Rocha Ferreira, E; (2014) The role of infection/inflammation, the TNF family of cytokines and myeloid cells in perinatal hypoxia-ischaemia brain injury. Doctoral thesis , UCL (University College London).

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Abstract

Synergy between materno-foetal infection and hypoxic-ischaemic insult around the time of birth is a known contributing factor to perinatal brain damage. This is a common precursor to cerebral palsy and other neurological deficits, affecting 2 to 5 per 1000 live births. Endotoxin up-regulates several molecules, including the TNF cluster of pro-inflammatory cytokines. Our group has explored the role of this cluster and shown that its deletion abolishes LPS sensitization to neonatal hypoxic-ischaemic insult. In this study we wanted to first investigate the effects of LPS-mediated sensitization across multiple wild type strains (C57BL/5, 129SVJ, BALB/c, CD1 and FVB) in order to then further characterize the TNF cluster, by studying the individual effects of TNFα, LTα and LTβ members of this cluster, using either global gene deletion, or peripheral myeloid/macrophage-specific deletion of the floxed TNFα allele with MLys::Cre (MLys+). Additionally, we decided to also look at the acquired cellular immune system, using the athymic nude mouse model of T cell deficiency (nu). At P7, littermates for each of the wild type strains, wild-type and homozygous knock-out offspring of heterozygous animals listed above underwent hypoxic-ischaemic insult, consisting of unilateral carotid occlusion followed 2 hours recovery before being placed in a hypoxic chamber for 30min with continuous 8% oxygen exposure. 12 hours prior, animals received a single intraperitoneal injection of 0.6µg/g LPS or saline as a control. 1/3 of animals in the wild type strains group underwent hypoxia-ischaemia alone as a control for saline treatment. LPS pre-treatment resulted in substantial increase inflammation, neuronal injury and infarct in all wild type strains, as well as in the phenotypically wild type littermates of the homozygous mutant animals. Mice lacking both copies of the LTα gene revealed a clear reduction in LPS-mediated sensitization. In reverse, global deletion of LTβ had a detrimental effect, with significant increase in brain damage. Global deletion of TNFα showed a trend towards greater damage, but deletion just in MLys+ macrophages was strongly protective, pointing to a dual role for the TNFα gene depending on in which cell-type it is expressed. Finally, nude animals (nu/nu) demonstrated a complete lack of LPS-mediated sensitization to subsequent hypoxic-ischaemic insult, suggesting that LPS sensitization may require T cell function.

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