Hakan Yaren

Molecular targets against mustard toxicity: implication of cell surface receptors, peroxynitrite production, and PARP activation

Despite many years of research into chemical warfare agents, cytotoxic mechanisms induced by mustards are not well understood. Reactive oxygen and nitrogen species (ROS and RNS) are likely to be involved in chemical warfare agents induced toxicity. These species lead to lipid peroxidation, protein oxidation, and DNA injury, and trigger many pathophysiological processes that harm the organism. In this article, several steps of pathophysiological mechanisms and possible ways of protection against chemical warfare agents have been discussed. In summary, pathogenesis of mustard toxicity is explained by three steps: (1) mustard binds target cell surface receptor, (2) activates intracellular ROS and RNS leading to peroxynitrite (ONOO−) production, and (3) the increased ONOO− level damages organic molecules (lipids, proteins, and DNA) leading to poly(adenosine diphosphate-ribose) polymerase (PARP) activation. Therefore, protection against mustard toxicity could also be performed in these ways: (1) blocking of cell surface receptor, (2) inhibiting the ONOO− production or scavenging the ONOO− produced, and (3) inhibiting the PARP, activated by ONOO− and hydroxyl radical (OH•) induced DNA damage. As conclusion, to be really effective, treatment against mustards must take all molecular mechanisms of cytotoxicity into account. Combination of several individual potent agents, each blocking one of the toxic mechanisms induced by mustards, would be interesting. Therefore, variations of combination of cell membrane receptor blockers, antioxidants, nitric oxide synthase inhibitors, ONOO− scavengers, and PARP inhibitors should be investigated.

Beneficial Effects of N-Acetylcysteine and Ebselen on Renal Ischemia/Reperfusion Injury

Abstract Introduction: It has been demonstrated that peroxynitrite accompanies acute renal ischemia and contributes to the pathophysiology of renal damage. Therefore, we aimed to investigate the roles of N-acetylcysteine (NAC), a well-known powerful antioxidant, and ebselen (E), a scavenger of peroxynitrite, on renal injury induced by renal ischemia/reperfusion injury (IRI) of rat kidney. Materials and methods: Forty male Sprague–Dawley rats were divided into five groups: sham, renal IRI, renal IRI+NAC, renal IRI+E, and renal IRI+NAC+E. IR injury was induced by 60 min of bilateral renal ischemia followed by 6 h of reperfusion. After reperfusion, kidneys and blood samples were obtained for histopathological and biochemical evaluations. Results: Renal IR resulted in increased malondialdehyde and nitrite/nitrate levels suggesting increased lipid peroxidation and peroxynitrite production and decreased superoxide dismutase and glutathione peroxidase activities. Both NAC and E alone significantly decreased malondialdehyde and nitrite/nitrate levels and increased superoxide dismutase and glutathione peroxidase activities. Additionally in the renal IRI+NAC+E group, all biochemical results were quite close to those of sham group. Histopathologically, the kidney injury in rats treated with combination of NAC and E was found significantly less than the other groups. Conclusions: Both NAC and E are able to ameliorate IRI of the kidney by decreasing oxidative and nitrosative stresses and increasing free radical scavenger properties. Additionally, combination of NAC and E prevents kidney damage more than when each drug is used alone, suggesting that scavenging peroxynitrite nearby antioxidant activity is important in preventing renal IRI.

High nitrate intake impairs liver functions and morphology in rats; protective effects of α-tocopherol.

The aim of this study was to determine the effect of high dose nitrate ingested in drinking water, on liver enzymes and histopathology, liver weight/body weight (lw/bw) ratio, serum and liver malondialdehyde (MDA) levels and osmotic fragility in Sprague-Dawley rats. These parameters were compared on 40 rats divided into four groups; control animals (group A) drank filtered tap water containing maximum 10mg/L nitrate while treatment groups drank 200mg/L (group B), 400mg/L (group C) and α-tocopherol plus 400mg/L (group D) nitrate containing water ad libitum for 60 days. As a result, lw/bw ratio increased significantly (p<0.05) among rats that consumed water with 400mg/L nitrate. Osmotic fragility increased significantly in treatment groups (p<0.05 versus control). Liver but not serum MDA levels increased in group C (p<0.05 versus control). Group A showed normal hepatic lobular architecture and histology. After nitrate administration, there was hepatocellular degeneration with increased intercellular space of the liver cells in groups B and C. Liver MDA, osmotic fragility and liver histology have returned to nearly normal in group D. These findings show clearly that high nitrate ingestion can cause pathological changes in liver histology and functions. Moreover, α-tocopherol can prevent these effects, possibly through antioxidant properties.

Oxidative Stress Levels in Rats Following Exposure to Oxygen at 3 atm for 0 - 120 min

BACKGROUND Hyperbaric oxygen (HBO) is known to cause oxidative stress in several organs and tissues. We previously defined the pressure-related oxidative effects of HBO in several tissues of rats. This study was performed to elucidate the relationship of HBO exposure time to its oxidative effects. METHODS A total of 49 rats were randomly divided into 5 groups. Study groups were subjected to 3 atm HBO for 30, 60, 90, and 120 min except the control group. Their blood and lungs were removed immediately after exposure and used for analysis. Thiobarbituric acid reactive substances (TBARS), superoxide dismutase (SOD), and glutathione peroxidase (GSH-Px) levels were determined to reflect oxidant and antioxidant status. RESULTS TBARS levels were found to increase in a time-dependent manner in both erythrocytes [median (min-max); from 0.65 (0.39-0.84) with 30 min HBO exposure up to 1.26 (1.00-1.44) nmol x g(-1) hemoglobin after 120 min] and lung tissue [from 2140 (1550-2510) up to 5465 (5090-5950) nmol x g(-1) protein]. Similarly, SOD activity also presented a dose-dependent course from 0.06 (0.05-0.10) to 0.18 (0.14-0.26) U x g(-1) hemoglobin in erythrocytes and from 16,660 (3479-25,994) to 52,522.5 (41,362-65,799) U x g(-1) protein in lung tissue. In contrast, GSH-Px activity reflected an irregular trend; its levels were mostly found to be increased, but they were decreased at one stage (in the erythrocytes of 30-min exposed rats). CONCLUSIONS The results of this study exhibited a clear relationship of HBO-induced oxidative action to exposure time. This action was most pronounced from 90 to 120 min of exposure.

Simultaneous determination of cyclosporine A, tacrolimus, sirolimus, and everolimus in whole-blood samples by LC-MS/MS.

Objectives. Cyclosporine A (CyA), tacrolimus (TRL), sirolimus (SIR), and everolimus (RAD) are immunosuppressive drugs frequently used in organ transplantation. Our aim was to confirm a robust sensitive and selective liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for determination of CyA, TRL, SIR, and RAD in whole-blood samples. Materials and Methods. We used an integrated online solid-phase extraction-LC-MS/MS system and atmospheric pressure ionization tandem mass spectrometry (API-MS/MS) in the multiple reaction monitoring (MRM) detection mode. CyA, TRL, SIR, and RAD were simultaneously analyzed in whole blood treated with precipitation reagent taken from transplant patients. Results. System performance parameters were suitable for using this method as a high-throughput technique in clinical practice. The high concentration of one analyte in the sample did not affect the concentration of other analytes. Total analytical time was 2.5 min, and retention times of all analytes were shorter than 2 minutes. Conclusion. This LC-MS/MS method can be preferable for therapeutic drug monitoring of these immunosuppressive drugs (CyA, TRL, SRL, and RAD) in whole blood. Sample preparation was too short and simple in this method, and it permits robust, rapid, sensitive, selective, and simultaneous determination of these drugs.

Epigenetic perturbations in the pathogenesis of mustard toxicity; hypothesis and preliminary results.

Epigenetic perturbations in the pathogenesis of mustard toxicity; hypothesis and preliminary results Among the most readily available chemical warfare agents, sulfur mustard (SM), also known as mustard gas, has been the most widely used chemical weapon. SM causes debilitating effects that can leave an exposed individual incapacitated for days to months; therefore delayed SM toxicity is of much greater importance than its ability to cause lethality. Although not fully understood, acute toxicity of SM is related to reactive oxygen and nitrogen species, oxidative stress, DNA damage, poly(ADP-ribose) polymerase (PARP) activation and energy depletion within the affected cell. Therefore several antioxidants and PARP inhibitors show beneficial effects against acute SM toxicity. The delayed toxicity of SM however, currently has no clear mechanistic explanation. One third of the 100,000 Iranian casualties are still suffering from the detrimental effects of SM in spite of the extensive treatment. We, therefore, made an attempt whether epigenetic aberrations may contribute to pathogenesis of mustard poisoning. Preliminary evidence reveals that mechlorethamine (a nitrogen mustard derivative) exposure may not only cause oxidative stress, DNA damage, but epigenetic perturbations as well. Epigenetic refers to the study of changes that influence the phenotype without causing alteration of the genotype. It involves changes in the properties of a cell that are inherited but do not involve a change in DNA sequence. It is now known that in addition to mutations, epimutations contribute to a variety of human diseases. Under light of preliminary results, the current hypothesis will focus on epigenetic regulations to clarify mustard toxicity and the use of drugs to correct possible epigenetic defects.

Protective effects of melatonin and S-methylisothiourea on mechlorethamine induced nephrotoxicity.

BACKGROUND In this study, we aimed to investigate the protective effects of melatonin (MEL) and S-methylisothiourea (SMT) on mechlorethamine (MEC) induced nephrotoxicity. MATERIALS AND METHODS A total of 36 male Sprague-Dawley rats were divided into four groups: control, MEC, MEC+MEL, and MEC+SMT. Three groups received single dose of MEC (3.5 mg/kg) via transdermal route. Control animals were given saline only via transdermal route. MEL (100 mg/kg) was administered intraperitoneally 30 min after the application of MEC, and after the same dose of MEL was given every 12 h for a total of six doses. SMT (50 mg/kg) was also given intraperitoneally 30 min after the application of MEC. RESULTS The tissue TNF-α, IL-1β, and NOx levels were found significantly different for all groups (P < 0.001). MEC application resulted in severe histopathological changes. Melatonin showed meaningful protection against kidney damage. But protection by SMT was weaker. TNF-α and IL-1β levels increased significantly with MEC application, and MEL and SMT ameliorated these increases in kidney tissue. MEC also elevated NOx levels in kidney tissue. CONCLUSIONS Both inflammation and oxidative stress may have an important role in the MEC induced nephrotoxicity. MEL and SMT may also have anti-inflammatory properties, as well as anti-oxidant properties.

Comparative Sporicidal Effects of Disinfectants after Release of a Biological Agent

Because of spore formation, Bacillus anthracis is considered the most resistant biological warfare agent known. The present study aimed to assess and compare well-known decontamination routes to inactivate the spores on daily-use environmental tools contaminated previously. To simulate the agent, Bacillus atrophaeus was used. Various environmental samples (such as tile, fabric clothing, wood, protective suit, glass, paper, soil, water, plastic, and metal) that may be contaminated after a biological incident were used as test carriers and inoculated with B. atrophaeus. Sodium hypochlorite, free chlorine, autoclaving, ethylene oxide, hydrogen peroxide, ultraviolet irradiation, and boiling decontaminated the samples. Glutaraldehyde (2%) and free chlorine solution (10,000 mg/L) were also found to be effective in decontaminating the samples and are recommended as alternatives to the use of sodium hypochlorite solution. Soil, tile, paper, and metal were determined to be the most difficult materials to decontaminate. It was concluded that 5% hypochlorite adjusted with acetic acid might also be used for decontamination. Decontamination strategies to reduce contamination of the environment by biological warfare agents need to be applied to mitigate the number of victims, in terms of prominent characteristics like cost-effectiveness and user-friendliness.

The protective effect of melatonin and S-methylisothiourea treatments in nitrogen mustard induced lung toxicity in rats

OBJECTIVES Mustard is highly toxic to the lung. Its toxic effects are associated with inflammatory cell accumulation and increased pro-inflammatory cytokines as well as reactive oxygen and nitrogen species. In this study, we aimed to investigate the efficiency of melatonin (MEL) and S-methylisothiourea (SMT) on mechlorethamine (MEC) induced lung toxicity. METHODS Thirty-six male rats were randomly divided into four groups: control, MEC, MEC+MEL, and MEC+SMT. Control group was given saline only via transdermal route. Other groups were exposured to a single dose of MEC (3.5 mg/kg) via transdermal route. MEL (100 mg/kg) was administered intraperitoneally 30 min after the application of MEC, and after the same dose of MEL was given every 12 h for a total of six doses. SMT (50 mg/kg) was also given intraperitoneally 30 min after the application of MEC. RESULTS MEC injection resulted in alveolar epithelial injury, hemorrhage, inflammation, edema and interalveolar septal thickening in the lung tissues. The tissue TNF-α, IL-1β, and nitrate/nitrite (NOx) levels were found significantly different for all groups (p<0.001). TNF-α and IL-1β levels increased significantly with MEC exposure, and MEL and SMT ameliorated these increases in lung tissues. MEC also elevated NOx levels in lung tissue. Melatonin showed meaningful protection against lung injury. But protection of SMT was weaker. CONCLUSION Inflammation plays an important role in the MEC induced lung toxicity as well as oxidative and nitrosative stress. Melatonin has also anti-inflammatory properties similar to SMT, as well as anti-oxidant properties. But melatonin treatment was found more efficient than SMT treatment.

Could rice be used as an anticaking agent in table salt

The aim of the study was to examine whether powdered rice could be used as an anticaking agent in table salt. Salts free from anticaking food additives were selected and powdered rice was added at different concentrations (0%, 1%, 2%, 5%, 10% and 20%). The samples were placed away from direct light and airflow, and left exposed in the laboratory at room temperature and below 30% humidity. The weight of the samples was measured using an electronic laboratory balance and recorded daily for 4 days. At the end of the experiment, all the samples were dried at 105°C in order to determine exact dryness. All the salt samples containing powdered rice lost weight (0.56±0.08%), while the other salt samples free of powdered rice gained weight (10.31±0.63%) (P<0.001). A statistically significant difference was found between the first two (1% and 2%) and last three concentrations (5%, 10% and 20%) (P<0.001). Adding powdered rice to salt as an anticaking agent during salt manufacturing at a concentration of 1% could take the place of other anticaking food additives used in table salt production.