Unconjugated bilirubin activates and damages microglia

Abstract

Microglia are the resident immune cells of the brain and are the principal source of cytokines produced during central nervous system inflammation. We have previously shown that increased levels of unconjugated bilirubin (UCB), which can be detrimental to the central nervous system during neonatal life, induce the secretion of inflammatory cytokines and glutamate by astrocytes. Nevertheless, the effect of UCB on microglia has never been investigated. Hence, the main goal of the present study was to evaluate whether UCB leads to microglial activation and to the release of the cytokines tumor necrosis factor (TNF)-alpha, interleukin (IL)-1beta, and IL-6. Additionally, we investigated the effects of UCB on glutamate efflux and cell death. The results showed that UCB induces morphological changes characteristic of activated microglia and the release of high levels of TNF-alpha, IL-1beta, and IL-6 in a concentration-dependent manner. In addition, UCB triggered extracellular accumulation of glutamate and an increased cell death by apoptosis and necrosis. These results demonstrate, for the first time, that UCB is toxic to microglial cells and point to microglia as an important target of UCB in the central nervous system. Moreover, they suggest that UCB-induced cytokine production, by mediating cell injury, can further contribute to exacerbate neurototoxicity. Interestingly, microglia cells are much more responsive to UCB than astrocytes. Collectively, these data indicate that microglia may play an important role in the pathogenesis of encephalopathy during severe hyperbilirubinemia.

Biological risks for neurological abnormalities associated with hyperbilirubinemia

“…Interestingly, the secretion of IL-6 that was suppressed by UCB in astrocytes was slightly increased in neurons (7 pg/10 5 cells) and markedly enhanced in microglia (203 pg/10 5 cells). 18,38 Collectively, our data support the conclusion that microglia are the neural cells most responsive to UCB-induced release of cytokines.…”

“…Microglia were also more vulnerable to UCB-induced apoptosis, revealing an increase of B2.6-and B4.5-fold when compared with astrocytes and neurons, and achieving levels of 11% over control values. 18,38 Recent studies have indicated that some molecules released by microglia may be harmful in acute brain insults and neurodegenerative diseases. 39 Therefore, we also looked for the ability of the three cell types to release glutamate, in particular microglia.…”

“…The release of glutamate by microglia was B4-and B8-fold over the values obtained for astrocytes and neurons, respectively (Figure 3). 18,38 Thus, microglia are the neural cells most susceptible to UCB-induced cell death and release of glutamate. We also have observed that cytokines are mainly Figure 1 Schematic representation of the signaling pathways involved in astrocyte reactivity to unconjugated bilirubin (UCB).…”

Gunn rats with glial activation in the hippocampus show prolonged immobility time in the forced swimming test and tail suspension test

“…A comprehensive behavioral test battery is required in further studies to clarify whether Gunn rats have symptoms of the other psychiatric disorders. Unconjugated bilirubin has been shown to activate both microglia (Gordo et al., 2006) and astrocytes (Fernandes et al., 2011) in vitro. In accordance with, microglial activation and astrocytic activation in Gunn rats appear to be caused by high levels of unconjugated bilirubin, which can enter the brain as the free fraction (Ostrow, Pascolo, Shapiro, & Tiribelli, 2003).…”

Astrocyte reactivity to unconjugated bilirubin requires TNF‐α and IL‐1β receptor signaling pathways

“…This situation occurs following deposition of unconjugated bilirubin (UCB) in the central nervous system (CNS) owing to conditions leading to extreme UCB serum levels in the newborn (Hansen, 2002). UCB elicits nerve cell injury through interaction with cellular membranes (Rodrigues et al, 2002a), by triggering oxidative and nitrosative stress (Brito et al 2008a,b), and accumulation of extracellular glutamate Fernandes et al, 2004;Gordo et al, 2006;Silva et al, 1999), producing a widespread of structural and functional alterations that culminate in cell death Rodrigues et al, 2002b;Silva et al, 2002). We have shown that UCB elicits the release of pro-inflammatory cytokines, such as TNF-a and IL-1b, by neurons , astrocytes (Fernandes et al, 2004, and microglia , through the activation of MAPK (i.e., p38, JNK1/2 and ERK1/2) and NF-jB signaling pathways at the intracellular level (Brites et al, 2009;Fernandes et al, 2006Fernandes et al, , 2007a.…”