Constitutive androstane receptor weakens the induction of panaxytriol on CYP3A4 by repressing the activation of pregnane X receptor

Nuclear receptors pregnane X receptor (PXR; NR1I2) and constitutive androstane receptor (CAR; NR1I3) play a vital role in regulating CYP3A4. Our previous studies have demonstrated that panaxytriol (PXT) upregulates the expression of CYP3A4 via the PXR regulatory pathway. This study aimed to explore how CAR mediates the regulation of CYP3A4 in the presence of PXT using HepG2 cell, hCAR-overexpressing HepG2 cell and hCAR-silenced HepG2 cell models. In HepG2 cells, PXT induced the expression of CYP3A4 in a concentration-dependent manner (10-80 μM) and the high concentration of PXT (80 μM) upregulated the expression of CAR. The concentrations of PXT (10-40 μM) had no impact on the expression of CAR, but could significantly induce the expression of CYP2B6 target gene by activating CAR. The dual-luciferase reporter gene assay also showed that CAR-mediated CYP3A4 luciferase activity can be promoted by 80 μM of PXT (1.54-fold), while 5, 10, 20, and 40 μM of PXT had no influence on CAR-mediated CYP3A4 luciferase activity. In hCAR-overexpressing HepG2 cells, PXT concentrations (10-40 μM) that significantly induced PXR and CYP3A4 in HepG2 cells had no impact on the expression of CYP3A4, CAR and PXR, whereas a high concentration of PXT (80 µM) could weakly induce the mRNA and protein levels of CAR and CYP3A4. Moreover, the expression of PXR and CYP3A4 in hCAR-silenced HepG2 cells was markedly elevated compared with the blank control or with normal HepG2 cells treated with 10-80 μM of PXT. In conclusion, CAR significantly weakens the ability of PXT to induce CYP3A4 expression by repressing the activation of PXR. There may be a cross-talk mechanism between PXR and CAR on the regulation of CYP3A4 in the presence of PXT. Additionally, a high concentration of PXT (80 μM) induced CYP3A4 via the CAR regulatory pathway.