NEONATAL SOCIETY ABSTRACTS
The sensitising effects of inflammation on hypoxic-ischaemic brain injury
Presented at the Neonatal Society 2005 Summer Meeting (programme).
Kendall G, Clements A, Varley H, Peebles D, Raivich G
Centre for Perinatal Brain Protection and Repair, Department of Obstetrics and Gynaecology, University College London, London WC1E 6HX, UK
Introduction: The fetal brain is protected from the effects of hypoxia-ischaemia (HI) by haemodynamic and metabolic compensatory mechanisms, so that pure HI is a relatively unusual cause of perinatal brain injury. A growing body of epidemiological and experimental animal data suggests that infection may sensitise the developing brain to mild HI1, reducing the threshold at which HI leads to brain injury. The aims of this study were to investigate the interaction between a bacterial product (endotoxin) mediated inflammation, and hypoxia-ischaemia on neuronal cell death and glial activation in a mouse model of neonatal HI.
Methods: Animal procedures were approved by the Home Office, and were carried out according to the UK Animals (Scientific Procedures) Act 1986. Experiments were performed on C57/Bl6 mice at postnatal day 7. An inflammatory reaction was provoked by i.p. injection of 0.3μg/g endotoxin (LPS) from E.coli, 055:B5. Mild HI was induced by left-sided carotid occlusion under isoflurane anaesthesia followed by exposure to 8% oxygen for 30 minutes. These insults were performed independently and in combination. Histological brain injury after HI was assessed at 48 hours by measuring infarct volume (expressed as a percentage of the contralateral hemisphere). The effect of LPS was assessed after 12 and 48h using histology, TUNEL and immunohistochemistry for microglial (M2-integrin) & astrocyte (GFAP) activation.
Results: Mild HI resulted in minimal histological brain jury with neuronal loss confined to the cortex and hippocampus. LPS alone (0.3μg/g) without subsequent HI did not result in any cell death (Nissl, TUNEL). LPS alone did, however, activate microglia in all areas of the forebrain at 12h following injection (F1,25=65.6 p<0.01); this activation was still present at 48h. LPS alone did not result in astrocyte activation (GFAP) over controls at 12h, but there was increased GFAP immunoreactivity by 48h. LPS given 4h before HI resulted in an overall increase in brain injury compared with animals exposed to HI alone (F3,19=6.37, p<0.01 n=10). This sensitising effect was evident when LPS was given within a window of 4-12 hours before the onset of HI. No significant sensitisation was observed when LPS was given immediately or 24 hours prior to the onset of HI (Fig 1,2).
Discussion: These data support the hypothesis that inflammation sensitises the brain to subsequent HI; further, the magnitude of this interaction depends on the time interval between exposure to LPS and HI. Interestingly, endotoxin alone already resulted in microglial activation within the critical 12 hour period. Activation of the innate immune system by LPS may therefore be an important mediating factor in the synergistic interaction between LPS and HI2. In comparison, astrocyte activation only occurs relatively late and may be secondary to microglial activation, following infectious and/or hypoxic stimuli.
1. Peebles DM, Wyatt JS. Synergy between antenatal exposure to infection and intrapartum events in causation of perinatal brain injury at term. BJOG. 2002 Jul;109(7):737-9.
2. Lehnardt S et al. Activation of innate immunity in the CNS triggers neurodegeneration through a Toll-like receptor 4-dependent pathway. Proc Natl Acad Sci. 2003 Jul 8;100(14):8514-9.