The cytotoxic effect of unconjugated bilirubin in human neuroblastoma SH-SY5Y cells is modulated by the expression level of MRP1 but not MDR1
Résumé
In vitro and in vivo studies have demonstrated that unconjugated bilirubin (UCB) is neurotoxic. Although previous studies suggested that both MRP1 and MDR1 proteins can protect cells against accumulation of UCB, direct comparison of their role in UCB transport was never performed. To this end, we modulated expression of MRP1 and MDR1 in human neuroblastoma SH-SY5Y cells using an inducible siRNA expression system. The effects of in vitro exposure to clinically-relevant levels of UCB were compared between cells with similar reductions in the expression of MRP1 or MDR1; cells expressing endogenous levels of the two proteins were used as controls. MRP1 deficient cells accumulated a significantly higher amount of UCB, while UCB uptake of MDR1 deficient cells was comparable to controls. Cell function, assessed by MTT assay, was significantly reduced in MRP1 deficient cells, even at a normally non-toxic unbound bilirubin concentration of 40 nM; no difference in toxicity was observed between MDR1 deficient and control cells. MTT assays, performed at intermediate levels of MRP1 or MDR1 silencing, showed that the increased susceptibility to UCB toxicity was closely correlated with the suppression of the expression of MRP1 but not of MDR1. These data indicate that limitation of cellular UCB accumulation, due to UCB export mediated by MRP1, may play an important role in prevention of bilirubin encephalopathy in the newborn.
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