Annals of Occupational and Environmental Medicine

Effect of Glutathione on Apoptosis Induced by Methyl Mercury Chloride: Jung Ho Youm, Dai Ha Koh, Keun Sang Kwon, Me Yae Lee; Korean Journal of Occupational and Environmental Medicine 2002;14(4):377-391. Published online December 31, 2002; DOI: https://doi.org/10.35371/kjoem.2002.14.4.377

Abstract PDF: OBJECTIVES
This study was performed to evaluate the critical role of glutathione(GSH) in methyl mercury chloride(MeHgCl)induced cell apoptosis.
METHODS
The effect of GSH in MeHgCl induced cell apoptosis was observed in mouse macrophage-derived RAW 264.7 cells in vitro. The cells were cultured in Dulbecco's modified Eagle's medium(DMEM).
RESULTS
MeHgCl exerted a dose dependent cytotoxicity,as demonstrated by the MTT assay, which is an assay dependent partially on the mitochondrial function. Moreover, in the presence of NAC, a GSH precursor, the MeHgCl induced cytotoxicity was significantly decreased whereas BSO, a specific GSH synthesis inhibitor,increased the MeHgCl induced cytotoxicity.The MeHgCl induced DNA fragmentation and chromatin condensation was consistent with the morphological alterations. The MeHgCl treated cells exhibited increasing annexin V-FITC binding to the phos-phatidylserine(PS)translocated from the inner to the outer leaflet of the plasma membrane and those cells with NAC pretreatment significantly exhibited decreasing annexin V-FITC binding compared to the cells treated with MeHgCl only. However BSO pretreatment markedly exhibited the increasing annexin V-FITC binding. The MeHgCl treated cells generated ROS, which was evidenced by the oxidation of dihydroethidine and the generation of the fluorescent product, ethidium. In addition, BSO pretreatment further enhanced the extent of ROS generation caused by MeHgCl whereas NAC pretreatment decreased the amount of ROS generation. MeHgCl led to a dose dependent decrease in the GSH content. Although MeHgCl exposure significantly reduced the GSH level, those cells that had a NAC pretreatment contained a higher level of GSH compared to the cells treated with MeHgCl only. In contrast, BSO pretreatment futher enhanced the extent of GSH depletion caused by MeHgCl.
CONCLUSIONS
These results indicate that MeHgCl reduced the GSH content and impaired the defense against oxidative damage caused by ROS formation in RAW 264.7 cells. It is possible that these factors leads to the activation of the apoptosis signaling pathway. Ultimately these results suggest that GSH plays a crucial role in protecting the activity against MeHgCl induced apoptosis.

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A Study on Factors Related to NO Synthesis by Mercurial Compounds in the EMT-6 cell: Jung Ho Youm; Korean Journal of Occupational and Environmental Medicine 1997;9(1):122-130. Published online February 28, 1997; DOI: https://doi.org/10.35371/kjoem.1997.9.1.122

Abstract PDF: The effects of several factors on the nitrite synthesis were observed in the culture of EMT-6 cells which are originated from mammary adenocarcinoma of Balb/c mouse. The cells were cultured in Dulbecco's Modified Eagle's Medium containing 10% fetal calf serum. Amounts of nitrite in the culture media after 24 and 36 hours of culture were about 2 fold, and 3-fold of those measured after 12 hours respectively. There were very close associations between the amounts of nitrite measured in the culture media. A significant nitrite synthesis by EMT-6 cells occurred when IL-1 was added to the culture medium with other cytokines as IFN gamma or TNF alpha . One of each cytokines were less effective as an inducer of nitrite than the combinations of cytokines. When mercury chloride or cinnamate was added in the culture medium, the nitrite synthesis was dose-dependently decreased by the concentration of these materials. The viability of EMT-6 cells were kept on 95% or above in 36 hours after beginning of culture without any specific additives except cytokines. While after 48 hours it went down to 85% or less. These viability were decreased by the prolongation of culture time (48 hours or more), the addition of TNF alpha to cytokine mixture, and the higher concentrations of mercury chloride or cinnamate to culture medium. Simultaneous addition of the equimolar dose of selenium completely prevented mercurial compounds-induced inhibitions of nitrite syntheses. But the single addition of selenium neither influenced the viability of cells nor the productions of nitrite. These results suggests that the disorder of cell mediated immunity by mercurial compounds could be related to the inhibition of nitric oxide synthesis and selenium decreased the cytotoxicity of mercurial compounds.

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A Study of Cytotoxical Mechanism of Mercurial Compounds in RAW264.7 Cell Line: Kong Ho Kirn, Byoung Yul Soh, Dai Ha Koh; Korean Journal of Occupational and Environmental Medicine 1996;8(3):560-569. Published online December 31, 1996; DOI: https://doi.org/10.35371/kjoem.1996.8.3.560

Abstract PDF: The effects of glucose on the productions of ATP and nitrite which are inhibited by mercury compounds, were examined in a cell culture system of RAW 264.7 cells. The cells were cultured in Dulbecco's Modified Eagle's medium (DMEM) with cytokines, IL-1 and TNF for 24 hours. The viablility of RAW 264.7 cells at the end of culture was significantly decreased by mercury chloride or methylmercury chloride added into the media in dose-dependent manner, however the viability of RAW 264.7 cells were influenced in the concentrations lese than 0.8micrometer of mercury chloride or 0.4micrometer of methylmercury chloride. The addition of 4.5 g/l glucose to normal DMEM lowered the pH of media to the range of 6.7-6.8 after 48 hours of culture, but not for the cell survivals. This supplement of glucose to the media also prevented the inhibitions of ATP and nitrite syntheses which were caused by mercurial compounds. These results suggest that the disorder of cell mediated immunity by mercurials could be related to the inhibition of nitric oxide synthesis which seams to be caused by the inhibition of ATP synthesis, especially related to the citric acid cycle.

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Effect of Mercury Chloride on Peritoneal Macrophage or EMT-6 cell from Balb/c mice: Dai Ha Koh, Jung Ho Youm, No Suk Ki, Gyung Jae Oh, Kuen Sang Kwon, Sung Yeup Kim, Nam Song Kim; Korean Journal of Occupational and Environmental Medicine 1996;8(2):201-209. Published online September 30, 1996; DOI: https://doi.org/10.35371/kjoem.1996.8.2.201

Abstract PDF: The effect of treatment of mercury chloride on the nitrite and nitrate synthesis was observed in peritoneal macrophages from Balb/c mice and EMT-6 cells in vitro. The cells were cultured in Dulbecco's modified Eagle's medium (DMEM) with cytokines. Amounts of nitrite and nitrate in the culture media after 24 and 36 hours of culture were about 2-fold, and 3-fold of those measured after 12 hours respectively. There were very close associations between the amounts of nitrite and nitrate measured in the culture media, according to culture time. The survival rate of peritoneal macrophages was significantly decreased by mercury chloride added into the media in dose-dependent manner, however the survivals of EMT-6 cells were not influenced by mercury chloride concentration in media. Nitrite and nitrate syntheses were dose-dependently decreased by mercury chloride added in culture media. These results reported here suggest that the disorder of cell mediated immunity by mercurials could be related to the inhibition of nitric oxide synthesis which seems to be caused by the inhibition of metabolism of cells.

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