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ABSTRACT: Glutaminase C (GAC), a splicing variant of the kidney-type glutaminase (KGA) gene, is a vital mitochondrial enzyme protein that catalyzes glutamine to glutamate. Earlier studies have shown that GAC proteins in the human hepatocarcinoma cell line, HepG2, were down-regulated by diphenylarsinic acid (DPAA), but the mechanism by which DPAA induced GAC protein down-regulation remained poorly understood. Here, we showed that DPAA promoted GAC protein degradation without affecting GAC transcription and translation. Moreover, DPAA-induced GAC proteolysis was mediated by mitochondrial Lon protease. DPAA insolubilized 0.5% Triton X-100-soluble GAC protein and promoted the accumulation of insoluble GAC in Lon protease knockdown cells. DPAA destroyed the native tetrameric GAC conformation and promoted an increase in the unassembled form of GAC when DPAA was incubated with cell extracts. Decreases in the tetrameric form of GAC were observed in cells exposed to DPAA, and decreases occurred prior to a decrease in total GAC protein levels. In addition, decreases in the tetrameric form of GAC were observed independently with Lon protease. Mitochondrial heat shock protein 70 is known to be an indispensable protein that can bind to misfolded proteins, thereby supporting degradation of proteins sensitive to Lon protease. When cells were incubated with DPAA, GAC proteins that can bind with mtHsp70 increased. Interestingly, the association of mtHsp70 with GAC protein increased when the tetrameric form of GAC was reduced. These results suggest that degradation of native tetrameric GAC by DPAA may be a trigger in GAC protein degradation by Lon protease.
Journal of Biological Chemistry 04/2012; 287(22):18163-72. · 4.77 Impact Factor
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ABSTRACT: Trivalent inorganic arsenite [iAs(III)] is known to alter the expression of a number of genes associated with transcription and cell proliferation, which was thought to be one of the possible mechanisms of arsenical carcinogenesis. However, the detailed mechanisms underlying iAs(III) induction of changes in gene expression are not fully understood. Here we examine the role of histone H3 phosphorylation at serine 10 (Ser(10) ) in gene regulation when the cells were treated with iAs(III). Among the 34 genes tested, iAs(III) induced mRNA expression of JUN, FOS, EGR1, HMOX1, HSPA1A, IL8, GADD45A, GADD45B and GADD153. Phosphorylation of histone H3 Ser(10) was induced by iAs(III) in interphase cells, and was effectively blocked by the ERKs pathway inhibitor (U0126). U0126 treatment significantly reduced constitutive mRNA expression of FOS and EGR1, and dramatically suppressed the induction of FOS, EGR1 and IL8 mRNA in iAs(III)-treated cells. The other genes, which were induced by iAs(III), were not affected by U0126 treatment. When the histone H3 nonphosphorylatable mutant of serine 10 (S10A) was overexpressed in cells, iAs(III) induction of FOS, EGR1and IL8 expression was significantly decreased as compared with wild-type cells. The other genes induced by iAs(III) were not changed in S10A cells nor by U0126 treatment. In addition, S10A cells were more resistant to iAs(III) cytotoxicity. These results indicated that the phosphorylation of histone H3 at Ser(10) through the ERKs pathway in interphase cells is an important regulatory event for iAs(III)-mediated gene expression. Aberrant gene expression seems to be an important cause of cytotoxicity and may have some relation to iAs(III) carcinogenicity. Copyright © 2012 John Wiley & Sons, Ltd.
Journal of Applied Toxicology 02/2012; · 2.48 Impact Factor
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ABSTRACT: Citizens in an area of Kamisu City, Ibaraki, Japan had exhibited unusual health problems, and pollution of well water by diphenylarsinic acid (DPAA) was found in the area. We examined the effects of DPAA on various behaviors in mice. DPAA was administered to mice through free intake of drinking water for 27 weeks (subchronic exposure) or 57 weeks (chronic exposure), and behavior was examined during exposure. DPAA at 30-100 ppm increased ambulatory activity and the response rate of the shuttle type discrete conditioned avoidance response of mice. DPAA reduced coordination ability on the fixed rod at 100 ppm. DPAA at 7.5-15 ppm also reduced coordination on the rotating rod, although these doses of DPAA did not affect coordination on the fixed rod. Chronic exposure to 7.5-15 ppm of DPAA produced anti-anxiety-like effects in the elevated plus maze test, whereas subchronic exposure to 100 ppm of DPAA produced anxiogenic-like effects. Neither subchronic nor chronic exposure to 7.5-100 ppm of DPAA affected learning ability and/or memory, as evaluated using the passive avoidance response. Exposure to 15-30 ppm of DPAA for 52 weeks did not alter weights of the cerebrum and cerebellum or amounts of neuron marker protein TUJ-1 or astrocyte marker protein glial fibrillary acidic protein in the cerebellum of mice. Behavioral effects observed in mice seem relevant to symptoms observed in patients from Kamisu City.
Neurotoxicology and Teratology 08/2011; 34(1):143-51. · 2.98 Impact Factor
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ABSTRACT: Trivalent dimethylarsinous acid [DMA(III)] has been shown to induce mitotic abnormalities, such as centrosome abnormality, multipolar spindles, multipolar division, and aneuploidy, in several cell lines. In order to elucidate the mechanisms underlying these mitotic abnormalities, we investigated DMA(III)-mediated changes in histone H3 phosphorylation and localization of Aurora B kinase, which is a key molecule in cell mitosis. DMA(III) caused the phosphorylation of histone H3 (ser10) and was distributed predominantly in mitotic cells, especially in prometaphase cells. By contrast, most of the phospho-histone H3 was found to be localized in interphase cells after treatment with inorganic arsenite [iAs(III)], suggesting the involvement of a different pathway in phosphorylation. DMA(III) activated Aurora B kinase and slightly activated ERK MAP kinase. Phosphorylation of histone H3 by DMA(III) was effectively reduced by ZM447439 (Aurora kinase inhibitor) and slightly reduced by U0126 (MEK inhibitor). By contrast, iAs(III)-dependent histone H3 phosphorylation was markedly reduced by U0126. Aurora B kinase is generally localized in the midbody during telophase and plays an important role in cytokinesis. However, in some cells treated with DMA(III), Aurora B was not localized in the midbody of telophase cells. These findings suggested that DMA(III) induced a spindle abnormality, thereby activating the spindle assembly checkpoint (SAC) through the Aurora B kinase pathway. In addition, cytokinesis was not completed because of the abnormal localization of Aurora B kinase by DMA(III), thereby resulting in the generation of multinucleated cells. These results provide insight into the mechanism of arsenic tumorigenesis.
Toxicology and Applied Pharmacology 09/2009; 241(3):275-82. · 4.45 Impact Factor
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ABSTRACT: Diphenylarsinic acid [DPAA(V)] was detected in ground water used as drinking water after a poisonous incident in Kamisu, Japan. An approach to define the target molecules of DPAA(V) with a high throughput analysis of proteins from cultured human cells demonstrated down-regulation of glutaminase C (GAC). GAC is a splicing variant of the kidney-type glutaminase (KGA) gene and has the enzyme activity of phosphate-activated glutaminase (PAG). To gain some insights into the mechanism of arsenic intoxication in Kamisu, the effects of various arsenic compounds, including arsenicals that were detected in ground water ([DPAA(V)], phenylarsonic acid [PAA(V)] and bis(diphenylarsine)oxide [BDPAO(III)]) and rice (phenylmethylarsinic acid [PMAA(V)]), were investigated for the expression of GAC and PAG activity. When cultured human HepG2 cells were incubated with arsenicals for 24h, the pentavalent phenylarsenic form of PAA(V) and PMAA(V) as well as DPAA(V) suppressed the expression of GAC protein and PAG activity in a concentration-dependent manner. On the other hand, the trivalent phenylarsenic form of BDPAO(III) had no suppressive effect on GAC and PAG. In addition, trivalent phenylarsenic compounds, such as the glutathione (GSH) conjugate of DPAA(V) [DPA-GS (III)] and triphenylarsine [TPA(III)], and the inorganic arsenics, iAs(V) and iAs(III), and methylated metabolites of inorganic arsenics, dimethylarsinic acid [DMA(V)] and dimethylarsinous acid [DMA(III)], had no suppressive effect on glutaminase. Likewise, methyl substituents of the hydroxyl groups of DPAA(V), PAA(V) and PMAA(V), diphenylmethylarsine oxide [DPMAO(V)] and phenyldimethylarsine oxide [PDMAO(V)], did not have any suppressive effects. These results suggest that pentavalent arsenic compounds with both phenyl groups and hydroxyl groups are effective in the suppression of glutaminase. In addition, the fact that it was only the arsenicals detected in Kamisu that were effective in suppressing glutaminase provides insights into the cause of the arsenic intoxication at Kamisu.
Toxicology 05/2009; 258(2-3):157-63. · 3.68 Impact Factor
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Ping Gao,
Irina Tchernyshyov,
Tsung-Cheng Chang,
Yun-Sil Lee,
Kayoko Kita, Takafumi Ochi,
Karen I Zeller,
Angelo M De Marzo,
Jennifer E Van Eyk,
Joshua T Mendell,
Chi V Dang
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ABSTRACT: Altered glucose metabolism in cancer cells is termed the Warburg effect, which describes the propensity of most cancer cells to take up glucose avidly and convert it primarily to lactate, despite available oxygen. Notwithstanding the renewed interest in the Warburg effect, cancer cells also depend on continued mitochondrial function for metabolism, specifically glutaminolysis that catabolizes glutamine to generate ATP and lactate. Glutamine, which is highly transported into proliferating cells, is a major source of energy and nitrogen for biosynthesis, and a carbon substrate for anabolic processes in cancer cells, but the regulation of glutamine metabolism is not well understood. Here we report that the c-Myc (hereafter referred to as Myc) oncogenic transcription factor, which is known to regulate microRNAs and stimulate cell proliferation, transcriptionally represses miR-23a and miR-23b, resulting in greater expression of their target protein, mitochondrial glutaminase, in human P-493 B lymphoma cells and PC3 prostate cancer cells. This leads to upregulation of glutamine catabolism. Glutaminase converts glutamine to glutamate, which is further catabolized through the tricarboxylic acid cycle for the production of ATP or serves as substrate for glutathione synthesis. The unique means by which Myc regulates glutaminase uncovers a previously unsuspected link between Myc regulation of miRNAs, glutamine metabolism, and energy and reactive oxygen species homeostasis.
Nature 03/2009; 458(7239):762-5. · 36.28 Impact Factor
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Ping Gao,
Irina Tchernyshyov,
Tsung-Cheng Chang,
Yun-Sil Lee,
Kayoko Kita, Takafumi Ochi,
Karen I. Zeller,
Angelo M. De Marzo,
Jennifer E. Van Eyk,
Joshua T. Mendell,
Chi V. Dang
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ABSTRACT: Altered glucose metabolism in cancer cells is termed the Warburg effect, which describes the propensity of most cancer cells to take up glucose avidly and convert it primarily to lactate, despite available oxygen
Nature 02/2009; 458(7239):762-765. · 36.28 Impact Factor
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ABSTRACT: New methods involving high-performance liquid chromatography/inductively coupled plasma mass spectrometry were examined for the determination of phenylarsenic compounds derived from chemical warfare agents. Several methods were examined for the separation of diphenylarsinic acid (DPAA), phenylarsonic acid, phenylmethylarsinic acid (PMAA), phenyldimethylarsine oxide, and diphenylmethylarsine oxide. Analysis of the urine samples of the patients exposed to phenylarsenic compounds indicated that the main phenylarsenic components were DPAA and PMAA; moreover, some unknown arsenicals, which were also found in contaminated groundwater and rice samples, were also detected.
Journal of Chromatography B 06/2008; 867(2):179-88. · 2.89 Impact Factor
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ABSTRACT: Thio-dimethylarsinate (thio-DMA), a recently discovered urine metabolite in humans, was investigated for its cytotoxic, genotoxic and cell-cycle disruptive effects in the cultured human hepatocarcinoma cell line, HepG2, and Syrian hamster embryo cells. In addition, the role of glutathione (GSH) on the cytotoxic effects of thio-DMA was investigated in terms of the effects of GSH depletion and the effects of exogenously added GSH. LC50 values of arsenicals for cells incubated for 48 h were 0.026 mM for thio-DMA, 0.343 mM for DMA and 3.66 mM for dithio-DMA. Depletion of cell GSH reduced the cytotoxic effects of thio-DMA. The cytotoxic effects of 0.02 mM and 0.05 mM thio-DMA were enhanced markedly when used in combination with 1 to 3 mM GSH, but decreased again when combined with 5 mM GSH. These results suggested that cytotoxic intermediates were generated by the interaction of thio-DMA with GSH, while an excessive amount of GSH suppressed the generation of these intermediates. Flow-cytometry showed that thio-DMA was an inducer of cells with 4N DNA and hypo 2N DNA. The results also demonstrated that cells arrested in the mitotic phase had abnormalities in their spindle organization and centrosome integrity. In addition, cells arrested in mitosis by thio-DMA had chromosome structural aberrations, such as chromatid gaps, chromatid breaks and chromatid exchanges. Moreover, the cytotoxic effects of thio-DMA may in part be associated with an apoptotic mode of cell death that was evaluated by the appearance of nucleosome level DNA fragmentations and an 85-kDa cleavage fragment of poly (ADP-ribose) polymerase. These findings suggest that the presence of thio-DMA in human urine has implications for human health in terms of arsenic metabolism and toxicity.
Toxicology and Applied Pharmacology 05/2008; 228(1):59-67. · 4.45 Impact Factor
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ABSTRACT: Over the last 6 years, much work on arsenic species in urine samples has been directed toward the determination of the reduced dimethylated arsenic species, DMA(III), because of its high toxicity and perceived key role in the metabolism of inorganic arsenic. Recent work, however, has suggested that DMA(III) may at times have been misidentified because its chromatographic properties can be similar to those of thio-dimethylarsinate (thio-DMA). We analyzed by HPLC-ICPMS (inductively coupled plasma mass spectrometry) urine samples from 75 arsenic-exposed women from Bangladesh with total arsenic concentrations ranging from 8 to 1034 microg As/L and found that thio-DMA was present in 44% of the samples at concentrations ranging mostly from trace amounts to 24 microg As/L (one sample contained 123 microg As/L). Cytotoxicity testing with HepG2 cells derived from human hepatocarcinoma indicated that thio-DMA was about 10-fold more cytotoxic than dimethylarsinate (DMA). The widespread occurrence of thio-DMA in urine from these arsenic-exposed women suggests that this arsenical may also be present in other urine samples and has so far escaped detection. The work highlights the need for analytical methods providing specific determinations of arsenic compounds in future studies on arsenic metabolism and toxicology.
Toxicology and Applied Pharmacology 09/2007; 222(3):374-80. · 4.45 Impact Factor
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ABSTRACT: In a poisonous incident in Kamisu, Japan, it is understood that diphenylarsinic acid (DPAA) was a critical contaminant of ground water. Most patients showed dysfunction of the central nervous system. To understand the overall mechanism of DPAA toxicity and to gain some insight into the application of a remedy specific for intoxication, the molecular target must be clarified. As an approach, a high throughput analysis of cell proteins in cultured human hepatocarcinoma HpG2 exposed to DPAA was performed by two-dimensional electrophoresis (2-DE). Four proteins, which were up- and down-regulated by exposure of cultured HepG2 cells to DPAA, were identified. They were chaperonin containing TCP-1 (CCT) beta subunit, aldehyde dehydrogenase 1 (ALDH1), ribosomal protein P0 and glutaminase C (GAC). Of these, GAC was the only protein that was down-regulated by DPAA exposure, and cellular expression levels were reduced by DPAA in a concentration- and time-dependent manner. Decrease in cellular GAC levels was accompanied by decreased activity of the enzyme, phosphate-activated glutaminase (PAG). Decreased expression of GAC by DPAA was also observed in human cervical carcinoma HeLa and neuroblastoma SH-SY5Y cells. By contrast, no significant changes in GAC protein expression were observed when cells were incubated with arsenite [iAs (III)] and trivalent dimethylarsinous acid [DMA (III)]. In the central nervous system, GAC plays a role in the production of the neurotransmitter glutamic acid. Selective inhibition of GAC expression by DPAA may be a cause of dysfunction of glutamatergic neuronal transmission and the resultant neurological impairments.
Toxicology and Applied Pharmacology 06/2007; 220(3):262-70. · 4.45 Impact Factor
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ABSTRACT: The role of glutathione (GSH) in the cytotoxicity of diphenylarsinic acid [DPAA(V)], which was detected in drinking well water after a poisoning incident in Kamisu, Japan, was investigated in cultured human HepG2 cells. DPA-GS(III), which is the GSH adduct of DPAA, was synthesized and analyzed by HPLC/ESI-MS. DPA-GS(III) was highly toxic to cells and the potency was about 1000 times that of DPAA(V). DPAA(V) was stable in culture medium, while DPA-GS(III) was unstable and changed to protein-bound As (protein-As). By contrast, DPA-GS(III) remained stable with the addition of exogenous GSH, thereby reducing transformation to protein-As. In addition, DPA-GS(III) was transformed to bis(diphenylarsine)oxide [BDPAO(III)], which was observed under serum-free conditions. BDPAO(III) was very unstable and disappeared conversely with an increase in protein-As. In contrast, the presence of GSH suppressed the transformation of BDPAO(III) to protein-As while it enhanced the transformation of BDPAO(III) to DPA-GS(III). Depletion of cell GSH enhanced the cytotoxic effects of DPA-GS(III) and BDPAO(III). Moreover, exogenously-added GSH suppressed the cytotoxic effects of DPA-GS(III) and BDPAO(III). The dynamic behavior of arsenicals in the culture medium and the resultant cytotoxic effects suggested that GSH played a role in regulating the formation of toxic intermediates, such as DPA-GS(III) and BDPAO(III). Moreover, the results suggested that the formation of protein-As in culture medium was compatible with the cytotoxic effects and that GSH was a factor capable of regulating the formation of protein-As from either DPA-GS(III) or BDPAO(III).
Toxicology 09/2006; 225(2-3):142-9. · 3.68 Impact Factor
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ABSTRACT: The mechanism underlying enhancement of the cytotoxic effects of diphenylarsinic acid (DPAA) by sulfhydryl (SH) compounds, such as glutathione (GSH) and dimercaptopropane sulfonate (DMPS), was investigated in terms of not only the effects of SH compounds on DPAA uptake by cells, but also the cytotoxic effects of the GSH adduct of DPAA, DPA-GS. In addition, the cytotoxic effects of DPA-GS and cellular uptake were investigated in conjunction with the effects of GSH depletion. Cells took up DPAA in a time- and temperature-dependent manner for up to 2 h, then the uptake leveled off for 6 h. Arsenic species other than DPAA were not detected in the cells. The presence of GSH and DMPS did not influence the rate of uptake of DPAA by the cells. By contrast, when the cytotoxic potential of DPA-GS was compared with that of DPAA, DPA-GS was about 1,000 times more toxic than DPAA, suggesting that enhancement of DPAA toxicity by SH compounds might be due to the formation of adducts in the culture medium. The cytotoxic effects of DPA-GS were suppressed markedly by the presence of GSH and DMPS, and the suppression was attributed to an inhibition of more than 90% by the SH compounds of DPA-GS uptake. Depletion of cell GSH enhanced the cytotoxic effects of DPA-GS by two to three times and the enhancement attributed to an increased cellular uptake of DPA-GS. These results suggest that GSH plays a role in regulating the formation of DPA-GS and cellular uptake.
Archive für Toxikologie 09/2006; 80(8):486-91. · 4.67 Impact Factor
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ABSTRACT: The presence of arsenic-containing carbohydrates, arsenosugars, in many seafoods raises questions of human health concerning the ingestion and metabolism of these compounds. A previous study investigating the metabolites in human urine after the ingestion of a common arsenosugar 2',3'-dihydroxypropyl 5-deoxy-5-dimethylarsinoyl-beta-d-riboside (oxo-arsenosugar) showed that the arsenic was rapidly excreted in the urine and was present as at least 12 metabolites, only three of which could be identified. In this repeat study with oxo-arsenosugar and using high-performance liquid chromatography/inductively coupled plasma mass spectrometry, we report the identification of seven arsenic metabolites, which together accounted for 88% of the total urinary arsenic collected over 61 h. The metabolites included previously reported human urinary arsenicals dimethylarsinate (DMA), oxo-dimethylarsenoethanol (oxo-DMAE), and trimethylarsine oxide, in addition to new human metabolites oxo-dimethylarsenoacetate (oxo-DMAA), thio-dimethlyarsenoacetate (thio-DMAA), thio-dimethylarsenoethanol (thio-DMAE), and the thio-arsenosugar. Cytotoxicity testing of the major metabolites DMA, oxo-DMAE, thio-DMAE, oxo-DMAA, and thio-DMAA showed that they were nontoxic even at 10 mM, except for DMA, which showed some toxic effects at 1 mM.
Chemical Research in Toxicology 10/2005; 18(9):1444-50. · 3.78 Impact Factor
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ABSTRACT: To obtain quantitative information on human metabolism of selenium, we have performed selenium speciation analysis by HPLC/ICPMS on samples of human urine from one volunteer over a 48-hour period after ingestion of selenium (1.0 mg) as sodium selenite, L-selenomethionine, or DL-selenomethionine. The three separate experiments were performed in duplicate. Normal background urine from the volunteer contained total selenium concentrations of 8-30 microg Se/L (n=22) but, depending on the chromatographic conditions, only about 30-70% could be quantified by HPLC/ICPMS. The major species in background urine were two selenosugars, namely methyl-2-acetamido-2-deoxy-1-seleno-beta-D-galactopyranoside (selenosugar 1) and its deacylated analog methyl-2-amino-2-deoxy-1-seleno-beta-D-galactopyranoside (selenosugar 3). Selenium was rapidly excreted after ingestion of the selenium compounds: the peak concentrations (approximately 250-400 microg Se/L, normalized concentrations) were recorded within 5-9 hours, and concentrations had returned to close to background levels within 48 hours, by which time 25-40% of the ingested selenium, depending on the species ingested, had been accounted for in the urine. In all experiments, the major metabolite was selenosugar 1, constituting either approximately 80% of the total selenium excreted over the first 24 hours after ingestion of selenite or L-selenomethionine or approximately 65% after ingestion of DL-selenomethionine. Selenite was not present at significant levels (<1 microg Se/L) in any of the samples; selenomethionine was present in only trace amounts (approximately 1 microg/L, equivalent to less than 0.5% of the total Se) following ingestion of L-selenomethionine, but it constituted about 20% of the excreted selenium (first 24 hours) after ingestion of DL-selenomethionine, presumably because the D form was not efficiently metabolized. Trimethylselenonium ion, a commonly reported urine metabolite, could not be detected (<1 microg/L) in the urine samples after ingestion of selenite or selenomethionine. Cytotoxicity studies on selenosugar 1 and its glucosamine isomer (selenosugar 2, methyl-2-acetamido-2-deoxy-1-seleno-beta-D-glucosopyranoside) were performed with HepG2 cells derived from human hepatocarcinoma, and these showed that both compounds had low toxicity (about 1000-fold less toxic than sodium selenite). The results support earlier studies showing that selenosugar 1 is the major urinary metabolite after increased selenium intake, and they suggest that previously accepted pathways for human metabolism of selenium involving trimethylselenonium ion as the excretionary end product may need to be re-evaluated.
Analytical and Bioanalytical Chemistry 09/2005; 383(2):235-46. · 3.78 Impact Factor
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Applied Organometallic Chemistry 01/2005; 19(2):287 - 293. · 2.06 Impact Factor
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ABSTRACT: The cytotoxicity and lipid peroxidative potency of the organochlorine fungicides captan (N-(trichloromethylthio)-4-cyclohexene-1,2-dicarboximide), dichlofluanid (N-dichlorofluoromethylthio-N'N'-dimethyl-N-phenylsulfamide) and chlorothalonil (2,4,5,6-tetrachloro-isophthalonitrile) were studied in isolated rat hepatocytes. These fungicides induced cytotoxicity and lipid peroxidation in a dose- and time-dependent manner. Considerable cytotoxicity and lipid peroxidation occurred after cells were treated with 25 microM and more of fungicide. The phosphatidylcholine hydroperoxide (PCOOH) content increased more than 300 times by captan (250-1000 microM), 400 times by dichlofluanid (250-1000 microM) and 20 times by chlorothalonil (25-1000 microM) after 1h of incubation, as compared with untreated control. Significant cytotoxicity occurred after 20 min (captan), 30 min (dichlofluanid) and 60 min (chlorothalonil) of incubation and lipid peroxidation was induced prior to cytotoxicity. The antioxidant alpha-tocopherol and cytochrome P450 inhibitor SKF-525A effectively prevented cytotoxicity and lipid peroxidation. Our results suggest that metabolites of these fungicides produced by the microsomal cytochrome P450 system, induced membrane phospholipid peroxidation that caused cytotoxicity.
Toxicology 12/2004; 204(2-3):97-107. · 3.68 Impact Factor
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ABSTRACT: The abilities of dimethylarsine iodide (DMI), a model compound of trivalent dimethylarsenicals, to induce cellular transformation, aneuploidy, centrosome abnormality, and multipolar spindle formations were investigated using the Syrian hamster embryo (SHE) cell model. Cellular growth was decreased in a concentration-dependent manner by treatment with DMI at concentrations over 0.1 microM. Treatment with DMI at concentrations from 0.1 to 1.0 microM induced morphological transformation in SHE cells. The transforming activity of DMI, determined by the frequency of morphologically transformed colonies, was approximately 30 times higher than that induced by treatment with the same concentration of sodium arsenite. Flow cytometry suggested an increase in the aneuploid population caused by DMI, as shown by the appearance of hypo-2N, hypo-4N and hypo-8N. DMI also caused abnormal staining of gamma-tubulin, indicating loss of centrosome integrity and a resultant induction of multipolar spindles in mitotic cells. Mitotic cells with centrosomes that coalesced partly at the cell periphery, not the cell center, were detected as early changes that resulted in multipolar spindles. These findings indicate that DMI has transforming activity in SHE cells. Moreover, the results suggest the importance of centrosome abnormalities as a causal change of DMI-induced aneuploidy.
Toxicology 11/2004; 203(1-3):155-63. · 3.68 Impact Factor
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ABSTRACT: Diphenylarsinic acid [DPAs(V)], a degradation product of diphenylcyanoarsine or diphenylchloroarsine, both of which were developed as chemical warfare agents, was investigated in terms of its capacity to induce cytotoxic effects, numerical and structural changes of chromosomes, and abnormalities of centrosome integrity and spindle organizations in conjunction with the effects of glutathione (GSH) depletion. DPAs(V) had toxic effects on cultured human hepatocarcinoma HepG2 cells at concentrations more than 0.5 mM. Depletion of GSH reduced the toxic effects of DPAs(V) as well as dimethylarsinic acid [DMAs(V)] toxicity, while toxicity by arsenite [iAs(III)] was enhanced. Exogenously added sulfhydryl (SH) compounds, such as dimercapropropane sulfonate (DMPS), GSH, and dithiothreitol (DTT), enhanced the toxic effects of DPAs(V) while they suppressed iAs(III) toxicity. DPAs(V) caused an increase in the mitotic index, and also structural and numerical changes in chromosomes in V79 Chinese hamster cells. Abnormality of centrosome integrity in mitotic V79 cells and multipolar spindles was also induced by DPAs(V) in a time- and concentration-dependent manner. These results suggested that highly toxic chemicals were generated by the interaction of DPAs(V) with SH compounds. Moreover, enhancements of toxicity by a combination of DPAs(V) and SH compounds suggested a risk in the use of SH compounds as a remedy for intoxication by diphenylarsenic compounds. Investigations on the effects of SH compounds on animals intoxicated with DPAs(V) are warranted.
Toxicology and Applied Pharmacology 11/2004; 200(1):64-72. · 4.45 Impact Factor
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ABSTRACT: Dimethylarsine iodide (DMI) was used as a model compound of trivalent dimethylarsenicals [DMA(III)], and the biological effects were extensively investigated in cultured Chinese hamster V79 cells. When the cytotoxic effects of DMA(III) were compared with those of inorganic arsenite and dimethylarsinic acid [DMA(V)], DMA(III) was about 10,000 times more potent than DMA(V), and it was even 10 times more toxic than arsenite. Depletion of cell glutathione (GSH) did not influence the cytotoxic effects of DMA(III), whereas it enhanced the cytotoxicity of arsenite. Chromosome structural aberrations, such as gaps, breaks and pulverizations, and numerical changes, such as aneuploidy, hyper- and hypo-tetraploidy, were induced by DMA(III) in a concentration-dependent manner. Mitotic index increased 9-12h after the addition of DMA(III), and then declined. By contrast, the incidence of multinucleated cells increased conversely with the decrease in mitotic index at and after 24h of exposure. The mitotic cell-specific abnormality of centrosome integrity and multipolar spindles were induced by DMA(III) in a time- and concentration-dependent manner. Moreover, DMA(III) caused abnormal cytokinesis (multipolar division) at concentrations that were effective in causing centrosome abnormality, multipolar spindles and aneuploidy. These results showed that DMA(III) was genotoxic on cultured mammalian cells. Results also suggest that DMA(III)-induced multipolar spindles and multipolar division may be associated with the induction of aneuploidy. In addition, the centrosome may be a primary target for cell death via multinucleated cells.
Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis 10/2003; 530(1-2):59-71. · 2.85 Impact Factor
