Acute cardiovascular effects of inhaled ambient particulate matter: Chemical composition-related oxidative stress, endothelin-1, blood pressure, and ST-segment changes in Wistar rats.

Short-term increases in particulate matter (PM) are associated with heightened morbidity and mortality from cardiovascular causes. Inhalation of PM is known to increase endothelin (ET)-1 levels. Yet, less is known about particle composition-related changes at the molecular level including the endothelinergic system and relationship with cardiovascular function changes. In this work, adult Wistar male rats were exposed for 4 h by nose-only inhalation to clean air, Ottawa urban particles (EHC-93, 48 mg/m3) and water-leached (EHC-93L, 49 mg/m3) particles, to examine the effect of particle compositional changes on oxidative stress, circulating ETs, blood pressure, and heart electrophysiology. Particle deposition in the respiratory compartment was estimated at 85 µg (25 ng/cm2). Lung cell proliferation was low in both treatment groups, indicating absence of acute injury. Inhalation of EHC-93 caused statistically significant elevations (p < 0.05) of oxidative stress markers, ET-1, ET-3, blood pressure, and a decrease of ST-segment duration in the ECG at 1.5 days post-exposure. Leached particles (EHC-93L) caused rapid but transient elevation (p < 0.05) of oxidative stress, ET-1, ET-2, and ET-3 at earlier time points, with no changes in blood pressure or ST-segment. These results demonstrate that inhalation of urban particles at an internal dose inadequate to cause acute lung injury can induce oxidative stress, enhance vasoactive endothelins, leading to vasopressor response, affecting cardiac electrophysiology in Wistar rats, consistent with the cardiovascular impacts of ambient particles in human populations. Change in particle potency after removal of soluble species, notably cadmium, zinc and polar organics suggests that the toxicodynamics of cardiovascular effects can be modified by physicochemical properties of particles.

The Value of Live Tissue Training for Combat Casualty Care: A Survey of Canadian Combat Medics With Battlefield Experience in Afghanistan.

INTRODUCTION: The optimum method for training military personnel for combat casualty care is unknown. In particular, there is debate regarding the incremental benefit of live animal tissue training (LTT) over inanimate human patient simulators (HPSs). Although both LTT and HPS are currently used for predeployment training, the efficacy of these models has not been established. MATERIALS AND METHODS: Canadian Armed Forces combat medics, deployed to Afghanistan between 2006 and 2011, were surveyed retrospectively regarding their experience with combat casualty care and predeployment training. HPSs were used to prepare these combat medics for early rotations. In later years, personnel received a combination of training modalities including HPS and LTT, using anaesthetized porcine models in accordance with appropriate animal care standards. Among those deployed on multiple rotations, there was a cohort who was prepared for deployment using only HPS training, and who later were prepared using mixed-modality training, which included LTT. We asked these medics to compare their predeployment training using HPS only versus their mixed-modality training in how each training package prepared them for battlefield trauma care. RESULTS: Thirty-eight individuals responded, with 20 respondents deployed on multiple rotations. Respondents performed life-saving skills during 89% of the rotations. Self-perceived competence and preparedness were notably higher after incorporation of LTT than after HPS alone. Of 17 respondents deployed on both early and late rotations, the majority felt the latter training was more worthwhile. In addition, almost all individuals felt that LTT should be added to HPS training. Narrative comments described multiple benefits of adding LTT to other types of training. CONCLUSIONS: Among many experienced Canadian Armed Forces personnel, LTT is considered essential predeployment preparation. Individuals who experienced only HPS training before active duty on their first combat deployment reported feeling more competent on subsequent combat deployments after the addition of live tissue models. IMPACT: There has been a movement away from the use of LTT in preparing combat medics for deployment. This article suggests that we should reconsider any decision to completely exclude Live Tissue Training as part of our training plan for combat medics. RECOMMENDATIONS: Military medical organizations should consider judiciously incorporating LTT with human patient simulation training to prepare combat medics for treating battlefield trauma.

Nitrative stress, oxidative stress and plasma endothelin levels after inhalation of particulate matter and ozone.

BACKGROUND: While exposure to ambient air contaminants is clearly associated with adverse health outcomes, disentangling mechanisms of pollutant interactions remains a challenge. OBJECTIVES: We aimed at characterizing free radical pathways and the endothelinergic system in rats after inhalation of urban particulate matter, ozone, and a combination of particles plus ozone to gain insight into pollutant-specific toxicity mechanisms and any effect modification due to air pollutant mixtures. METHODS: Fischer 344 rats were exposed for 4 h to a 3 × 3 concentration matrix of ozone (0, 0.4, 0.8 ppm) and EHC-93 particles (0, 5, 50 mg/m(3)). Bronchoalveolar lavage fluid (BALF), BAL cells, blood and plasma were analysed for biomarkers of effects immediately and 24 h post-exposure. RESULTS: Inhalation of ozone increased (p < 0.05) lipid oxidation products in BAL cells immediately post-exposure, and increased (p < 0.05) total protein, neutrophils and mature macrophages in the BALF 24 h post-exposure. Ozone increased (p < 0.05) the formation of reactive oxygen species (ROS), assessed by m-, p-, o-tyrosines in BALF (Ozone main effects, p < 0.05), while formation of reactive nitrogen species (RNS), indicated by 3-nitrotyrosine, correlated with dose of urban particles (EHC-93 main effects or EHC-93 × Ozone interactions, p < 0.05). Carboxyhemoglobin levels in blood exhibited particle exposure-related increase (p < 0.05) 24 h post recovery. Plasma 3-nitrotyrosine and o-tyrosine were increased (p < 0.05) after inhalation of particles; the effect on 3-nitrotyrosine was abrogated after exposure to ozone plus particles (EHC-93 × Ozone, p < 0.05). Big endothelin-1 (BET-1) and ET-1 were increased in plasma after inhalation of particles or ozone alone, but the effects appeared to be attenuated by co-exposure to contaminants (EHC-93 × Ozone, p < 0.05). Plasma ET levels were positively correlated (p < 0.05) with BALF m- and o-tyrosine levels. CONCLUSIONS: Pollutant-specific changes can be amplified or abrogated following multi-pollutant exposures. Oxidative and nitrative stress in the lung compartment may contribute to secondary extra-pulmonary ROS/RNS formation. Nitrative stress and endothelinergic imbalance emerge as potential key pathways of air pollutant health effects, notably of ambient particulate matter.

Non-cholinergic intervention of sarin nerve agent poisoning.

The protective effects of selected anesthetic regimens on sarin (GB) were investigated in domestic swine. At 30% oxygen, the toxicity of this agent in isoflurane anesthetized animals (LD(50)=10.1µg/kg) was similar to literature sited values in awake swine (LD(50)=11.8µg/kg) and slightly higher than that of both ketamine (LD(50)=15.6µg/kg) and propofol (LD(50)=15.3µg/kg) anesthetized swine. Use of 100% oxygen in ketamine anesthetized animals resulted in three-fold protective effects compared to 30% oxygen. Use of 100% oxygen in both isoflurane and propofol anesthetized animals, compared to 30% resulted in profound protection against GB poisoning (>33×). There were no differences in the severity of the poisoning or recovery time in animals treated over dose ranges of 10-350µg/kg (isoflurane) or 15-500µg/kg GB (propofol). Survivors of high GB challenges that were revived from propofol anesthetic exhibited no signs of cognitive impairment seven days later. Protective treatments did not attenuate cholinesterase (ChE) inhibition; survivors of otherwise supralethal GB concentrations exhibited very low blood ChE activities. This work indicates that propofol has protective effects against GB, and that oxygen tension may have an important role in treating nerve agent casualties. More importantly, it demonstrates that non-cholinergic protective mechanisms exist that may be exploited in the future development of medical countermeasures against organophosphorous nerve agents.

Ionic dependence of sulphur mustard cytotoxicity.

The effect of ionic environment on sulphur mustard (bis 2-chloroethyl sulphide; HD) toxicity was examined in CHO-K1 cells. Cultures were treated with HD in different ionic environments at constant osmolar conditions (320 mOsM, pH 7.4). The cultures were refed with fresh culture medium 1h after HD exposure, and viability was assessed. Little toxicity was apparent when HD exposures were carried out in ion-free sucrose buffer compared to LC(50) values of approximately 100-150 microM when the cultures were treated with HD in culture medium. Addition of NaCl to the buffer increased HD toxicity in a salt concentration-dependent manner to values similar to those obtained in culture medium. HD toxicity was dependent on both cationic and anionic species with anionic environment playing a much larger role in determining toxicity. Substitution of NaI for NaCl in the treatment buffers increased HD toxicity by over 1000%. The activity of the sodium hydrogen exchanger (NHE) in recovering from cytosolic acidification in salt-free and in different chloride salts did not correlate with the HD-induced toxicity in these buffers. However, the inhibition by HD of intracellular pH regulation correlated with its toxicity in NaCl, NaI and sucrose buffers. Analytical chemical studies and the toxicity of the iodine mustard derivative ruled out the role of chemical reactions yielding differentially toxic species as being responsible for the differences in HD toxicity observed. This work demonstrates that the early events that HD sets into motion to cause toxicity are dependent on ionic environment, possibly due to intracellular pH deregulation.

pH-dependent toxicity of sulphur mustard in vitro.

The dependence of sulphur mustard (HD) toxicity on intracellular (pH(i)) and extracellular pH was examined in CHO-K1 cells. HD produced an immediate and significant concentration-dependent decline in cytosolic pH, and also inhibited the mechanisms responsible for restoring pH(i) to physiological values. The concentration-response of HD-induced cytosolic acidification, closely paralleled the acidification of the extracellular buffer through HD hydrolysis. A viability study was carried out in order to assess the importance of HD-induced cytosolic acidification. Cultures were exposed to HD for 1 h in media that were adjusted through a pH range (pH 5.0-10), and the 24 h LC(50) values were assessed using the viability indicator dye alamarBlue. The toxicity of HD was found to be dependent on extracellular pH, with a greater than eight-fold increase in LD(50) obtained in cultures treated with HD at pH 9.5, compared to those treated at pH 5.0. Assays of apoptotic cell death, including morphology, soluble DNA, caspase-3 activity and TUNEL also showed that as pH was increased, much greater HD concentrations were required to cause cell death. The modest decline in HD half-life measured in buffers of increasing pH, did not account for the protective effects of basic pH. The early event(s) that HD initiates to eventually culminate in cell death are not known. However, based on the data obtained in this study, we propose that HD causes an extracellular acidification through chemical hydrolysis and that this, in both a concentration and temporally related fashion, results in cytosolic acidification. Furthermore, HD also acts to poison the antiporter systems responsible for maintaining physiological pH(i), so that the cells are unable to recover from this insult. It is this irreversible decline in pH(i) that initiates the cascade of events that results in HD-induced cell death.

Alteration in aromatic hydroxylation and lipid oxidation status in the lungs of rats exposed to ozone.

Fischer 344 rats were exposed to ozone by inhalation to identify sensitive indices of acute exposure. 5-Aminosalicylic acid (5-ASA) hydroxylation in bronchoalveolar lavage (BAL), an indicator of hydroxyl radical (*OH) formation, and lipid oxidation in various regions of airways, representing oxidative stress, were measured to verify whether they can function as markers of exposure. BAL cells and supernatants taken from rats that received saline or 5-ASA (ip, 50 mg/kg) prior to ozone exposure (0, 0.4, or 0.8 ppm for 4 h) were analyzed for products of lipid oxidation. *OH formation was assessed by analysis of the BAL supernatant for 5-aminotetrahydroxybenzoic acid (5-ATHBA), a hydroxylation product of 5-ASA. The tetrahydroxy derivative of 5-ASA was higher in the BAL of ozone-treated rats than in air controls, reaching significance (p <. 05) at 0.8 ppm of ozone, The products of lipid oxidation propanal and hexanal were higher in BAL cells taken from rats exposed to ozone, reaching significance (p <. 05) at a 0.8 ppm ozone level, compared to air control animals, irrespective of whether they received saline or 5-ASA prior to ozone exposure. Increases in cholesterol levels were also seen in BAL cells after rats were exposed to ozone. However, there were no significant dose-related changes in the lipid oxidation products in BAL supernatants after exposure to ozone. Lipid oxidation products in BAL cells and 5-ATHBA in lavage exhibited the potential to serve as markers of ozone exposure. This work was supported by Health Canada (#4320105) and Toxic Substances Research Initiatives (TSRI #60).