Abstract
BackgroundHypoxic-ischemic (HI) brain injury remains a major problem in newborns, resulting in increased risk of neurological disorders. Neonatal HI triggers a broad inflammatory reaction in the brain, including activation of the innate immune system. Toll-like receptors (TLRs), which are key components of the innate immune system, are believed to play a role in adult cerebral ischemic injury. The expression of TLRs in the neonatal brain and their regulation after HI is unknown.MethodsWild type C57BL/6, TLR 1 knockout (KO) and TLR 2 KO mice were subjected to HI at postnatal day 9 and sacrificed 30 min, 6 h, 24 h or 5 days after HI. TLR mRNA expression was determined by RT-qPCR and protein and cell type localisation by immunohistochemistry (IHC). To evaluate brain injury, infarct volume was measured in the injured hemisphere.ResultsmRNA expression was detected for all investigated TLRs (TLR1-9), both in normal and HI exposed brains. After HI, TLR-1 was down-regulated at 30 min and up-regulated at 6 h and 24 h. TLR-2 was up-regulated at 6 h and 24 h, and TLR-7 at 24 h. Both TLR-5 and TLR-8 were down-regulated at 24 h and 30 min respectively. IHC showed an increase of TLR-1 in neurons in the ipsilateral hemisphere after HI. TLR-2 was constitutively expressed in astrocytes and in a population of neurons in the paraventricular nucleus in the hypothalamus. No changes in expression were detected following HI. Following HI, TLR-2 KO mice, but not TLR-1 KO, showed a decreased infarct volume compared to wild type (p = 0.0051).ConclusionsThis study demonstrates that TLRs are regulated after HI in the neonatal brain. TLR-1 protein was up-regulated in injured areas of the brain but TLR-1 KO animals were not protected from HI. In contrast, TLR-2 was constitutively expressed in the brain and TLR-2 deficiency reduced HI injury. These data suggest that TLR-2, but not TLR-1, plays a role in neonatal HI brain injury.
Highlights
Hypoxic-ischemic (HI) brain injury remains a major problem in newborns, resulting in increased risk of neurological disorders
Results mRNA expression of Toll-like receptors (TLRs) in normal brain and after hypoxia-ischemia mRNA expression was detected for all TLRs present on the array in the postnatal day (PND) 9 mouse brain (Table 1)
Following HI, TLR-1 mRNA was down-regulated at 30 min up-regulated at 6 h and 24 h; TLR-2 was up-regulated at 6 h and 24 h and TLR-7 was up-regulated at 24 h
Summary
Hypoxic-ischemic (HI) brain injury remains a major problem in newborns, resulting in increased risk of neurological disorders. The precise aetiology of brain injury in the newborn is sometimes unclear, hypoxia-ischemia (HI) is well accepted as a contributing factor [1] Both infection and intrapartum asphyxia is associated with inflammation in the brain [2] and there are increased levels of cytokines in the cerebral spinal fluid in term infants that have suffered birth asphyxia [3]. MyD88 is an adaptor protein, which upon recruitment to the activated receptor initiates a signalling cascade leading to activation of different transcription factors, e.g. nuclear factor B (NFB) and activator protein-1 (AP1). This activation gives rise to a generation of pro-inflammatory cytokines such as interleukin (IL)-6 and tumour necrosis factor-a (TNFa). Recruitment of TRIF leads to the activation of the transcription factor interferon regulatory factor (IRF) -3 and -7 and the generation of antiviral molecules such as interferon (IFN)-b
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