Validation of a mouse model of chemical-induced asthma using trimellitic anhydride, a respiratory sensitizer, and dinitrochlorobenzene, a dermal sensitizer

doi:10.1016/j.jaci.2006.01.027

Journal of Allergy and Clinical Immunology

Volume 117, Issue 5, May 2006, Pages 1090-1097

https://doi.org/10.1016/j.jaci.2006.01.027 Get rights and content

Background

Occupational asthma can be caused by chemicals. Previously, we established a murine model of immunologically mediated chemical-induced asthma using toluene diisocyanate.

Objective

We sought to verify this model using trimellitic anhydride (TMA), a respiratory sensitizer, and 1-chloro-2,4-dinitrobenzene (DNCB), a dermal sensitizer.

Methods

BALB/c mice received dermal applications (vehicle or chemical) on days 1 and 7. On day 10, they received an intranasal instillation (vehicle or chemical). Whole-body plethysmography (enhanced pause) was used to monitor changes in ventilatory function and methacholine reactivity. Pulmonary inflammation was assessed by using bronchoalveolar lavage (cells, TNF-α levels, and macrophage inflammatory protein 2 levels). Immunologic parameters included total serum IgE levels, lymphocyte distribution in auricular and cervical lymph nodes, and IL-4 and IFN-γ levels in supernatants of lymph node cells incubated with or without concanavalin A.

Results

Mice dermally treated and intranasally challenged with TMA experienced markedly increased enhanced pause immediately after intranasal challenge and increased methacholine reactivity (24 hours later). Mice similarly treated with DNCB did not show any ventilatory changes. Neutrophil influx and increased macrophage inflammatory protein 2 and TNF-α levels were found in bronchoalveolar lavage fluid in both TMA- and DNCB-treated mice. The proportion of CD19⁺ B cells was increased in auricular and cervical lymph nodes of TMA-treated mice. IL-4 and IFN-γ levels were increased in supernatants of concanavalin A–stimulated auricular and cervical lymph node cells of TMA- or DNCB-treated mice; however, the relative proportions of IL-4 and IFN-γ levels differed between TMA- and DNCB-treated mice. Serum total IgE levels were increased in TMA-treated mice only.

Conclusion

Both compounds induce a mixed T_H1-T_H2 response, but only TMA induced ventilatory changes.

Clinical implications

In the workplace avoiding skin contact with chemical sensitizers might be advised to prevent chemical-induced asthma.

Section snippets

Methods

A complete description of the materials and methods used is available in the Online Repository at www.jacionline.org.

TMA and DNCB were dissolved in a vehicle (acetone olive oil [AOO]) consisting of a mixture of 2 volumes of acetone and 3 volumes of olive oil. Concentrations of TMA and DNCB are given as percentages (wt/vol) in AOO.

Male BALB/c mice (6 weeks old) received dermal applications of 20 μL of vehicle or 20 μL of chemical (5% TMA or 0.2% DNCB) on each ear on days 1 and 7. On day 10, they

Ventilatory function

Fig 1 shows the early ventilatory responses immediately after intranasal instillation. Mice that had previously received dermal applications of TMA, either once on day 1 (T/0/T) or twice on days 1 and 7 (T/T/T), showed significant increases in Penh after intranasal instillation with TMA, whereas no significant changes in Penh were observed in the 0/0/T control group.

Mice dermally treated with DNCB did not show significant changes in ventilatory function after challenge with DNCB compared with

Discussion

In this study we demonstrated that intranasal instillation of TMA to dermally sensitized mice led to an early ventilatory response compatible with airway obstruction, increased methacholine responsiveness, and pulmonary inflammation, as we had shown previously with TDI.18, 19 In contrast, dermal application and intranasal instillation of DNCB did not lead to ventilatory changes and only resulted in a low inflammatory response. These ventilatory changes were accompanied by increased production

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With the increasing morbidity and mortality of asthma, asthma aggravated by environmental pollution has drawn more attention. This study investigated the exacerbating effects of trimellitic anhydride (TMA), a typical pollutant, in ovalbumin (OVA)-induced asthmatic mice and the gene and protein expressions of TRPA1, V1, V2 in lung tissue. Female BALB/c mice were respectively administered for 42 days as follow: sensitized and challenged with OVA, sensitized and challenged with TMA, sensitized with OVA and challenged with OVA plus TMA, as well as sensitized and challenged with OVA plus TMA. 24 h after the last challenge, the changes in airway resistance (RI) and lung dynamic compliance (Cdyn) were tested. The levels of the inflammatory cells in blood and bronchoalveolar lavage fluid (BALF) were determined. The gene and protein expressions of TRPA1, V1, V2 in lung tissue were examined, and levels of interleukin (IL)-4, −13, substance P (SP), prostaglandin D₂ (PGD₂), nerve growth factor (NGF) in BALF and the supernatant of lung homogenate were measured. The results indicated that OVA plus TMA significantly increased the amount of inflammatory cells in blood and BALF, enhanced RI while decreased Cdyn, and aggravated lung injury. Increased gene and protein expressions of TRPA1, V1, V2 in lung tissue, level of IL-4 in the supernatant of lung homogenate, levels of IL-13, SP, PGD₂, NGF in BALF and the supernatant of lung homogenate were observed. It was suggested that exacerbating effects of TMA in OVA-induced asthma might be related to the regulation of TRPA1, V1, V2 and relevant neurokines.
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Symptoms of exposure to DNFB include irritation of the skin, the eyes, and the mucous membranes (PubChem, CID = 6264). Although some researchers suggest that nitro derivatives of benzene (e.g. dinitrochlorobenzene-DNCB) are typical dermal irritants, which are not able to induce respiratory sensitization (Dearman and Kimber, 1991; Vanoirbeek et al., 2006), it has been proven that epicutaneous sensitization with DNFB and local airway challenge result in non-atopic asthma response (van Houwelingen et al., 2002). Van der Kleij et al. (2003) developed a model in which mice were dermally sensitized with DNFB and intranasally challenged with water-soluble, cognate compound dinitrobenzene sulphonic acid (DNS).
Asthma is a chronic inflammatory disorder of the respiratory tract that is characterized by reversible airflow obstruction and airway hyperresponsiveness. The non-atopic variant of asthma that appears later in life has no allergic background and is more severe and resistant to standard treatment. Hapten-induced asthma models can be utilized to investigate mechanisms behind the development of non-atopic and occupational asthma, in which non-allergic processes seems to play significant role. The development of adequate animal models of non-allergic asthma is a necessary prerequisite both for understanding the pathophysiology of non-allergic asthma and for the possibility of testing new therapies. Still, there is no ideal model that represents all the hallmarks of this complex disease. In this review, we examine the most popular hapten-induced murine models of occupational and non-atopic asthma. For this reason, we describe the most popular sensitizing haptens, sensitization and challenge protocols, symptoms produced by asthma, and advantages and disadvantages of the models.
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The pathogenesis of atopic dermatitis (AD) involves T helper 2 (Th2) cells, and effective therapies remain elusive due to the paucity of animal models. We aimed to develop a mouse model of an immune system aberration caused by allergen. Experiments were conducted in two phases. In experiment 1, BALB/c mice were sensitized with one of four chemical allergens – toluene diisocyanate (TDI), hexamethylene diisocyanate (HDI), trimellitic anhydride (TMA), or 2,4-dinitrochlorobenzene (DNCB) – for 3 weeks. Based on results of experiment 1, immunological features were compared between TMA-sensitized BALB/c mice and NC/Nga mice, after exposure to mite extracts, harmful chemicals and detergents in experiment 2. Sensitization by allergen caused a large number of pathological changes in the skin, and an increase in mast cell number. TMA-sensitized BALB/c mice models showed higher sensitivity to an environmental allergen than NC/Nga mice did. Overall, the initial sensitization with TMA leads to disturbances in Th2-mediated immunity.
Chemical- and Drug-Induced Asthma
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Each year, approximately 300 million people worldwide are diagnosed with asthma, and more than 250,000 deaths annually are attributed to asthma. However, the sources and mechanisms behind the development of asthma are diverse. High-molecular-weight proteins, including detergent enzymes and common allergens, contribute to an immunoglobulin E (IgE)-mediated atopic asthma response. Low-molecular-weight chemicals typically are haptens that form hapten–protein complexes resulting in IgE-mediated or IgE-independent immune-mediated asthma. Hapten–protein exposures typically occur at the workplace and can include acid anhydrides, isocyanates, as well as others. Irritant-induced asthma typically results from a single, accidental exposure to a highly concentrated irritant in the workplace that induces pulmonary damage by releasing proinflammatory mediators that lead to asthma. Additionally, ingestion of certain medications including aspirin, β-blockers, or angiotensin-converting enzyme inhibitors can cause drug-induced asthma by enzymatic reactions leading to the production of proinflammatory mediators and modulation of smooth muscle function. Overall, the diversity of asthma makes this prevalent lung disease difficult to diagnose and manage. This article compares and contrasts phenotypes, mechanisms, and possible treatments for these differing types of drug- and chemical-induced asthma that are currently understudied.
Methylisothiazolinone: Dermal and respiratory immune responses in mice
2015, Toxicology Letters
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In this model of chemical-induced asthma, applications of typical respiratory sensitizers, such as toluene diisocyanate (TDI) (De Vooght et al., 2010a; Tarkowski et al., 2007), trimellitic anhydride (Vanoirbeek et al., 2006) and ammonium persulfate (De Vooght et al., 2010b), induce increased numbers of T- and B-lymphocytes, increased production of predominantly Th2 (IL-4 and IL-13), but also Th1 cytokines, increased levels of total serum IgE, AHR to methacholine and neutrophil inflammation in BAL 24 h following a respiratory challenge (Vanoirbeek et al., 2004, 2006). Typical contact sensitizers, such as dinitrochlorobenzene, on the other hand, induce only a slight increase in the number of B-lymphocytes, an increased production of predominantly Th1 (IFN-γ), but also Th2 cytokines, but no increased levels of total serum IgE, neutrophil inflammation in BAL, nor ventilatory changes (Vanoirbeek et al., 2006). This indicates again that the cytokine production profile can contribute to the discrimination between respiratory and contact sensitizers (De Jong et al., 2009; Dearman et al., 1995).
Methylisothiazolinone (MI), a widely used chemical preservative in industrial and household products, and cosmetics, has been associated with allergic contact dermatitis. However, the asthmogenic capacity of MI is currently unknown. In this study, we investigated the capacity of MI to elicit asthma-like responses in a validated mouse model. On days 1 and 8, mice (C57Bl/6 and BALB/c) were dermally treated with MI or vehicle on each ear. On day 15, mice received a single intranasal challenge with MI or vehicle. Immediately after the challenge, the early ventilatory response was measured using a double chamber plethysmograph. One day later, airway hyperreactivity, pulmonary inflammation and immune-related parameters were assessed. Dermal treatment with MI in both C57Bl/6 and BALB/c mice induced increased T- and B-cell proliferation in the auricular lymph nodes, along with IFN-γ production and limited increases in total serum IgE, confirming dermal sensitization. An airway challenge with MI led to an early ventilatory response (decreased breathing frequency), indicative for acute sensory irritation. However, 24 h later no allergic respiratory response (no airway hyperreactivity (AHR) nor pulmonary inflammation) was found in either mouse strains. Our study indicates that MI can be classified as a strong dermal sensitizer and irritant, but not an asthmogen after initial dermal sensitization, followed by an airway challenge.
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Treatment with EP dramatically decreased airway hyperresponsiveness in TDI-challenged mice, as well as numbers of neutrophils in BAL fluid and peribronchovascular regions. Both the TDI-induced raised protein level and abnormal distribution of HMGB1 were significantly recovered by EP in a dose-dependent manner. The concentration of MIP-2 in TDI-induced asthma mice was significantly higher than that of the control ones, while EP had few effects on MIP-2. The mRNA expression of CXCR2 didn't change significantly, and TNF-α was not detected in BAL fluids.
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Supported by the Belgian Federal Science Policy Office (PS/01/43) and “FWO-Flanders” (project 7.0024.00).

Disclosure of potential conflict of interest: The authors have declared that they have no conflict of interest.

View full text

Environmental and occupational respiratory disordersValidation of a mouse model of chemical-induced asthma using trimellitic anhydride, a respiratory sensitizer, and dinitrochlorobenzene, a dermal sensitizer

Background

Objective

Methods

Results

Conclusion

Clinical implications

Section snippets

Methods

Ventilatory function

Discussion

J Allergy Clin Immunol

J Invest Dermatol

Toxicology

Toxicology

J Allergy Clin Immunol

Toxicology

Food Chem Toxicol

Fundam Appl Toxicol

Environ Toxicol Pharmacol

Toxicol Appl Pharmacol

Toxicol Appl Pharmacol

Toxicol In Vitro

Occupational allergens

Curr Allergy Asthma Rep

Proceedings of the first Jack Pepys Occupational Asthma Symposium

Am J Respir Crit Care Med

Occupational asthma: a review

Environ Health Perspect

Occupational asthma: “the big picture”

Occup Med

Asthma in the workplace

A chemical dataset for evaluation of alternative approaches to skin-sensitization testing

Contact Dermatitis

The local lymph node assay: past, present and future

Contact Dermatitis

Environmental and occupational respiratory disorders
Validation of a mouse model of chemical-induced asthma using trimellitic anhydride, a respiratory sensitizer, and dinitrochlorobenzene, a dermal sensitizer