Schisandra Chinensis Regulates Drug Metabolizing Enzymes And Drug Transporters Through Nrf2-mediated Signaling Pathway

ObjectiveNuclear factor erythriod2-related factor2(Nrf2) is a transcription factor that play an important role in the cellular defense mechanisms against xenobiotic toxicity or carcinogenicity. As stimulated by oxidative stress or electrophile, Nrf2disassociates from Keap1and translocates into the nucleic where it deterizes with some cofactor like small Maf protein and binds to ARE. To investigate the mechanism of Nrf2pathway mediated Phase Ⅱ and Drug Transporter in HepG2cells by Schisandrae Chinese Extracts (SCE), our research:①The activation of the ARE-luciferase reporter by SCE treatment, and if SCE act as an Nrf2activator.②The induction effect of Phase Ⅱ and drug transporter such as HO-1, MRP2, P-gp et al.③The regulation of cellular GST activity and GSH level by SCE treatment.④The role of Nrf2pathway during the inducing.⑤The effect of protein kinese such as ERK, p38MAPK in the induction by SCE.MethodThe HepG2cells were treated by SCE which extracted from Wu Wei Zi according to the literature method. To verify if SCE act as an Nrf2activator or not, ARE-luciferase activity was measured by treatment of HepG2cells with SCE.The regulation of downstream target genes such as HO-1, GCLM, MRPs, OATPs and Nrf2, Keap1were investigated as HepG2treated with or without pretreatment of siRNA by SCE. On the other hand, we also identify the regulation of cellular GST activity and GSH level, and the effect of regulation of HO-1, Nrf2et al with pretreatment of protein kinese inhibitor by SCE.Results:①Wu Wei Zi were refluxed with ethanol, followed by macroporous resin (AB-8) purification. The lignanoid content of SCE is39.77mg/ml, SchA is1.577mg/g and SchB is 0.763mg/g②SCE activated the activity of ARE-luciferase reporter, SCE is an Nrf2activator.③SCE up-regulate the phase Ⅱ and Efflux transporter such as HO-1, MRP2and P-gp; And down-regulate the organic anion transporting polyoeotude such as OATP1A2, OATP1B1.④Cellular GST level and GST activity are as a parabolic increase with growing concentration of SCE in HepG2cells⑤Nrf2mediated the regulation of phase Ⅱ and drug transporters by SCE.⑥SCE up-regulated the translocation of Nrf2to nucleus and down-regulate Keapl⑦SCE increased Nrf2stability, the t1/2of Nrf2protein can be increased from13min to26min.⑧The induction of HO-1, P-gp et al by SCE could be inhibited by pretreatment of protein kinese inhibitor, while the effect of Nrf2were difference.Conclusion:The Nrf2activator SCE up-regulate cellular GSH level and GST activity, leading enhanced expression of phase Ⅱ gene protein expression of HO-1and GCLM via Nrf2signal pathway and stabilize Nrf2protein, and the mechanism by which SCE have an influence on Nrf2activation may be relevant to ERK and p38MAPK have an effect of Nrf2phosphorylation directly elevated nuclear levels of Nrf2and translocation. Collectively, nucleic translocation of Nrf2and activation of Nrf2/ARE pathway may contribute to the mechanism of hepatic protective of Wu Wei Zi.