Accelerated Urinary Excretion of Methylmercury following Administration of Its AntidoteN-Acetylcysteine Requires Mrp2/Abcc2, the Apical Multidrug Resistance-Associated Protein

Abstract

N-Acetylcysteine (NAC) is a sulfhydryl-containing compound that produces a dramatic acceleration of urinary methylmercury (MeHg) excretion in poisoned mice, but the molecular mechanism for this effect is poorly defined. MeHg readily binds to NAC to form the MeHg-NAC complex, and recent studies indicate that this complex is an excellent substrate for the basolateral organic anion transporter (Oat)-1, Oat1/Slc22a6, thus potentially explaining the uptake from blood into the renal tubular cells. The present study tested the hypothesis that intracellular MeHg is subsequently transported across the apical membrane of the cells into the tubular fluid as a MeHg-NAC complex using the multidrug resistance-associated protein-2 (Mrp2/Abcc2). NAC markedly stimulated urinary [14 C]MeHg excretion in wild-type Wistar rats, and a second dose of NAC was as effective as the first dose in stimulating MeHg excretion. In contrast with the normal Wistar rats, NAC was much less effective at stimulating urinary MeHg excretion in the Mrp2-deficient (TRϪ) Wistar rats. The TRϪ rats excreted only ϳ30% of the MeHg excreted by the wild-type animals. To directly test whether MeHg-NAC is a substrate for Mrp2, studies were carried out in plasma membrane vesicles isolated from livers of TRϪ and control Wistar rats. Transport of MeHg-NAC was lower in vesicles prepared from TRϪ rats, whereas transport of MeHg-cysteine was similar in control and TRϪ rats. These results indicate that Mrp2 is involved in urinary MeHg excretion after NAC administration and suggest that the transported molecule is most likely the MeHg-NAC complex.

Substrates and Inhibitors of Human Multidrug Resistance Associated Proteins and the Implications in Drug Development

“…However, the amino acid derivative NAC has more recently been shown to be effective in increasing urinary MeHg excretion in mice [329]. The MeHg-NAC complex appears to be an excellent substrate for the renal Oat1/Slc22a6, with an apparent K m value of 31 [330,331] and Mrp2/Abcc2 [324].…”

“…Mrp2/Abcc2 also participates in the urinary excretion of methylmercury (MeHg) in conjugation with the amino acid derivative Nacetyl-L-cysteine (NAC, Fig. (42)) [324]. MeHg is a common environmental toxicant that is from eating fish with bioaccumulated MeHg [325].…”

Mechanisms involved in the transport of mercuric ions in target tissues

“…MRP2, which is capable of transporting GSH, has since been identified in the canalicular membrane of hepatocytes (Ballatori and Dutczak 1994; Ballatori and Truong 1995; Fernandez-Checa et al 1992, 1993; Garcia-Ruiz et al 1992), and thus, it probably plays an important role in the export of CH 3 Hg + . Indeed, recent studies in TR − rats indicate that MRP2 is involved in the hepatobiliary elimination of mercuric ions following exposure to CH 3 Hg + (Madejczyk et al 2007; Zalups and Bridges 2009). …”

“…However, the mechanisms by which this extraction occurs have been identified only recently. Data from in vivo studies using TR − rats exposed to CH 3 Hg + and treated subsequently with NAC, DMPS, or DMSA indicate that MRP2 plays an important role in the transport of CH 3 Hg + from within proximal tubular cells into the tubular lumen (Madejczyk et al 2007; Zalups and Bridges 2009). Even in the absence of a metal chelator, MRP2 has been shown to play a role in the renal handling of Hg 2+ (Bridges et al 2011).…”

<i>S</i>-Mercuration of ubiquitin carboxyl-terminal hydrolase L1 through Cys152 by methylmercury causes inhibition of its catalytic activity and reduction of monoubiquitin levels in SH-SY5Y cells

“…Under certain conditions, MeHg-mediated toxicity involves the modification of cellular proteins through the S-mercuration reaction (Rabenstein and Saetre, 1977). Once MeHg invades cells, this environmental electrophile undergoes glutathione (GSH) conjugation, resulting in the excretion of a polar metabolite into the extracellular spaces through multidrug resistance-associated proteins (MRPs) (Toyama et al, 2007;Kumagai et al, 2013;Ballatori, 2002;Madejczyk et al, 2007). We reported previously that MeHg activates transcription factor Nrf2 through S-mercuration of its negative regulator Keap1 and up-regulates its downstream genes, such as glutamate-cysteine ligase (GCL), a latelimiting enzyme for GSH synthesis, GSH S-transferases and MRPs, resulting in detoxification of the organomercury compound (Toyama et al, 2007(Toyama et al, , 2011; Itoh et al, 1997;Vollrath et al, 2006).…”