The first 2(IB),3(IA)-heterodifunctionalized ß-cyclodextrin derivatives as artificial enzymes.

Novel 2,3-heterodisubstituted ß-cyclodextrin derivatives were designed as artificial enzymes to degrade chemical warfare agents. One of them reduced the acetylcholinesterase inhibitory potential by soman faster than its monosubstituted analog.

Effects of repeated low-dose exposure of the nerve agent VX on monoamine levels in different brain structures in mice.

In a previous report, alterations of the serotonin metabolism were previously reported in mice intoxicated with repeated low doses of soman. In order to better understand the effects induced by repeated low-dose exposure to organophosphorus compounds on physiological and behavioural functions, the levels of endogenous monoamines (serotonin and dopamine) in different brain areas in mice intoxicated with sublethal dose of (O-ethyl-S-[2(di-isopropylamino) ethyl] methyl phosphonothioate) (VX) were analysed by HPLC method with electrochemical detection. Animals were injected once a day for three consecutive days with 0.10 LD50 of VX (5 µg/kg, i.p). Neither severe signs of cholinergic toxicity nor pathological changes in brain tissue of exposed animals were observed. Cholinesterase (ChE) activity was only inhibited in plasma (a maximum of 30% inhibition 24 h after the last injection of VX), but remained unchanged in the brain. Serotonin and dopamine (DA) metabolism appeared significantly modified. During the entire period of investigation, at least one of the three parameters investigated (i.e. DA and DOPAC levels and DOPAC/DA ratio) was modified. During the toxic challenge, an increase of the serotonin metabolism was noted in hippocampus (HPC), hypothalamus/thalamus, pons medulla and cerebellum (CER). This increase was maintained 4 weeks after exposure in HPC, pons medulla and CER whereas a decrease in cortex 3 weeks after the toxic challenge was observed. The lack of correlation between brain ChE activity and neurochemical outcomes points out to independent mechanisms. The involvement in possibly long-lasting behavioural disorders is discussed.

Cutaneous challenge with chemical warfare agents in the SKH-1 hairless mouse. (I) Development of a model for screening studies in skin decontamination and protection.

Exposure to lethal chemical warfare agents (CWAs) is no longer only a military issue due to the terrorist threat. Among the CWAs of concern are the organophosphorus nerve agent O-ethyl-S-(2[di-isopropylamino]ethyl)methyl-phosphonothioate (VX) and the vesicant sulfur mustard (SM). Although efficient means of decontamination are available, most of them lose their efficacy when decontamination is delayed after exposure of the bare skin. Alternatively, CWA skin penetration can be prevented by topical skin protectants. Active research in skin protection and decontamination is thus paramount. In vivo screening of decontaminants or skin protectants is usually time consuming and may be expensive depending on the animal species used. We were thus looking for a suitable, scientifically sound and cost-effective model, which is easy to handle. The euthymic hairless mouse Crl: SKH-1 (hr/hr) BR is widely used in some skin studies and has previously been described to be suitable for some experiments involving SM or SM analogs. To evaluate the response of this species, we studied the consequences of exposing male anaesthetized SKH-1 mice to either liquid VX or to SM, the latter being used in liquid form or as saturated vapours. Long-term effects of SM burn were also evaluated. The model was then used in the companion paper (Taysse et al.(1)).

Cutaneous challenge with chemical warfare agents in the SKH-1 hairless mouse (II): effects of some currently used skin decontaminants (RSDL and Fuller's earth) against liquid sulphur mustard and VX exposure.

Using the hairless mouse screening model presented in the companion paper(1) the aim of this study was to assess two skin decontaminating systems: Fuller's earth (FE) and Reactive Skin Decontamination Lotion (RSDL) against two extremely toxic chemical warfare agents that represent a special percutaneous hazard, sulphur mustard (SM) and O-ethyl-S-(2[di-isopropylamino]ethyl)methyl-phosphonothioate (VX). Five minutes after being exposed on the back to either 2 µL of neat sulphur mustard or 50 µg.kg(-1) of diluted VX, mice were decontaminated. Both systems were able to reduce blisters 3 days after SM exposure. However, RSDL was found to be more efficient than FE in reducing the necrosis of the epidermis and erosion. In the case of VX exposure, RSDL, whatever the ratio of decontaminant to toxicant used (RSDL 10, 20, 50), was not able to sufficiently prevent the inhibition of plasma cholinesterases taken as a surrogate marker of exposure and toxicity. Only FE reduced significantly the ChE inhibition. Some of these observations are different from our previous results obtained in domestic swine and these changes are thus discussed in the perspective of using SKH-1 hairless mice for the initial in vivo screening of decontaminants.

Effects of repeated low-dose soman exposure on monoamine levels in different brain structures in mice.

In order to better understand the effects of repeated low-dose exposure to organophosphorus (OPs) on physiological and behavioural functions, we analysed the levels of endogenous monoamines (serotonin and dopamine) in different brain areas after repeated exposure of mice to sublethal dose of soman. Animals were injected once a day for 3 days with 0.12 LD50 of soman (47 microg/kg, i.p.). They did not show either severe signs of cholinergic toxicity or pathological changes in brain tissue. 24 h after the last injection of soman, inhibition of cholinesterase was similar in plasma and brain (32% and 37% of inhibition respectively). Afterwards, recovery of cholinesterase activity was faster in the plasma than in the brain. Dopamine levels were not significantly modified. On the other hand, we observed a significant modification of the serotoninergic system. An increase of the 5-HIAA/5-HT ratio was maintained for 2 and 4 weeks after exposure in the hippocampus and the striatum respectively. This study provides the first evidence of a modification of the 5-HT turnover in the hippocampus and the striatum after repeated low-dose intoxication with a nerve agent. Further experiments are necessary to evaluate the relationship between these modifications and the unexpected neuropsychological disorders usually reported after chronic exposure of organophosphorus.

Skin decontamination of mustards and organophosphates: comparative efficiency of RSDL and Fuller's earth in domestic swine.

Research in skin decontamination and therapy of chemical warfare agents has been a difficult problem due to the simultaneous requirement of rapid action and non-aggressive behaviour. The aim of this study was to compare the performance of two decontaminating systems: the Canadian Reactive Skin Decontaminant Lotion (RSDL) and the Fuller's Earth (FE). The experiment was conducted with domestic swine, as a good model for extrapolation to human skin. RSDL and FE were tested against sulphur mustard (SM), a powerful vesicant, and VX, a potent and persistent cholinesterase inhibitor. When used 5 min after contamination, the results clearly showed that both systems were active against SM (10.1 mg/cm(2)) and VX (0.06 mg/cm(2)). The potency of the RSDL/sponge was statistically better than FE against skin injury induced by SM, observed 3 days post-exposure. RSDL was rather more efficient than FE in reducing the formation of perinuclear vacuoles and inflammation processes in the epidermis and dermis. Against a severe inhibition (67%) of plasmatic cholinesterases induced by VX poisoning, the potencies of the RSDL/sponge and FE were similar. Both systems completely prevented cholinesterase inhibition, which indirectly indicates a prevention of toxic absorption through the skin.

Protection against soman-induced neuropathology and respiratory failure: a comparison of the efficacy of diazepam and avizafone in guinea pig.

The purpose of this study was to compare the efficacy of diazepam and the pro-diazepam avizafone in preventing the severity of soman-induced pathology in guinea pig. Survival, respiration and seizures of experimental animals were investigated with on-line monitoring of respiratory and EEG parameters. Guinea pigs were pretreated with pyridostigmine (0.1mg/kg i.m.) and 30 min later challenged with 1 or 2 LD50 soman. One minute after intoxication they were treated with atropine (3 or 33.8 mg/kg), pralidoxime chloride (32 mg/kg) and either diazepam (2 mg/kg), avizafone (3.5 mg/kg) or saline solution. The highest dose of atropine (33.8 mg/kg) gave a protective effect in groups treated without anticonvulsants by reducing the severity of clinical signs and death within 24 h but also by decreasing seizure occurrence and brain injuries. When injected at the similar molar dose of 7 micromoles/kg, the protection of anticonvulsants against soman neurotoxicity was higher with the atropine/pralidoxime/avizafone combination than with atropine/pralidoxime/diazepam. Indeed, when atropine was used at the lowest dose, avizafone was found to prevent early mortality and seizures occurrence with better efficacy than diazepam. On the other hand, when added to the therapy, the both anticonvulsants did not prevent the moderate EEG depression (reduction of amplitude by 30-52%) observed under 2 LD50 soman. Moreover, the number of animals suffering from respiratory distress (defined as a decrease of minute ventilation of more than 20% from the baseline value) was enhanced when diazepam or avizafone were used in the therapy. This effect was dependent on the atropine dose and the nature of the anticonvulsant. The beneficial effects of the different therapeutics tested were assessed and compared to the previous data obtained with the same therapies against sarin and from the pharmacokinetics properties of the atropine/diazepam mixture.

Induction of acute lung injury after intranasal administration of toxin botulinum a complex.

The inhalation of aerozolized botulinum toxin may represent a potential significant hazard to both military and civilian personnel. Since the lung is the primary target organ for inhaled toxin, the investigation reported herein was conducted to examine lung function in mice exposed to botulinum toxin A complex by intranasal route. Data includes lethality, symptomatology, measurement of respiratory function (minute ventilation, respiratory frequency, and tidal volume), and histopathology of the lungs. The clinical signs of intoxication are similar to those observed in foodborne botulism. Plethysmography revealed severe impairment of all respiratory parameters tested from 7 hours postexposure. Severe lung lesions, possibly secondary to the intoxication, were observed in mice who survived 14 days after the toxin challenge. These included intra-alveolar hemorrhage and interstitial edema. Mice immunized by the pentavalent (ABCDE) toxoid were protected against the neurotoxin (4 LD50) as revealed by the decrease of lethality and severity of nervous signs of intoxication, but not against histopathological changes in the lungs. These effects are nonspecific and require further experiments in order to specify the relationships between the pathology and the inflammatory process in the lung due to mediators such as cytokines,and possibly permanent physiological sequelae.