Dermal Exposure to Vesicating Nettle Agent Phosgene Oxime: Clinically Relevant Biomarkers and Skin Injury Progression in Murine Models.

Phosgene oxime (CX), categorized as a vesicating chemical threat agent, causes effects that resemble an urticant or nettle agent. CX is an emerging potential threat agent that can be deployed alone or with other chemical threat agents to enhance their toxic effects. Studies on CX-induced skin toxicity, injury progression, and related biomarkers are largely unknown. To study the physiologic changes, skin clinical lesions and their progression, skin exposure of SKH-1 and C57BL/6 mice was carried out with vapor from 10 µl CX for 0.5-minute or 1.0-minute durations using a designed exposure system for consistent CX vapor exposure. One-minute exposure caused sharp (SKH-1) or sustained (C57BL/6) decrease in respiratory and heart rate, leading to mortality in both mouse strains. Both exposures caused immediate blanching, erythema with erythematous ring (wheel) and edema, and an increase in skin bifold thickness. Necrosis was also observed in the 0.5-minute CX exposure group. Both mouse strains showed comparative skin clinical lesions upon CX exposure; however, skin bifold thickness and erythema remained elevated up to 14 days postexposure in SKH-1 mice but not in C57BL/6 mice. Our data suggest that CX causes immediate changes in the physiologic parameters and gross skin lesions resembling urticaria, which could involve mast cell activation and intense systemic toxicity. This novel study recorded and compared the progression of skin injury to establish clinical biomarkers of CX dermal exposure in both the sexes of two murine strains relevant for skin and systemic injury studies and therapeutic target identification. SIGNIFICANCE STATEMENT: Phosgene oxime (CX), categorized as a vesicating agent, is considered as a potent chemical weapon and is of high military and terrorist threat interest since it produces rapid onset of severe injury as an urticant. However, biomarkers of clinical relevance related to its toxicity and injury progression are not studied. Data from this study provide useful clinical markers of CX skin toxicity in mouse models using a reliable CX exposure system for future mechanistic and efficacy studies.

Metabolomics for identifying pathways involved in vesicating agent lewisite-induced corneal injury.

Lewisite (LEW) is an arsenical vesicant that can be a potentially dangerous chemical warfare agent (CWA). Eyes are particularly susceptible to vesicant induced injuries and ocular LEW exposure can act swiftly, causing burning of eyes, edema, inflammation, cell death and even blindness. In our previous studies, we developed a LEW exposure-induced corneal injury model in rabbit and showed increased inflammation, neovascularization, cell death, and structural damage to rabbit corneas upon LEW exposure. In the present study, we further assessed the metabolomic changes to delineate the possible mechanisms underlying the LEW-induced corneal injuries. This information is vital and could help in the development of effective targeted therapies against ocular LEW injuries. Thus, the metabolomic changes associated with LEW exposures in rabbit corneas were assessed as a function of time, to delineate pathways from molecular perturbations at the genomic and proteomic levels. New Zealand white rabbit corneas (n = 3-6) were exposed to LEW vapor (0.2 mg/L; flow rate: 300 ml/min) for 2.5 min (short exposure; low dose) or 7.5 min (long-exposure; high dose) and then collected at 1, 3, 7, or 14 days post LEW exposure. Samples were prepared using the automated MicroLab STAR® system, and proteins precipitated to recover the chemically diverse metabolites. Metabolomic analysis was carried out by reverse phase UPLC-MS/MS and gas chromatography (GC)-MS. The data obtained were analyzed using Metabolon's software. The results showed that LEW exposures at high doses were more toxic, particularly at the day 7 post exposure time point. LEW exposure was shown to dysregulate metabolites associated with all the integral functions of the cornea and cause increased inflammation and immune response, as well as generate oxidative stress. Additionally, all important metabolic functions of the cells were also affected: lipid and nucleotide metabolism, and energetics. The high dose LEW exposures were more toxic, particularly at day 7 post LEW exposure (>10-fold increased levels of histamine, quinolinate, N-acetyl-ß-alanine, GMP, and UPM). LEW exposure dysregulated integral functions of the cornea, caused inflammation and heightened immune response, and generated oxidative stress. Lipid and nucleotide metabolism, and energetics were also affected. The novel information about altered metabolic profile of rabbit cornea following LEW exposure could assist in delineating complex molecular events; thus, aid in identifying therapeutic targets to effectively ameliorate ocular trauma.

Ocular injury progression and cornea histopathology from chloropicrin vapor exposure: Relevant clinical biomarkers in mice.

Ocular tissue is highly sensitive to chemical exposures. Chloropicrin (CP), a choking agent employed during World War I and currently a popular pesticide and fumigating agent, is a potential chemical threat agent. Accidental, occupational, or intentional exposure to CP results in severe ocular injury, especially to the cornea; however, studies on ocular injury progression and underlying mechanisms in a relevant in vivo animal model are lacking. This has impaired the development of effective therapies to treat the acute and long-term ocular toxicity of CP. To study the in vivo clinical and biological effects of CP ocular exposure, we tested different CP exposure doses and durations in mice. These exposures will aid in the study of acute ocular injury and its progression as well as identify a moderate dose to develop a relevant rodent ocular injury model with CP. The left eyes of male BALB/c mice were exposed to CP (20% CP for 0.5 or 1 min or 10% CP for 1 min) using a vapor cap, with the right eyes serving as controls. Injury progression was evaluated for 25 days post-exposure. CP-exposure caused a significant corneal ulceration and eyelid swelling which resolved by day 14 post exposure. In addition, CP-exposure caused significant corneal opacity and neovascularization. Development of hydrops (severe corneal edema with corneal bullae) and hyphema (blood accumulation in the anterior chamber) was observed as advanced CP effects. Mice were euthanized at day 25 post-CP-exposure, and the eyes were harvested to further study the corneal injury. Histopathological analyses showed a significant CP-induced decrease in corneal epithelial thickness and increased stromal thickness with more pronounced damage, including stromal fibrosis, edema, neovascularization, trapped epithelial cells, anterior and posterior synechiae, and infiltration of inflammatory cells. Loss of the corneal endothelial cells and Descemet's membrane could be associated with the CP-induced corneal edema and hydrops which could lead to long term term pathological conditions. Although exposure to 20% CP for 1 min caused more eyelid swelling, ulceration, and hyphema, similar effects were observed with all CP exposures. These novel findings following CP ocular exposure in a mouse model outline the corneal histopathologic changes that associate with the continuing ocular clinical effects. The data are useful in designing further studies to identify and correlate the clinical and biological markers of CP ocular injury progression with acute and long-term toxic effects on cornea and other ocular tissues. We take a crucial step towards CP ocular injury model development and in pathophysiological studies to identify molecular targets for therapeutic interventions.

IRF3 Signaling within the Mouse Stroma Influences Sepsis Pathogenesis.

IFN regulatory factor 3 (IRF3) is a transcription factor that is activated by multiple pattern-recognition receptors. We demonstrated previously that IRF3 plays a detrimental role in a severe mouse model of sepsis, induced by cecal ligation and puncture. In this study, we found that IRF3-knockout (KO) mice were greatly protected from sepsis in a clinically relevant version of the cecal ligation and puncture model incorporating crystalloid fluids and antibiotics, exhibiting improved survival, reduced disease score, lower levels of serum cytokines, and improved phagocytic function relative to wild-type (WT) mice. Computational modeling revealed that the overall complexity of the systemic inflammatory/immune network was similar in IRF3-KO versus WT septic mice, although the tempo of connectivity differed. Furthermore, the mediators driving the network differed: TNF-α, IL-1ß, and IL-6 predominated in WT mice, whereas MCP-1 and IL-6 predominated in IRF3-KO mice. Network analysis also suggested differential IL-6-related inflammatory programs in WT versus IRF3-KO mice. We created bone marrow chimeras to test the role of IRF3 within leukocytes versus stroma. Surprisingly, chimeras with IRF3-KO bone marrow showed little protection from sepsis, whereas chimeras with IRF3-KO stroma showed a substantial degree of protection. We found that WT and IRF3-KO macrophages had a similar capacity to produce IL-6 and phagocytose bacteria in vitro. Adoptive transfer experiments demonstrated that the genotype of the host environment affected the capacity of monocytes to produce IL-6 during sepsis. Thus, IRF3 acts principally within the stromal compartment to exacerbate sepsis pathogenesis via differential impacts on IL-6-related inflammatory programs.

Effect of dexamethasone treatment at variable therapeutic windows in reversing nitrogen mustard-induced corneal injuries in rabbit ocular in vivo model.

Nitrogen mustard (NM) is an analogue of the potent vesicating agent sulfur mustard, with well-established ocular injury models in rabbit eyes to study vesicant-induced ocular toxicity. The effects of NM-exposure to eyes may include irritation, redness, inflammation, fibrosis, epithelial degradation, blurred vision, partial/complete blindness, which may be temporary or permanent, depending on the route, duration, and dosage of exposure. Effective countermeasures against vesicant exposure are presently not available and are warranted in case of any terrorist activity or accidental leakage from stockpiles. Herein, our focus was to evaluate whether dexamethasone (DEX), an FDA approved potent corticosteroid with documented anti-inflammatory activities, could be an effective treatment modality. Accordingly, utilizing NM-induced corneal injuries in rabbit ocular in vivo model, we examined and compared the efficacy of DEX treatments when administration was started at early (2 h), intermediate (4 h), and late (6 h) therapeutic windows of intervention after NM-exposure and administered every 8 h thereafter. The effects of NM-exposure and DEX treatments were evaluated on clinical (corneal opacity, ulceration, and neovascularization), biological (epithelial thickness, epithelial-stromal separation, blood vessels density, and inflammatory cell and keratocyte counts) and molecular (COX-2 and VEGF expression) parameters, at day 1, 3, 7 and 14. Results indicated that DEX treatment markedly and effectively reversed the NM-induced injury markers in rabbit corneas. Early administration of DEX at 2 h was found to be most effective in reversing NM-induced corneal injuries, followed by DEX 4 h and DEX 6 h administration initiation, indicating that DEX has best efficacy at the early therapeutic window in our study model.

A review of chemical warfare agents linked to respiratory and neurological effects experienced in Gulf War Illness.

Over 40% of veterans from the Persian Gulf War (GW) (1990-1991) suffer from Gulf War Illness (GWI). Thirty years since the GW, the exposure and mechanism contributing to GWI remain unclear. One possible exposure that has been attributed to GWI are chemical warfare agents (CWAs). While there are treatments for isolated symptoms of GWI, the number of respiratory and cognitive/neurological issues continues to rise with minimum treatment options. This issue does not only affect veterans of the GW, importantly these chronic multisymptom illnesses (CMIs) are also growing amongst veterans who have served in the Afghanistan-Iraq war. What both wars have in common are their regions and inhaled exposures. In this review, we will describe the CWA exposures, such as sarin, cyclosarin, and mustard gas in both wars and discuss the various respiratory and neurocognitive issues experienced by veterans. We will bridge the respiratory and neurological symptoms experienced to the various potential mechanisms described for each CWA provided with the most up-to-date models and hypotheses.

mPR-Specific Actions Influence Maintenance of the Blood-Brain Barrier (BBB).

Cerebral cavernous malformations (CCMs) are characterized by abnormally dilated intracranial microvascular sinusoids that result in increased susceptibility to hemorrhagic stroke. It has been demonstrated that three CCM proteins (CCM1, CCM2, and CCM3) form the CCM signaling complex (CSC) to mediate angiogenic signaling. Disruption of the CSC will result in hemorrhagic CCMs, a consequence of compromised blood-brain barrier (BBB) integrity. Due to their characteristically incomplete penetrance, the majority of CCM mutation carriers (presumed CCM patients) are largely asymptomatic, but when symptoms occur, the disease has typically reached a clinical stage of focal hemorrhage with irreversible brain damage. We recently reported that the CSC couples both classic (nuclear; nPRs) and nonclassic (membrane; mPRs) progesterone (PRG)-receptors-mediated signaling within the CSC-mPRs-PRG (CmP) signaling network in nPR(-) breast cancer cells. In this report, we demonstrate that depletion of any of the three CCM genes or treatment with mPR-specific PRG actions (PRG/mifepristone) results in the disruption of the CmP signaling network, leading to increased permeability in the nPR(-) endothelial cells (ECs) monolayer in vitro. Finally, utilizing our in vivo hemizygous Ccm mutant mice models, we demonstrate that depletion of any of the three CCM genes, in combination with mPR-specific PRG actions, is also capable of leading to defective homeostasis of PRG in vivo and subsequent BBB disruption, allowing us to identify a specific panel of etiological blood biomarkers associated with BBB disruption. To our knowledge, this is the first report detailing the etiology to predict the occurrence of a disrupted BBB, an indication of early hemorrhagic events.

Acute corneal injury in rabbits following nitrogen mustard ocular exposure.

Sulfur mustard (SM), a potent vesicating chemical warfare agent, and its analog nitrogen mustard (NM), are both strong bi-functional alkylating agents. Eyes, skin, and the respiratory system are the main targets of SM and NM exposure; however, ocular tissue is most sensitive, resulting in severe ocular injury. The mechanism of ocular injury from vesicating agents' exposure is not completely understood. To understand the injury mechanism from exposure to vesicating agents, NM has been previously employed in our toxicity studies on primary human corneal epithelial cells and ex vivo rabbit cornea organ culture model. In the current study, corneal toxicity from NM ocular exposure (1%) was analyzed for up to 28 days post-exposure in New Zealand White male rabbits to develop an acute corneal injury model. NM exposure led to conjunctival and eyelid swelling within a few hours after exposure, in addition to significant corneal opacity and ulceration. An increase in total corneal thickness and epithelial degradation was observed starting at day 3 post-NM exposure, which was maximal at day 14 post-exposure and did not resolve until 28 days post-exposure. There was an NM-induced increase in the number of blood vessels and inflammatory cells, and a decrease in keratocytes in the corneal stroma. NM exposure resulted in increased expression levels of cyclooxygenase-2, Interleukin-8, vascular endothelial growth factor and Matrix Metalloproteinase 9 indicating their involvement in NM-induced corneal injury. These clinical, biological, and molecular markers could be useful for the evaluation of acute corneal injury and to screen for therapies against NM- and SM-induced ocular injury.

Cutaneous exposure to vesicant phosgene oxime: Acute effects on the skin and systemic toxicity.

Phosgene Oxime (CX), an urticant or nettle agent categorized as a vesicant, is a potential chemical warfare and terrorist weapon. Its exposure can result in widespread and devastating effects including high mortality due to its fast penetration and ability to cause immediate severe cutaneous injury. It is one of the least studied chemical warfare agents with no effective therapy available. Thus, our goal was to examine the acute effects of CX following its cutaneous exposure in SKH-1 hairless mice to help establish a relevant injury model. Results from our study show that topical cutaneous exposure to CX vapor causes blanching of exposed skin with an erythematous ring, necrosis, edema, mild urticaria and erythema within minutes after exposure out to 8h post-exposure. These clinical skin manifestations were accompanied with increases in skin thickness, apoptotic cell death, mast cell degranulation, myeloperoxidase activity indicating neutrophil infiltration, p53 phosphorylation and accumulation, and an increase in COX-2 and TNFα levels. Topical CX-exposure also resulted in the dilatation of the peripheral vessels with a robust increase in RBCs in vessels of the liver, spleen, kidney, lungs and heart tissues. These events could cause a drop in blood pressure leading to shock, hypoxia and death. Together, this is the first report on effects of CX cutaneous exposure, which could help design further comprehensive studies evaluating the acute and chronic skin injuries from CX topical exposure and elucidate the related mechanism of action to aid in the identification of therapeutic targets and mitigation of injury.

Clinical progression of ocular injury following arsenical vesicant lewisite exposure.

Ocular injury by lewisite (LEW), a potential chemical warfare and terrorist agent, results in edema of eyelids, inflammation, massive corneal necrosis and blindness. To enable screening of effective therapeutics to treat ocular injury from LEW, useful clinically-relevant endpoints are essential. Hence, we designed an efficient exposure system capable of exposing up to six New-Zealand white rabbits at one time, and assessed LEW vapor-induced progression of clinical ocular lesions mainly in the cornea. The right eye of each rabbit was exposed to LEW (0.2 mg/L) vapor for 2.5, 5.0, 7.5 and 10.0 min and clinical progression of injury was observed for 28 days post-exposure (dose-response study), or exposed to same LEW dose for 2.5 and 7.5 min and clinical progression of injury was observed for up to 56 days post-exposure (time-response study); left eye served as an unexposed control. Increasing LEW exposure caused corneal opacity within 6 h post-exposure, which increased up to 3 days, slightly reduced thereafter till 3 weeks, and again increased thereafter. LEW-induced corneal ulceration peaked at 1 day post-exposure and its increase thereafter was observed in phases. LEW exposure induced neovascularization starting at 7 days which peaked at 22-35 days post-exposure, and remained persistent thereafter. In addition, LEW exposure caused corneal thickness, iris redness, and redness and swelling of the conjunctiva. Together, these findings provide clinical sequelae of ocular injury following LEW exposure and for the first time establish clinically-relevant quantitative endpoints, to enable the further identification of histopathological and molecular events involved in LEW-induced ocular injury.

Situational analysis of the quality of palliative care services across India: a cross-sectional survey.

Objective: Palliative care services in India were established in the 1980s but there is no detailed up-to-date knowledge about the quality-of-service provision nationally. We aim to describe the current quality of palliative care provision in India, as measured against nationally adopted standards. Method: A digital survey adapted from the Indian Association of Palliative Care Standards Audit Tool was administered to 250 palliative care centres. Results: Two hundred and twenty-three (89%) palliative care centres participated - 26.4% were government-run, while the rest include non-governmental organisations, private hospitals, community-led initiatives and hospices. About 200 centres 'often' or 'always' fulfilled 16/21 desirable criteria; however, only 2/15 essential criteria were 'often' or 'always' fulfilled. Only 5.8% provide uninterrupted access to oral morphine. Significance of the results: Palliative care centres in India are falling short of meeting the essential quality standards, indicating the urgent need for new initiatives to drive national change.

Detection of Blood Cell Surface Biomarkers in Septic Mice.

Sepsis arises when an infection induces a dysregulated immune response, resulting in organ damage. New methods are urgently needed to diagnose patients in the early stages of sepsis, and identify patients with a poor disease prognosis. One promising approach is to identify the rapid changes in cell surface antigens (biomarkers) that occur during sepsis, as a consequence of leukocyte mobilization and activation. This chapter describes the method for staining whole blood with fluorescently conjugated antibodies that detect cell surface biomarkers, and performing flow cytometry analysis to quantify biomarker-positive cells. Our protocol is designed to detect blood cell surface biomarkers in septic mice, but could also be applied to study potential biomarkers in blood obtained from human patients with sepsis and other medical conditions.

Aged IRF3-KO Mice are Protected from Sepsis.

PURPOSE: Sepsis is a leading cause of hospital admissions and deaths. Older adults (>65 years) are particularly susceptible to sepsis and experience higher morbidity and mortality rates than younger people. We previously showed that interferon regulatory factor 3 (IRF3) contributes to sepsis pathogenesis in young mice subject to cecal ligation and puncture (CLP). In this study, we investigated if IRF3 contributes to sepsis in the context of aging. METHODS: Sepsis was induced in aged wild-type (WT) and IRF3-knock-out (KO) mice, using a clinically-relevant CLP-sepsis model including fluids and antibiotics. Animal survival, disease score and hypothermia were evaluated as indicators of sepsis pathogenesis. Serum cytokines and serum enzymes indicative of organ damage were also measured. RESULTS: Aged WT mice were highly susceptible to sepsis (90% mortality). In comparison, aged IRF3-KO mice were significantly protected (20% mortality). Aged IRF3-KO mice showed a lower disease score and reduced hypothermia following CLP, compared to WT mice. Serum cytokines interleukin (IL)-6, IL-12/23p40 and macrophage chemoattractant protein (MCP)-1, and creatinine kinase (CK) were lower in aged IRF3-KO septic mice compared to WT counterparts. Aged male mice were found to be more susceptible to sepsis compared to females. Female mice, however, produced higher levels of serum cytokines and CK. CONCLUSION: These results demonstrate that IRF3 plays a detrimental role in sepsis in aged mice and highlight the impact of biological sex.

Cellular uptake of polylactide particles induces size dependent cytoskeletal remodeling in antigen presenting cells.

Phagocytosis of particulate vaccine delivery systems is a critical immune mechanism involved in antigen capture and processing by macrophages and dendritic cells. The internalization and degradation of the particles involve a complex sequence of events. This process coordinates lipids, signaling proteins, and the cytoskeleton. Dynamic changes in the actin cytoskeleton are essential for phagocytosis and antigen presentation. Knowledge regarding the correlation of surface properties, attached ligand density and geometric size of particles with the efficiency of phagocytosis may facilitate their design and application. To investigate this, polylactide biodegradable particles with different diameters (2-4 µm and 200-300 nm) were exposed to murine macrophages and dendritic cells and the effect of size on a series of cellular responses was evaluated. Cellular uptake studies using microscopy and flow cytometry showed size dependent internalization of particles, with nanoparticles accumulating in cells at a faster rate. The particles induced homoaggregation of cells and also showed cytoskeletal remodeling that could be inhibited by cytochalasin-D. Scanning electron microscopy images showed the time dependent formation of phagocytic cups and invaginations that promote particle uptake. The particles were observed to co-localized with the endo-lysosomal compartments after phagocyotosis. In our experiments, particle mediated immunoactivation, antigen processing and cytokine secretion have shown a good correlation with the uptake process. These findings would allow a better understanding of the process of particle uptake and may be instrumental in the rational design of optimal vaccine delivery systems.

Large Peritoneal Macrophages and Transitional Premonocytes Promote Survival during Abdominal Sepsis.

Monocytes and macrophages are early sentinels of infection. The peritoneum contains two resident populations: large and small peritoneal macrophages (LPMs and SPMs). While LPMs self-renew, circulating monocytes enter the peritoneum and differentiate into SPMs. We lack information on the dynamics of monocyte-macrophage trafficking during abdominal sepsis, reflecting an important knowledge gap. In this study, we characterize the presence of LPMs, SPMs, and monocytes in the peritoneum of mice following cecal ligation and puncture (CLP)-induced sepsis and sham surgery. LPMs rapidly disappeared from the peritoneum and were scarce at 18-66 h after CLP or sham surgery. By 14 d, LPMs returned for sham mice, but they remained scarce in CLP mice. Depletion of LPMs from the peritoneum of CD11b-DTR mice greatly increased animal mortality. These data imply that LPMs are critical for sepsis survival. Monocytes rapidly infiltrated the peritoneum and were abundant at 18-66 h after CLP or sham surgery. Surprisingly, SPMs only increased at 14 d post-CLP. Therefore, monocytes may defend hosts from acute sepsis mortality without generating SPMs. More monocytes were present in mice predicted to survive sepsis versus mice predicted to die. However, altering monocyte numbers via CCR2 deficiency or adoptive transfer did not significantly affect animal survival. We reasoned that animals destined to survive sepsis may exhibit a different monocyte phenotype, rather than merely enhanced numbers. Indeed, mice predicted to survive possessed more CD31+, CXCR4hi transitional premonocytes in their abdomen. Inhibition of CXCL12-CXCR4 signaling via AMD3100 exacerbated sepsis. These data imply that recruitment of transitional premonocytes to the abdomen promotes sepsis survival.

Evaluating the Timeliness and Specificity of CD69, CD64, and CD25 as Biomarkers of Sepsis in Mice.

ABSTRACT: Sepsis occurs when an infection induces a dysregulated immune response, and is most commonly bacterial in origin. This condition requires rapid treatment for successful patient outcomes. However, the current method to confirm infection (blood culture) requires up to 48 h for a positive result and many true cases remain culture-negative. Therefore, new diagnostic tests are urgently needed. Recent clinical studies suggest that CD69, CD64, and CD25 may serve as useful biomarkers of sepsis. In this study, we evaluated the cecal ligation and puncture and cecal slurry mouse models as tools to study these biomarkers in young and aged mice, and elucidate the timeliness and specificity of sepsis diagnosis. Fluorescence-activated cell sorting analysis revealed that all three biomarkers were elevated on blood leukocytes during sepsis. CD69 was specifically upregulated during sepsis, while CD64 and CD25 were also transiently upregulated in response to sham surgery. The optimal biomarker, or combination of biomarkers, depended on the timing of detection, mouse age, and presence of surgery. CD69 demonstrated an excellent capacity to distinguish sepsis, and in some scenarios the diagnostic performance was enhanced by combining CD69 with CD64. We also analyzed biomarker expression levels on specific cell populations (lymphocytes, monocytes, and neutrophils) and determined the cell types that upregulate each biomarker. Elevations in blood biomarkers were also detected via microfluidic analyses; in this case CD64 distinguished septic mice from naive controls. Our results suggest that CD69 and CD64 are valuable biomarkers to rapidly detect sepsis, and that mouse models are useful to study and validate sepsis biomarkers.

Effect of supersaturated oxygen emulsion treatment on chloropicrin-induced chemical injury in ex vivo rabbit cornea.

With a possibility for the use of chemical weapons in battlefield or in terrorist activities, effective therapies against the devastating ocular injuries, from their exposure, are needed. Oxygen plays a vital role in ocular tissue preservation and wound repair. We tested the efficacy of supersaturated oxygen emulsion (SSOE) in reducing ex vivo corneal and keratocyte injury from chloropicrin (CP). CP, currently used as a pesticide, is a chemical threat agent like the vesicating mustard agents and causes severe corneal injury. Since our previous study in human corneal epithelial cells showed the treatment potential of SSOE (55 %), we further tested its efficacy in an ex vivo CP-induced rabbit corneal injury model. Corneas were exposed to CP (700 nmol) for 2 h, washed and cultured with or without SSOE for 24 h or 96 h. At 96 h post CP exposure, SSOE treatment presented a healing tendency of the corneal epithelial layer, and abrogated the CP-induced epithelial apoptotic cell death. SSOE treatment also reduced the CP induced DNA damage (H2A.X phosphorylation) and inflammatory markers (e.g. MMP9, IL-21, MIP-1ß, TNFα). Further examination of the treatment efficacy of SSOE alone or in combination with other therapies in in vivo cornea injury models for CP and vesicants, is warranted.

Pathophysiology and inflammatory biomarkers of sulfur mustard-induced corneal injury in rabbits.

Sulfur mustard (SM) is a cytotoxic, vesicating, chemical warfare agent, first used in 1917; corneas are particularly vulnerable to SM exposure. They may develop inflammation, ulceration, neovascularization (NV), impaired vision, and partial/complete blindness depending upon the concentration of SM, exposure duration, and bio-physiological conditions of the eyes. Comprehensive in vivo studies have established ocular structural alterations, opacity, NV, and inflammation upon short durations (<4 min) of SM exposure. In this study, detailed analyses of histopathological alterations in corneal structure, keratocytes, inflammatory cells, blood vessels, and expressions of cyclooxygenase (COX)-2, matrix metalloproteinase (MMP)-9, vascular endothelial growth factor (VEGF), and cytokines were performed in New Zealand white rabbits, in a time-dependent manner till 28 days, post longer durations (5 and 7 min) of ocular SM exposure to establish quantifiable endpoints of injury and healing. Results indicated that SM exposure led to duration-dependent increases in corneal thickness, opacity, ulceration, epithelial-stromal separation, and epithelial degradation. Significant increases in NV, keratocyte death, blood vessels, and inflammatory markers (COX-2, MMP-9, VEGF, and interleukin-8) were also observed for both exposure durations compared to the controls. Collectively, these findings would benefit in temporal delineation of mechanisms underlying SM-induced corneal toxicity and provide models for testing therapeutic interventions.

Oncology-Based Palliative Care Development: The Approach, Challenges, and Solutions From North-East Region of India, a Model for Low- and Middle-Income Countries.

BACKGROUND: Access to palliative care within healthcare systems of low- and middle-income countries (LMICs) has never been more pronounced than in current times. The Lancet Commission Report (2018) estimates that 80% of global serious health-related suffering (SHS), which demands access to palliative care for its relief, are in LMICs. Cancer is a major contributor to SHS and a rapidly growing burden in LMICs. Similar to many LMICs, cancer is a leading cause of death in India. The North-East Region (NER) of India has a high prevalence of cancer and paucity of services for cancer and palliative care. OBJECTIVES: To describe the strategies used to initiate and strengthen palliative care services integrated with the comprehensive cancer care initiatives in the state of Assam in NER. METHODS: After an initial assessment of the status of palliative care in the NER, a multipronged strategy was adopted that aligned with the WHO framework recommended for initiating palliative care services. A core team working with a government and private collaborative strategized and activated supportive policies, education, and training and improved access and availability to essential drugs, while implementing the components synchronously within the state. SIGNIFICANCE: This project demonstrates an informed regional adaptation of the WHO model. It highlights the strengths of integrating palliative care within cancer care program right from its inception. It emphasizes the sustainability of services activated across public healthcare systems, as compared with the donor- or champion-driven initiatives. The outcome of this project underlines the relevance of this model for LMIC regions with similar health systems and sociocultural and economic contexts.

A Supersaturated Oxygen Emulsion for the Topical Treatment of Ocular Trauma.

INTRODUCTION: Roughly 13% of all battlefield injuries include some form of ocular trauma. Ocular tissue preservation is critical for wound healing for warfighters with ocular injuries. Our team hypothesized that oxygen plays a vital role in ocular tissue preservation and wound healing and has developed a supersaturated oxygen emulsion (SOE) for the topical treatment of ocular trauma. MATERIALS AND METHODS: The partial pressure of oxygen (PO2) was measured in the SOE. Safety and efficacy studies were carried out in primary human corneal epithelial (HCE) cells, as the outermost layer is the first barrier to chemical and mechanical injury. Western blot, scratch assay, and MTT assays were conducted to determine the effect of the SOE on various molecular markers, the rate of scratch closure, and cellular viability, respectively. RESULTS: Data indicate that the SOE releases oxygen in a time-dependent manner, reaching a partial pressure within the emulsion over four times atmospheric levels. Studies in HCE cells indicate that application of the SOE does not lead to DNA damage, promote cell death, or hinder the rate of scratch closure and enhances cellular viability. Preliminary studies were carried out with chloropicrin (CP; developed as a chemical warfare agent and now a commonly used pesticide) as a chemical agent to induce ocular injury in HCE cells. CP exposures showed that SOE treatment reverses CP-induced DNA damage, apoptotic cell death, and oxidative stress markers. CONCLUSIONS: Maintaining adequate tissue oxygenation is critical for tissue preservation and wound repair, especially in avascular tissues like the cornea. Further studies examining the application of the SOE in corneal injury models are warranted.