St. John's Wort (Hypericum perforatum) induces overexpression of multidrug resistance protein 2 (MRP2) in rats: a 30-day ingestion study
Introduction
St. John's Wort (Hypericum perforatum, SJW) is a herbal medicine that is frequently used for depression. SJW extract is usually used for humans at doses of 900–1050 mg/day (Johne et al., 2002, Philipp et al., 1999, Sugimoto et al., 2001). It was reported that 1050 mg SJW extract (350 mg three times daily) was an effective dose in patients with moderate depression. The dose was more effective than the daily treatment with 100 mg imipramine in moderate depression (Philipp et al., 1999). In USA sales increased by 2,800% in one year, and in Europe total sales figures were approximately US$6 billion in 1998. In addition, a number of foods that contain SJW extracts have been sold as supplement foods for health care in Japan. However, the safety of SJW has recently been questioned. Several studies have raised the possibility of important interactions between SJW extracts and prescribed drugs such as cyclosporin and theophylline (Henderson et al., 2002). An in vitro study strongly suggested that the properties of SJW extracts induced cytochrome P450 (CYP) 3A4 and 1A2 (Ernst, 1999). Another in vivo study showed that SJW induced the expression of intestinal P-glycoprotein/MDR1 (in rats and humans), hepatic CYP3A2 (in rats), and intestinal and hepatic CYP3A4 (in humans) (Dürr et al., 2000).
P-glycoprotein (P-gp) is an ATP-dependent efflux pump for hydrophobic drugs, and is a member of ABC (ATP-binding cassette) superfamily (Sharom, 1997). In addition, multidrug resistance protein (MRP) family is a member of ABC superfamily. The membrane proteins mediating the ATP-dependent unidirectional transport of conjugates of lipophilic substances with glutathione, glucuronate or sulfate has been recognized as members of MRP family. Multidrug resistance protein 2 (MRP2), known as a canalicular multispecific organic anion transporter (cMOAT), functions in the terminal excretion of cytotoxic and carcinogenic substances, and plays an important role in detoxification and chemoprevention (Jörg et al., 1999).
Microsomal CYP proteins are key detoxification enzymes and catalyze the first step in the biotransformation process with xenobiotics being oxidized to forms that can then be further modified by conjugating enzymes (Satish et al., 1998). The members of CYP 1 gene family are induced by polycyclic aromatic hydrocarbons. Such chemicals include natural combustion products [benzo(a)pyrenes], dietary constituents (heterocyclic amines) and manufacturing by-products (dioxines) (Satish et al., 1998). On the other hand, glutathione S-transferase P (GST-P) is a conjugating enzyme known to be markedly increased during chemical hepatocarcinogenesis of the rat (Kitahara et al., 1984, Sugioka et al., 1985).
In the present study, we examined the effects of SJW on the overexpressions of P-gp, MRP1, MRP2, GST-P and CYP1A2 in rats. The rats were given SJW extract at a dose of 400 mg/kg/day, since it has been reported that the dose of SJW extract is effective antidepressant dose in rats (Rezvani et al., 1999). We performed also the ingestion experiments at a low dose of 40 mg/kg similar to human dose (usual human doses, 900–1050 mg/day) to determine whether the low dose ingestion lead to the overexpression of the enzymes and transporters.
Section snippets
Reagents
St John's Wort extract tablets (Brand name “Feel Balance”) were purchased from Rohto Pharmaceutical Co. Ltd. (Osaka, Japan). A mouse monoclonal antibody specific for anti-P-glycoprotein, mdr1 and mdr3 (C219) was purchased from DAKO Corporation (Carpinteria, CA, USA). A mouse monoclonal antibody specific for anti-multidrug resistance-related protein cMOAT/MRP2 (M2III-6) was purchased from SANBIO (AM Uden, The Netherlands). Monoclonal antibody M2III-6 was generated against amino acids 1340–1541
Effect of SJW treatment on the hepatic P-gp and MRP1 expression levels
Hepatic P-gp was detected as a 190-kDa protein; MRP1 was also detected as a 170-kDa protein (Fig. 1). However, SJW treatment showed no effect on the expressions of P-gp and MRP1 (Table 1).
Effect of SJW treatment on the hepatic MRP2 expression levels
MRP2 was detected as a 190-kDa protein (Fig. 2a). The expression levels of MRP2 are expressed as a percentage of the corresponding CMC treated group (Fig. 2b). The low dose of SJW treatment (40 mg/kg/day) for 10 consecutive days showed no expression levels of MRP2 in the liver. High dose SJW treatment (400
Discussion
We examined the effect of oral administration of SJW on the endogenous expressions of drug transporters and enzymes in rats. Rats were treated with the high dose (400 mg/kg) or the low doses (40 mg/kg) of SJW. It has been reported that the high dose of SJW correlated with reduce alcohol intake, which is an index of the antidepressant effect, in high-alcohol intake rats (Rezvani et al., 1999). Thus, it is estimated that the high dose of SJW is an antidepressant effective dose in rats. In the
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