Effects of carbamazepine on the P-gp and CYP3A expression correlated with PXR or NF-κB activity in the bEnd.3 cells.

Drug-resistant epilepsy (DRE) is present in 20-30% of all patients who develop epilepsy. Growing evidences demonstrated that glutamate released during seizures to increase the brain P-glycoprotein (P-gp) expression. Carbamazepine (CBZ) is known to influence the P-gp and cytochrome P450 (CYP) expression. However, the exact molecular mechanism is still unknown. We investigated that the effects of NF-κB and pregnane X receptor (PXR) activity on P-gp and CYP3A expression in mouse brain endothelial (bEnd.3) cells treated with l-glutamate (mimicking the seizure conditions), CBZ (mimicking the AED treating conditions) or both (l-glutamate plus CBZ) through qPCR and Western blotting assay. Mean fluorescence intensity was used to observe P-gp efflux function by analysis of intracellular Rhodamine123 (Rho123) accumulation. P-gp, CYP3A, PXR and NF-κB p65 were elevated in bEnd.3 cells incubated with l-glutamate, CBZ or CBZ pretreated by l-glutamate for 30 min. Both the mRNA and protein levels of P-gp and CYP3A were remarkably reduced by PXR or NF-κB p65 knock-down by siRNA transfections. The decreased intracellular accumulation of Rho123 suggested that the expression of P-gp was enhanced in bEnd.3 cells. These data suggested that overexpression of P-gp and CYP3A during seizures and treated with CBZ may be regulated by PXR or NF-κB p65 activity and expression, which revealed a mechanism underlying the development of DRE.

Pregnane X Receptor Not Nuclear Factor-kappa B Up-regulates P-glycoprotein Expression in the Brain of Chronic Epileptic Rats Induced by Kainic Acid.

Drug-resistance epilepsy (DRE) is attributed to the brain P-glycoprotein (P-gp) overexpression. We previously reported that nuclear factor-kappa B (NF-κB) played a critical role in regulating P-gp expression at the brain of the acute seizure rats. This study was extended further to investigate the interaction effect of NF-κB and pregnane X receptor (PXR) on P-gp expression at the brain of chronic epileptic rats treated with carbamazepine (CBZ). The chronic epileptic models were induced by the micro-injection of kainic acid (KA) into rats' hippocampus. Subsequently, the successful models were treated with different intervention agents of CBZ; PMA(a non-specific PXR activity inhibitor) or PDTC(a specific NF-κB activity inhibitor) respectively. The expression levels of P-gp and its encoded gene mdr1a/b were significantly up-regulated on the brain of KA-induced chronic epilepsy rats or the epilepsy rats treated with CBZ for 1 week, meanwhile with a high expression of PXR. The treatment of PMA dramatically reduced both PXR and P-gp expressions at the protein and mRNA levels in the chronic epilepsy brain. By compared to the epilepsy model group, the P-gp expression was not markedly attenuated by the inhibition of NF-κB activity with PDTC treatment, nevertheless with a decrease of NF-κB expression in this intervention group. Higher levels of proinflammatory cytokines(IL-1ß, IL-6, TNF-α) were found both in the brain tissue and the serum in the epilepsy rats of each group. There was a declined trend of the pro-inflammatory cytokines expression of the PDTC treatment group but with no statistical significance. This study demonstrates for the first time that P-gp up-regulation is due to increase PXR expression in the chronic phase of epilepsy, differently from that NF-κB signaling may induce the P-gp expression in the acute seizure phase. Our results offer insights into the mechanism underlying the development of DRE using or not using CBZ treatment.