Mechanisms of isocyanate sensitisation. An in vitro approach
Introduction
Isocyanates are widely used in industry in the production of paints and polyurethanes and are one of the most common inducers of occupational asthma. There is an abundance of clinical evidence which suggests that inhalation of isocyanates can cause asthma, but these findings often do not correlate with the presence of specific IgE antibodies to isocyanate (Kennedy and Brown, 1992). A recent review (Tee et al., 1998) reported that the specificity of the RAST analysis for specific IgE antibodies to isocyanates is often 100%, although the maximum reported sensitivity was 39%, suggesting that specific IgE to isocyanates is a more specific than sensitive index of occupational asthma. It is presumed that isocyanates combine with body proteins to form isocyanate protein conjugates (Kochman et al., 1990), but Baur et al. (1996) suggested that the hydrolysis products of isocyanates in an aqueous solution (namely monomeric or oligomeric diamines) may contribute to the induction of respiratory health effects. Studies performed on patients with isocyanate induced asthma have also demonstrated increased numbers of eosinophils, T-cells (Redlich et al., 1996) and airway inflammation which may also contribute to the aetiology of symptoms described by exposed individuals.
Isocyanates may also have a pharmacological mode of action, since they have been shown to reduce the ability of beta-adrenergic receptors to produce cyclic adenosine monophosphate interfering with the maintenance of bronchial tone (Mapp et al., 1988). In vitro studies (Wass and Belin, 1989) have also demonstrated that toluene diisocyanate affects prostaglandin receptors which can result in hyper-reactivity of smooth muscle.
Previous research by this laboratory has demonstrated that colophony, a complex mix of low molecular weight chemicals (LMWC), induced an oxidative burst response in HL60 cells that had been differentiated into monocyte- and neutrophil-like cell types (Elms et al., 2000), which suggested that the inhalation of colophony could lead to the production of ROS, contributing to tissue damage in the lung. A further consequence of the induction of oxidative burst, specifically the production of hydrogen peroxide has shown to be the increase in ICAM-1 expression, which has been demonstrated in vitro using endothelial cell lines (Roebuck, Rahman, Lakshminarayanan, Janakidevi, & Malik, 1995, Lakshminarayanan, Beno, Costa, & Roebuck, 1997).
In this study, the isocyanates toluene diisocyanate (TDI), diphenylmethane diisocyanate (HDI) and hexamethylene diisocyanate (HDI) were dissolved in phosphate buffered saline to mimic the physiological conditions of the lung. The potential of the isocyanates and their corresponding amines to induce an increase in peroxide levels and ICAM-1 expression in the monocytic cell line mono-mac-6 was investigated.
Section snippets
Materials and methods
Unless stated otherwise, all reagents and chemicals were purchased from Sigma (Dorset, UK).
Challenge with isocyanates
Treated and untreated cell lines exhibited >85% viability when challenged with isocyanates, amines or glycerol at 10 mm for up to 18 h.
Endpoints—measurement of oxidative burst
An initial time course experiment performed in singlet, demonstrated that following challenge with MDI, intracellular peroxide levels rose between a 5-min and 20-min incubation period (data not included); therefore for subsequent challenge assays 5-min and 20-min time points were used. Peroxide levels were significantly increased (P<0.05) in mono-mac-6 cells
Discussion
Isocyanates are often used in the production of paints and polyurethanes, and are one of the most commonly reported inducers of occupational asthma (Meredith et al., 1991). Although there is an abundance of clinical evidence to support this observation, the mechanistic basis for this is not well defined.
In this study, we have demonstrated a significant increase in the levels of intracellular peroxide in a monocytic cell line challenged with the isocyanates TDI and MDI, above cells challenged
References (13)
- et al.
Toluene diisocyanate-induced conformational changes of serum albumina study on repeated inhalation in guinea-pigs
Toxicology Letters
(1990) - et al.
Differential regulation of interleukin-8 and intercellular adhesion molecule 1 by H2O2 and tumour necrosis factor-α in endothelial and epithelial cells
Journal of Biological Chemistry
(1997) - et al.
Immunologic responses to isocyanates in sensitised asthmatic subjects
Chest
(1996) - et al.
H2O2 and tumor necrosis factor-α activate Intercellular adhesion molecule 1 (ICAM-1) gene transcription through distinct cis-regulatory elements within the ICAM-1 promoter
Journal of Biological Chemistry
(1995) - et al.
Specific IgE to isocyanates: a useful diagnostic role in occupational asthma
Journal of Allergy and Clinical Immunology
(1998) - et al.
Immunologic specificity of isocyanate-induced IgE antibodies in serum from sensitised workers
Journal of Allergy and Clinical Immunology
(1989)
Cited by (33)
Toxicological analysis of limonene reaction products using an in vitro exposure system
2013, Toxicology in VitroCitation Excerpt :Different models can be utilized depending on the health effect of interest (Verstraelen et al., 2008a). Inflammation and irritation of the lower respiratory tract is often evaluated in bronchial epithelial cells (NHBE, BEAS-2B) (Pichavant et al., 2005; Persoz et al., 2012) or alveolar epithelial cells (A549) (Krakauer, 2000) while respiratory sensitization is often evaluated in monocyte/macrophage (Mono-Mac-6, THP-1) cell lines (Elms et al., 2001; Verstraelen et al., 2008b). Other advances in the field also include the use of primary cell lines and the development of highly differentiated three dimensional human airway tissue samples, such as (EpiAirway™ Tissue Model (Mattek, Ashland, MA) and MucilAir™ Epithelix (Geneva, Switzerland).
Serum ferritin and transferrin levels as serologic markers of methylene diphenyl diisocyanate-induced occupational asthma
2008, Journal of Allergy and Clinical ImmunologyCitation Excerpt :Increased epithelial production of reactive oxygen species is accepted as a major pathogenic mechanism of isocyanate-induced OA. It was previously shown in vitro study that exposure to isocyanates, including TDI, hexamethylene diisocyanate, and MDI, increased the level of intracellular peroxide,34 whereas in vivo antioxidants were found to decrease TDI-induced airway inflammation.35 On the basis of these findings, reactive oxygen species production may be one pathogenic mechanism of MDI-OA.
Guidelines for occupational asthma
2006, Archivos de BronconeumologiaFerrocenoyl piperazide as derivatizing agent for the analysis of isocyanates and related compounds using liquid chromatography/electrochemistry/ mass spectrometry (LC/EC/MS)
2004, Journal of the American Society for Mass Spectrometry