Abstracts

Rationale: Nuclear receptors and cytochrome P450 together regulates majority of drugs in the liver. In past years, we found a significant contribution of cytochrome P450 to drug metabolism in human epileptic brain. In the present study, we explored whether nuclear receptors such as pregnane X (PXR), constitutive androstane (CAR) and glucocorticoid (GR) receptors, all involved in the control of P450 expression in the liver are also responsible for abnormal expression of P450 enzymes in human epileptic brain. Methods: Primary cultures of human drug resistant brain endothelial cells (EPI-EC, n=7), control brain endothelial cells (HBMEC, n=7) and hepatocytes (n=3) were used. Nuclear receptor mRNA levels were assessed by cDNA microarrays. The expression of nuclear receptors, PXR, CAR and GR was also evaluated by western blotting. PXR and key drug regulated CYPs (CYP3A4, 2C9, 2C19, 2D6 and 2E1) involved in hepatic AED metabolism were studied by western blot and siRNA approaches. Results: Increased mRNA levels of nuclear receptors were found in epileptic brain endothelial cells compared to control. CYP3A4, 2C9, 2E1 were overexpressed in majority of EPI-ECs compared to control; in contrast, CYP2D6, 2C19 were down-regulated or absent in EPI-EC. Increased PXR and GR expression were found in EPI-ECs (*p<0.05) whereas CAR expression were variable across the samples. Hepatocytes consistently showed an elevated levels of PXR, CAR and GR (*p<0.01). The levels of CYPs directly correlated with PXR and GR expression in EPI-ECs and controls, although such effect could not be demonstrated for CAR. Levels of PXR were positively correlated with CYP3A4 and CYP2E1 whereas CYP2C9 remained unaltered when PXR was silenced. Conclusions: The study suggests that at steady-state PXR transcriptional activity is elevated in drug resistant epileptic BBB cells; this is supported by increased expression of PXR regulated CYP. Among other nuclear receptors GR followed a similar trend as PXR in EPI-EC whereas CAR did not, thereby suggesting a different pattern of P450-nuclear receptor regulation in epileptic brain endothelial cells. Acknowledgements: Supported by R01NS078307 (N.M. and D.J.), R01NS43284, R41MH093302, R21NS077236, R42MH093302, UH3TR000491, and R21HD057256 (D.J.). AHA-SDG 13SDG13950015 and NARSAD Brain-Behavior Research Foundation (C.G).