Purpose: Selenite (H₂SeO₃) reacts with glutathione resulting in the formation of selenodiglutathione (GSSeSG) during absorption processes. GSSeSG has been reported to show cytotoxic effect in cancer cells, however the reaction mechanisms remained unclear. In this study, we investigated cytotoxicity and DNA oxidation properties of GSSeSG as well as the mechanisms of action.
Methods: Human breast cancer cell (MCF-7) viability, living cell specific fluorescence reagent; Apoptosis, annexin V staining; Oxidative DNA damage, 8-oxo-7,8-dihydro-2’-deoxyguanosine (8-oxodG) formation; DNA strand break, agarose gel electrophoresis; Superoxide anion radical (O･₂-) detection, hydroxylammonium/sulfanilic acid/N-(1-naphtyl) ethylenediamine dihydrochloride.
Results and discussion: GSSeSG decreased the viability and increased apoptotic cells. In the DNA prepared from the GSSeSG-treated cells, 8-oxodG level was increased in a dose-dependent fashion, indicating that GSSeSG causes oxidative stress in the cells. GSSeSG induced strand breaks in purified calf thymus DNA in the presence of GSH. O･₂- generation was confirmed in the reaction system. GSSeSG disappeared immediately after addition into culture media. We assume that membrane permeable hydrogen selenite (H₂Se) produced via further reduction of GSSeSG could be taken into the cells and react with cellular GSH resulting in the generation of O･₂-. GSSeSG would be a clue to unveil cellular effects of Se including anticancer activity.

抄録全体を表示

Purpose: Platinum(IV) [Pt(IV)] complexes exert anti-cancer activity via endogenous reductants-mediated reduction to Pt(II) complexes. The reduction efficiency of Pt(IV) complexes seems to be fluctuating depending on ligand structure of Pt(IV) complexes. In this study, we determined that cytotoxicity and DNA damaging potency, reduction characteristic of three Pt(IV) complexes, cis-(NH₃)₂-Cl₄-Pt(IV) [cis-Pt(IV)], trans-(NH₃)₂-Cl₄-Pt(IV) [trans-Pt(IV)], and cis,cis,trans-(NH₃)₂-Cl₂-(OH)₂-Pt(IV) (oxoplatin).
Methods: Human ovarian cancer cell lines (A2780 and cisplatin resistant subline A2780cis) were used for cytotoxicity assay. Oxidative DNA damage was determined base on the amount of 8-oxo-7,8-dihydro-2’-deoxyguanosine (8-oxodG). DNA-crosslink formation was measured as ethidium bromide elimination in agarose gel electrophoresis.
Results and discussion: cis-Pt(IV) showed the most potent cytotoxicity in both cell lines compared to the other Pt-complexes. Since there is no detectable effect in A2780cis, trans-Pt(IV) moderately decreased cell viability in A2780 cells. However, trans-Pt(IV) induced comparable DNA-crosslink formation with cis-Pt(IV) does. Thiol reductants such as gluthatione efficiently inhibited DNA-crosslinking by trans-Pt(IV) as compared with cis-Pt(IV), suggesting that trans-Pt(IV) readily reacts with biomolecules after reduction, resulting in an inactivation. Oxoplatin decreased cell viability effectively in A2780 cells and slightly in A2780cis cells. Oxoplatin needed 5 times more AsA than cis-Pt(IV) for DNA-crosslinking. These results indicate cytotoxic effect of Pt(IV) complexes is strongly associated with coodination structure and physioligical property of ligands. Therefore, a structure-activity relationship study could clarify critical rules of reaction between ligands and reductants, expecting to contribute further development of much better Pt(IV) complexes.

抄録全体を表示

Arsenic is known to be an environmental toxic metalloid that causes various cancers and skin disorders, but its mechanism of toxicity is not yet clearly understood. Many chemical substances, such as arsenic comounds, might induce cytotoxicity by impairing the functions essential for cell growth. In this study, we performed a screen for essential cell growth related-genes involved in protection against toxicity of arsenite, an arsenic compound, using budding yeast as a eukaryotic cell model. We identified several genes (FAP7, ERB1 and SPT16 etc.) as arsnite-resistant gene. Most of these factors had not previously been suggested to be related the arsenite toxicity. Fap7 is a putative P-loop NTPase essential for the processing of 20S Pre-rRNA in ribosome biogenesis. To clarify relationship between P-loop NTPase and arsenite toxicity, we examined sensitivity of yeast cells overexpressing one of other P-loop NTPases. Overexpression of P-loop NTPases, expect for Fap7, did not affect sensitivity of yeast cells to arsenite, suggesting that not all P-loop NTPases confer arsenite resistance on yeast cells. Furthermore, when a Fap7(K20R) mutant with decreased activity in processing of 20S Pre-rRNA was overexpressed, the sensitivity to arsenite of the yeast cells was similar to that of the control strain. Our result suggest that the processing of 20S Pre-rRNA by Fap7 in ribosome biogenesis might be involved in protection against arsenite toxicity.

抄録全体を表示

Cisplatin (cis-diamminedichloroplatinum, CDDP), used widely in cancer chemotherapeutic agent, is a well-known kidney toxicant to caused a proximal tubular apoptosis and necrosis. Previous studies have been reported that age-related differences in CDDP-induced nephrotoxicity were controversy in clinical or animal studies. The objective of this study is to investigate the age-dependent sensitivity to nephrotoxicity induced by cisplatin. In the present study, young (3 weeks old) and old (20 weeks old) rats were single intraperitoneally injected with CDDP (10 mg/kg). After 3 days, all animals were sacrificed and serum, urine, and kidney tissues were collected. In urine analysis, lactate dehydrogenase (LDH), total protein, and glocose levels were markedly increased in young rats compared to old rats. Blood urea nitrogen (BUN) and creatinine levels were significantly decreased in CDDP-treated rats compared to control. In serum analysis, BUN, creatinine, AST, and glucose levels were significantly increased, but age-related difference was not observed. In the case of kidney toxicity biomarkers, NAGL, VEGF, clusterin levels were significantly increased in in CDDP-treated rats, whereas calbindin levels were markedly decreased. The histological changes of kidney after cisplatin treatment revealed acute tubular necrosis, which is more severe in young rats compared to old rats. The results indicate that young rats may be most sensitive, whereas old rats may be resistant to CDDP-induced nephrotoxicity, which may be related to the accumulation of CDDP.

抄録全体を表示

Methylmercury (MeHg) is a widely distributed environmental toxicant that causes disorders in central nervous system (CNS). However, the mechanism of MeHg toxicity remains unclear. Our previous study indicated that MeHg increases expressions of several chemokines (CCL2, 4, 7, etc.) in the cerebellum of mice. Chemokines are classified into four subfamilies, CC, CXC, C and CX3C chemokines. In this study, we exhaustively investigated effects of MeHg on the expressions of all chemokines in various tissues (cerebrum, cerebellum, kidney, liver and spleen) of MeHg-treated mice by using the quantitative real-time PCR (qRT-PCR). Significantly increased levels of expressions of CC chemokines (CCL2, 3, 4, 7, 9, 11, 12 and 17) and CXC chemokines (CXCL5, 10 and 16) were observed in the brain (cerebrum and cerebellum) of MeHg-treated mice. On the other hand, there were no significant effects of methylmercury on expression levels of C and CX3C chemokines in the brain of MeHg-treated mice. Among chemokines with increased expression in the brain of MeHg-treated mice, CCL3 and CCL4 expression levels were not changed in other tissues (kidney, liver and spleen). Our results suggest the possibility that CCL3 and CCL4 may play a crucial role in MeHg-induced CNS disorders.

抄録全体を表示

The objective of this study was to develop PBPK modeling to assess exposure of cadmium to healthy Korean. Cadmium (Cd) is believed to induce kidney disease and nephrotoxicity becomes the major target concerned in public health to Cd. Cd absorption is relatively low (<10 %) after dietary exposure in humans with a very long biological half-life up to 30 years. The primary route of cadmium exposure is known to be through intake of food. The previous 8-compartment PBPK model (Nordberg-Kjellstrom, 1979) for Cd was modified according to parameters for Korean. PBPK modeling for Cd exposure was validated based on the biomonitoring data of urinary Cd excretion and Cd concentration in blood for Korean from Korea Food and Drug Administration. The PBPK model can be applicable to assess dietary Cd exposure to Korean.

抄録全体を表示

Gadolinium (Gd) is widely used for various industrial and medical purposes, particularly in magnetic resonance imaging (MRI) contrast agent. There has been a large volume of studies about nephrotoxicity and neurotoxicity of Gd, whereas the research on detailed mechanism of cytotoxicity in lymphocytes is insufficient yet. And it is important to investigate the effect of ELF-EMF on the cyto- and geno-toxicity of Gd, since patients are co-exposed to Gd and extremely low-frequency electromagnetic fields (ELF-EMF) generated by MRI scanner. The aims of this study is to investigate the cyto- and geno-toixcity of Gd and the possible enhancing effect of ELF-EMF on the toxicity of Gd in cultured human lymphocytes by performing micronuclei (MN) assay, trypan blue dye-exclusion assay, single cell gel electrophoresis assay and apoptosis analysis using flow cytometry. MN frequencies were 13.0, 14.0, 15.3 and 17.7 per 1000 cells in Gd treatment cells at concentrations of 0, 0.4, 0.8 and 1.2 mM. ELF-EMF exposure (0.8 mT) increased the frequencies of MN induced by Gd treatment alone; 13.3, 17.0, 20.3 and 24.0 per 1000 cells at each Gd concentrations. ELF-EMF exposure also increased cell death, olive tail moment (OTM) and apoptosis induced by Gd treatment alone. These results suggest that Gd induces DNA damage and apoptotic cell death in human lymphocytes and ELF-EMF can enhance the cyto- and geno-toxicity of Gd.

抄録全体を表示

Selenium (Se) is an essential trace element for animals and appears in various forms such as selenoproteins, selenoamino acids and low-molecular metabolites. Compared to the many studies of the physiological and toxicological effects of Se in mammals, avian Se metabolism is still an unexplored topic. Some birds are useful as poultry for human nutrition. Moreover, birds belong to higher trophic levels in the biosphere and thus, may play an important role in Se circulation in the ecosystem in the same way as mammals. In this study, we analyzed the distribution and metabolism of Se in an experimental bird, the Japanese quail, which was fed drinking water containing sodium selenite or selenomethionine (SeMet). The highest concentration of Se was detected in pancreas, followed by down feather, liver, and kidney. SeMet was more efficiently incorporated into quail than selenite. In the same manner as mammals, selenosugar and trimethylselenonium were the major metabolites in quail excreta. Three unknown Se metabolites were detected by HPLC-ICP-MS. Although part of the metabolic pathway of Se in Japanese quail fed selenite and SeMet was the same as that observed in mammals, the bird also showed certain avian-specific metabolic process for Se.

抄録全体を表示

The severe injury of the cerebrum cortex in the patients with Minamata disease is localized around deep sulci. An experimental study suggested that the neuronal damage induced by methylmercury is a secondary manifestation of edematous changes around deep sulci. The purpose of the present study was to investigate the molecular mechanism of cytotoxic edema from the viewpoint of the toxic effects of methylmercury on the polyol pathway in brain microvascular endothelial cells and pericytes. Human brain microvascular endothelial cells and pericytes were incubated with methylmercury and morphologically observed. The expression of aldose reductase (AR) and sorbitol dehydrogenase (SDH), the key enzymes of polyol pathway, were determined at mRNA and protein levels by real-time RT-PCR and western blot analysis, respectively. In endothelial cells, neither the shape of the cells nor the expression of AR and SDH changed after exposure to methylmercury. In contrast, in pericytes exposed to methylmercury, the cell morphology changed from spindle to cobblestone appearance. At this time, the expression of AR was elevated whereas that of SDH was reduced by methylmercury. The morphological change with a higher expression of AR was also observed in pericytes cultured under a high glucose condition; the change in morphology was suppressed by knockdown of AR. These results suggest that methylmercury cause a cytotoxic edematous change in pericytes by swelling due to elevation of the intercellular osmotic pressure and water content, which would be caused by excess accumulation of sorbitol via induction of AR expression and reduction of SDH expression.

抄録全体を表示

This study evaluated the feasibility of transgenic Arabidopsis engineered to express the bacterial heavy metal transporter MerC for the phytoremediation of mercury pollution. MerC, MerC-SYP121, or MerC-AtVAM3 proteins were found to be expressed in leaf segments of transgenic plants using an anti-MerC antibody immunostaining method. Transgenic Arabidopsis plants expressing merC, merC-SYP121, and merC-AtVAM3 were more resistant to mercury and accumulated significantly more of this metal compared with wild-type Arabidopsis. These results demonstrated that the expression of the bacterial heavy metal transporter MerC can promote the transport and accumulation of mercury in transgenic Arabidopsis, which represents a method of improving plants for the phytoremediation of mercury pollution.

抄録全体を表示

Diurnal susceptibility to medical drugs have often been demonstrated and known as chronotherapy. However, such observations especially about heavy metals are still scarce. We report the diurnal susceptibility to cadmium toxicity in mice. Male C57BL/6J mice kept in cages on 8:00-20:00 L/D cycle were administrated intraperitoneally (i.p.) with cadmium chloride at 6 different clock time (10:00, 14:00, 18:00, 22:00, 2:00 and 6:00 h), describing as zeitgeber time (ZT); ZT2, ZT6, ZT10, ZT14, ZT18 and ZT22. In case of dark period (ZT14, ZT18 and ZT22), administrations were performed under red light. Mortalities were monitored until 14 days after the injection. Surprisingly, all mice were dead at ZT2 injection; conversely, all mice were survived at ZT18 injection. The hepatotoxicity was estimated 24 hr after the Cd injection using the plasma GPT values as an index. The GPT value was markedly elevated at ZT6 injection while not any changed at ZT18 injection. In ZT6 and ZT18, there were no difference in hepatic Cd accumulation, metallothionein (MT) induction level, further, both basal hepatic MT level and GSH level. When hepatic GSH level was depressed by L-buthionine-[S,R]-sulfoximine, however, the difference in hepatotoxic susceptibility was clearly disappeared. We showed the clear diurnal variation of Cd-induced toxicity and one of the candidate factors for determination of this variation was GSH.

抄録全体を表示

Selenium has been shown to reduce the risk of several forms of human cancer. However, the mechanisms underlying its chemopreventive effects remain largely unresolved. In the present study, we found that methylseleninic acid (MSeA), a monomethylated selenium derivative, induced expression of NQO-1, an important cytoprotection enzyme against oxidative and electrophilic stresses. Nrf2 through interaction with antioxidant response element (ARE), induces expression of many antioxidant enzymes. MSeA treatment resulted in an increased nuclear translocation, subsequent ARE binding and transactivation of Nrf2. Silencing Nrf2 using dominant negative Nrf2 or Nrf2 siRNA markedly reduced the MSeA-induced NQO-1 expression. Moreover, MSeA treatment induced expression of NQO-1 to a much less extent in Nrf2^-/- MEF than in Nrf2^+/+ MEF. Furthermore, MSeA reduced the level of Keap1, a repressor of Nrf2, at a post-translational level. Ubiquitination of Keap1 was markedly increased in cells exposed to MSeA, which resulted in decreased steady-state levels of Keap1 in parallel with inhibition of keap1-dependent ubiquitination of Nrf2. In addition, Oral administration of MSeA (1 mg/kg) by gavage to mice also induced NQO-1 protein expression in the liver. Methylselenol generated from selenomethionine (SeMet) by methioninase (METase) activity induced expression of NQO-1. In contrast to the treatment with SeMet and METase, SeMet itself failed to induce NQO-1 expression, suggesting that methylselenol, not its metabolite, is directly responsible for NQO-1 induction. Taken together, these results suggest that MSeA, the immediate precursor of methylselenol, upregulates the expression of NQO-1 via the Keap1-Nrf2 signaling, preferentially by targeting cysteine thiol(s) present in Keap1.

抄録全体を表示

Background: Arsenic contamination in water, especially groundwater, has been recognized as a major environmental problem in Bangladesh. It acts as a carcinogen possibly due to the generation of reactive oxygen species (ROS). Gender disparity has been reported in many arsenic-exposure related diseases like cardiovascular diseases, hypertension, liver diseases, as well as skin lesions or cancers. However, no study had been reported about gender disparity in arsenic-induced oxidative stress in Bangladeshi population. Objective: The objective of the study was to examine the levels of urinary 8-OHdG and F2α –isoprostane, two established oxidative stress biomarkers, in relation to the urinary arsenic. Methods: Data were obtained from 219 healthy men (96) and women (126), aged 18-45 years, giving informed consent, from two communities of south-western part of Bangladesh in August 2009. Water of the tube wells that they were using and urine samples were collected and analyzed for total arsenic (by ICP-MS) and for oxidative stress (ELISA). Results: Urinary arsenic (u-As) showed positive association with urinary oxidative stress markers- 8-OHdG (r= 0.60; p<0.001) or 15-F2t-IsoP (r= 0.46; p<0.001). Arsenic-exposed women had significantly higher 15-F2t-IsoP (adjusted mean (95% CI) 6.1 (5.0, 7.2) vs 4.8 (3.9, 5.8) ng/mg Cr) and arsenic methylation capacity (%DMA) (p<0.05) than men. Gender difference in u-As and urinary 8-OHdG was not found in this study group. Conclusions: The results suggest that oxidative DNA damage and lipid peroxidation were associated with arsenic exposure, which were sex-dependent to some extent.

抄録全体を表示

Metallothionein (MT) is a small, cysteine-rich protein active in zinc homeostasis, cadmium detoxification, and protection against reactive oxygen species. Mouse MT-I gene transcription is regulated by metal response element-binding transcription factor-1 (MTF-1), which is recruited to the promoter by zinc. We examined alterations in the chromatin structure of the MT-I promoter associated with enhanced transcriptional activation and effect of the alteration on the MT-I gene transcriptional activation. MTF-1 proved essential for zinc-induced epigenetic changes in the MT-I promoter. Chromatin immunoprecipitation assays demonstrated that zinc treatment rapidly decreased total histone H3 but not histone H3.3. Micrococcal nuclease sensitivity of the MT-I promoter was increased by zinc. Thus, the chromatin structure in the promoter may be locally disrupted by zinc-induced nucleosome removal. Without MTF-1 these changes were not observed, and an MTF-1 deletion mutant recruited to the MT-I promoter by zinc that did not recruit the coactivator p300 or activate MT-I transcription did not affect histone H3 in the MT-I promoter in response to zinc. Interleukin-6, which induces MT-I transcription independently of MTF-1, did not reduce histone H3 levels in the promoter. Rapid disruption of nucleosome structure at the MT-I promoter is mediated by zinc-responsive recruitment of an active MTF-1-coactivator complex. Furthermore, when zinc was removed, the chromatin did not rapidly revert to its resting state. We also demonstrated that nucleosome-disrupted MT-I promoter showed enhanced transcriptional activation. The exclusion of the histone might act as a memory of the first zinc-induced MT-I transcription.

抄録全体を表示

[PURPOSE] The present study was designed to evaluate reproductive and neurobehavioural effects of brilliant blue FCF in mice throughout two generations.
[METHODS] Brilliant blue FCF was given to mice in the diet at levels of 0 (control), 0.08%, 0.24%, and 0.72% from 5 weeks of age of the F₀ generation to 11 weeks of age of the F₁ generation, and selected reproductive and neurobehavioural parameters were measured.
[RESULTS] In exploratory behaviour of adult females of the F₀ generation, movement time (s) showed a significant tendency to be increased and average time of rearing (s) showed a significant tendency to be decreased in the treatment groups. In the F₁ generation, surface righting at postnatal day (PND) 4 was affected significantly in male and female offspring in a dose-related manner. In exploratory behaviour of adult females of the F₁ generation, number of horizontal activities showed a significant tendency to be decreased in the treatment groups. After weaning, the body weight gain of females was significantly affected during 10−11 weeks of age in a dose-related manner.
[CONCLUSION] In the present study, brilliant blue FCF showed a few significant adverse effects on neurobehavioural parameters. The high dose level was based on the current ADI of brilliant blue FCF. The actual dietary intake of brilliant blue FCF in Japan is presumed to be much lower approximately 0.28−0.56 μg/kg/day. It would therefore appear that the levels of actual dietary intake of brilliant blue FCF are unlikely to produce any adverse effects in humans.

抄録全体を表示

Many studies have shown that lipoxygenase (LOX) can mediate the co-oxidation and activation of xenobiotics including carcinogens in vitro enzyme system. To the authors' knowledge, however, there is almost no evidence that the oxidation can be conducted in cells, tissues and mammals. It has also been unclear whether 4-aminobiphenyl (4-ABP) can be activated by LOX in living organisms. This study was carried out to provide the evidence that LOX is a metabolic pathway of activation of precarcinogen. 4-ABP reacted with soybean lipoxygenase (SLO) in vitro enzyme system and the products were tested by spectrophotometry. The livability of human bronchial epithelial (HBE) cells was tested by MTT assay, the protein expression of 5-lipoxygenase (5-LOX) by western blot and DNA damage by single cell gel electrophoresis. Results showed that SLO could oxidize 4-ABP in the presence of hydrogen peroxide. A LOX inhibitor nordihydroguaiaretic acid could suppress the oxidation. The expression of 5-LOX in HBE cells was strengthened by 4-ABP but not affected by AA-861. The DNA of HBE cells treated by 400μmol•L^-1 4-ABP was obviously damaged and 47.7% of the cells displayed a comet shape. AA-861 and naproxen inhibited the DNA damage and the maximum protection rates were 58.1% and 21.7% respectively. These results suggest that the expression of 5-LOX can be influenced by 4-ABP, and the co-oxidation of 4-ABP by 5-LOX in the cells may produce electrophilic free radicals which can combine with DNA, thus causing DNA damage. This may be one of the carcinogenic mechanism of 4-ABP.
KEY WORDS: 5-lipoxygenase; co-oxidation; 4-aminobiphenyl; protein expression; DNA damage

抄録全体を表示

Prioritized assessment chemicals will be evaluated for their risks, and then, they will be designated to the second specific chemicals in the amended Kashin-law from April 2011. In the genotoxic risk evaluation, in vivo test data will be needed if in vitro test showed positive. However, there is no consensus on what case or which test should be requested. Therefore, prioritization of request of in vivo micronucleus assay (MN) data was investigated in case of positive for in vitro chromosomal aberration test (CA). There are 66 chemicals which are positive for in vitro CA tests with Ames and MN data among 277 existing chemicals in Kashin-law database. Lowest effective concentration (LEC) in the CA data and results of Ames and MN test were compared. There were tendencies that Ames-positive chemicals show MN-positive regardless of LEC value and CA-positive chemicals with low LEC show MN-positive even if Ames-negative. Following criterion values are set for LEC: 1) ≤ 0.05 mg/mL, 2) 0.05-0.5 mg/mL, and 3) > 0.5 mg/mL. MN-positives (Ames-negatives, within) and Ames-positives are 8 (3), 6 among 19 chemicals in LEC 1), respectively, and are 1 (0), 4 among 21 chemicals in LEC 2), and 3 (1), 7 among 26 chemicals in LEC 3). These results indicate that priorities of request of in vivo MN data are high, middle, and low in LEC 1), 2) and 3), respectively. Ames-positives are also high priority. This approach will be effective in risk communication for the request of MN data under the Kashin-law.

抄録全体を表示

The cell models of SiO₂ activating macrophages were set up in order to explore the roles of Annexin A5 in the activation. SiO₂ produced an increase in the expression of Annexin A5, IL-1α and TGFβ1 in macrophages. When Annexin A5 knock-down by siRNA, the expression of IL-1α and TGFβ1 were decreased, and the secretive levels of TGFβ1 were also down-regulate. These foundings suggested that the expression of Annexin A5 activated macrophages and influenced the producing and relieving inflammatory cytokines and fibrotic factors. Recent years it was found that Fas-ligand cell signaling has a key role in the initial phase of lung fibrosis, so we studied on the protein expressions of Fas signal pathway. When SiO₂ activited the macrophages, the expressions of Fas/FasL signal pathway molecules including FasL, were increased. However, when FasL knock-down by siRNA, the expression of IL-1α and TGFβ1 were decreased and the protein expressions in down stream, Fas, Casp8, Casp3, decreased, too, and the secretion of TGFβ1 from macrophages was also down-regulate. At last we explored the interaction between Annexin A5 and FasL in SiO₂ activited macrophage models. It was found that Annexin A5 knock-down could decrease the expressiond of FasL, but Annexin A5 was up regulate when FasL knock-down. Those foundings suggested that Annexin A5 could effect on the activation of macrophage and has the interaction with Fas/FasL signal pathway.

抄録全体を表示

Azo dye precursors and some aromatic amine metabolites such as 3,3'-Dichlorobenzidine (DCB) produced through biotransformation of azo dye compounds have been shown to be carcinogenic, especially bladder cancer and lymphohematopoietic cancer. Epidemiological studies have suggested a relationship between workers long term exposed to DCB-based dyes in the dyeing industry and increased the risks of cancer development. In this study, the signaling pathways of DCB-induced cytotoxicity in HepG2 cells were investigated. We found that DCB could increase the expression level of γ-H2AX, a sensitive molecular marker of DNA double-strand breaks, in a dose-dependent manner and reach a maximum effect in cell treated with 10 μM of DCB. DCB at concentration ranges of 25-100 μM showed a 20~40% inhibition of HepG2 cell viability. Cell cycle analysis revealed G2/M phase arrest and accumulation of sub-G1 phase in HepG2 cells following 24 h exposure to DCB. We found Bcl-2/Bax ratio and mitochondrial membrane potential were decreased in cell treated with DCB. In addition, DCB also could enhance caspase-3 and caspase-9 activities in a dose-dependent manner. These results indicated that DCB could induce HepG2 cell apoptosis through a mitochondria/caspases pathway. To determine the signaling pathways of DCB-induced cytotoxicity in HepG2 cells, we found that DCB significantly increased the phosphorylation levels of JNK and ERK but not p38 in a time- and dose-dependent manner. To further determine whether ERK and JNK activation were required for DCB-induced apoptosis, the pharmacological inhibitors were used. We found JNK inhibitor, SP600125, could attenuate DCB-induced caspase-3 activity. Taken together, DCB could induce cell apoptosis in HepG2 cells via MAPK (JNK) and mitochondria/caspases pathways.

抄録全体を表示

1-Bromopropane (1-BP) was introduced as one of new substitutes for ozone depleting compounds. 1-BP is used in multiple manufacturing processes, including vapor and immersion degreasing operations to clean metal, precision instruments, electronics, optical instruments and ceramics and used as a solvent vehicles for aerosol applied adhesives in cushion manufacture industries. Several human case studies have reported polyneuropathy and neurotoxicity in 1-BP-exposed workers. Animal studies have shown reproductive/developmental toxicity as well as central neurotoxicity. However, developmental neurotoxicity remains unclear. Here we report that 1-BP affects hippocampal excitability and sensitivity to excitotoxic compound pentylenetetrazole (PTZ) during developmental period. Pregnant Wistar rats were inhalationally exposed to 1-BP vapor from day 1 to 20 (6 h/day) with the concentrations of 0, 400 and 700 ppm. On the days of PND 13, 14 and 15, hippocampal slices were prepared from the offspring. At PND 14, stimulation/response curves of the field EPSP and the population spike (PS) were enhanced and paired-pulse ratios of PS were decreased in the hippocampal CA1 region in the 1-BP exposed groups. Systemic injection of PTZ to the PND14 offspring revealed a significant decrease the occurrence of wild running episode, tonic convulsion and status epilepticus in the 1-BP exposed groups compared to the control group. Our results suggest that prenatal exposure to 1-BP modifies the development of neuronal networks and changes excitability of the brain during the lactation period.

抄録全体を表示

2,5-hexanedione is the metabolic end product of n-hexane, which is an organic solvent that has been known to alter different reproductive functions in mammals. The aim of this study was to determine the effects of 2,5-hexanedione on reproductive functions in mouse, especially the maturation of mouse oocytes. We observed that extrusion of the first polar body in mouse oocytes were significantly inhibited by 2,5-hexanedione. Furthermore, mitochondrial membrane potentials (ΔΨm) were also altered in mouse oocytes, leading to apoptosis of the oocytes. Our results suggest that the 2,5-hexanedione might have affected the maturation of oocytes causing the ΔΨm to alter leading to apoptosis which maybe one of the most important mechanisms.

抄録全体を表示

This study aimed to evaluate the expression of the complex APC, GSK-3β, Axine and β-Catenin, which are components from canonical Wnt signaling pathway during chemically induced oral carcinogenesis by 4-nitroquinoline 1-oxide in rats. Male Wistar rats were distributed into three groups of 10 animals each and treated with 50 ppm 4NQO via drinking water, during 4, 12 and 20 weeks. A total of 10 animals were used as controls. No histopathological changes were noticed to control group and that exposed to 4NQO during 4 weeks. The animals exposed to 4NQO for 12 and 20 weeks pointed out leukoplakia, exophytic and ulcerated lesions, as well as hyperplasia and hyperkeratosis with epithelial dysplasia and squamous cell carcinoma of well differentiated type, respectively. A high immunopositivity for APC and axine was noticed to experimental groups exposed to 4NQO during 12 and 20 weeks, when compared to negative control. Nevertheless, β-catenin pointed out an up-regulation in pre-neoplastic lesions and oral squamous cell carcinomas. GSK-3β did not show remarkable changes along a time course experiment. Taken together, a downregulation of APC and Axine as well as an up-regulation of β-catenin play a crucial role during oral cancer progression in rats.

抄録全体を表示

Deoxycorticosterone acetate salt hypertension (DSH) model is commonly used for investigating hypertension-associated renal fibrosis. Sesame oil is a natural product with potent anti-fibrosis property. We investigate the protective effect of sesame oil on deoxycorticosterone acetate salt-induced kidney damage in rats. Chronic renal injury in uninephrectomized rats was induced by deoxycorticosterone acetate and salty drinking water. After 4 weeks, rats were ingested with sesame oil (0.5 or 1 ml/kg) for 7 days. Renal injury, histopathological examination, collagen levels and osteopontin expression were assessed 24 h after the last dose of sesame oil. Serum blood urea nitrogen, creatinine levels as well as urine volume were significantly increased in DSH rats. Sesame oil significantly decreased all these parameters compared with them in DSH rats. Sesame oil significantly increased creatinine clearance rate in DSH rats. Further, sesame oil significantly decreased DSH-induced renal collagen and osteopontin over-expressions. Therefore, we summarize that sesame oil ameliorates chronic renal fibrosis through inhibiting renal osteopontin expression in DSH rats.

Keywords: deoxycorticosterone acetate salt hypertension (DHS), mineralocorticoid, chronic renal fibrosis, sesame oil, osteopontin, rat

抄録全体を表示

Objectives: The aim of this study is to evaluate the anti-androgenic effects of Diethyl-O-phthalate in the reproductive development of immature male rats in vivo. Methods: Immature Sprague-Dawley rats were dosed daily with corn oil or Diethyl-O-phthalate ( 100, 200 or 500 mg/kg ) by gavage from postnatal day(PND) 12 and through PND 35 respectively, Flutamide and Testosterone propionate were used as control. Sex accessory tissues were weighed and anogenital distances (AGD) were measured. Plasma levels of Testosterone(T) and luteinizing hormone(LH) were also evaluated with ELISA kits. Results: The weights of seminal vesicle, ventral prostate, drosolateral prostate, lecator ani/bulbocavernosus (LABC) muscle and AGD distances decreased significantly in response to Diethyl-O-phthalate in a dose-dependent manner, but no significant in the other accessory sex tissues. Testosterone levels were significantly decreased in Diethyl-O-phthalate treated groups, whereas luteinizing hormone levels were not altered. Conclusion: our results demonstrated that Diethyl-O-phthalate shows toxicological effects on the male reproductive system in immature male Sprague-Dawley rats resulted from its anti-androgenic effects.

抄録全体を表示

In India, cancer of the breast is the most common cancer among women in many regions and has overtaken cervix cancer. Organochlorine pesticides are one of the risk factors of this disease because of their potential estrogenic activity, immunosuppressive and tumor promoting properties. Organochlorine pesticides including, Dichlorodiphenyltrichloroethane (DDT) and Hexachlorocyclohexane (HCH) have been shown to enhance oxidative stress and lipid peroxidation in various tissues. Redox status of breast tumors and surrounding tumor free breast tissues in association with organochlorine pesticides levels was investigated to find any possible association between them. Ten newly diagnosed breast cancer patients, mean age of (50.75±8.16) years from King George Medical University, Lucknow, were chosen for the study to determine antioxidant enzyme activities namely superoxide dismutase (SOD), catalase (CAT), glutathione (GSH) and lipid peroxidation (MDA) was assayed. In case of blood MDA and SOD activity were found significantly higher in malignant group however, GSH level was higher in control group (p<0.05 each). In case of tissue, tumor levels of MDA, CAT and SOD activity were detected higher in comparison to adjacent disease free tissue. On the other hand, GSH level was significantly higher found in disease free tissue than those of tumor (p<0.05 each). In our opinion, enhanced antioxidant capacity in form of enzyme activity is an adaptive measure to counteract organochlorine pesticides mediated free radical generation, as evident by higher MDA level in tumor as well as in blood. It shows that organochlorine pesticides mediated oxidative stress is, at least partly, associated with breast cancer.

抄録全体を表示

Microsomal prostaglandin E synthase-1 (mPGES-1) and prostacyclin synthase (PGIS) are prostaglandin (PG) terminal synthases that function downstream of inducible cyclooxygenase (COX-2) in the PGE₂ and PGI₂ biosynthetic pathway, respectively. Accumulating evidence indicates that COX-2-derived PGE₂ participates in the development of various tumors, but the precise contribution of mPGES-1 and PGIS to multiple process of chemical carcinogenesis is not yet fully understood. Here, we examined the role of mPGES-1 and PGIS in chemical carcinogen-induced colon carcinogenesis using a mouse model. mPGES-1 knockout (KO), PGIS KO, mPGES-1/PGIS double KO and mPGES-1 overexpressing mice and control mice were intraperitoneally injected with colonotropic carcinogen, azoxymethane (AOM), once weekly for 6 weeks. As the results, we found that at 6 weeks after the last injection, mPGES-1 deficiency significantly reduced the number of aberrant crypt foci (ACF) compared to control mice, while mPGES-1 transgenic overexpression and PGIS deletion increased the number. Furthermore, genetic deletion of mPGES-1 significantly reduced both the total number and size of colorectal polyps at 18 weeks after AOM administration with reduced nuclear translocation of β-catenin. On the other hand, the number of large polyps was increased in PGIS KO mice. In mPGES-1/PGIS DKO mice, both ACF and polyp formation tended to be reduced but the reducing effect was smaller than in mPGES-1 KO mice. These results indicate that mPGES-1-derived PGE₂ and PGIS-derived PGI₂ have opposing effects on chemical-induced colon carcinogenesis. Namely, the former PGE₂ promotes and exacerbates chemical carcinogenesis, while the latter PGI₂ suppresses it.

抄録全体を表示

It has been reported that occupational exposure to 1-bromopropane (1BP), a volatile organic solvent used for degreasing agents and spray adhesives as a substitute for chlorofluorocarbons, could cause central and peripheral neurotoxicity. Here, we investigated direct effects of 1BP on neurotransmitter receptors, and behavioral phenotypes of rat subchronic and prenatal exposure models of 1BP. In vitro experiments using Xenopus oocyte expression system showed that 1BP directly affected the function of neurotransmitter-gated ion channels, especially GABA-A and neuronal nicotinic acetylcholine receptors, which is likely to be responsible for an anesthetic-like effect. On the other hand, adult male rats exposed to 1BP (700 ppm) for six weeks (from 8 to 13 weeks of age) showed a significant increase in distance moved and duration of time on the following day after the exposure period in open-field test. Interestingly, rat male offspring prenatally exposed to 1BP (700 ppm, from gestational day 1 to 20) showed a tendency toward hyperlocomotion at young period (4 weeks of age), but hypolocomotion at adult period (12 weeks of age). Prenatally-exposed offspring also exhibited memory impairment in novel objective recognition and passive avoidance tasks at 6 weeks of age. Our results provide the evidence that neurobehavioral effects of exposure to 1BP could appear not only as a subchronic effect in adult, but also as a next-generation effect induced by prenatal exposure.

抄録全体を表示

Renal papillary necrosis (RPN) is a type of kidney injury induced by diabetes or nonsteroidal anti-inflammatory drugs or anticancer drugs. However, no biomarkers (BMs) capable of detecting prognostic or early stages of RPN in humans have been identified at present. We therefore explored novel BMs suitable for the early detection of RPN using toxicoproteomic (TPx) and toxicogenomic (TGx) techniques. Urine and kidney papilla from rat models of RPN induced by 2-bromoethylamine hydrobromide were used for TPx and TGx, respectively, and we also conducted histopathological examinations for comparisons with BM candidates. In addition, our present results were compared with those obtained using model rats with glomerular or proximal tubular injury induced by puromycin, cisplatin, or gentamicin. As a result of the TPx analysis of urine from RPN model rats, 94 proteins were identified as BM candidates, which included a number of acute phase proteins. From the TGx analysis, changes in the expression of mRNAs involved in the enhancement of IL-1 signaling, apoptosis, and oxidative stress were observed. In particular, elevation of fibrinogen and C3 were detected by both TPx and TGx analyses and were considered to be BMs associated with RPN. However, the levels of fibrinogen and C3 were also increased in urine by proximal tubular injury. Thus, fibrinogen and C3 do not only detect RPN, but also serve as BMs of proximal tubular injury. The increase in acute phase proteins is thought to be due to the enhancement of inflammation-related signaling. We are presently screening for other RPN-specific BMs among the remaining 92 BM candidates.

抄録全体を表示

Benzophenones are organic chemicals widly used as stabilizers to UV light irradiation in plastic surface coatings and food packaging. They have been known that thier derivatives appear to have cytotoxic effects. 2-Hydroxy-4-methoxybenzophenone (Benzophenone-3, BP-3) and its metabolite, 2,4-dihydroxybenzophenone (Benzophenone-1, BP-1), are used mostly in the formulation of nail polishes, enamels, bath products, sunscreens and skin care products. In this study, the estrogenic effects of BP-1 were examined in an ovarian cancer BG-1 cells expressing high levels of estrogen receptors (ERs) by an cell viability assay, semi-quantitative reverse-transcription PCR (semi-quantitative RT-PCR) and Western blot analysis. The treatment of BG-1 cells with BP-1 (10^-8~10^-5 M) resulted in an increase in their cell proliferation as 17-beta estradiol (E2) did. But, the cell growth stimulation by BP-1 was reversed by cotreatmwnt with ICI 182,780, an ERs antagonist, suggesting that the proliferation of BG-1 cells is mediated by an ER-dependent pathway. In addition, BP-1 upregulated the expression levels of cell-cycle regulating genes, i.e., cyclin D1, which is a downstream target of ER, at 6 h after treatment. But, the expression of p21 gene was not altered by BP-1 at any time points. In translational levels, BP-1 also upregulated the expression level of cyclin D1 at 24 h after its treatment. But, upregulated expression of cyclin D1 by BP-1 was reversed by cotreatmwnt with ICI 182,780, at both mRNA and protein levels. Taken together, these results suggest that BP-1 is one of EDCs with have apparent estrogenic activities by stimulating cell proliferation of BG-1 cells and by inducing the expression of cyclin D1. Our results can support that BP-1 may have a potency to disrupt endocrine system and to stimulate cell growth in ER-positive cancer cells. [This work was supported by a National Research Foundation of Korea (NRF) grant funded by the Ministry of Education, Science and Technology (MEST) of Korea government (no. 2011-0015385).]

抄録全体を表示

Iodotyrosine deiodinase (IYD) has an ability to salvage iodide from iodotyrosine that are generated as byproducts of thyroid hormone biosynthesis. This enzyme has also been discovered to act as a dehalogenase of bromo- and chlorotyrosine. Halogenated phenolic chemicals may disrupt thyroid hormone metabolism through the inhibition of IYD activity. The objective of this study was to clarify the effect of halogenated environmental chemicals on thyroid hormone metabolism, we investigated the inhibitory effects of these chemicals on IYD activity using HEK293T cells recombinant human IYD. Inhibition of IYD activity was detected by two flame retardants, 3,3’,5,5’-tetrachlorobisphenol A and 3,3’,5,5’-tetrabromobisphenol A, by the antimicrobial agent, triclosan, by antihyperuricemic drug, benzbromarone, by two antiparasitic drugs, bithionol and oxyclozanide, and by three food colorants, phloxine B, erythrosine B and rose bengal. These results suggest that the halogen substitution adjacent to hydroxyl group is structural requirement for the inhibition of IYD activity.

抄録全体を表示

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) is known to cause Parkinson-like symptoms, and neuronal nitric oxide synthase (nNOS) exacerbates MPTP-induced neurotoxicity. However, the detailed mechanisms underlying this neurotoxicity have not ever been clarified. Recently, 8-nitroguanosine 3',5'-cyclic monophosphate (8-nitro-cGMP) was identified as a novel second messenger that is endogenously formed on excess production of NO coupled with reactive oxygen species (ROS). NO/ROS signaling mechanisms are currently under study as a novel signaling pathway. In this study, to clarify the mechanisms underlying neurotoxicity in Parkinson's disease, we investigated the relationship between NO/ROS signaling mechanisms and neurotoxicity induced by 1-methyl-4-phenylpyridinium ion (MPP⁺), an active metabolite of MPTP.
We prepared transfected PC12 cells stably expressing nNOS and examined cytotoxicity and the production of ROS and 8-nitro-cGMP in nNOS-expressing cells treated with MPP⁺. To reveal the molecular mechanisms underlying MPP⁺-induced cytotoxicity, we analyzed the involvement of active the Ras/Erk signaling pathway by Western blotting.
MPP⁺-induced cytotoxicity was more severe in nNOS-expressing cells than in control cells, and the production of ROS and 8-nitro-cGMP was higher in nNOS-expressing cells treated with MPP⁺ than in control cells. We also found that ROS scavenger significantly ameliorated MPP⁺-induced cytotoxicity in nNOS-expressing cells. These results indicate that ROS produced by nNOS are involved in MPP⁺-induced cytotoxicity. Additionally, nNOS-expressing cells treated with MPP⁺ showed activation of Ras and Erk. Consequently, these findings show that ROS produced by nNOS are responsible for MPP⁺-induced neurotoxicity and suggest that 8-nitro-cGMP, a downstream molecule in NO/ROS signaling, induces neuronal cell death via an active Ras/Erk signaling pathway.

抄録全体を表示

Toluene inhalation is a common form of substance abuse. Chronic toluene use can cause persistent psychiatric symptoms even after cessation of toluene use. Our previous studies have shown that adolescent toluene exposure causes cognitive impairment and social dysfunction in mice, which can be reversed by atypical antipsychotics, such as clozapine and aripiprazole. This investigation was designed to extend the previous findings and identify the role of 5-HT_1A receptors in the beneficial effects of atypical antipsychotics. Male NMRI mice received injection per day of either toluene (750 mg/kg) or oil at postnatal day P35-P39, and P42-P46. Behavioral tests were conducted after 7-day washout period to confirm the toluene-treated animals with long-lasting behavioral impairment. Thereafter, toluene-treated mice were administered with the antipsychotics clozapine (20 mg/kg), aripiprazole ( 0.1 and 0.3 mg/kg) or buspirone (1 and 5 mg/kg), a 5-HT_1A receptor partial agonist, alone or co-administered with WAY100635 (0.5 mg/kg), a serotonin 5-HT_1A receptor antagonist, 1 hr prior to behavioral tests. The results showed that acute treatment of clozapine, aripiprazole and buspirone could effectively ameliorate the toluene-induced social and cognitive impairment and these effects were blocked by co-treatment with WAY100635. Our findings suggest that an interaction with 5-HT_1A receptors might be at least in part, responsible for the beneficial effects of clozapine and aripiprazole on toluene-induced social and cognitive deficits and 5-HT_1A receptor agonists might be novel therapeutic agents for treatment of behavioral disorders related to toluene abuse.

抄録全体を表示

Metabonomics, the global analysis of metabolites, have been widely used to identify endogenous metabolites that correlate with a specific physiological state of an organism. Therefore metabonomic profiling is thought to be a powerful tool for following systematic changes associated with metabolic reaction to xenobiotics. In this study, we observed the effects of typical CYP inducers on urinary metabonomic profiles to clarify the effects of CYP induction using mice. And we used two different diets, one was a regular diet for rats and mice and the other was the purified diet, for comparing the effects of chemicals. These diets were fed to C57BL/6JJcl mice before mating. Their offspring were kept on each diet. Dexamethasone (DEX) or 1,4-Bis[2-(3,5-dichloropyridyloxy)]benzene (TCPOBOP) was dosed once daily for 3 days to mice at dosage of 100 or 0.3 mg/kg, respectively. Urine of each 8-week-old mouse was collected over a 24-h period. Phosphate buffer solution (pH 7.2) was added to the urine, and the ¹H-NMR spectrum was acquired and submitted to Principal Component Analysis (PCA). The results of urinary metabolic profiling showed different classification of treatment in PCA scores plots indicating that there are metabolites variable between control and chemicals treated. PCA loading plots suggested that the level of urinary metabolites including taurine, glucose and trimethylamine were changed. In addition, difference of metabolic profile was observed between control and treated group. Variable levels of these endogenous biomolecules in urine might provide an indirect indicator of hepatic CYP induction in mice.

抄録全体を表示

This study was designed to investigate the effect of boiled Moringa oleifera LAM pod (bMO) on azoxymethane (AOM)-initiated and dextran sodium sulfate (DSS)-promoted two stage mouse colon carcinogenesis in the promotion stage. Male ICR mice were divided into 8 Groups. Group 1 was served as a negative control group. Group 2 was received AOM (10 mg/kg BW, i.p.), followed by DSS (2% in drinking water for one week), and served as a positive control group. Groups 3-5 were fed with 1.5%, 3.0%, and 6.0% bMO, respectively supplemented in a basal diet after AOM/DSS administration, for 15 weeks. Groups 6-8 were fed with only 1.5%, 3.0%, 6.0% bMO, respectively in basal diets for 15 weeks. At the end of the study we found that the incidences and multiplicities of tumor in Groups 3-5 were decreased from positive control but significantly only in Group 3. PCNA index in Groups 3 and 4 was also significantly decreased, compared with those of the positive control group, while iNOS expression was significantly decreased in all doses of bMO. However, there was loss of COX-2 expression in all doses of bMO. The results suggest that M. oleifera LAM pod has potentially suppressive effect against colitis-related colon carcinogenesis induced by AOM/DSS. The lower dose is more effective at the promotion stage than at higher doses.

抄録全体を表示

Regular exercise protects against harmful consequences of reactive oxygen species (ROS) and represents an effective and cost-efficient strategy for the prevention of a wide variety of diseases, which are often caused by oxidative stress. ROS generated during inflammation can lead to an increase in cell proliferation, survival, and inhibition of pro-apoptotic pathway, ultimately resulting in carcinogenesis. Exercise has been considered to reduce colon carcinogenesis by potentiating cellular defense mechanisms that protect against detrimental effects mediated by ROS. The present study was aimed to examine a possible protective effect of regular exercise against oxidative stress and inflammation induced by dextran sulfate sodium (DSS) in mouse colon. Administration of 3% DSS in drinking water to male ICR mice for 7 days resulted in a decreased body weight, bloody stool, diarrhea and a shortened colon length. The expression of COX-2 and iNOS, two representative pro-inflammatory enzymes, was elevated. The expression of COX-2 and iNOS is regulated by NF-κB. Short-term treadmill running exercise prior to administration of 3% DSS attenuated colitis and reduced the expression of COX-2 and iNOS by blocking NF-κB signaling, while the levels of glutathione, the principal cellular non-enzymatic antioxidant, were increased. Long-term exercise further decreased the expression of inflammatory factors and elevated GSH levels. In conclusion, regular exercise protected against DSS-induced inflammation and ROS-mediated oxidative stress in mouse colon by targeting NF-κB pathways, suggesting potential for prevention of inflammation-associated colon carcinogenesis.

抄録全体を表示

1-Bromopropane (1-BP) is neurotoxic in both experimental animals and humans. Previous proteomic analysis of rat hippocampus implicated alteration of protein expression in oxidative stress, suggesting that oxidative stress plays a role in 1-BP-induced neurotoxicity. To understand this role at the protein level, we exposed male F344 rats to 1-BP at 0, 400, or 1000 ppm for 8 h/day for 1 week or 4 weeks by inhalation and quantitated changes in hippocampal protein carbonyl using a protein carbonyl assay, two-dimensional gel electrophoresis (2-DE), immunoblotting, and matrix-assisted laser-desorption ionization time-of-flight mass spectrometry (MALDI-TOF-TOF/MS). Hippocampal reactive oxygen species and protein carbonyl were significantly increased, demonstrating 1-BP-associated induction of oxidative stress. MALDI-TOF-TOF/MS identified 10 individual proteins with increased carbonyl modification (p<0.05; fold-change≥1.5). The identified proteins were involved in diverse biological processes including glycolysis, ATP production, tyrosine catabolism, GTP binding, guanine degradation, and ethanol metabolism. Hippocampal triosephosphate isomerase (TPI) activity was significantly reduced and negatively correlated with TPI carbonylation (p<0.001; r=0.83). Advanced glycation end-product (AGE) levels were significantly elevated both in the hippocampus and plasma, and hippocampal AGEs correlated negatively with TPI activity (p<0.001; r=0.71). 1-BP-induced neurotoxicity in the rat hippocampus seems to involve oxidative damage of cellular proteins, decreased TPI activity, and elevated AGEs. High plasma levels of protein carbonyl and AGEs are potential biomarkers of 1-BP-related neurotoxicity.

抄録全体を表示

1-Bromopropane (1-BP) has been used in the workplace as an alternative to ozone-depleting solvents. However, the effect of 1-BP on inflammation is unclear. In the present study, we examined the effect of 1-BP on cyclooxygenase-2 (COX-2) expression and analyzed the molecular mechanism of its activity in murine RAW 264.7 macrophages. 1-BP significantly induced COX-2 protein and mRNA levels, as well as COX-2 promoter-driven luciferase activity and the production of prostaglandin E₂ (PGE₂), a major COX-2 metabolite, in macrophages. Transfection experiments with several human COX-2 promoter constructs revealed that 1-BP activated the transcription factors nuclear factor-κB (NF-κB) and CCAAT/enhancer-binding protein (C/EBP), but not AP-1 or the cyclic AMP response element binding protein. Phosphorylation level of PI3K/Akt, extracellular signal-regulated kinase (ERK) and p38 MAPK phosphorylation were also significantly activated by 1-BP. Taken together, 1-BP has the potential to provide a possibility of inflammation-associated disorders and these findings provide further insight into the signal transduction pathways involved in the inflammatory effects of 1-BP.

抄録全体を表示

Purpose: 1-Bromoprane (1-BP) is used in workplaces as a cleaning and adhesive solvent. Patients intoxicated with 1-BP showed depression, unstable mood and impair of cognitive function. On the other hand, recent studies suggest suppression of neurogenesis (neuron proliferation) in adult hippocampus is involved in depression. Based on the above backgrounds, the present study was designed to investigate effect of exposure to 1-BP on neuron proliferation.
Method: Forty eight male Wistar rats were divided into four groups of twelve and were exposed to 1-BP at 0, 400, 800 and 1000 ppm in inhalation chambers 8 h/day for 7 days. Six rats of each group were perfused for confirming cell proliferation. Cell proliferation was detected with 5-bromo-2'-deoxy-uridine (BrdU) immunostaining. Another six rats of each group were decapitated for measurement of mRNA of brain-derived neurotrophic factor (BDNF) with quantitative real time-polymerase chain reaction.
Result: Cell proliferation didn’t show any change after one-week exposure but decreased at 800ppm after four-week exposure. BDNF mRNA decreased dose-dependently after four-week exposure.
Conclusion: Exposure to 1-BP decreases neuron proliferation. Down-regulation of BDNF might partially contribute to the decrease in neuron proliferation.

抄録全体を表示

Naphtofuran compounds have been known to have cytotoxic activity and inhibited Hepatocyte nuclear transcription factors 4 alpha (HNF4alpha) which is associated human hepatocellular carcinoma (HCC) progression and induces the increase of proliferation rate, dedifferentiation and metastasis. In this study, we investigated whether a N-(3,5-bis(trifluoromethyl)phenyl)-5-hydroxy-1,2-dihydronaphtho[2,1-b]furan-2-carboxamide compound, naphtofuran compound, could modulate HNF4alpha activity and induced apoptotic cell death as well as expression of the apoptosis regulating genes of hepatocellular carcinoma cell. Treatment with different concentrations(1-5μg/ml) of N-(3,5-bis(trifluoromethyl)phenyl)-5-hydroxy-1,2-dihydronaphtho[2,1-b]furan-2-carboxamide compound for various periods (0-72h) inhibited liver cancer cell (HepG2, Hep3B) growth followed by induction of apoptosis in a concentration dependent manner. We also found that this compound induced apoptotic cell death as well as expression of the apoptosis regulating genes. And Pull down assay proved that this compound directly binds to HNF4alpha and inhibit its activity. These results suggest that a novel naphtofuran compound inhibited liver cancer cell growth through induction of apoptotic cell death by modulating of HNF4alpha.

抄録全体を表示

DMAC are widely used industrially as a solvent. DMAC is taken into the body via the dermal and respiratory tract to cause dermatitis and liver dysfunction. However, the mechanism for the onset of liver failure has been reported is still unknown. In this study, we confirmed the occurrence of liver failure due to exposure, and thereby to elucidate the mechanism for its onset. ICR male mice were divided into four groups of six each and exposed to DMAC at 0, 10, 50, 250ppm for 6 h/day for 14 days by inhalation. At the end of exposure period, histopathological changes were investigation by H&E staining in livers. The histopathological change was evaluated scoring of liver cell necrosis, the degree of inflammation. The plasma and part of the liver were used for biochemical analysis. Exposure by inhalation DMAC, in the exposed group compared with the control group, increased plasma ALT, AST. In the 250ppm exposure group were observed significant large area of liver necrosis, and also, the exposed groups of 10ppm and 50ppm were observed liver necrosis. In addition, the inflammatory cells were observed by DMAC inhalation. In the 50 ppm and 250 ppm exposed groups showed higher 8-OHdG level, and also, HO-1- and NQO1-mRNA were increased by DMAC inhalation. The results showed that mRNA level and protein level of CYP2E1 were increased in dose-dependent manner in livers of mice after DMAC inhalation. These results suggest that increase the oxidative stress in the process of oxidative metabolism of DMAC via CYP2E1.

抄録全体を表示

In Japan, health risk assessment of chemicals in ambient air should be based on information obtained from epidemiological studies for setting the Guideline Values and Environmental Quality Standards. However, toxicity data derived from epidemiological studies often are limited to the cases of occupational exposure.
Because protective measures against excess exposure have improved during recent years, the cases of overt toxicity due to high doses are expected to be quite rare. Consequently, future health risk assessment will rely on data of animal studies rather than those of epidemiological studies in humans.
To this end, we need to confirm the appropriateness of extrapolating to humans from animals by using the benchmark dose approach and uncertainty factors.
First, we assessed the cancer risk of vinyl chloride monomer and 1,3-butadiene on the basis of inhalation studies in animals, and compared these assessment values estimated from animal studies with the Guideline Values for these substances in ambient air in Japan, which previously were established on the basis of epidemiological studies in humans. The Guideline Values of these substances were within the ranges of the risk assessment values caculated from animal studies.
However, in the case of assessment of non-carcinogenic endpoint (e.g. adverse effects of trichloroethylene and tetrachloroethylene) our analysis showed that the assessment values based on animal studies might differ from those based on epidemiological studies, if large uncertainty factors are adopted.

抄録全体を表示

Gibberellic acid (GA₃), a plant growth regulator, has registered in Taiwan for shoot growth, floral development and fruit set in celery, spinach, grapes, wax apples and pears. It was determined to be safe as its low acute toxicity and specified usage. Several literatures, however, raised its concern about potential endocrine disrupting effects of GA₃ on estrogenic response in animal tests. Therefore, the aim of this study is to explore possible mechanism of GA₃ regarding endocrine disruption with two kinds of well-known in vitro estrogen activity-detecting systems, i.e. estrogen receptor (ER)-mediated transcriptional activation assay and MCF-7 cell proliferation assay (E-screen). The results indicated that significant transactive and cell proliferative responses were observed at 0.1 to 1000 uM of GA₃ in both assays. In the meantime, those responses were demonstrated that it can be inhibited with an ER antagonist, 4-hydroxytamoxifen. In conclusion, GA₃ can elicit estrogenic activity via receptor-mediated pathway in human cells.

抄録全体を表示

Oxidative injury can occur in the lung through the generation of reactive oxygen species (ROS) via redox cycling due to intentional or accidental ingestion of paraquat (PQ), a common herbicide. A wide array of phytochemicals has been shown to reduce cellular oxidative damage by modulating cytoprotective genes. Quercetin, a well-known flavonoid, has been reported to display cytoprotective effects by up-regulating certain cytoprotective genes. In this context, we investigated the effect of quercetin on PQ-induced cellular cytotoxicity in alveolar A549 cells, modulation of antioxidant genes, activation of transcription factor-Nrf2 and HO-1 (a gene target of Nrf-2). After 24 h of treatment, quercetin reduced PQ-induced cytotoxicity in A549 cells that was evaluated by both MTT and LDH assays. Modulation of antioxidant genes was compared when cells were treated with PQ, quercetin and both paraquat and quercetin by qRT-PCR. Activation of transcription factor-Nrf2 and induction of its target gene, HO-1 was demonstrated by Western blot analysis. A remarkable reduction in ROS as well as increases in cellular GSH occurred when PQ-exposed cells were treated with quercetin. Our findings suggest that quercetin may be used to mitigate or minimize oxidative stress via reducing the generation of reactive oxygen species but need further evaluation.

抄録全体を表示

Introduction-
The use of pesticide in agricultural fitioneld is most hazardous operation as these HEALTH RISK OF PESTICIDE APPLICATORS OF SENCHOWA AREA OF NOWGONG DISTRICT, ASSAM, INDIA workers. The formulator, manufactures and applicators are occupationally exposed to pesticides. They constitute high risk population group. Applicator are of special significance since they handle more than one pesticide for long time and exposed for entire life span . So a study was undertaken to assess the health risk of pesticide applicators exposed to combination of organ chlorine and organophophorus pesticides in Senchowa area of Nowgong district Assam India.
Material and Methods-
In the Nowgong district some places are selected randomly from Senchowa area such as Bebajia,kumargong, Alanisatra Gahi, KhutiKotia. The pesticide used widely Melathion(50%), BHC(10%dust), Dimethiate(90%EC,5%), Carboryl(20%EC,2-5%dust). There was no definite work schedule for the workers. We recorded 400 exposed workers. They Sprayed for 2-4 hours a day in a particular season. So, the exposure were recorded in terms of week. All the workers were male and with in age group of 28.38 ±5.46 and exposed for a period of 10-20 hours and above. A group of 108 subjects comparable to socio-economic status, but not occupationally exposed to pesticides were studied as control. No protective devices were used by the applicators. Of course, normal hygienic measure such as washing of hands and faces before and after meal are being practiced. All the exposed and control subjects were normally healthy with no clinical illness. (As recorded from personal interview)
Results and discussions-
Exposed applicators were divided in to three groups –mild exposure period up to one month, moderately exposed for exposure period with one month to one year and individuals. The highly effected age group was 25-30 years. From the clinical examination the type of acute toxic symptoms recorded were neurological including fatigue(29% RR=1.51),muscle weakness(22%,RR=1.170), dry/sore throat(29%RR=1.974), stomach pain(5%,RR=.743), skin redness(23%,RR.=1.17), weakness and psychological symptoms along with gastrointestinal problems were observed However, symptoms were disappearing when they stop using pesticide. A significant depression in Ache were observed in the applicators as compared to control. It was observed that a subjects with only psychological symptoms had less depressions of whole blood Che activity, then the subjects with the subjects with gastrointestinal and cardio respiratory symptoms.
Conclusion-
The result of overall observation showed that toxic symptoms in exposed individual were due to the effect of O C and O. P pesticide, neurotoxic, psychological along with gastrointestinal cardio respiratory symptoms were mainly due to Organophosphate pesticides The present Investigation revealed that the health risk of pesticide applicators

抄録全体を表示

The present study aimed to determine the role of paraquat (PQ) in the NF-E2-related factor-2 (Nrf2) / heme oxygenase 1 (HO-1) pathway activation and the possible neuroprotective effects of tert-butylhydroquinone (tBHQ) pretreatment on PQ-induced neurodegeneration in vivo and in vitro. Treatment with PQ caused decreased spontaneous locomotor activity, decreased tyrosine hydroxylase (TH)-positive neurons, increased terminal deoxynucleotidyl transferase-mediated dUTP biotin nick end-labeling (TUNEL)-positive cells in the substantia nigra, as well as increased protein levels of both nuclear Nrf2 and HO-1. In PQ-treated mice, pretreatment with tBHQ significantly attenuated behavioral performance impairment, the decrease in TH-positive neurons, and the increase in TUNEL-positive cells in the substantia nigra, as well as increased the protein expression of both nuclear Nrf2 and HO-1. In PC12 cells, pretreatment with tBHQ protected against PQ-mediated cytotoxicity. The dual-luciferase reporter gene also revealed the transcriptional activation of the HO-1 gene expression of antioxidant responsive element via Nrf2 as a consequence of PQ exposure. Collectively, these results clearly indicated for the first time that the Nrf2/HO-1 pathway in dopaminergic nigra was activated by PQ, and pretreatment with tBHQ conferred neuroprotection against PQ-induced Parkinsonism probably by increasing Nrf2 and HO-1 expression. This work was supported by National Natural Science Foundation in China (Grant 30800936, 81172715) and by Program for New Century Excellent Talents in Fujian Province University (NCETFJ).
Keywords: Paraquat; Neurodegeneration; tert-Butylhydroquinone (tBHQ); NF-E2-related factor 2 (Nrf2); Heme oxygenase 1 (HO-1)

抄録全体を表示

Capsaicin (CPS), a constituent of green and red peppers, has been linked with suppression of tumorigenesis and carcinogenesis. However, the influence of CPS on CYP3A4 expression and their mechanisms remain unclear. In this study, we examined the effect of capsaicin on CYP3A4 expression in human hepatoma HepG2 cells and the pharmacokinetic profiles of CYP3A4 substrates in male Sprague-Dawley rats. Capsaicin induced enzyme activity and expression of CYP3A4 in pregnane X receptor (PXR)-transfected HepG2 cells. Additionally, this induction was mediated by the transient receptor potential vanilloid type-1 receptor (TRPV1) in capsaicin-treated HepG2 cells. Capsaicin also induced the activation of CCAAT/enhancer-binding proteinβ (C/EBPβ). Furthermore, capsaicin elevated the level of CYP3A1 in rat liver and significantly increased the biotransformation of nifedipine to dehydronifedipine. Taken together, capsaicin induces CYP3A4 expression in vitro and in vivo and this induction was achieved by the induction of PXR and activation of C/EBPβ.

抄録全体を表示

Sesame oil has been used in traditional Chinese medicine to treat inflammatory pain and irritation of toothache, minor scrapes, cuts, and bruises. We investigated the preventive effect of sesame oil on monosodium urate monohydrate (MSU) crystal-induced acute gouty inflammation in rats. Air pouch, a pseudosynovial cavity, was established by injecting 24 mL of filtered sterile air subcutaneously in the backs of the rats. At day 0, inflammation in air pouch was induced by injecting MSU crystal (5 mg/rat, suspended in sterilized phosphate buffered saline, pH 7.4), while sesame oil (0, 1, 2, or 4 mL/kg, orally) was given 6 h before MSU crystal injection. Parameters in lavage and skin tissue from the air pouches were assessed 12 h after MSU. MSU increased the leukocyte counts in lavage, and increased the levels of tumour necrosis factor-α, interleukin (IL)-1β, and IL-6 in both lavage and pouch tissue. Pre-treated sesame oil prevents the increase in all the examined parameters. However, no change in lipid peroxidation levels was found in MSU alone group and sesame oil-treated groups. We summarized that sesame oil has a potent preventive effect against MSU crystal-induced acute inflammatory response in rat.

KEYWORDS: Gout, hyperuricemia, monosodium urate monohydrate, inflammation, arthritis, rats

抄録全体を表示

Progoitrin is one of the glucosinolates which are abundant in Cruciferous vegetables and broken down with myrosinase when plant tissues are damaged through food processing or mastication. Progoitrin is reported to be degraded to either one of the two cyanide compounds or an isothiocyanate. The isothiocyanate is spontaneously cyclized to goitrin, which have been demonstrated to have an adverse effect on the thyroid. In this study, degradation of progoitrin from Chinese cabbage and rape was investigated in different food processing pH (pH 4, pH 6, and fermentation) and temperature of grinding (room temperature, 37 °C, and 100 °C). The amounts of the breakdown products and the remaining amount of progoitrin were monitored by GC-MS and HPLC, respectively. The generation of cyanide compounds was relatively favored in boiling temperature, pH 4, and pH 6.5 groups reflecting typical non-enzymatic degradation. However the fermentation showed a significant increase in goitrin formation. Goitrin was found to be the main product in fermentation group, comprising 45 % and 39 % of total breakdown products in room temperature and 37 °C groups, respectively. Considering large consumption of fermented Cruciferous vegetables such as Kimchi in Korea, the result warrants further research on the effect of fermentation and risk assessment of progoitrin.

抄録全体を表示

Our Percellome Toxicogenomics Project aims at describing dynamic and comprehensive toxicobiological gene network for the development of predictive toxicology using time- and dose-dependent transcriptomic responses induced by a chemical. We consider that this approach meet demand for food safety assessment as well. There are 45 food-related chemicals among over 100 chemicals tested in the Percellome Project. The list includes, food additives, food components, functional health foods, agrochemicals, food contaminants including those could leach out from the package materials. As an example of our application of Percellome Toxicogenomics approach to food, we report a case study of a flavor, estragole, which is known as a major component of tarragon, and essential oils of basil.
Male mice (C57BL/6, 12 weeks old) were given a single oral dose of estragole at three doses (10, 30 and 100 mg/kg) as well as vehicle control of corn oil. We sampled the liver at 2, 4, 8 and 24 hr post-gavage (three animals from each dose group), and applied to Affymetrix GeneChip Mouse Genome 430 2.0 to obtain Percellome transcriptomic data.
Our data indicated that estragole is a PPAR-alpha agonist, with a potency comparable to clofibrate in mg/kg basis. Binding and signaling property of estragole to PPAR-alpha should be confirmed for further toxicological consideration of this food additive. If proven, its hepatocarcinogenic property should be reevaluated accordingly.

抄録全体を表示

Resveratrol (trans-3,4',5-trihydroxystilbene, RES), a phytoestrogen, exists in grape skin, peanuts, and red wine, known for beneficial material, due to its antioxidant, anti-inflammatory effects. Endocrine disrupting chemicals (EDCs) appear to promote the development and progression of the estrogen dependant cancers. In this study, we evaluated the inhibitory effect of RES on the cell growth and progression induced by various EDCs in BG-1 ovarian cancer cells expressing estrogen receptors (ERs). The various EDCs, i.e., bisphenol A (BPA), nonylphenol (NP), octylphenol (OP), methoxychlor (MXC), and hexabromocyclododecane (HBCD) were employed in this study. In the in vitro experiments, treatments of BG-1 cells with E2, BPA, NP, OP, MXC, or HBCD resulted in an increase of their growth. The treatment of BG-1 cells with ICI 182,780, a well known antagonist of ERs, reversed EDCs induced cell growth in these cells, indicating that their growth stimulatory effect is mediated through ERs. In addition, we evaluated the effect of RES in the presence of other EDCs by MTT assay. As a result, increased cell viability induced by these EDCs. On the other hand, cell viability of co-cultured with RES was decreased. In addition, we further examined the regulation of cell cycle dependent genes by RT-PCR in E2, BPA, or NP and co-treatment of RES and each EDC. Concretely, the treatment with each EDC only decreased the gene expression of p21 and increased the expression of cell cycle-dependent kinase 2 (CDK2). However, co-treatment with RES and one of EDCs resulted in the increased gene expressions of p21 and the decreased expression of CDK2. Cyclin D1 was increased by downregulating p21 when only treated with each EDC in the absence of RES, while co-treatment with RES and each EDC decreased the gene expression of cyclin D1 by upregulating p21. Taken together, these results indicate that RES appear to be Cyclin D1 and CDK2 inhibitor and is responsible for the cell cycle arrest at G₁ phase. In addition, when co-treated with each EDC, RES increased the expressions of p21 and resulted in the growth inhibition of BG-1 ovarian cancer cells. As a result, we confirmed the cell growth inhibitory effect of RES, a dietary phytoestrogen, on the estrogen-dependant ovarian cancer cells prompted by EDCs. [This work was supported by a National Research Foundation of Korea (NRF) grant funded by the Ministry of Education, Science and Technology (MEST) of Korea government (no. 2011-0015385).]

抄録全体を表示