Predicting clinical outcome of sulfur mustard induced ocular injury using machine learning model.

The sight-threatening sulfur mustard (SM) induced ocular injury presents specific symptoms in each clinical stage. The acute injury develops in all exposed eyes and may heal or deteriorate into chronic late pathology. Early detection of eyes at risk of developing late pathology may assist in providing unique monitoring and specific treatments only to relevant cases. In this study, we evaluated a machine-learning (ML) model for predicting the development of SM-induced late pathology based on clinical data of the acute phase in the rabbit model. Clinical data from 166 rabbit eyes exposed to SM vapor was used retrospectively. The data included a comprehensive clinical evaluation of the cornea, eyelids and conjunctiva using a semi-quantitative clinical score. A random forest classifier ML model, was trained to predict the development of corneal neovascularization four weeks post-ocular exposure to SM vapor using clinical scores recorded three weeks earlier. The overall accuracy in predicting the clinical outcome of SM-induced ocular injury was 73%. The accuracy in identifying eyes at risk of developing corneal neovascularization and future healed eyes was 75% and 59%, respectively. The most important parameters for accurate prediction were conjunctival secretion and corneal opacity at 1w and corneal erosions at 72 h post-exposure. Predicting the clinical outcome of SM-induced ocular injury based on the acute injury parameters using ML is demonstrated for the first time. Although the prediction accuracy was limited, probably due to the small dataset, it pointed out towards various parameters during the acute injury that are important for predicting SM-induced late pathology and revealing possible pathological mechanisms.

The use of aflibercept (VEGF trap) in mitigating sulfur mustard-induced corneal neovascularization in a rabbit model.

Sulfur mustard (SM)-induced ocular injury is characterized by an acute inflammatory response that may become chronic or enter a latent phase with delayed pathology. This study aimed to evaluate the efficacy of ziv-aflibercept and aflibercept in preventing and ameliorating corneal neovascularization (NV), respectively, following chemical eye exposure to SM vapor in a rabbit model. Chemical SM ocular insult was induced in the right eye of rabbits. A single application of ziv-aflibercept was administered 2 h or 9 days post-exposure. A single subconjunctival aflibercept treatment in an ocular formulation was administered 4 weeks after SM vapor exposure and subsequent to an initial 1-week treatment with 0.1 % dexamethasone. Clinical monitoring was performed 5-12 weeks post-exposure, and digital corneal pictures were taken to assess the extent of NV. The rabbits were euthanized and the corneas were processed for histological assessment. Treatment with ziv-aflibercept 2 h and 9 days post-exposure moderately reduced insult severity and partially delayed or prevented corneal NV. Aflibercept application 4 weeks post-exposure significantly reduced the extent of NV for 8 weeks. The substantial decrease in existing corneal NV in this group was confirmed by histology. These results reveal the powerful anti-angiogenic efficacy of the VEGF-trap for ameliorating existing NV as opposed to preventing NV development, revealing the ability of this treatment to mitigate corneal NV.

Limbal stem cell deficiency (LSCD) in rats and mice following whole body exposure to sulfur mustard (SM) vapor.

Ocular injuries following sulfur mustard (SM) exposure are characterized by an acute phase expressed by corneal erosions and inflammation of the anterior segment that after a clinically silent period may be followed by irreversible corneal injuries. The latter includes epithelial defects, chronic inflammation and neovascularization (NV), and were defined in rabbits and in humans as Limbal Stem Cell Deficiency (LSCD), that derived from a delayed loss of corneal epithelial stem cells (ESC), due to secondary processes most likely in the epithelial stem cell (SC) niche. The present study expands our research on SM-induced ocular injury to rodents (rats and mice) following whole body vapor exposure, aiming to define whether the delayed development of LSCD is a general characteristic of SM ocular toxicity. Freely moving rats and mice were exposed to SM vapor (155 µg/l, 10 min). Clinical examination was carried out in rats and included a slit-lamp bio-microscopy, up to 6 months. Eyes were taken for histology at different time points following exposure and evaluation included hematoxylin and eosin (H&E) staining for general morphology, PAS for identification of goblet cells and p63 immunohistochemistry for progenitor epithelial cells. Whole body exposure to SM vapor in rats and mice resulted in acute ocular injury characterized by corneal erosions and ocular inflammation. Following a brief recovery period, 80-90% of the exposed eyes developed corneal NV associated with abnormal corneal epithelium, stromal inflammation and endothelial damage. The late injury was accompanied by migration of conjunctival goblet cells to the cornea and a loss of limbal epithelial progenitor cells, indicating LSCD. The long-term ocular injury shown hereby in rats and mice was consistent with the lesions described in rabbits and in human casualties and demonstrated the general phenomenon of limbal epithelial stem cells deficiency in SM ocular toxicity. The delayed manifestation of this pathology points towards a therapeutic window for the development of medical countermeasures in small animals following exposure in a real life scenario.

Mice with induced pulmonary morbidities display severe lung inflammation and mortality following exposure to SARS-CoV-2.

Mice are normally unaffected by SARS coronavirus 2 (SARS-CoV-2) infection since the virus does not bind effectively to the murine version of the angiotensin-converting enzyme 2 (ACE2) receptor molecule. Here, we report that induced mild pulmonary morbidities rendered SARS-CoV-2-refractive CD-1 mice susceptible to this virus. Specifically, SARS-CoV-2 infection after application of low doses of the acute lung injury stimulants bleomycin or ricin caused severe disease in CD-1 mice, manifested by sustained body weight loss and mortality rates greater than 50%. Further studies revealed markedly higher levels of viral RNA in the lungs, heart, and serum of low-dose ricin-pretreated mice compared with non-pretreated mice. Furthermore, lung extracts prepared 2-3 days after viral infection contained subgenomic mRNA and virus particles capable of replication only when derived from the pretreated mice. The deleterious effects of SARS-CoV-2 infection were effectively alleviated by passive transfer of polyclonal or monoclonal antibodies generated against the SARS-CoV-2 receptor binding domain (RBD). Thus, viral cell entry in the sensitized mice seems to depend on viral RBD binding, albeit by a mechanism other than the canonical ACE2-mediated uptake route. This unique mode of viral entry, observed over a mildly injured tissue background, may contribute to the exacerbation of coronavirus disease 2019 (COVID-19) pathologies in patients with preexisting morbidities.

Therapeutic measures for sulfur mustard-induced ocular injury.

The use of sulfur mustard (SM) in global terrorism is still a relevant threat to both civilian population and military personnel. Casualties exposed to SM may present mild, moderate or severe acute ocular lesions followed by a complete ocular resolution, chronic lesions or re-emerged ocular pathologies after a latent period. Current treatment for SM-induced ocular injury is based mainly on the clinical manifestation at the different stages of the injury and includes pharmaceutical and surgical interventions. These therapeutic measures are beneficial but not sufficient, and the ocular injury remains a continuous challenge for medical professionals. This review focuses on treatment experience carried out in humans and studied in animal models, for both SM-induced ocular acute injury and late pathology. In general, therapeutic measures are based on clinical features of the ocular injury or on the involvement of specific factors during the ocular injury that point out towards potential treatments. Anti-inflammatory treatments and limbal stem cell transplantation techniques were developed based on the clinical manifestation of the ocular injury. Optional therapies for impaired corneal innervation and endothelium are suggested for future research. Additionally, studies on potential treatments with anti-matrix metalloproteinase (MMP), anti-vascular endothelial growth factor (VEGF) and anti-IL-6 agents are discussed. Consequently, future studies may reveal the potential of additional pharmacological and biological treatments or advanced cellular and molecular biology methods to serve as novel therapeutic measures and techniques for this complicated ocular injury.

Skin decontamination efficacy of sulfur mustard and VX in the pig model: A comparison between Fuller's earth and RSDL.

Skin decontamination following exposure to chemical agents is a most important component of the individual defense doctrine, removing the agent, ceasing its penetration and preventing secondary contamination of the first responders. The goal of the current study was to compare the efficacy of Reactive Skin Decontaminant Lotion (RSDL) and Fuller's Earth (FE) following exposure to sulfur mustard (SM) and VX, aiming to find the optimal procedure for mass casualty decontamination protocol. Decontamination efficacy was evaluated in pigs by measurement of lesion area and erythema (SM) and cholinesterase inhibition and clinical symptoms (VX). FE and RSDL were highly effective against both agents. Following SM exposure, the two decontaminants demonstrated a significant decrease in lesions' size together with the decrease in exposure duration. Likewise, skin decontamination following exposure to VX with either FE or RSDL resulted in reduction in clinical symptoms and prevention of death. Decontamination was worthwhile even if postponed, up to 30 min (SM) and 2 h (VX). In conclusion, both decontamination products were efficient in ameliorating the toxic effects even though in a different mechanism. Finally, for mass casualty scenario, FE is preferred as a universal decontaminant, considering its safety, ease of use and longer shelf life.

Ocular surface histopathological insult following sarin and VX exposure and potential treatments in the rat model.

Eye exposure to organophosphate (OP) chemical warfare irreversible acetylcholinesterase inhibitors, results in long-term miosis and impaired visual function. In contrast to the well-documented miotic and ciliary muscle spasm observed following chemical warfare, OP ocular exposure, little is known regarding the ocular surface histopathological insult. The aim of the present study was to determine the degree of the ocular surface insult following sarin or VX ocular exposure and to evaluate potential anti-cholinergic treatments in counteracting this insult. Rats that were whole body exposed to various sarin concentrations (0.049-43 µg/L; 5 min exposure), showed a dose-dependent miotic response and light reflex impairment. Following whole body sarin exposure, a dose dependent ocular surface histopathological insult was developed. A week following exposure to a low concentration of 0.05 µg/L, conjunctival pathology was observed, while corneal insult was noticed only following exposure to a concentration of 0.5 µg/L and above. Both tissues presented poorer outcomes when exposed to higher sarin concentrations. In contrast, eyes topically exposed to 1 µg sarin demonstrated no ocular insult a week following exposure. On the contrary, topical exposure to 1 µg VX resulted in a significant corneal insult. Anticholinergic treatments such as 0.1% atropine or 2% homatropine, given shortly following VX exposure, counteracted this insult. The results of this study show that not only do anti-cholinergic treatments counteract the miotic response, but also prevent the histopathological insult observed when given shortly following OP exposure.

A comprehensive analysis of corneal mRNA levels during sulfur mustard induced ocular late pathology in the rabbit model using RNA sequencing.

Exposure to sulfur mustard (SM) may result in severe ocular injuries. While some of the eyes show a clinical resolution of the injury (defined as clinically non-impaired), part of the eyes develop irreversible late ocular pathologies (defined as clinically impaired) that may lead to corneal blindness. Understanding the pathological mechanisms underlying the development of the late pathology may lead to improved treatment options. Therefore, this study aimed to investigate the mRNA expression profiles of corneas from clinically impaired, clinically non-impaired and naïve eyes. Rabbit eyes were exposed to SM vapor and a clinical follow-up was carried out up to 4 weeks using a slit lamp microscope. At this time point, corneal tissues from clinically impaired, clinically non-impaired and naïve eyes were processed for RNA sequencing (RNA-seq) and differential expression analyses. The differential expression profiles were further subjected to pathway enrichment analysis using Ingenuity Pathway Analysis (IPA). Real-time PCR was used for RNA-seq validation. The late pathology developed in 54%-80% of the eyes following ocular exposure to SM, clinically manifested by inflammation, corneal opacity and neovascularization. RNA-seq results showed significant differences in mRNA levels of hundreds of genes between clinically impaired, clinically non-impaired and naïve corneas. Pathway enrichment analysis showed common pathways that were activated in all of the exposed eyes, such as Th1 and Th2 activation pathway, in addition to pathways that were activated only in the clinically impaired eyes compared to the clinically non-impaired eyes, such as IL-6 and ERK5 signaling. Corneal mRNA expression profiles for the clinically impaired, clinically non-impaired and naïve eyes generated a comprehensive database that revealed new factors and pathways, which for the first time were shown to be involved in SM-induced late pathology. Our data may contribute to the research on both the pathological mechanisms that are involved in the development of the late pathology and the protective pathways that are activated in the clinically non-impaired eyes and may point out towards novel therapeutic strategies for this severe ocular injury.

Whole body exposure of rats to sulfur mustard vapor.

Sulfur mustard (SM) is an incapacitating chemical warfare agent used in numerous conflicts around the world and it is still a major threat for both, army troops and civilians. To evaluate its multiple targets effects in experimental setup, a model of whole body exposure (WBE) to SM vapor was established in rats and its simultaneous effects on lungs and eyes as well as on general wellbeing were examined. Rats were exposed to SM vapor. Evaluation (up to 10 weeks post-exposure) included body weight, general observation, blood counts and histological analysis. Results showed that following a latency-period of several hours, rats typical symptoms developed over a period of more than one week. The initial symptoms, characterized by swollen and erythematic nose, deteriorated into extensive rhinorrhea, eye closure, excessive lacrimation as well as rhonchi, wheezing and breathing difficulties. Alopecia and behavioral abnormality were also recorded. A weight loss of up to 40% was measured within one week with spontaneous recovery to baseline level within three weeks after exposure. Blood counts revealed leukopenia during the first three days post-exposure. Histological evaluation revealed a long lasting damage to the trachea, lungs and eyes. Thus, WBE to SM, was found to closely mimic the deleterious effects of SM on the sensitive tissues previously described in human victims during WWI and the Iran-Iraq war. The use of this animal model will enable comprehensive characterization of changes in biological processes that may lead to the development of therapeutic measures to ameliorate SM induced multi-system injuries.

Differential expression of corneal and limbal cytokines and chemokines throughout the clinical course of sulfur mustard induced ocular injury in the rabbit model.

PURPOSE: The sight threatening sulfur mustard (SM) induced ocular injury presents specific symptoms for each clinical stage. The acute injury develops in all of the exposed eyes and is characterized by erosions and severe inflammation. The irreversible late pathology develops only in part of the eyes, and is clinically expressed by chronic inflammation and corneal neovascularization (NV). The mechanisms underlying this injury are still in research and treatment is insufficient. Aiming to shed light on pathological mechanisms and improve the therapeutic measures, we studied the expression pattern of various cytokines and chemokines at different clinical stages of the ocular injury. METHODS: Rabbit right eye was exposed to SM vapor and a clinical follow-up was carried out up to 4 weeks. Corneal and limbal tissues were collected at 48 h, 1w and 4w post exposure and IL-1α, IL-1ß, IL-6, TNFα, macrophage chemotactic protein (MCP)-1 and IL-8 levels were measured by commercial ELISA kits. RESULTS: SM exposed eyes presented an acute injury that was partially resolved within a week in all of the exposed eyes, and was followed by an irreversible late pathology in 50%-80% of the eyes, beginning at 2w. A significant elevation was seen in levels of the studied factors, however each factor presented a unique expression pattern. At the peak of the acute injury, at 48 h, significantly higher levels of corneal IL-1α, IL-8, and TNFα and limbal IL-1α and MCP-1 were found compared to naïve eyes. At 1w, corneal IL-1ß, IL-6, IL-8 and TNFα and limbal IL-8 and MCP-1 levels were significantly higher compared to naïve eyes. During the late pathology, at 4w, elevated levels of corneal IL-1ß, IL-6 and MCP-1 and limbal MCP-1 and IL-8 were found only in eyes presenting NV. CONCLUSIONS: The levels of the studied factors changed throughout the dynamic course of the ocular injury. The prolonged increased levels of limbal MCP-1 and IL-8 may contribute to the continuous recruitment of inflammatory cells, characterizing the symptoms of the late pathology. The significantly elevated IL-1ß and IL-6 at 1w, after the resolution of the acute injury but before the clinical manifestation of the late pathology suggests a therapeutic window for intervention with prevention therapy. Mapping the expression pattern of these cytokines and chemokines points out towards stage-specific therapeutic options.

Successful single treatment with ziv-aflibercept for existing corneal neovascularization following ocular chemical insult in the rabbit model.

PURPOSE: To evaluate the efficacy of ziv-aflibercept as a treatment for established corneal neovascularization (NV) and to compare its efficacy to that of bevacizumab following ocular chemical insult of sulfur mustard (SM) in the rabbit model. METHODS: Chemical SM burn was induced in the right eye of NZW rabbits by vapor exposure. Ziv-aflibercept (2 mg) was applied once to neovascularized eyes by subconjunctival injection while subconjunctival bevacizumab (5 mg) was administered twice a week, for 3 weeks. Non-treated exposed eyes served as a control. A clinical follow-up employed by slit-lamp microscope, was performed up to 12 weeks following exposure and digital photographs of the cornea were taken for measurement of blood vessels length using the image analysis software. Eyes were taken for histological evaluation 2, 4 and 8 weeks following treatment for general morphology and for visualization of NV, using H&E and Masson Trichrome stainings, while conjunctival goblet cell density was determined by PAS staining. RESULTS: Corneal NV developed, starting as early as two weeks after exposure. A single subconjunctival treatment of ziv-aflibercept at 4 weeks post exposure, significantly reduced the extent of existing NV already one week following injection, an effect which lasted for at least 8 weeks following treatment, while NV in the non-treated exposed eyes continued to advance. The extensive reduction in corneal NV in the ziv-aflibercept treated group was confirmed by histological evaluation. Bevacizumab multiple treatment showed a benefit in NV reduction, but to a lesser extent compared to the ziv-aflibercept treatment. Finally, ziv-aflibercept increased the density of conjunctival goblet cells as compared to the exposed non-treated group. CONCLUSIONS: Subconjunctival ziv-aflibercept single treatment presented a highly efficient long-term therapeutic benefit in reducing existing corneal NV, following ocular sulfur mustard exposure. These findings show the robust anti-angiogenic efficacy of ziv-aflibercept and demonstrate the advantage of this treatment over the other anti-angiogenic therapies in ameliorating corneal NV and protecting the ocular surface.

Ocular Surface Changes After Sulfur Mustard Exposure in Rabbits, Monitored by Impression Cytology.

PURPOSE: Ocular injuries after exposure to sulfur mustard (SM) are characterized by acute corneal erosion and inflammation of the anterior segment that may be followed by delayed corneal neovascularization and epithelial defects, associated with limbal stem cell deficiency in part of the exposed eyes. This study aimed to further clarify the mechanism of the late injury by monitoring SM-induced cytological alterations in the ocular surface, in relation to the clinical symptoms, using impression cytology (IC). METHODS: Rabbit eyes were exposed to SM vapor (n = 20) and were clinically observed up to 4 weeks. Samples for IC were collected simultaneously from the upper bulbar conjunctiva, limbus, and cornea and then fixed and stained with periodic acid-Schiff and hematoxylin. At 1 month, animals were killed and eyes dissected and processed for histology. RESULTS: Concomitant with clinical symptoms of SM ocular toxicity, IC showed significant long-term loss of conjunctival goblet cells shortly after exposure, followed by abnormal differentiation toward squamous metaplasia. Simultaneously with corneal erosion, apoptotic bodies and cellular debris were seen in the corneal epithelium, followed by regeneration at 1 week. Migration of conjunctival goblet cells toward the cornea was noted in neovascularized eyes, as early as 1 week, indicating limbal stem cell deficiency. The IC findings were supported by histological evaluation. CONCLUSIONS: Continuous monitoring of the ocular surface after SM exposure by IC enables earlier detection of pathology and therapeutic intervention, therefore, is recommended for routine follow-up of casualties. Prolonged loss of goblet cells may point toward the role of mucin in the pathogenesis.

Dermostyx (IB1) - High efficacy and safe topical skin protectant against percutaneous toxic agents.

Prevention of the penetration of toxic agents through the skin is crucial for both military troops and civilian populations. We have developed a novel topical skin protectant (TSP), coded as IB1 and commercially available as Dermostyx protective solution (Rekah Pharm, Israel). The formulation afforded significant protection against chemical warfare agents such as sulfur mustard (SM) and VX (2LD50), pesticides such as parathion and irritants such as acrolein. The efficacy of the protectant was evaluated in the pig model using clinical, histological and biochemical monitoring. A single topical application prior to exposure to the toxic agents reduced significantly the size and severity of skin lesions and ameliorated or prevented systemic clinical symptoms. The barrier properties of IB1 are immediate upon application and remain effective for at least 12 h. It is absorbed into the stratum corneum of the skin and remains there until rinsing with water, yet the ingredients are not absorbed into the body. The formulation is a hydrophilic water-based solution, composed of magnesium sulfate and glycerin that are widely used in cosmetic and medicine, and was shown to be safe in preclinical and in Phase I clinical studies. The suggested mode of action is based on the unique interaction of glycerin with the stratum corneum, changing its properties to hydrophilic and on the "salting out" effect of magnesium sulfate. The expected use of the TSP is by application on exposed skin areas and sensitive skin sites (e.g. armpits, groin, waist), when necessary. A quantity of 10 ml is sufficient for one application covering approximately 20% of the body surface area. The formulation was approved for human use by the Israel Ministry of Health and a CE mark certificate in Europe has been recently issued (Class I). Dermostyx has been adopted by the IDF and first responders as a skin protectant for special needs.

Using the skin protective lotion IB1 as a substitute for chemical protective gloves.

We aimed to evaluate the performance of medical personnel in using the IB1 topical protective lotion on their hands and wrists together with standard disposable medical gloves, compared to standard-issued medical chemical protective gloves. This randomized cross-over study included 144 medical personnel. Primary endpoints were time-to-completion of autoinjection; success rate, number of attempts, and time-to-achieve successful endotracheal intubation; time-to-achieve satisfactory tube fixation; time-to-draw and inject the content of an ampoule; and the total time-to-perform all medical procedures. Secondary endpoints included the subjective assessment of convenience to perform these four procedures with each protective measure. Mean time was significantly shorter using IB1 compared to chemical protective gloves for tube fixation, ampoule drawing, and the total time-to-perform all procedures (58.6±22.7 seconds vs. 71.7±29.7; 31.5±21.8 vs. 38.2±19.4; 137.4±56.1 vs. 162.5±63.6, respectively; P<.001 for all). For all medical procedures, the use of IB1 was reported as significantly more convenient than the use of chemical protective gloves (P<.001 for all comparisons). IB1 with standard medical gloves significantly shorten the time-to-perform medical procedures requiring fine motor dexterities and is subjectively more convenient than chemical protective gloves. IB1 should be considered as an appropriate alternative for medical teams in a chemical event.

Sarin-induced brain damage in rats is attenuated by delayed administration of midazolam.

Sarin poisoned rats display a hyper-cholinergic activity including hypersalivation, tremors, seizures and death. Here we studied the time and dose effects of midazolam treatment following nerve agent exposure. Rats were exposed to sarin (1.2 LD50, 108 µg/kg, im), and treated 1 min later with TMB4 and atropine (TA 7.5 and 5 mg/kg, im, respectively). Midazolam was injected either at 1 min (1 mg/kg, im), or 1 h later (1 or 5 mg/kg i.m.). Cortical seizures were monitored by electrocorticogram (ECoG). At 5 weeks, rats were assessed in a water maze task, and then their brains were extracted for biochemical analysis and histological evaluation. Results revealed a time and dose dependent effects of midazolam treatment. Rats treated with TA only displayed acute signs of sarin intoxication, 29% died within 24h and the ECoG showed seizures for several hours. Animals that received midazolam within 1 min survived with only minor clinical signs but with no biochemical, behavioral, or histological sequel. Animals that lived to receive midazolam at 1h (87%) survived and the effects of the delayed administration were dose dependent. Midazolam 5 mg/kg significantly counteracted the acute signs of intoxication and the impaired behavioral performance, attenuated some of the inflammatory response with no effect on morphological damage. Midazolam 1mg/kg showed only a slight tendency to modulate the cognitive function. In addition, the delayed administration of both midazolam doses significantly attenuated ECoG compared to TA treatment only. These results suggest that following prolonged seizure, high dose midazolam is beneficial in counteracting adverse effects of sarin poisoning.

Early brain magnetic resonance imaging can predict short and long-term outcomes after organophosphate poisoning in a rat model.

INTRODUCTION: Magnetic resonance (MR) imaging is a sensitive modality for demonstrating in vivo alterations in brain structure and function after acute organophosphate (OP) poisoning. The goals of this study were to explore early imaging findings in organophosphate-poisoned animals, to assess the efficacy of centrally acting antidotes and to find whether early MR findings can predict post-poisoning cognitive dysfunction. METHODS: Sprague-Dawley rats were poisoned with the agricultural OP paraoxon and were treated with immediate atropine and obidoxime (ATOX) to reduce acute mortality caused by peripheral inhibition of acetylcholinesterase. Animals were randomly divided into three groups based on the protocol of centrally acting antidotal treatment: group 1 - no central antidotal treatment (n=10); group 2 - treated with midazolam (MID) at 30 min after poisoning (n=9), group 3 - treated with a combination of MID and scopolamine (SCOP) at 30 min after poisoning (n=9) and controls (n=6). Each animal had a brain MR examination 3 and 24 h after poisoning. Each MR examination included the acquisition of a T2 map and a single-voxel (1)H MR spectroscopy (localized on the thalami, to measure total creatine [Cr], N-acetyl-aspartate [NAA] and cholines [Cho] levels). Eleven days after poisoning each animal underwent a Morris water maze to assess hippocampal learning. Eighteen days after poisoning, animals were euthanized, and their brains were dissected, fixed and processed for histology. RESULTS: All paraoxon poisoned animals developed generalized convulsions, starting within a few minutes following paraoxon injection. Brain edema was maximal on MR imaging 3 h after poisoning. Both MID and MID+SCOP prevented most of the cortical edema, with equivalent efficacy. Brain metabolic dysfunction, manifested as decreased NAA/Cr, appeared in all poisoned animals as early as 3h after exposure (1.1 ± 0.07 and 1.42 ± 0.05 in ATOX and control groups, respectively) and remained lower compared to non-poisoned animals even 24h after poisoning. MID and MID+SCOP prevented much of the 3h NAA/Cr decrease (1.22 ± 0.05 and 1.32 ± 0.1, respectively). Significant correlations were found between imaging findings (brain edema and spectroscopic changes) and clinical outcomes (poor learning, weight loss and pathological score) with correlation coefficients of 0.4-0.75 (p<0.05). CONCLUSIONS: MR imaging is a sensitive modality to explore organophosphate-induced brain damage. Delayed treatment with midazolam with or without scopolamine provides only transient neuroprotection with some advantage in adding scopolamine. Early imaging findings were found to correlate with clinical consequences of organophosphate poisoning and could be potentially used in the future to predict long-term prognosis of poisoned casualties.

VEGF Mediates ApoE4-Induced Neovascularization and Synaptic Pathology in the Choroid and Retina.

Apolipoprotein E4 (apoE4), the most prevalent genetic risk factor for Alzheimer's disease (AD), is associated with neuronal and vascular impairments. Recent findings suggest that retina of apoE4 mice have synaptic and functional impairments. We presently investigated the effects of apoE4 on retinal and choroidal vasculature and the possible role of VEGF in these effects. There were no histological differences between the retinal and choroidal vasculatures of naïve apoE3 and apoE4 mice. In contrast, laserdriven choroidal injury induced higher levels of choroidal neovascularization (CNV) in apoE4 than in apoE3 mice. These effects were associated with an inflammatory response and with activation of the Muller cells and asrocytic markers gluthatione synthetase and GFAP, all of which were more pronounced in the apoE4 mice. CNV also induced a transient increase in the levels of the synaptic markers synaptophysin and PSD95 which were however similar in the apoE4 and apoE3 naive mice. Retinal and choroidal VEGF and apoE levels were lower in naïve apoE4 than in corresponding apoE3 mice. In contrast, VEGF and apoE levels rose more pronouncedly following laser injury in the apoE4 than in apoE3 mice. Taken together, these findings suggest that the apoE4-induced retinal impairments, under basal conditions, may be related to reduced VEGF levels in the eyes of these mice. The hyper-neovascularization in the apoE4 mice might be driven by increased inflammation and the associated surge in VEGF following injury. Retinal and choroidal VEGF and apoE levels were lower in naïve apoE4 than in corresponding apoE3 mice. In contrast, VEGF and apoE levels rose more pronouncedly following laser injury in the apoE4 than in apoE3 mice. Taken together, these findings suggest that the apoE4-induced retinal impairments, under basal conditions, may be related to reduced VEGF levels in the eyes of these mice. The hyper-neovascularization in the apoE4 mice might be driven by increased inflammation and the associated surge in VEGF following injury.

Acute and long-term ocular effects of acrolein vapor on the eyes and potential therapies.

OBJECTIVE: Acrolein is a potent irritant and a vesicant that was used by the French during WWI as the warfare agent named: "papite". Nowadays, it is produced in large amounts all over the world in the industry for the production of acrylic acid and for agriculture use as herbicide. The aim of this study was to characterize the effects of acute eye exposure to acrolein vapor and to evaluate the efficacy of a topical post-exposure combination treatment with a local anesthetic and a steroid. METHODS: Rabbit eyes were exposed to three doses of acrolein vapor (low, intermediate and high) and treated topically with either 0.4% benoxinate hydrochloride (localin, for 2 h) or dexamethasone (dexamycin, for 6 days) or a combination of both drugs. Clinical follow-up using slit lamp examinations and histological evaluation was performed 4 weeks post-exposure. RESULTS: Acrolein vapor caused immediate eye closure with excess tearing, corneal erosions and severe inflammation of the anterior chamber. This was followed by corneal neovascularization (NV) starting as early as 1 week post-exposure. The damage to the eyes was long lasting, and at 4 weeks following exposure, significant pathological changes were observed. Immediate post-exposure application of the local anesthetic, localin, prevented the eye closure, and the dexamycin treatment reduced significantly the initial inflammation as well as the extent and incidence of corneal NV. CONCLUSIONS: Short-term eye exposure to the irritant acrolein may result in immediate eye closure and long lasting pathologies that could lead to blindness. An anti-inflammatory treatment combined with short-term application of a local anesthetic prevents incapacitation and might minimize significantly the extent of eye injuries.

Beneficial effects of activated macrophages on sulfur mustard-induced cutaneous burns, an in vivo experience.

OBJECTIVE: Macrophages are known to have key functions in almost every stage of wound healing and there is evidence for their beneficial effects in treating decubital ulcers and deep sternal wound infections in human. This study aimed to investigate the efficacy of a treatment with activated macrophages on ameliorating acute and long-term sulfur mustard (SM) induced skin injuries in the hairless guinea pig (HGP) model. METHODS: HGP were exposed to SM vapor and treated with either a single or multiple intra-dermal injections of human activated macrophages in suspension (hAMS) into the wound bed. Clinical and histological evaluations were conducted up to 4 weeks post-exposure. RESULTS: A single treatment with hAMS early after exposure (15 min and 6 h) resulted in a reduction in the number of damaged cells and vesications in the epidermis at 24 h. A substantial increase in cellular infiltration, mostly polymorphonuclears, was taking place in the hAMS-treated animals starting as early as 1 h after exposure. This flow of inflammatory cells continued, in the treated group, for at least 4 weeks, long after the injected macrophages were not detected. Repeated injections of hAMS (15 min, 48 h and 7 d post-exposure) decreased significantly the area of the wounds and improved the integrity of the barrier function as expressed by measuring trans-epidermal water loss up to 10 d. CONCLUSIONS: Our results indicate that the role of macrophages in wound healing is complex; their efficacy may depend on the timing of administration. Further investigation is required to determine whether they are required during the early phase of wound development and/or during the late phase of scar formation and remodeling.