Constitutive and Inducible Levels of Cytochrome P450 enzymes related to Bilirubin-Induced Encephalopathy

GAMBARO SABRINA ELIANA; GAZZIN SILVIA; ROBERT MARIA CELESTE; TIRIBELLI CLAUDIO

Torino

Simposio; 11th International Symposium on Cytochrome P450; 2012

Cytochrome P450

Bilirubin encephalopathy is the severe, permanent neurological disorder caused by the toxic effect of unconjugated bilirubin (UCB) to neuronal cells. There is a strong evidence of different neurotoxicity depending on the region (basal ganglia, hippocampus and cerebellum) and cell type (neurons more sensitive than glia). The Gunn rat, unable to conjugate bilirubin due to a genetic absence of glucuronidation enzyme, is the animal model used to study this pathology. The homozygote Gunn rat accumulates bilirubin rapidly after birth, with a peak at 9 days after the birth (12mg/dL). Interestingly, during the third week level decreases till a plateau (5mg/dL), indicating the existence of alternative mechanisms of elimination. The hepatic cytochrome P450 enzymes (Cyp1A1, Cyp1A2 and Cyp2A3) have been shown to be involved in the oxidation of bilirubin leading to elimination of the pigment. Since the role of Cyps in bilirubin clearance is unkown. To investigate if specific regional expression of Cyps may selectively reduce accumulation and neurotoxicity, we analysed the Cyps mRNA expression in primary culture of cerebellar (UCB damaged region) and cortex (not damaged region) astrocytes of Wistar rat. To valutate the inducibility of the brain Cyps, cells were induced by B-Naphtophlavone (BNF) at different concentrations and times (2.5; 5; 10; 20µM and 1; 6 and 24H, respectively). In addition, to analyse if bilirubin is able to upregulate Cyps expression cells were treated with UCB (140nM;2H). mRNA expression and Cyp1A1 activity were assessed. BNF induce a higher Cyp1A1 expression in astrocytes obtained from cerebellum than cortex, while Cyp1A2 present the opposite trend; Cyp2a3 is not induced. Of notice UCB induce Cyp1A1 and Cyp1A2 more in cortex than in cerebellar astocytes. This study confirms that constitutive and inducible expression of Cyps differs in different brain regions with a good correlation between Cyps mRNA levels and the regional specific bilirubin neurotoxicity.