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(Stroke. 2008;39:2129.)
© 2008 American Heart Association, Inc.

Original Contributions

Strong Neuroprotection by Inhibition of NF-B After Neonatal Hypoxia-Ischemia Involves Apoptotic Mechanisms but Is Independent of Cytokines

Cora H.A. Nijboer, MSc; Cobi J. Heijnen, PhD; Floris Groenendaal, MD, PhD; Michael J. May, PhD; Frank van Bel, MD, PhD Annemieke Kavelaars, PhD

From the Laboratory of Psychoneuroimmunology (C.H.A.N., C.J.H., A.K.) and the Department of Neonatology (C.H.A.N., F.G., F.v.B.), University Medical Center Utrecht, The Netherlands; and the Department of Animal Biology (M.J.M.), University of Pennsylvania, School of Veterinary Medicine, Philadelphia.

Correspondence to Cobi J. Heijnen, University Medical Center Utrecht, Laboratory of Psychoneuroimmunology, Lundlaan 6, KC 03.068.0, 3584 EA Utrecht, The Netherlands. E-mail C.Heijnen{at}umcutrecht.nl

Background and Purpose— Interactions between excitotoxic, inflammatory, and apoptotic pathways determine outcome in hypoxic-ischemic brain damage. The transcription factor NF-B has been suggested to enhance brain damage via stimulation of cytokine production. There is also evidence that NF-B activity is required for neuronal survival. We used the NF-B inhibitor NBD, coupled to TAT to facilitate cerebral uptake, to determine the neuroprotective capacity of NF-B inhibition in neonatal hypoxia-ischemia (HI) and to identify its contribution to cerebral inflammation and damage.

Methods— Brain damage was induced in neonatal rats by unilateral carotid artery occlusion and hypoxia and analyzed immunohistochemically; NF-B activity was analyzed by EMSA. We analyzed cytokine mRNA levels and activation of apoptotic pathways by Western blotting. In vitro effects of TAT-NBD were determined in a neuronal cell line.

Results— Inhibition of cerebral NF-B activity by TAT-NBD had a significant neuroprotective effect; brain damage was reduced by more than 80% with a therapeutic window of at least 6 hours. In contrast to earlier suggestions, the protective effect of TAT-NBD did not involve suppression of early cytokine upregulation after HI. Moreover, NF-B inhibition prevented HI-induced upregulation and nuclear as well as mitochondrial accumulation of p53, prevented mitochondrial cytochrome-c release and activation of caspase-3. Finally, TAT-NBD could directly increase neuronal survival because TAT-NBD was sufficient to inhibit death in a neuronal cell line. A nonactive mutant peptide did not have any effect.

Conclusions— Inhibition of NF-B has strong neuroprotective effects that involve downregulation of apoptotic molecules but are independent of inhibition of cytokine production.

Key Words: nuclear factor–kappa-B • inflammation • neonatal • neuroprotection