Role of brain cytochrome P450 mono-oxygenases in bilirubin oxidation-specific induction and activity

Abstract

In the Crigler–Najjar type I syndrome, the genetic absence of efficient hepatic glucuronidation of unconjugated bilirubin (UCB) by the uridine 5′-diphospho-glucuronosyltransferase1A1 (UGT1A1) enzyme produces the rise of UCB level in blood. Its entry to central nervous system could generate toxicity and neurological damage, and even death. In the past years, a compensatory mechanism to liver glucuronidation has been indicated in the hepatic cytochromes P450 enzymes (Cyps) which are able to oxidize bilirubin. Cyps are expressed also in the central nervous system, the target of bilirubin toxicity, thus making them theoretically important to confer a protective activity toward bilirubin accumulation and neurotoxicity. We therefore investigated the functional induction (mRNA, EROD/MROD) and the ability to oxidize bilirubin of Cyp1A1, 1A2, and 2A3 in primary astrocytes cultures obtained from two rat brain region (cortex: Cx and cerebellum: Cll). We observed that Cyp1A1 was the Cyp isoform more easily induced by beta-naphtoflavone (βNF) in both Cx and Cll astrocytes, but oxidized bilirubin only after uncoupling by 3, 4,3′,4′-tetrachlorobiphenyl (TCB). On the contrary, Cyp1A2 was the most active Cyp in bilirubin clearance without uncoupling, but its induction was confined only in Cx cells. Brain Cyp2A3 was not inducible. In conclusion, the exposure of astrocytes to βNF plus TCB significantly enhanced Cyp1A1 mediating bilirubin clearance, improving cell viability in both regions. These results may be a relevant groundwork for the manipulation of brain Cyps as a therapeutic approach in reducing bilirubin-induced neurological damage.

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Acknowledgments

Sabrina Eliana Gambaro and Maria Celeste Robert were supported by Ministero degli Affari Esteri grants (Foreign Affairs Ministry). Silvia Gazzin was partly supported by an internal grant from the Italian liver Foundation, partly by the Telethon grant GGP10051. Claudio Tiribelli did not received support for his contribution to this work. We are grateful to Miguel Mano for the help in the setup of EROD/MROD tests. We thank Paola Zarattini (Animal Facility, Università degli Studi di Trieste) and Andrea Lorenzon (Consorzio per il Centro di Biomedicina Molecolare S.c.r.l.-CBM) for their help in animal management.

Ethical standards

Animal care and procedures were conducted according to the guidelines approved by the Italian Law (decree 116-92), and the European Communities Council Directive 86-609-ECC. Regular communication to the National Ethic Committee was done (2009–2011).

Conflict of interest

The supporting agencies did not play any role in the study.

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Correspondence to Silvia Gazzin.

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Gambaro, S.E., Robert, M.C., Tiribelli, C. et al. Role of brain cytochrome P450 mono-oxygenases in bilirubin oxidation-specific induction and activity. Arch Toxicol 90, 279–290 (2016). https://doi.org/10.1007/s00204-014-1394-4

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Keywords

  • Cytochrome P450 enzymes (Cyp)
  • Bilirubin oxidation
  • Kernicterus
  • Astrocytes
  • βNF
  • EROD/MROD