In vitro and in vivo studies have demonstrated that UCB (unconjugated bilirubin) is neurotoxic. Although previous studies suggested that both MRP1 (multidrug resistance-associated protein 1) and MDR1 (multidrug resistance protein 1) may protect cells against accumulation of UCB, direct comparison of their role in UCB transport was never performed. To this end, we used an inducible siRNA (small interfering RNA) expression system to silence the expression of MRP1 and MDR1 in human neuroblastoma SH-SY5Y cells. The effects of in vitro exposure to clinically-relevant levels of unbound UCB were compared between unsilenced (control) cells and cells with similar reductions in the expression of MRP1 or MDR1, documented by RT–PCR (reverse transcription–PCR) (mRNA), immunoblotting (protein), and for MDR1, the enhanced net uptake of a specific fluorescent substrate. Cytotoxicity was assessed by the MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide] test. MRP1-deficient cells accumulated significantly more UCB and suffered greater cytotoxicity than controls. By contrast, MDR1-deficient cells exhibited UCB uptake and cytotoxicity comparable with controls. At intermediate levels of silencing, the increased susceptibility to UCB toxicity closely correlated with the decrease in the expression of MRP1, but not of MDR1. These data support the concept that limitation of cellular UCB accumulation, due to UCB export mediated by MRP1, but not MDR1, plays an important role in preventing bilirubin encephalopathy in the newborn.
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Research Article|
December 12 2008
The cytotoxic effect of unconjugated bilirubin in human neuroblastoma SH-SY5Y cells is modulated by the expression level of MRP1 but not MDR1
Lucia Corich;
Lucia Corich
*Centro Studi Fegato, Bld Q, AREA Science Park, Basovizza Campus, Ss 12 km 163.5, c 3412 Trieste, Italy
†Department of Life Sciences, University of Trieste, 34012 Trieste, Italy
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Alejandro Aranda;
Alejandro Aranda
*Centro Studi Fegato, Bld Q, AREA Science Park, Basovizza Campus, Ss 12 km 163.5, c 3412 Trieste, Italy
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Laura Carrassa;
Laura Carrassa
‡Laboratory of Molecular Pharmacology, Department of Oncology, Istituto di Ricerche Farmacologiche Mario Negri, Milan, Italy
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Cristina Bellarosa;
Cristina Bellarosa
*Centro Studi Fegato, Bld Q, AREA Science Park, Basovizza Campus, Ss 12 km 163.5, c 3412 Trieste, Italy
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J. Donald Ostrow;
J. Donald Ostrow
§GI/Hepatology Division, Department of Medicine, University of Washington, Seattle, WA, 98195, U.S.A.
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Claudio Tiribelli
Claudio Tiribelli
1
*Centro Studi Fegato, Bld Q, AREA Science Park, Basovizza Campus, Ss 12 km 163.5, c 3412 Trieste, Italy
1To whom correspondence should be addressed (email ctliver@csf.units.it).
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Publisher: Portland Press Ltd
Received:
May 09 2008
Revision Received:
August 14 2008
Accepted:
August 19 2008
Accepted Manuscript online:
August 19 2008
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© The Authors Journal compilation © 2009 Biochemical Society
2009
Biochem J (2009) 417 (1): 305–312.
Article history
Received:
May 09 2008
Revision Received:
August 14 2008
Accepted:
August 19 2008
Accepted Manuscript online:
August 19 2008
Citation
Lucia Corich, Alejandro Aranda, Laura Carrassa, Cristina Bellarosa, J. Donald Ostrow, Claudio Tiribelli; The cytotoxic effect of unconjugated bilirubin in human neuroblastoma SH-SY5Y cells is modulated by the expression level of MRP1 but not MDR1. Biochem J 1 January 2009; 417 (1): 305–312. doi: https://doi.org/10.1042/BJ20080918
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