Cyclooxygenase-2 mediates the sensitizing effects of systemic IL-1-beta on excitotoxic brain lesions in newborn mice
Introduction
Despite a reduction in neonatal mortality and morbidity in the last 40 years, rates of cerebral palsy (CP) remain significant in Western countries (Hagberg et al., 1996, Himmelmann et al., 2005). Recently hypothesized etiologies of CP have gone beyond hypoxic–ischemic mechanisms to include multiple preconceptional and prenatal factors such as hypoxia/perfusion failure, genetic components, growth-factor deficiency, and maternal infection and inflammation leading to the production of excess cytokines (Nelson and Willoughby, 2000, Volpe, 2001, Dammann et al., 2002, Gressens et al., 2002).
The potential deleterious role of perinatal inflammation has been proposed for both preterm and full term neonates at risk for the development of brain lesions and CP (Murphy et al., 1995, Zupan et al., 1996, Nelson and Willoughby, 2000, Volpe, 2001, Dammann et al., 2002, Gressens et al., 2002). According to the mechanism hypothesized, in utero infection and/or inflammation induce a transplacental inflammatory response associated with the production of excess circulating cytokines capable of harming the developing brain. Findings from several studies support an association between maternal–fetal infection, circulating cytokines (such as interleukin or IL-1-beta, IL-6, and tumor necrosis factor or TNF-alpha), and periventricular white matter damage (PWMD) in preterm infants (Romero et al., 1990, Greig et al., 1993, Singh et al., 1996, Yoon et al., 1996, Yoon et al., 1997, Martinez et al., 1998). In addition, a striking association between increased levels of perinatal circulating cytokines, including IL-1-beta, IL-6, IL-8, IL-9, and TNF-alpha, and the subsequent occurrence of CP in full-term infants has been reported (Nelson et al., 1998).
Using a murine model of neonatal excitotoxic brain lesions based on the intracerebral administration of ibotenate, a glutamate analog acting on N-methyl-d-aspartate (NMDA) and metabotropic receptors, we have previously shown that pups pretreated with IL-1-beta, IL-6, or TNF-alpha develop significantly greater ibotenate-induced cortical and white matter damage than controls (Dommergues et al., 2000). The precise molecular mechanisms by which circulating mediators of inflammation have a deleterious effect on perinatal brain lesions remain a matter for debate (Hagberg and Mallard, 2005). Circulating cytokines do not seem to cross the intact blood–brain barrier (BBB). However, three alternative pathways have been proposed to link serum cytokine levels with brain lesion sizes. Firstly, circulating cytokines could alter the permeability of the BBB to inflammatory mediators and cells. Secondly, circulating cytokines could act directly on parts of the brain lacking the BBB such as the circumventricular organs, meninges and choroid plexus or, as demonstrated in the adult brain, indirectly through the activation of the vagal nerve. Thirdly, cytokine effects could be mediated by cyclooxygenase (Cox) located on the BBB. In particular, cytokines could activate the inducible isoform Cox-2 to enhance the local production of prostaglandin E2 (PGE2) that could have deleterious effects on the developing brain. Some of these deleterious effects could involve an autocrine/paracrine loop leading to the excess production of inflammatory cytokines by brain cells.
In the present study, using the murine model of neonatal excitotoxic brain lesions described above, we explored the mechanisms by which systemically injected inflammatory cytokines sensitize the developing brain. We investigated the impact of systemically administered IL-1-beta on the production of cytokines by brain cells, focusing on the potential role of Cox-2 in this process. To do this, we used indomethacin and nimesulide, two non-steroidal anti-inflammatory drugs (NSAIDs) that inhibit Cox activity. Our data support the previously reported epidemiological association between high levels of circulating pro-inflammatory cytokines and an elevated risk of developing CP and/or PWMD.
Section snippets
Materials and methods
Experimental protocols were approved by the institutional review committee and meet Inserm guidelines as well as the Guide for the Care and use of Laboratory Animals as promulgated and adopted by the National Institutes of Health, USA.
Effects of IL-1-beta on ibotenate-induced lesions: reversal by Cox inhibitors
Control pups injected i.p. with vehicle between P1 and P5 and intracerebrally with ibotenate on P5 developed cortical lesions and periventricular white matter cysts (Figs. 1A and D). The cortical lesion was typical of ibotenate-induced lesions, with severe neuronal loss in all neocortical layers and the almost complete disappearance of neuronal cell bodies along the axis of ibotenate injection. There was no significant effect of gender on the size of the lesions (Fig. 1D).
IL-1-beta (40 ng)
Discussion
The most salient finding of this study is that the deleterious effects of pre-treatment with systemic IL-1-beta on excitotoxic brain lesions in newborn mice were totally abolished by the co-administration of nimesulide or indomethacin, two Cox inhibitors. In addition, the IL-1-beta-induced increase in the production of brain PGE2 was totally abolished by the Cox inhibitor indomethacin. The blockade of Cox activity also reversed some of the effects of the systemic administration of IL-1-beta on
Conclusion
Our data support the hypothesis that, in newborn mice, the sensitizing effects of circulating inflammatory cytokines on excitotoxic brain lesions are mediated by the Cox located on the BBB. In particular, cytokines activate the inducible isoform Cox-2, resulting in the enhanced local production of PGE2 and other derivatives that exacerbate excitotoxic brain lesions. Some of these deleterious effects could involve an autocrine/paracrine loop leading to a disruption in the balance between
Acknowledgments
We thank Marc Laburthe’s laboratory for their help with prostaglandin measurement. This work was supported by the INSERM, Université Paris 7, the Fondation pour la Recherche Médicale, and the Fondation Grace de Monaco. This article was prepared with editorial help from Gap Junction, www.gap-junction.com.
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