Induction of oxidative stress and TNF-α secretion by Dichloroacetonitrile, a water disinfectant by-product, as possible mediators of apoptosis or necrosis in a murine macrophage cell line (RAW)

A. E. Ahmed, Judith Aronson, S. Jacob

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25 Citations (Scopus)

Abstract

The water disinfectant by-product dichloroacetonitrile (DCAN) is a direct-acting mutagen and induces DNA strand breaks in cultured human lymphoblastic cells. Cellular activation by environmental agents may exert detrimental effects to the cells. Activated macrophages produce reactive oxygen intermediates such as H2O2, -OH and O2. Therefore, the effect of various concentrations of DCAN (100-400 μM) on the activity macrophage cells (RAW 264.7) was studied. In these cells, DCAN-induced oxidative stress was characterized by the production of reactive oxygen intermediates (ROI). Also, the ratios of intracellular GSH/GSSG was assessed and used as a biomarker for oxidative stress. The secretion of TNF-α was assessed since macrophages are known to secrete TNF-α as a result of cellular oxidative stress. Electrophoretic detection of DNA degradation and light microscopy was utilized for the characterization of DCAN-induced apoptosis. Lactate dehydrogenase (LDH) leakage and trypan blue exclusion were used as markers of cellular necrosis. Following exposure to DCAN (200 μM and 400 μM), intracellular GSSG was increased (2.5-fold of control, P<0.05). DCAN activation of RAW cells was detected by elevated levels of intracellular ROI (1.9-2.5-fold than control, P<0.05) and increased secretion of TNF-α (4.5 fold-than control, P <0.05). Elecrophoresis of genomic DNA of treated cells indicated a dose-dependent increase in degradation of genomic DNA. Morphological studies also indicated that exposure of RAW cells to 100 μM or 200 μM DCAN incites apoptic cell death. At higher concentrations (400 μM), however, significant (P<0.05) increase in LDH leakage and decrease in cell viability (55% of control) indicative of cellular necrosis, were observed. These studies indicate that DCAN induces dose-dependent apoptosis or necrosis in RAW cells that could be due to the disturbance in intracellular redox status and initiation of ROI-mediated oxidative mechanisms of cellular damage. Copyright (C) 2000 Elsevier Science Ltd.

Original languageEnglish (US)
Pages (from-to)199-210
Number of pages12
JournalToxicology in Vitro
Volume14
Issue number3
DOIs
StatePublished - Jun 2000

Fingerprint

Oxidative stress
Disinfectants
Macrophages
Byproducts
Oxidative Stress
Necrosis
Cells
Apoptosis
Cell Line
Water
Oxygen
Glutathione Disulfide
DNA
L-Lactate Dehydrogenase
Chemical activation
Degradation
DNA Breaks
Trypan Blue
dichloroacetonitrile
Mutagens

Keywords

  • Apoptosis
  • DCAN
  • Glutathione
  • GSH oxidation
  • HAN
  • Macrophage activation
  • Necrosis
  • Oxidative stress
  • Peritoneal macrophages
  • TNF-α

ASJC Scopus subject areas

  • Toxicology

Cite this

@article{62927f5e299b4cafac45cdac36199a6d,
title = "Induction of oxidative stress and TNF-α secretion by Dichloroacetonitrile, a water disinfectant by-product, as possible mediators of apoptosis or necrosis in a murine macrophage cell line (RAW)",
abstract = "The water disinfectant by-product dichloroacetonitrile (DCAN) is a direct-acting mutagen and induces DNA strand breaks in cultured human lymphoblastic cells. Cellular activation by environmental agents may exert detrimental effects to the cells. Activated macrophages produce reactive oxygen intermediates such as H2O2, -OH and O2. Therefore, the effect of various concentrations of DCAN (100-400 μM) on the activity macrophage cells (RAW 264.7) was studied. In these cells, DCAN-induced oxidative stress was characterized by the production of reactive oxygen intermediates (ROI). Also, the ratios of intracellular GSH/GSSG was assessed and used as a biomarker for oxidative stress. The secretion of TNF-α was assessed since macrophages are known to secrete TNF-α as a result of cellular oxidative stress. Electrophoretic detection of DNA degradation and light microscopy was utilized for the characterization of DCAN-induced apoptosis. Lactate dehydrogenase (LDH) leakage and trypan blue exclusion were used as markers of cellular necrosis. Following exposure to DCAN (200 μM and 400 μM), intracellular GSSG was increased (2.5-fold of control, P<0.05). DCAN activation of RAW cells was detected by elevated levels of intracellular ROI (1.9-2.5-fold than control, P<0.05) and increased secretion of TNF-α (4.5 fold-than control, P <0.05). Elecrophoresis of genomic DNA of treated cells indicated a dose-dependent increase in degradation of genomic DNA. Morphological studies also indicated that exposure of RAW cells to 100 μM or 200 μM DCAN incites apoptic cell death. At higher concentrations (400 μM), however, significant (P<0.05) increase in LDH leakage and decrease in cell viability (55{\%} of control) indicative of cellular necrosis, were observed. These studies indicate that DCAN induces dose-dependent apoptosis or necrosis in RAW cells that could be due to the disturbance in intracellular redox status and initiation of ROI-mediated oxidative mechanisms of cellular damage. Copyright (C) 2000 Elsevier Science Ltd.",
keywords = "Apoptosis, DCAN, Glutathione, GSH oxidation, HAN, Macrophage activation, Necrosis, Oxidative stress, Peritoneal macrophages, TNF-α",
author = "Ahmed, {A. E.} and Judith Aronson and S. Jacob",
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T1 - Induction of oxidative stress and TNF-α secretion by Dichloroacetonitrile, a water disinfectant by-product, as possible mediators of apoptosis or necrosis in a murine macrophage cell line (RAW)

AU - Ahmed, A. E.

AU - Aronson, Judith

AU - Jacob, S.

PY - 2000/6

Y1 - 2000/6

N2 - The water disinfectant by-product dichloroacetonitrile (DCAN) is a direct-acting mutagen and induces DNA strand breaks in cultured human lymphoblastic cells. Cellular activation by environmental agents may exert detrimental effects to the cells. Activated macrophages produce reactive oxygen intermediates such as H2O2, -OH and O2. Therefore, the effect of various concentrations of DCAN (100-400 μM) on the activity macrophage cells (RAW 264.7) was studied. In these cells, DCAN-induced oxidative stress was characterized by the production of reactive oxygen intermediates (ROI). Also, the ratios of intracellular GSH/GSSG was assessed and used as a biomarker for oxidative stress. The secretion of TNF-α was assessed since macrophages are known to secrete TNF-α as a result of cellular oxidative stress. Electrophoretic detection of DNA degradation and light microscopy was utilized for the characterization of DCAN-induced apoptosis. Lactate dehydrogenase (LDH) leakage and trypan blue exclusion were used as markers of cellular necrosis. Following exposure to DCAN (200 μM and 400 μM), intracellular GSSG was increased (2.5-fold of control, P<0.05). DCAN activation of RAW cells was detected by elevated levels of intracellular ROI (1.9-2.5-fold than control, P<0.05) and increased secretion of TNF-α (4.5 fold-than control, P <0.05). Elecrophoresis of genomic DNA of treated cells indicated a dose-dependent increase in degradation of genomic DNA. Morphological studies also indicated that exposure of RAW cells to 100 μM or 200 μM DCAN incites apoptic cell death. At higher concentrations (400 μM), however, significant (P<0.05) increase in LDH leakage and decrease in cell viability (55% of control) indicative of cellular necrosis, were observed. These studies indicate that DCAN induces dose-dependent apoptosis or necrosis in RAW cells that could be due to the disturbance in intracellular redox status and initiation of ROI-mediated oxidative mechanisms of cellular damage. Copyright (C) 2000 Elsevier Science Ltd.

AB - The water disinfectant by-product dichloroacetonitrile (DCAN) is a direct-acting mutagen and induces DNA strand breaks in cultured human lymphoblastic cells. Cellular activation by environmental agents may exert detrimental effects to the cells. Activated macrophages produce reactive oxygen intermediates such as H2O2, -OH and O2. Therefore, the effect of various concentrations of DCAN (100-400 μM) on the activity macrophage cells (RAW 264.7) was studied. In these cells, DCAN-induced oxidative stress was characterized by the production of reactive oxygen intermediates (ROI). Also, the ratios of intracellular GSH/GSSG was assessed and used as a biomarker for oxidative stress. The secretion of TNF-α was assessed since macrophages are known to secrete TNF-α as a result of cellular oxidative stress. Electrophoretic detection of DNA degradation and light microscopy was utilized for the characterization of DCAN-induced apoptosis. Lactate dehydrogenase (LDH) leakage and trypan blue exclusion were used as markers of cellular necrosis. Following exposure to DCAN (200 μM and 400 μM), intracellular GSSG was increased (2.5-fold of control, P<0.05). DCAN activation of RAW cells was detected by elevated levels of intracellular ROI (1.9-2.5-fold than control, P<0.05) and increased secretion of TNF-α (4.5 fold-than control, P <0.05). Elecrophoresis of genomic DNA of treated cells indicated a dose-dependent increase in degradation of genomic DNA. Morphological studies also indicated that exposure of RAW cells to 100 μM or 200 μM DCAN incites apoptic cell death. At higher concentrations (400 μM), however, significant (P<0.05) increase in LDH leakage and decrease in cell viability (55% of control) indicative of cellular necrosis, were observed. These studies indicate that DCAN induces dose-dependent apoptosis or necrosis in RAW cells that could be due to the disturbance in intracellular redox status and initiation of ROI-mediated oxidative mechanisms of cellular damage. Copyright (C) 2000 Elsevier Science Ltd.

KW - Apoptosis

KW - DCAN

KW - Glutathione

KW - GSH oxidation

KW - HAN

KW - Macrophage activation

KW - Necrosis

KW - Oxidative stress

KW - Peritoneal macrophages

KW - TNF-α

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DO - 10.1016/S0887-2333(00)00019-9

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VL - 14

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EP - 210

JO - Toxicology in Vitro

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