Effects of Shikonin on Drug-Metabolizing System in Rat Primary Hepatocytes

• Main Authors: Tzu-Yu Lin, 林諮榆
• Other Authors: Chong-Kuei Lii
• Format: Others
• Language: zh-TW
• Published: 2014
• Online Access: http://ndltd.ncl.edu.tw/handle/cx4ps8

碩士 === 中國醫藥大學 === 營養學系碩士班 === 102 === In general, people exposed to a wide variety of environmental pollutants, toxicants, drugs, and food additives that may cause toxicity. To minimize chemical insults, converting these substances into hydrophilic metabolites by drug-metabolizing system is necessary. Shikonin, a naphthoquinone pigment, is rich in the root of Chinese herbal plant Lithospermum erythrorhizon. Evidence indicates that shikonin displays multiple physiological functions including anti-microbes, anti-inflammation, and anti-oxidation. In this study, we intend to examine the effect of shikonin on hepatic Phase I and Phase II drug-metabolizing enzymes as well as Phase III membrane transport proteins expression and the possible mechanism involved. Twenty-four h post plating, rat primary hepatocytes were treated with 0-2 μM shikonin for 16 or 24 h. Immunoblotting assay and RT-PCR revealed that shikonin dose-dependently increases cytochrome P450 (CYP) 1A1/2, CYP3A2, CYP2D1, and CYP2C6) and pi class of glutathione S-transferase (PGST), UDP glucuronosyltransferase 1A1 (UGT1A1), and NADP(H) quinone oxidoreductase 1 (NQO1) protein and RNA expression. Moreover, the protein and mRNA levels of Phase III membrane transport proteins organic anion-transporting polypeptide (OATP) 1B1, OATP2B1, P-glycoprotein, and multidrug resistance-associated protein (MRP) 2/3 were also increased by shikonin. In addition, nuclear translocation of aryl hydrocarbon receptor (AhR) and NF-E2-related factor 2 (Nrf2) was augmented by shikonin. EMSA confirmed that shikonin increases AhR, pregnane X receptor, and Nrf2 binding to dioxin-responsive element, direct repeat 4 (DR4), and antioxidant-responsive element, respectively.Shikonin time-dependently increased ERK1/2, JNK, and p38 phosphorylation up to 30 min. ERK inhibitor (PD98059), JNK inhibitor (SP600125), and p38 inhibitor (SB2035801) pretreatment suppressed shikonin-induced Nrf2 nuclear translocation. Moreover, AhR and Nrf2 knockdown attenuated shikonin-induced increase in CYP1A1/2 induction and NQO1 and PGST expression, respectively. Taken together, results suggest that shikonin effectively upregulates the transcription of Phase I and II drug-metabolizing enzyme and Phase III membrane transporters at least partially through activation of AhR, PXR, and Nrf2 and Nrf2-dependent activation is MAPK-dependent.