-2 upregulation promotes intestinal injury and alters tissue repair mechanisms in rodent NEC. III) To determine the effects of scavengers of NO or inhibitors of NO production on the development of experimental rodent NEC.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
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Special Emphasis Panel (ZRG1-SSS-W (39))
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Rothermel, Annette L
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Children's Hospital of Los Angeles
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