Subcellular Distribution of Inorganic and Methylated Arsenic Compounds in Human Urothelial Cells and Human Hepatocytes

Abstract

ABSTRACT:Epidemiological studies have indicated that exposure of humans to inorganic arsenic in drinking water is associated with the occurrence of bladder cancer. The mechanisms by which arsenic induces this malignancy are still uncertain; however, arsenic metabolites are suspected to play a pivotal role. The aim of the present study was the investigation of uptake capabilities of human urothelial cells (UROtsa) compared with primary human hepatocytes (phH) as well as the intracellular distribution of the arsenic species. Additionally, we were interested in the cyto-and genotoxic potential (comet assay, radical generation) of the different arsenic compounds in these two cell types. Our results show that UROtsa cells accumulate higher amounts of the arsenic species than the phH. Differential centrifugation revealed that the arsenic compounds are preferentially distributed into nuclei and ribosomes. After 24-h exposure, arsenic is mainly found in the ribosomes of UROtsa cells and in the nuclei and mitochondria of phH. In contrast to the pentavalent arsenic species, the trivalent species induced a 4-to 5-fold increase of DNA damage in hepatocytes. Radical generation, measured by thiobarbituric acid reactive substances, was more pronounced in hepatocytes than in urothelial cells. In summary, the uptake of arsenic compounds appears to be highly dependent upon cell type and arsenic species. The nonmethylating urothelial cells accumulate higher amounts of arsenic species than the methylating hepatocytes. However, cyto-and genotoxic effects are more distinct in hepatocytes. Further studies are needed to define the implications of the observed accumulation in cellular organelles for the carcinogenic activity of arsenic.The association between arsenic exposure and urinary bladder cancer, typically transitional cell carcinomas, has been observed in the same endemic areas of the world in which skin cancer populations have been identified (Chiou et al., 1995). In addition to bladder and skin cancer, chronic arsenic exposure causes several malignant and nonmalignant human diseases [for review, see Tseng (2007) (Challenger, 1945) consists of a series of reductions and oxidative methylations. In the sequence of reactions, the ϩ5 oxidative arsenic species are always formed before the analogous ϩ3 arsenic species.Recently, Hayakawa et al. (2005) proposed a new metabolic pathway for arsenic biotransformation in which the ϩ3 arsenic species are formed before the respective ϩ5 species. The trivalent metabolites are oxidized by hydrogen peroxide or other agents to the pentavalent species, which are considered end products of arsenic metabolism.It is generally accepted that the ϩ3 methylated arsenic species are more cyto-and genotoxic (e.g., Styblo et al., 2000;Mass et al., 2001;Aposhian et al., 2003;Kligerman et al., 2003;Dopp et al., 2004) and are more potent enzyme inhibitors (e.g., Styblo et al., 1997aStyblo et al., , 2002Schuliga et al., 2002;Chang et al., 2003) than the pentavalent counThis work was kindly supported by the Ge...

CHAPTER 17. Arsenic, Antimony and Bismuth

“…Dopp et al showed that inorganic and methylated arsenicals generate reactive oxygen radicals in cultured human hepatocytes cultures and that the action was cell specific. 199,200 In each case, As(III) ion was more effective in inducing cytotoxic change and ROS induction than As(V) ion. In mouse lymphoma cell lines, As(III) compounds such as sodium arsenite and arsenic trioxide invoked a clear dose-related pattern of gene mutation as a basis for carcinogenic change.…”

“…Whereas, in human studies, the As(III) ion appears to target epithelial cells in the skin and urinary bladder, hepatocytes and pneumocytes, cellular responses in cultured cells are markedly different. 164,199,200 This is attributed to the greater ability of cytotoxic As(III) cation to traverse cell membranes, methylate MMA and DMA and to accumulate in and bind to cytoplasmic proteins. Human urothelial cells in vitro accumulated As(III) more readily than hepatocytes but methylated complexes migrated more readily to ribosomes, nuclei and mitrochondria in hepatocytes.…”

Methylarsenic compounds [MAK Value Documentation, 2014]

“…In most studies, the toxicity of methylarsonous acid and dimethylarsinous acid was greater than that of inorganic arsenite or methylarsonic acid or dimethylarsinic acid, as has been found in studies of cytotoxicity in Chang hepatocytes (ATCC CCL-13) (Petrick et al 2000;Styblo et al 2000), or that of the formation of radicals in human urothelial cells and primary human hepatocytes (Dopp et al 2008), or of DNA damage in plasmids (Andrewes et al 2003) or HeLa-S3 cells or that of the clastogenic effects in human lymphocytes (Kligerman et al 2003), or the induction of apoptosis in leukaemia cells (Chen et al 2003) or the proliferation of keratinocytes (Mudipalli et al 2005).…”

Intracellular Arsenic Speciation and Quantification in Human Urothelial and Hepatic Cells

“…Especially the trivalent arsenicals are known to exhibit strong genotoxic effects (Dopp et al, 2010a). These effects can either be detected as DNA damage using the micronucleus test or the comet assay (Dopp et al, 2008), as oxidative damage by assaying the amount of 8-oxo-dG (8-Oxo-2'-deoxyguanosine) or in form of chromosomal aberrations (Dopp et al, 2004). In our study we determined the Alkaline Comet Assay to analyse single and double strand breaks of the DNA.…”