Countermeasures against Pulmonary Threat Agents

Abstract

Inhaled toxicants are used for diverse purposes, ranging from industrial applications such as agriculture, sanitation, and fumigation to crowd control and chemical warfare, and acute exposure can induce lasting respiratory complications. The intentional release of chemical warfare agents (CWAs) during World War I caused life-long damage for survivors, and CWA use is outlawed by international treaties. However, in the past two decades, chemical warfare use has surged in the Middle East and Eastern Europe, with a shift toward lung toxicants. The potential use of industrial and agricultural chemicals in rogue activities is a major concern as they are often stored and transported near populated areas, where intentional or accidental release can cause severe injuries and fatalities. Despite laws and regulatory agencies that regulate use, storage, transport, emissions, and disposal, inhalational exposures continue to cause lasting lung injury. Industrial irritants (e.g., ammonia) aggravate the upper respiratory tract, causing pneumonitis, bronchoconstriction, and dyspnea. Irritant gases (e.g., acrolein, chloropicrin) affect epithelial barrier integrity and cause tissue damage through reactive intermediates or by direct adduction of cysteine-rich proteins. Symptoms of CWAs (e.g., chlorine gas, phosgene, sulfur mustard) progress from airway obstruction and pulmonary edema to acute lung injury (ALI) and acute respiratory distress syndrome (ARDS), which results in respiratory depression days later. Emergency treatment is limited to supportive care using bronchodilators to control airway constriction and rescue with mechanical ventilation to improve gas exchange. Complications from acute exposure can promote obstructive lung disease and/or pulmonary fibrosis, which require long-term clinical care.

SIGNIFICANCE STATEMENT

Inhaled chemical threats are of growing concern in both civilian and military settings, and there is an increased need to reduce acute lung injury and delayed clinical complications from exposures. This minireview highlights our current understanding of acute toxicity and pathophysiology of a select number of chemicals of concern. It discusses potential early-stage therapeutic development as well as challenges in developing countermeasures applicable for administration in mass casualty situations.

Introduction

Over two hundred chemicals of concern (CoC) have been identified by the US Department of Homeland Security as high-consequence public health threats, and one-fourth of these toxicants affect the lungs. Chemical warfare agents (CWAs) are high-priority health concerns with short exposure windows that cause immediate casualties and lung injury and can induce progressive lung disease. Industrial and agricultural chemicals (irritants) are of significant concern as accidental or intentional exposures affect civilian populations. Lung irritants cause reversible nonimmunologic reactions after direct contact with the nose, throat, and respiratory tract.

Lung tissue in the lower respiratory tract is composed of numerous alveoli that mediate gas exchange and is highly sensitive to inhaled toxicants due its extensive surface area. Toxicants are inhaled through absorption of chemical vapors, particulates, and incomplete combustion byproducts into the lungs. Clinical symptoms and severity depend on dose and proximity, water solubility, and toxicant size. Highly water-soluble (hydrophilic, e.g., ammonia) and larger compounds (>10 μm) mainly affect the upper airways, stimulating the trigeminal nerves of the nasal passages, with rapid symptom onset that presents as burning, irritation, and cough. Those of intermediate size and solubility (5–10 μm, e.g., chlorine gas) cause greater parenchymal injury, and small, poorly water-soluble (hydrophobic) toxicants <5 μm (e.g., phosgene gas) penetrate to the respiratory bronchioles and alveoli of the lower respiratory tract to cause detrimental lung injury (Parkes, 1994; Muskat, 2008; Nelson and Hoffman, 2014). Oxidant gases (e.g., chlorine, phosgene, ammonia) react initially with antioxidants in the epithelial lining fluid, and during acute, high-dose toxicant exposures, this first line of defense can be overwhelmed (Addis et al., 2021). Understudied chemicals and some of the acute toxicities that lead to lung complications are described in Table 1.

TABLE 1.

Acute toxicity of pulmonary chemicals of concern

Chemical	Ammonia	Acrolein	Chloropicrin	Chlorine	Phosgene
Formula	NH3	C3H4O	CCl3NO2	Cl2	COCl2
Deposition	URT	URT	URT	URT/LRT	LRT
Industrial/agricultural use	Refrigerant Fertilizer	Biocide Livestock feed	Disinfectant Soil fumigant	Disinfectant Sanitation Plastics Paper	Chemicals Adhesives Plastics
Reactive by-products	NH4OH	C3H4O2, COx	COCl2, Cl2, NOx, (intermediates: HOCl, CH3NO2)	HCl, HOCl	HCl
Pathophysiology	Irritant, exothermically reacts with water, damages tissue	Irritant, strong electrophile, damages tissue	Irritant, induces ER stress, mutagen	Reacts with epithelial lining, lyses cells, causes fibrin deposition	Hydrolyses with mucus layer, acylates biomolecules, depletes surfactant
Acute toxicity	Bronchiectasis, BO, COPD, ILD, respiratory failure	Bronchial obstruction, LRT necrosis	Bronchitis, pulmonary edema, respiratory failure	ALI/ARDS, RADS, pulmonary fibrosis, respiratory failure	Pulmonary edema, hypoxemia, ALI/ARDS, respiratory failure

AKG, α-ketoglutarate; C₃H₄O, acrolein; CCl₃NO₂, chloropicrin; CH₃NO₂, nitromethane; Cl₂, chlorine gas; COCl₂, phosgene; COx, carbon oxide; ER, endoplasmic reticulum; HCl, hydrogen chloride; HOCl, hypochlorous acid; LRT, lower respiratory tract; NH₃, anhydrous ammonia; NH₄OH, ammonium hydroxide; NOx, nitrogen oxide; URT, upper respiratory tract.

Mechanisms of early lung injury are shared among irritants and CWAs, whereas the development and progression of delayed lung complications are toxicant specific. CWAs cause respiratory distress, rapid airway constriction, and pulmonary edema that can lead to respiratory failure. Similarly, CWAs can cause bronchoconstriction and airway smooth muscle activation that accelerates cardiopulmonary injury. Exposure induces two-phase effects, which initially present as pneumonitis and bronchitis, with latent symptoms of pulmonary edema, bronchiolitis, and acute lung injury (ALI)/acute respiratory distress syndrome (ARDS) days to weeks later (Muskat, 2008). High-dose irritants initially cause persistent rhinosinusitis, airway hyperresponsiveness (AHR), and accelerated lung function decline. Complications from exposure may proceed from airway obstruction to persistent reactive airways disease (RADS) and, at high doses, from pulmonary edema to ALI/ARDS (Leduc et al., 1992; Lu et al., 2017; Pesonen and Vähäkangas, 2020).

ARDS develops as a cascading consequence of ALI and often progresses to respiratory failure. Current research is aimed at studying ALI mechanisms and intervention strategies that can be applied before ARDS develops. Symptoms of ALI and ARDS overlap, with clinical distinction based on the degree of hypoxemia present [PaO₂/FiO₂ ratio, or oxygenation index, is the ratio of partial pressure of oxygen (PaO₂) to fraction of inspired oxygen (FiO₂): ALI ≤ 300 mm Hg; ARDS ≤ 200 mm Hg]. The term ALI/ARDS is now widely used (Butt et al., 2016), and diagnosis pairs deteriorating oxygenation levels with evidence of bilateral lung infiltrates on chest X-ray. ALI/ARDS represents a continuum of physiologic changes that disrupt the integrity of the endothelial and epithelial barriers of the lung, causing diffuse alveolar damage and increased lung permeability that impairs gas exchange. It is characterized by three phases: exudative (days 1–6), proliferative (days 7–14), and fibrotic (>14 days), and diverse chemicals included on the CoC list may induce phenotypically diverse effects along the trajectory of lung injury (Radbel et al., 2020).

The exudative phase is indicated by inflammation, surfactant deficiency, and tissue factor dysfunction, which contribute to intravascular and intra-alveolar coagulation and hypoxemia. During this phase, lymphatic drainage from the lung cannot remove fluid that filters from the lung vasculature, and potentially fatal pulmonary edema develops (Lindsay, 2011). The proliferative phase is largely mediated by proresolution macrophages, which recruit specialized proresolving mediators to initiate resolution. In this phase, alveolar type II cells proliferate due to Wnt/β-catenin signaling and differentiate into alveolar type I cells to repair the damaged epithelial barrier (Radbel et al., 2020). Impaired resolution prompts the debilitating fibrotic phase, which is characterized by collagen deposition and persistent thickening of the alveolar interstitium that impedes gas exchange. This promotes obstructive lung diseases [e.g., bronchiolitis obliterans (BO), chronic obstructive pulmonary disease (COPD)] that require long-term clinical management. For comprehensive reviews of ALI/ARDS, see Radbel et al. (2020) and Butt et al. (2016).

In CWA-induced lung injury, the additional extravascular coagulation and fibrin formation in the alveolar compartment may promote the development of pulmonary fibrosis. Early symptoms include unproductive cough, weight loss, and fatigue, largely due to hypoxia (Savin et al., 2022). Diagnosis is based on lung function measures, and blood and imaging tests are used to rule out other lung-related illnesses. Pulmonary fibrosis is a heterogeneous disease with distinct tissue pathology, encompassing multiple chronic lung outcomes, including interstitial lung disease (ILD) and idiopathic pulmonary fibrosis. Left unresolved, pulmonary fibrosis progressively destroys the alveolar structure of the lung and can lead to respiratory failure. For a comprehensive review of fibrosis, see Savin et al. (2022).

The National Institutes of Health Chemical Countermeasures Research Program is focused on toxicant-induced ALI/ARDS and delayed effects of lung injury. Its mission is to understand molecular and cellular perturbations and identify common mechanisms of lung injury to develop safe and effective medical countermeasures that reduce acute mortality and complications of chronic lung disease.

Recent Advances in Inhalational Research

Irritants: Ammonia.

Anhydrous ammonia (NH₃) is a compressed liquid used as an agricultural fertilizer and refrigerant and is widely transported through highly populated areas. Upon release, ammonia reacts with water to form ammonium hydroxide, which irritates the mucosa and, at high concentrations, causes severe upper airways irritation and respiratory failure within 2–5 minutes of exposure (Saeed et al., 2018). Tissue damage occurs through exothermic reactions with body tissues, causing alkali skin burns and liquefactive necrosis (Pangeni et al., 2022). In some cases, survivors may develop complications such as bronchiectasis, AHR, BO, COPD, ILD, and end-stage disease that requires lung transplantation. Diagnosis is based on exposure and by physical examination and body-system focused laboratory tests. α-Ketoglutarate (AKG) and fluticasone propionate (Flonase) have been shown to reduce injury in animal and cell culture models (Ali et al., 2012, 2013); however, there are currently no antidotes or tests for the extent of toxicity.

Recent animal studies have established an LD₅₀ value and mechanisms by which ammonia induces ALI/ARDS and systemic changes (Perkins et al., 2017; Elfsmark et al., 2019). Intratracheal ammonia instillation causes severe lung injury and increased vascular resistance in mice, with persistent respiratory acidosis and alveolar damage at day 1 and interstitial hemorrhage and mortality within 7 days of exposure (Elfsmark et al., 2022). Early activation of surfactant protein D (SP-D), which regulates pulmonary innate immunity and defense against xenobiotics (Madsen et al., 2000), and sustained activation of coagulation factors such as plasminogen activator inhibitor 1 (PAI-1) and fibrinogen are observed. Ammonia induces biphasic injury, with an acute inflammatory phase that deteriorates to obstructive lung disease within days. Comparable symptoms of dyspnea, acute neutrophilic airway inflammation, and pulmonary edema are seen in ammonia-induced ALI/ARDS. Case reports show that some patients develop delayed lung injuries without initial upper respiratory tract obstruction (Close et al., 1980) and persistent RADS after a single high-dose exposure (Brooks et al., 1985; Brooks, 2008).

Irritant Gases: Acrolein and Chloropicrin.

Acrolein (C₃H₄O) and chloropicrin (CCl₃NO₂) were deployed as CWAs in World War I and are now used for industrial and agricultural applications. Acrolein is a volatile aldehyde formed during incomplete combustion of fossil fuels, and large quantities are synthesized for industrial use as a biocide (Conklin, 2016). It reacts with the respiratory lining fluid and cellular macromolecules, altering gene regulation and mucociliary transport and compromising endothelial barrier integrity. Acrolein induces AHR and mucus hypersecretion, which obstructs the lumen and leads to airspace enlargement with loss of lung elasticity. At high concentrations, it promotes epithelial cell hyperplasia with lower respiratory tract necrosis that induces pulmonary edema and ALI/ARDS if not resolved (Xiong et al., 2018). Acrolein-exposed lungs have histologic features of squamous cell differentiation and dysfunctional tissue remodeling (Beauchamp et al., 1985).

Mechanistic studies show that acrolein affects energy balance, with profound effects on lipid and mitochondrial metabolism (Fabisiak et al., 2011; Agarwal et al., 2013; Snow et al., 2017). RNA-sequencing analyses indicate sex-dependent differences in injury severity and increased neutrophil extracellular trap formation (Bein et al., 2021). In vitro studies using acrolein vapor suggest that interleukin (IL)-1β, IL-22, and IL-17 pathway genes play a central role in toxicity (Johanson et al., 2020). Potential therapies include chemosensory transient receptor potential (TRP)-A1 antagonists (Conklin, 2016), antioxidants (Hochman et al., 2014), and targeting claudin 5 (CLDN5) to preserve endothelial barrier integrity (Jang et al., 2011). Investigators show increased perivascular edema and reduced CLDN5 expression in acrolein-sensitive mice (Jang et al., 2011), and CLDN5 is being studied in other models of chemically induced ALI/ARDS (Geng et al., 2018, 2020).

Chloropicrin is a halogenated fumigant and disinfectant that causes respiratory complications when inhaled. Decomposition releases toxic gases such as phosgene, chlorine, and nitrogen oxides, and it can dehalogenate in aqueous systems to form nitromethane. Although initial exposure irritates the lungs, at higher concentrations chloropicrin induces vomiting and breathing difficulties and airway damage that triggers bronchitis and pulmonary edema. This can progress to respiratory failure or, in some instances, life-long complications such as COPD. Histologic lesions including ulceration and necrosis are seen in animal models, and chloropicrin is toxic to other organs (Pesonen et al., 2017).

Mechanisms of chloropicrin-induced lung damage are not well understood, and animal studies have mainly focused on ocular injury. Chloropicrin-exposed primary human bronchial epithelial cells show elevated levels of extracellular signal-regulated kinase 1 and 2 that induce ER stress (Pesonen et al., 2015) and increase cytoplasmic vacuolization (Pesonen et al., 2017). Human bronchial epithelial cells show deformed cytosketal ultrastructure with weakened cell attachments and mitochondrial dysfunction. This suggests that chloropicrin targets β-tubulin and interferes with the tubulin network, inducing apoptosis. Chloropicrin is mutagenetic and modifies sulfhydryl groups on cysteines and enzymes important for energy metabolism (Sparks et al., 2000). There are no specific biomarkers of exposure, and therapy is limited to supportive care and the use of antioxidants. N-acetylcysteine acts as a thiol-reducing agent to scavenge reactive oxygen species/reactive nitrogen species and stimulate glutathione synthesis and prevents chloropicrin-induced cytotoxicity and vacuolization in cell culture models (Pesonen et al., 2014).

Chemical Warfare Agents: Chlorine, Phosgene, and Sulfur Mustard.

Chlorine gas (Cl₂) is used industrially for sanitation and water purification and is also used as a CWA. It reacts with the mucosal lining to form hydrochloric and hypochlorous acids and the oxidative byproduct hypochlorite, which mediates cytotoxic effects in the lungs (Achanta and Jordt, 2021). Chlorine initially causes symptoms of bronchospasm and chest pain and, at high concentrations, destroys lung structure, inducing hemorrhage, pulmonary edema, and respiratory collapse within 1 hour of exposure. Survivors of acute exposure may develop symptoms of pulmonary fibrosis and RADS, with cardiovascular complications that last for years (Zaky et al., 2015; Carlisle et al., 2016).

Animal models of chlorine injury show histologic features of epithelial desquamation, leukocyte infiltration, atelectasis, and necrosis (Balakrishna et al., 2014). Biologic changes observed include protease activation, coagulopathies, compromised barrier integrity, and fibrin deposition (Mo et al., 2015; Musah et al., 2017) as well as increased susceptibility to infection (Gessner et al., 2013; Song et al., 2015). Potential therapeutics to counteract chlorine toxicity include antioxidants, corticosteroids, and combined therapies. TRP inhibitors, nitrite administration, heparin, and autophagy activators also show limited benefits (Achanta and Jordt, 2021). The recent identification of chlorinated lipids as biomarkers of exposure provides a potential diagnostic measure of chlorine toxicity (Spickett, 2007; Ford et al., 2016).

Phosgene (COCl₂; CG) is a toxic, colorless gas used for chemical synthesis and as a CWA. CG is hydrophobic and slowly reacts with water to form carbon dioxide and hydrochloric acid, which directly damages the respiratory tract (Pauluhn, 2021; Cao et al., 2022). CG bypasses chemosensory perception through limited retention in the upper airways, and reflex bradypnea is largely absent (Pauluhn, 2021). CG collects in the lower respiratory tract, and noncardiogenic pulmonary edema often goes unnoticed, with development of secondary hypoxemia and deterioration of the blood-gas barrier (Pauluhn et al., 2007). Phosgene-induced ALI is preceded by persistent respiratory depression and cardiogenic pulmonary edema within 15–20 hours of exposure (Pauluhn, 2021). This occult or asymptomatic period is not evident on clinical examination. Fluid shifts from the systemic to the pulmonary circulation induce hypovolemia and hypotension that parallel pulmonary edema onset (Pauluhn, 2021). In late-stage phosgene-induced acute lung injury, survivors often develop fibrosis and chronic obstructive lung disease (Cao et al., 2022).

A consistent LC₅₀ in animal models has been difficult to determine (Hobson et al., 2021), and toxic load depends linearly on dose × exposure duration, such that acute high dose is no more toxic than chronic low levels of exposure. CG is electrophilic and acylates nucleophilic biomolecules (Holmes et al., 2016), inducing pulmonary edema as the blood-gas barrier is permeabilized. CG undergoes hemolytic cleavage to form highly reactive carbamoyl chloride derivatives (Arroyo et al., 1993; Holmes et al., 2016) that further alter surfactant levels. Increased nitrosative stress affects neuronal cells that innervate the lungs as well as epithelial, endothelial, and blood cells. Ongoing debate on the role of inflammation in vasopermeability and pulmonary edema (Russell et al., 2006; Chen et al., 2013; Holmes et al., 2016) has impacted agreement on which biomarkers and therapeutics should be explored.

Anti-inflammatories show little benefit for CG-induced injury, and TRP antagonists are limited to reducing neurogenic inflammation (Pauluhn, 2021). Administration of antioxidants such as N-acetylcysteine increase glutathione availability (Ji et al., 2010), and melatonin attenuates CG-induced lung injury through Wnt/β-catenin signaling (Zhang et al., 2017). Strategies using ulinastatin and NOS-2 inhibitors show promise in small animal models (Shen et al., 2014; Filipczak et al., 2015; Zhang et al., 2017), but early treatment during the asymptomatic phase is critical to minimize the risk of cardiogenic edema (Lu et al., 2021). Mesenchymal stem cells that target miRNAs (Xu et al., 2019; Qu et al., 2020; Jiang et al., 2021) or overexpress angiogenic (Shao et al., 2018) or heat shock proteins (Jin et al., 2020) are shown to counteract toxicity in vitro, and administration of cardioprotective Fv-HSP72 may ameliorate injury (Hobson et al., 2021).

Sulfur mustard (C₄H₈Cl₂S; SM) is a toxic vesicant primarily used as a CWA, with short and long-term lung injury effects. It causes blistering and erythema of the respiratory tract (Schmidt et al., 2018) that destroys bronchial tissue and obstructs the airway minutes to hours after exposure. At high doses, this is followed by hemorrhagic pulmonary edema, secondary pneumonia, and respiratory failure 24 hours to 1 week later (Ghanei et al., 2005). Acute exposures can induce pathogenic lesions and fibrosis that promotes BO and/or ILD decades later (Ghanei et al., 2008). In addition, acute SM exposure causes systemic injury and increases cancer risk (Razavi et al., 2016).

Although the lungs are a major target of exposure, SM is highly lipophilic and enters the body through multiple routes. It eliminates chloride ions through intramolecular substitution to form cyclic sulfonium ions. These reactive intermediates alkylate DNA and other nuclear components, inducing genotoxicity and cytotoxic damage that suppresses the immune system and inducible nitric oxide synthase (iNOS) signaling (Tahmasbpour et al., 2019). Inhaled SM activates phagocytic leukocytes and inflammatory mediators that alter serum cytokines, C-reactive protein (CRP), and soluble proapoptotic fasciclin 1 (Fas1) expression. Vascular leakage and endothelial permeability are reported in animal models (Calvet et al., 1994; Rancourt et al., 2013; Malaviya et al., 2020) as well as fibrin cast formation that occludes the airways (White et al., 2016). Increased fibrinogen suppresses surfactant and can prolong macrophage activation and inflammation (Wygrecka et al., 2008).

There are currently no approved therapies for SM inhalation; however, combined administration of vitamin E and L-thiocitrulline (L-TC) helps replenish nitric oxide (NO) and manage respiratory symptoms. Therapies being explored include anti-inflammatories, antioxidants, and protease inhibitors, and some benefits from natural products are shown (Boskabady and Farhadi, 2008; Hossein et al., 2008). Studies show restoration of surfactant may protect against SM-induced lung injury (van Helden et al., 2004) and that SP-D, a nonspecific alveolar injury marker, may serve as an indicator of early respiratory disease (Starosta and Griese, 2006; Sorensen, 2018; Malaviya et al., 2020).

Early Mechanisms of Toxicant-Induced Lung Injury

Although ALI/ARDS progression and delayed respiratory complications are toxicant specific, toxicant-induced inflammation and injury share overlapping protective mechanisms to limit lung damage (Fig. 1). Acute exposure induces nitrosative stress and endothelial damage that occurs concurrently with inflammation (Radbel et al., 2020; Elfsmark et al., 2022). Reactive intermediates (e.g., reactive oxygen species/reactive nitrogen species) induce inflammatory cytokines/chemokines that activate iNOS, which generates excess NO that can magnify oxidative stress (Sun et al., 2010). Peripheral lung inflammation stimulates histamine release and endothelial NO production, which amplifies the inflammatory cascade (Branco et al., 2018; Radbel et al., 2020). In CWA models, the overwhelming magnitude of nitrosative stress depletes NO stores from alveolar macrophages (Malaviya et al., 2023) and aggravates histamine intolerance (Branco et al., 2018). It also reduces glutathione levels and alters phospholipid production, exhausting surfactant reserves in multiple models of toxicity (Tahmasbpour et al., 2019; Pesonen and Vähäkangas, 2020; Elfsmark et al., 2022).

Fig. 1.

Mechanisms by which inhaled toxicants affect the bronchioles and alveoli of the lung, leading to nitrosative stress and inflammation. Activation, elevation, and depletion of shared signaling pathways enhance injury. DNA damage, protein adduction, and lipid peroxidation, if left unresolved, can develop into bronchitis, pulmonary edema, and ultimately ALI/ARDS. Current therapies for symptoms include supportive care with bronchodilator use and mechanical ventilation.

Chemosensory TRP channels respond to a variety of inhaled toxicants and induce changes in intracellular calcium that alter alveolar and vascular permeability. Activation of TRP channels in the respiratory epithelium prompts membrane depolarization and inflammatory neuropeptide release (Bessac and Jordt, 2010; Achanta and Jordt, 2020; Pesonen and Vähäkangas, 2020). Irritation from chloropicrin and acrolein is mediated by TRPA1 (Conklin, 2016; Pesonen and Vähäkangas, 2020), stimulating afferent airway fibers that trigger reflex bradypnea and respiratory braking. Chlorine and sulfur mustard directly activate TRP channels (Stenger et al., 2015; Zellner and Eyer, 2020), and this complex interplay of inflammatory mediators and nitrosative stress prompts bronchoconstriction that restricts airflow.

For sulfur mustard and analogs, the heightened nitrosative burden desensitizes β-adrenergic receptors in the distal airways, affecting cAMP levels and impairing endothelial barrier function (Ghanei et al., 2007; Kabir et al., 2009; Rambacher and Moniri, 2020). This allows blood plasma to enter the interstitium and exacerbates pulmonary edema and inflammation (Radbel et al., 2020). Inhaled toxicants can induce lung hemorrhage and activation of the coagulation cascade that mediates fibrinolysis, exacerbating ALI/ARDS. Coagulation is elevated in ammonia and chlorine models (Zarogiannis et al., 2014; Elfsmark et al., 2022), and extravascular coagulation and impaired fibrinolysis is pronounced in SM injury (White et al., 2016). In phosgene models, hemolysis destroys red blood cells, which obstructs the pulmonary capillaries (Aggarwal et al., 2019).

Challenges to Medical Countermeasures Development

The development of intervention strategies to counteract acute toxicant exposures presents innumerable challenges. There are ample opportunities to advance our fundamental mechanistic understanding of toxicant-induced ALI and delayed lung effects such as pulmonary fibrosis. As discussed earlier, the resultant lung injury involves endothelial and epithelial damage, alveolar permeability dysfunction, and dysregulated lung inflammation, which reduce lung compliance and compromise respiration. An immediate challenge in mass casualty scenarios is to develop strategies to protect survivors without prior knowledge of the inhaled toxicant. In addition, animal studies using pregnant dams and neonates have shown that vulnerable subgroups are highly susceptible to pulmonary toxicants and may require population-specific care (Addis et al., 2020, 2021).

Immediate treatment of acute exposures involves personal protective equipment for first responders, removal of victims from the area, and decontamination. As there are few antidotes for inhaled toxicants, therapy largely depends on supportive care, with administration of warm, humidified oxygen, antitussives, and bronchodilators (Walker et al., 2015). In some instances, survivors with compromised airways need endotracheal intubation and low tidal volume mechanical ventilation. Those with severe epithelial injury and sloughing may require bronchoscopic lavage to maintain airflow. Inhaled and systemic corticosteroids can be used to reduce inflammation; however, demonstrations of efficacy are limited to small, uncontrolled studies from other airway diseases. Follow-on clinical therapies are based on long-term outcomes (e.g., ALI/ARDS, BO, RADS, idiopathic pulmonary fibrosis).

To date, several laboratories have explored therapeutic approaches to reduce toxicant-induced ALI using antioxidants, angiotensin-converting enzyme inhibitors, neutrophil elastase inhibitors, anti–tumor necrosis factor α therapy, β-agonists, heparin, and phosphodiesterase inhibitors, and some combinations of these drugs have shown limited success. However, the high attrition rate of ARDS drugs may be due to a missed therapeutic window between ALI development and progression to full-blown ARDS. The translation of findings from other lung disease models (e.g., lipopolysaccharide, bleomycin) to high-dose CWA exposures that involve rapid transition of ALI to ARDS is especially difficult.

Lung injury intensity is determined by multiple factors, including proximity to the exposure epicenter, duration, and population demographics (e.g., age, sex, preexisting disease status). The National Institutes of Health Chemical Countermeasures Research Program supports the development of therapeutic agents that can be administered in the field to treat ALI and in trauma/critical care centers to reduce the potential for delayed onset lung complications as well as uncontrolled pulmonary edema from low dose exposure. Current research efforts have focused on understanding the acute pathophysiological consequences of exposure to a limited number of CWAs and industrial toxicants. Given the number of potential CoCs, it is prohibitive to develop a countermeasure specific to each chemical.

Instead, research efforts toward medical countermeasure development should focus on identifying common molecular, cellular, and pathophysiological pathways involved in toxicant-induced ALI using available in vitro, in silico, and organoid systems as well as animal models. Ideally, genomic, epigenomic, and metabolomic approaches could be used to establish kinetic models that demonstrate the evolution and progression of disease and fully characterize specific cell types involved throughout the process. Defining mechanisms of ALI after industrial and agricultural toxicant exposures would be beneficial as we have limited understanding of their molecular pathophysiology. High-throughput screens could accelerate the identification of potential therapeutic targets that counteract acute and delayed effects from diverse toxicant insults.

There is compelling need for better mechanistic understanding of the temporal etiology of lung injury following exposure. Advances in the field are largely driven by findings in animal models and are complicated by variability due to species- and strain-specific differences and duration and routes of exposure. There is a renewed need to develop and refine animal models of exposure and to increase the efficacy and specificity of drug candidates. Significant challenges exist due to ethical constraints on human research and testing. Longitudinal population studies that follow large-scale chemical accidents could delineate long-term effects of toxicant exposures, and appropriate global infrastructure is needed to support these efforts. Ultimately, these comprehensive research efforts will further our understanding of toxicant-induced lung injury progression and support the development of countermeasures against pulmonary threats.

Abbreviations

AHR

airway hyperresponsiveness

ALI

acute lung injury

ARDS

acute respiratory distress syndrome

BO

bronchiolitis obliterans

CG

phosgene

CLDN5

claudin 5

CoC

chemical of concern

COPD

chronic obstructive pulmonary disease

CWA

chemical warfare agent

IL

interleukin

ILD

interstitial lung disease

NO

nitric oxide

RADS

reactive airways disease

SM

sulfur mustard

TRP

transient receptor potential

Authorship Contributions

Wrote or contributed to the writing of the manuscript: Marzec, Nadadur.