Role of ABCG2 in Transport of the Mammalian Lignan Enterolactone and its Secretion into Milk in Abcg2 Knockout Mice

Abstract

Lignans are phytoestrogens that are metabolized by the gut microbiota to enterodiol and enterolactone, the main biologically active enterolignans. Substantial interindividual variation in plasma concentration and urinary excretion of enterolignans has been reported, this being determined, at least in part, by the intake of lignan precursors, the gut microbiota, and the host's phase 2 conjugating enzyme activity. However, the role of ATP-binding cassette (ABC) transporters in the transport and disposition of enterolactone has not been reported so far. Active transport assays using parental and Madin-Darby canine kidney epithelial cells transduced with murine and human ABCG2 showed a significant increase in apically directed translocation of enterolactone in transduced cells, which was confirmed by using the selective ABCG2 inhibitor Ko143. In addition, enterolactone also inhibited transport of the antineoplastic agent mitoxantrone as a model substrate, with inhibition percentages of almost 40% at 200 mM for human ABCG2. Furthermore, the endogenous levels in plasma and milk of enterolactone in wild-type and Abcg2(2/2) knockout female mice were analyzed. The milk/ plasma ratio decreased significantly in the Abcg2 (2/2) phenotype, as compared with the wild-type mouse group (0.4 6 0.1 as against 6.4 6 2.6). This paper is the first to report that enterolactone is a transported substrate and therefore most probably a competitive inhibitor of ABCG2, which suggests it has a role in the interindividual variations in the disposition of enterolactone and its secretion into milk. The inhibitory activity identified provides a solid basis for further investigation in possible food-drug interactions. IntroductionThe ATP-binding cassette (ABC) subfamily G2 (ABCG2) protein is apically expressed ABC membrane transporter that mediates the active and outward transport of a wide range of anticancer drugs, dietary compounds, food carcinogens, and antibiotics from cells (van Herwaarden et al., 2003;Merino et al., 2005;Vlaming et al., 2009). In recent years, its role as a transporter of phytoestrogens and their conjugated metabolites has gained special relevance (Zhu et al., 2010;Alvarez et al., 2011;Tan et al., 2013).ABCG2 is found in tumor cells, but also in the cells of a variety of normal tissues involved in the uptake and elimination of compounds, such as enterocytes, hepatocytes, and cells of the proximal tubules of the kidney. The apical localization in these cells allows ABCG2 to mediate hepatobiliary and urinary elimination and to function as a barrier to uptake from the gut lumen (van Herwaarden and Schinkel, 2006). In addition, induced expression of ABCG2 in the lactating mammary gland supports its important role in the active secretion of several xenobiotics and beneficial compounds such as vitamins into milk Merino et al., 2006;van Herwaarden et al., 2007).Dietary intake of lignans and their derivatives, as part of a healthy diet, has been associated with protective effects against a number of chronic diseases (Hu e...

Effect of bovine ABCG2 polymorphism Y581S SNP on secretion into milk of enterolactone, riboflavin and uric acid

“…Our results show that the in vitro models can act as a valuable tool to test the potential role of the Y581S bovine polymorphism in the secretion of drugs (Gonzalez-Lobato et al, 2014) and dietary-derived compounds such as enterolactone (Miguel et al, 2014) Mean ratio values within a column not sharing the same superscripts differ significantly at P < 0.05.…”

“…Two hours before the start of the experiment, medium at both the apical (AP) and basolateral (BL) sides of the monolayer was replaced with 2 ml of transport medium without serum, and either with or without the specific ABCG2 inhibitor Ko143 (1 μM). The experiment started (t = 0) when the medium in either the AP or BL compartment was replaced with fresh medium containing physiological concentrations of the compounds studied (100 µM of uric acid; 10 µM of enterolactone; 0.1 µM of riboflavin; Larson and Moyes 2010; Koop et al, 2014;Miguel et al, 2014). In the case of enterolactone and uric acid, transport medium used was Opti-MEM I Reduced Serum medium (Life Technologies-Gibco 31985-047).…”

“…Mitoxantrone (MXR, 10 µM) was used as fluorescence substrate of ABCG2, and enterolactone (100 and 200 µM) was used to inhibit the transporter (Miguel et al, 2014). Cells were cultured in 12-well plates (20 × 10 3 cells/well) in complete medium for 36 h to subconfluence.…”

“…Chromatographic and mass spectrometry conditions were the same as those previously published for the analyses and quantification of enterolactone (Miguel et al, 2014). Enterolactone was identified in plasma and/or milk samples based on the accurate mass and isotopic pattern given in QTOF-MS acquisition mode, fragmentation information offered by targeted MS/MS experiments and direct comparison with an authentic standard.…”

“…Enterolactone possesses antioxidant activity and dietary intake of lignans, and their derivatives, as part of a healthy diet, have been associated with protection against cardiovascular disease, diabetes and metabolic syndrome, cancer, oxidative stress and inflammation (Adolphe et al, 2010). Recently, enterolactone has been identified as a substrate of the human and murine abcg2 (Miguel et al, 2014). Riboflavin deficiency can occur in populations fed with diets poor in dairy products.…”