Inflammatory signalling pathways involved in astroglial activation by unconjugated bilirubin

During neonatal hyperbilirubinaemia, astrocytes activated by unconjugated bilirubin (UCB) may contibute to brain toxicity through the production of cytokines. As a first step in addressing the signal transduction cascades involved in the UCB-induced astroglial immunological response, we tested whether tumour necrosis factor (TNF)-alpha receptor 1 (TNFR1), mitogen-activated protein kinase (MAPK) and nuclear factor kappa B (NF-kappa B) would be activated in astrocytes exposed to UCB, and examined the profile of cytokine production. Astrocyte cultures stimulated with UCB showed a rapid rise in TNFR1 protein levels, followed by activation of the MAPKs p38, Jun N-terminal kinase1/2 and extracellular signal-regulated kinase1/2, and NF-kappa B. Interestingly, the induction of these signal effectors preceded the early up-regulation of TNF-alpha and interleukin (IL)-1 beta mRNAs, and later secretion of TNF-alpha, IL-1 beta and IL-6. Treatment of astrocytes with UCB also induced cell death, with levels comparable to those obtained after exposure of astrocytes to recombinant TNF-alpha and IL-1 beta. Moreover, loss of cell viability and cytokine secretion were reduced when the NF-kappa B signal transduction pathway was inhibited, suggesting a key role for NF-kappa B in the astroglial response to UCB. These results demonstrate the complexity of the molecular mechanisms involved in cell injury by UCB during hyperbilirubinaemia and provide a basis for the development of novel therapeutic strategies.