Elsevier

Toxicology

Volume 231, Issue 1, 28 February 2007, Pages 30-39
Toxicology

Dose- and time-dependent effects of sulfur mustard on antioxidant system in liver and brain of rat

https://doi.org/10.1016/j.tox.2006.11.048Get rights and content

Abstract

This study investigates the dose- and time-dependent effects of sulfur mustard (SM) on antioxidant system and lipid peroxidation in liver and brain of rats. For this purpose, male Wistar rats were randomly divided into eight groups and treated as follows: group1 as control and groups 2–8 as experimental groups that received SM (1–80 mg/kg) through intraperitoneal injection. Rats were killed after 2, 7 and 14 days of exposure. SM dose-dependently decreased body weight. Superoxide dismutase (SOD), catalase (CAT) and glutathione S-transferase (GST) activities in liver were significantly increased at SM doses lower than 10 mg/kg after 2 and 7 days of exposure. However, the recovery of these parameters was observed after 14 days. At these concentrations, no significant change in glutathione (GSH) and malondialdehyde (MDA) levels were observed.

At doses higher than 10 mg/kg, SM significantly decreased SOD, CAT, glutathione peroxidase (GPX), and GST activities in liver and brain and decreased glutathione reductase (GR) activity in liver, which was associated with a depletion of GSH and increased MDA level. Present data indicate that the effect of SM is dose- and time-dependent and at higher doses (>10 mg/kg) induces an oxidative stress response by depleting the antioxidant defense systems and increasing lipid peroxidation in liver and brain of rats.

Introduction

Sulfur mustard (SM) or bis (2-chloroethyl) sulfide) is a strong alkylating agent with known mutagenic and suspected carcinogenic properties that is frequently used as a chemical warfare agent (Marrs et al., 1996, Balali-Mood and Hefazi, 2005). In aqueous solutions, SM rapidly hydrolyzes to form cyclic ethylene episulfonium intermediate, which reacts with compounds containing nucleophilic functional groups like amino, sulfhydryl, carboxylic and hydroxyl in proteins and nucleic acid (Papirmeister et al., 1991, Le and Knudsen, 2006).

The formation of reactive oxygen species (ROS) is a naturally occurring intracellular metabolic process. These harmful species are known to cause oxidative damage to a number of molecules in cells, including membrane lipids, proteins, and nucleic acids (Mates et al., 1999, Raha and Robinson, 2000). The potential harmful effects of these species are controlled by the cellular antioxidant defense system that includes enzymes such as superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPX), and glutathione reductase (GR), and antioxidants such as reduced glutathione (GSH) and Vitamins A, C, and E (Shull et al., 1991, McCord, 1993). Also, glutathione S-transferase (GST) protects cells against toxicants by conjugating them to GSH, thereby neutralizing their electrophilic sites, and increasing their solubility and aiding excretion from the cell (Habig et al., 1984). The imbalance between the rate of ROS production and the effect of protective antioxidatiants leads to oxidative damage, which is also known as oxidative stress (Southorn and Powis, 1988, McCord, 2000).

It is well known that liver is the first organ that absorbs a variety of toxic chemicals, e.g. SM and it is obliged to their metabolism by biotransformation. This process causes the decrease of NAD+:NADH ratio leads to manifesting oxidase activity, including xanthine oxidase, which is a principal source of superoxide anion formation. The superoxide can cross membranes via anion channels and may assume the peroxyl radical. The peroxyl radical is more reactive than the superoxide and is particularly dangerous to membrane proteins and lipids. A high concentration of lipid peroxides is toxic to cells (Papirmeister et al., 1991, Naghii, 2002, Elsayed and Omaye, 2004). For biochemical, physiological, and anatomical reasons, brain is especially vulnerable to oxygen mediated injury (Evans, 1999).

Although it's cytotoxic and vesicant properties have been documented extensively (Dacle and Goldman, 1996, Saladi et al., 2006), the molecular basis of its mechanism of action remains unknown. Understanding the underlying mechanisms of cell injury and death induced by SM will be extremely helpful in the development of effective countermeasures to this weapon of terror. This study was undertaken in order to evaluate the dose and time response of SM-induced changes in the endogenous antioxidant defense system such as the GSH level, antioxidant enzyme activities of SOD, CAT, GPX, GR and GST, and lipid peroxidation in rats.

Section snippets

Chemicals

Reduced glutathione, oxidized glutathione (GSSG), NADH, NADPH, 1-chloro-2, 4-dinitrobenzene (CDNB), glutathione reductase, and 5, 5′-dithiobis 2-nitrobenzoic acid (DTNB) were obtained from Sigma Chemical Company. Sephadex G-25 was purchased from Amersham Biosciences. All other chemicals used were of extra pure grad and obtained from Sigma and Merck. SM (purity>98%) was dissolved in dimethyl sulfoxide (DMSO) at a stock concentration of 160 mg/ml, and then diluted with 10 mM phosphate buffer pH 7.4

Results

The results show that SM decreased the body weight of rats (86, 82, 74 and 70% of control) at 10, 20, 40 and 80 mg/kg dose, respectively, 48 h after its administration. The significant decrease in body weight was observed at 5 mg/kg (88%), 10 mg/kg (74%) and 20 mg/kg (68%) dose of SM, on 7 days post-treatment. SM-exposed animals showed less food and water intake than the control animals. At higher than 10 mg/kg dose of SM, diarrhea and gastrointestinal bleeding were seen in rats and their eyes were

Discussion

Although mustard has been considered a major chemical weapon for years, there is still no clear understanding of its biochemical mechanism of action; therefore, no specific therapy for its effects exists (Sidell et al., 1997). Several hypotheses have been proposed to explain SM-induced toxicity. One theory suggested that SM reacts with intracellular GSH, resulting in a rapid inactivation of sulfhydryl groups which in turn leads to loss of protection against free radicals, specifically that

Acknowledgements

The author would like to thank R. Rezaie, H. Mahdavi, and J. Rasouli for their assistance. This work was supported by a grant from the Research Center of Chemical Injuries of Baqiyatallah Medical Sciences University.

References (56)

  • Y. Kono et al.

    Superoxide radical inhibits catalase

    J. Biol. Chem.

    (1982)
  • O. Kumar et al.

    Protective effect of various antioxidants on the toxicity of sulphur mustard administered to mice. By inhalation or percutaneous routes

    Chem. Biol. Interact.

    (2001)
  • J.M. Mates et al.

    Antioxidant enzymes and human diseases

    Clin. Biochem.

    (1999)
  • J.M. McCord

    Human disease, free radicals, and the oxidant antioxidant balance

    Clin. Biochem.

    (1993)
  • J.M. McCord

    The evolution of free radicals and oxidative sress

    Am. J. Med.

    (2000)
  • A. Meister

    Glutathione metabolism

    Methods Enzymol.

    (1995)
  • H. Ohkawa et al.

    Assay for lipid peroxides in animal tissues by thiobarbitoric acid reaction

    Anal. Biochem.

    (1979)
  • F. Paoletti et al.

    Determination of superoxide dismutase activity by purely chemical system based on NAD (P)H oxidation

    Methods Enzymol.

    (1990)
  • E. Pigeolet et al.

    Glutathione peroxidase, superoxide dismutase and catalase inactivation by peroxides and oxygen derived free radicals

    Mech. Aging Dev.

    (1990)
  • S. Raha et al.

    Mitochondria, oxygen free radicals, disease, and aging

    Trends Biochem. Sci.

    (2000)
  • R. Ray et al.

    Sulfur mustard-induced increase in intracellular free calcium level and arachidonic acid release from cell membrane

    Toxicol. Appl. Pharmacol.

    (1995)
  • D. Ritter et al.

    Development of a cell culture model system for routine testing of substances inducing oxidative stress

    Toxicol. In Vitro

    (1999)
  • S. Shull et al.

    Differential regulation of antioxidant enzymes in response to oxidants

    J. Biol. Chem.

    (1991)
  • P.A. Southorn et al.

    Free radicals in medicine. 1. Chemical nature and biology reactions

    Mayo. Clin. Proc.

    (1988)
  • R. Vijayaraghavan et al.

    Dermal intoxication of mice with with bis (2-chloroethyl) sulphide and the protective effect of flavonoids

    Toxicology

    (1991)
  • K.B. Atkins et al.

    N-acetylcysteine and endothelial cell injury by sulfur mustard

    J. Appl. Toxicol.

    (2000)
  • M. Balali-Mood et al.

    The pharmacology, toxicology, and medical treatment of sulphur mustard poisoning

    Fundam. Clin. Pharmacol.

    (2005)
  • I. Carlberg et al.

    Glutathion reductase

    Methods Enzymol.

    (1985)
  • Cited by (81)

    • Clinical and experimental research progress on neurotoxicity of sulfur mustard and its possible mechanisms

      2023, Toxicology
      Citation Excerpt :

      It has been reported that the effect of SM is dose- and time- dependent and oxidative stress response can induce at higher doses (>10 mg/kg) by depleting the antioxidant defense system and by raising the production of peroxides of liver and brain in rats. Moreover, SM can significantly reduce superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPX), and glutathione S-transferase (GST) activities in liver and brain (Jafari, 2007). Sharma et al. (2009) examined the effects of chronic SM poisoning on neurobehavioral impairments, oxidative stress of mitochondria in male Swiss Albino mice and its effect on causing apoptotic neuron death.

    • Evidence for the systemic diffusion of (2-chloroethyl)-ethyl-sulfide, a sulfur mustard analog, and its deleterious effects in brain

      2021, Toxicology
      Citation Excerpt :

      A widely proposed explanation to the toxicity of SM and CEES is their ability to deplete the GSH pool (Abel et al., 2011; Black et al., 1992a, b; Davison et al., 1961; Kinsey and Grant, 1947) and thereby inducing oxidative stress by reducing cellular antioxidant capacities. This effect is reflected by the modulation of a wide variety of markers of oxidative stress (Husain et al., 1996; Jafari, 2007; Mukhopadhyay et al., 2006; Paromov et al., 2007; Pohanka et al., 2011, 2013; Varmazyar et al., 2019). Evidence of the role of oxidative stress is confirmed by the ability of antioxidants at counteracting the toxic effects of SM (Mukherjee et al., 2009; Paromov et al., 2011, 2008; Rappeneau et al., 2000; Shohrati et al., 2008; Tewari-Singh et al., 2011).

    View all citing articles on Scopus
    View full text