The Influence of Selenium on Arsenic Hepatobiliary Transport

Abstract

Over 200 million people worldwide are exposed to the proven human carcinogen arsenic at levels exceeding the World Health Organization guideline of 10 μg/L. In animal models arsenic and selenium are mutually protective via the formation and biliary excretion of the seleno‐bis (S‐glutathionyl) arsinium ion [SeAs(GS)2]−, which allows for the fecal elimination of both compounds. Consistent with this, selenium deficiency in humans living in arsenic endemic regions is associated with an increased risk of arsenic‐induced disease. These observations have led to the initiation of human selenium supplementation trials. Despite these ongoing trials in arsenic endemic regions, the influence of selenium on human hepatic handling of arsenic is not adequately understood. Furthermore, supplementation trials have utilized different chemical forms of selenium with unknown influence on efficacy. In the liver, multidrug resistance protein 2 (MRP2/ABCC2) transports arsenic metabolites, including [SeAs(GS)2]− into bile, and the related MRP4 (ABCC4) transports other arsenic metabolites into sinusoids. We hypothesized that selenium will increase the biliary excretion of arsenic from HepaRG cells, an immortalized cell line used as a surrogate for primary human hepatocytes. To test this hypothesis, we studied the influence of selenite (SeIV), selenide (SeII), methylselenocysteine (MSC) and selenomethionine (SeMet) on arsenic efflux from HepaRG cells. The expression of genes involved in hepatic arsenic methylation (As3MT) and export (ABCC2 and ABCC4) were assessed. Crude membrane preparations subjected to immunoblots were used to evaluate the presence of MRP2 and MRP4 proteins. The influence of arsenic on the levels of these proteins was also assessed similarly. Fluorescence microscopy after treatment with 5(6)‐carboxy,2’,7’‐dichlorofluorescein (CDF) diacetate was performed to visualize the canalicular networks and assess MRP2 function. Transport across sinusoidal and canalicular membranes was measured after treatment of HepaRG cells with 1 μM 73AsIII ± selenium (in the forms SeIV, SeII, MSC or SeMet) using B‐CLEAR® technology. Biliary excretion indices (BEIs) were calculated to quantify the extent of arsenic export into bile. ABCC2, ABCC4, and As3MT are expressed by HepaRG Cells. MRP2 and MRP4 proteins were detected, and their expression was increased by the presence of AsIII. CDF accumulation in canalicular networks suggested that MRP2 was functional. At a 5 minute time point, the BEI of 73AsIII alone was 17%. The addition of SeII increased biliary excretion of 73AsIII to 30%. Biliary excretion of 73AsIII was lost in the presence of SeIV, SeMet and MSC. Arsenic underwent biliary excretion in HepaRG cells and this was stimulated by SeII, and inhibited by SeIV, SeMet and MSC. These data have implications for the form of selenium utilized for selenium supplementation trials in arsenic endemic regions.Support or Funding InformationCanadian Institutes of Health Research (CIHR)