Analyses of arsenic (As) species in tissues and body fluids of individuals chronically exposed to inorganic arsenic (iAs) provide essential information about the exposure level and pattern of iAs metabolism. We have previously described an oxidation state-specific analysis of As species in biological matrices by hydride-generation atomic absorption spectrometry (HG-AAS), using cryotrapping (CT) for preconcentration and separation of arsines. To improve performance and detection limits of the method, HG and CT steps are automated and a conventional flame-in-tube atomizer replaced with a recently developed multiple microflame quartz tube atomizer (multiatomizer). In this system, arsines from As(III)-species are generated in a mixture of Tris-HCl (pH 6) and sodium borohydride. For generation of arsines from both As(III)- and As(V)-species, samples are pretreated with L-cysteine. Under these conditions, dimethylthioarsinic acid, a newly described metabolite of iAs, does not interfere significantly with detection and quantification of methylated trivalent arsenicals. Analytical performance of the automated HG-CT-AAS was characterized by analyses of cultured cells and mouse tissues that contained mono- and dimethylated metabolites of iAs. The capacity to detect methylated As(III)- and As(V)-species was verified, using an in vitro methylation system containing recombinant rat arsenic (+3 oxidation state) methyltransferase and cultured rat hepatocytes treated with iAs. Compared with the previous HG-CT-AAS design, detection limits for iAs and its metabolites have improved significantly with the current system, ranging from 8 to 20 pg. Recoveries of As were between 78 and 117%. The precision of the method was better than 5% for all biological matrices examined. Thus, the automated HG-CT-AAS system provides an effective and sensitive tool for analysis of all major human metabolites of iAs in complex biological matrices.

No abstract

Background: Type 2 diabetes is characterized by glucose intolerance and insulin resistance. Obesity is the leading cause of type 2 diabetes. Growing evidence suggests that chronic exposure to inorganic arsenic (iAs) also produces symptoms consistent with diabetes. Thus, iAs exposure may further increase the risk of diabetes in obese individuals.Objectives: Our goal was to characterize diabetogenic effects of iAs exposure and high-fat diet (HFD) in weaned C57BL/6 mice.Methods: Mice were fed HFD or low-fat diet (LFD) while exposed to iAs in drinking water (25 or 50 ppm As) for 20 weeks; control HFD and LFD mice drank deionized water. Body mass and adiposity were monitored throughout the study. We measured glucose and insulin levels in fasting blood and in blood collected during oral glucose tolerance tests (OGTT) to evaluate the diabetogenic effects of the treatment.Results: Control mice fed HFD accumulated more fat, had higher fasting blood glucose, and were more insulin resistant than were control LFD mice. However, these diabetes indicators decreased with iAs intake in a dose-dependent manner. OGTT showed impaired glucose tolerance for both control and iAs-treated HFD mice compared with respective LFD mice. Notably, glucose intolerance was more pronounced in HFD mice treated with iAs despite a significant decrease in adiposity, fasting blood glucose, and insulin resistance.Conclusions: Our data suggest that iAs exposure acts synergistically with HFD-induced obesity in producing glucose intolerance. However, mechanisms of the diabetogenic effects of iAs exposure may differ from the mechanisms associated with the obesity-induced type 2 diabetes.

Previous epidemiologic studies found increased prevalences of type 2 diabetes mellitus in populations exposed to high levels of inorganic arsenic (iAs) in drinking water. Although results of epidemiologic studies in low-exposure areas or occupational settings have been inconclusive, laboratory research has shown that exposures to iAs can produce effects that are consistent with type 2 diabetes. The current paper reviews the results of laboratory studies that examined the effects of iAs on glucose metabolism and describes new experiments in which the diabetogenic effects of iAs exposure were reproduced in a mouse model. Here, weanling male C57BL/6 mice drank deionized water with or without the addition of arsenite (25 or 50 ppm As) for 8 weeks. Intraperitoneal glucose tolerance tests revealed impaired glucose tolerance in mice exposed to 50 ppm As, but not to 25 ppm As. Exposure to 25 and 50 ppm As in drinking-water resulted in proportional increases in the concentration of iAs and its metabolites in the liver and in organs targeted by type 2 diabetes, including pancreas, skeletal muscle and adipose tissue. Dimethylarsenic was the predominant form of As in the tissues of mice in both 25 and 50 ppm groups. Notably, the average concentration of total speciated arsenic in livers from mice in the 50 ppm group was comparable to the highest concentration of total arsenic reported in the livers of Bangladeshi residents who had consumed water with an order of magnitude lower level of iAs. These data suggest that mice are less susceptible than humans to the diabetogenic effects of chronic exposure to iAs due to a more efficient clearance of iAs or its metabolites from target tissues.

Metabolism of inorganic arsenic (iAs) is one of the key factors determining the character of adverse effects associated with exposure to iAs. Results of previous studies indicate that liver plays a primary role in iAs metabolism. This paper reviews these results and presents new data that link the capacity of human hepatocytes to metabolize iAs to the expression of specific membrane transporters. Here, we examined relationship between the expression of potential arsenic transporters (AQP9, GLUT2, P-gp, MRP1, MRP2, and MRP3) and the production and cellular retention of iAs and its methylated metabolites in primary cultures of human hepatocytes exposed for 24 h to subtoxic concentrations of arsenite. Our results show that the retention of iAs and methylarsenic metabolites (MAs) by hepatocytes exposed to sub-micromolar concentrations of arsenite correlates negatively with MRP2 expression. A positive correlation was found between MRP2 expression and the production of dimethylarsenic metabolites (DMAs), specifically, the concentration of DMAs in culture media. After exposures to high micromolar concentrations of arsenite which almost completely inhibited MAs and DMAs production, a positive correlation was found between the expression of GLUT2 and cellular retention of iAs and MAs. MRP3, AQP9, or P-gp expression had no effect on the production or distribution of iAs, MAs, or DMAs, regardless of the exposure level. Hepatocytes from seven donors used in this study did not contain detectable amounts of MRP1 protein. These data suggest that MRP2 plays an important role in the efflux of DMAs, thus, regulating kinetics of the methylation reactions and accumulation of iAs and MAs by human hepatocytes. The membrane transport of iAs by high-capacity GLUT2 transporters is not a rate-limiting step for the metabolism of arsenite at low exposure level, but may play a key role in accumulation of iAs after acute exposures which inhibit iAs methylation.