Background and Purpose
Germinal matrix hemorrhage-intraventricular hemorrhage (GMH-IVH) is the most common neurological problem of premature infants. Despite this, mechanisms of brain injury from IVH are elusive. We hypothesized that GMH-IVH, by induction of NAD(P)H oxidases, might cause oxidative/nitrosative stress contributing to brain injuries and that NAD(P)H oxidase inhibition could offer neuroprotection .
To test this hypothesis, we exploited our rabbit pup model of glycerol-induced GMH-IVH. We delivered rabbit pups prematurely (E29) by C-section and administered intra-peritoneal glycerol at 2h postnatal age. Free-radical adducts, including nitrotyrosine, 4-hyroxynonenal and 8-Hydroxy-deoxyguanosine [8-OHdG] as well as O2·− and H2O2 levels were measured in the forebrain. To determine the source of free-radical generation, we used inhibitors for NAD(P)H oxidase (apocynin), xanthine oxidase (allopurinol), COX-2 (indomethacin) or NO synthases (L-NAME). IVH pups were treated with apocynin and cell death was compared between apocynin-treated and vehicle-treated pups.
Nitrotyrosine, 4-hyroxynonenal and 8-OHdG levels were higher in pups with IVH than controls. Likewise, O2·− and H2O2 levels were significantly greater in both the periventricular area and cerebral cortex of pups with IVH than controls. In pups with IVH, reactive oxygen species (ROS) production was more in the periventricular area than in cortex. Apocynin, but not allopurinol, indomethacin or L-NAME, inhibited ROS generation. Importantly, apocynin reduced cell death in pups with IVH.
Activation of NAD(P)H oxidase was the predominant mechanism of free-radical generation in pups with IVH. NAD(P)H oxidase inhibition by apocynin might suppress ROS production and confer neuroprotection in premature infants with IVH.