In this study, higher autoantibody levels for the three epithelial CKs (CK8, CK18, and CK19) were detected in serum samples from patients with TDI-induced asthma than in asymptomatic exposed workers, in patients with allergic asthma and healthy unexposed control subjects. Among the three autoantibodies, those to CK18 and CK19 showed the highest sensitivities. Furthermore, the prevalence CK18 and CK19 autoantibodies were significantly higher in patients with TDI-induced asthma than in asymptomatic exposed workers, and their binding specificities were confirmed by ELISA inhibition tests. Based on the prevalence of anti-TDI-HSA antibodies observed in this study, we suggest that serum CK18 and CK19 autoantibodies might be used as serological markers for identifying patients with TDI-induced asthma among exposed workers, although the sensitivity was not high enough (26.2%).
Previous studies on the long-term prognosis of TDI-induced asthma suggest that early diagnosis of sensitized patients and immediate removal from the exposure could increase the likelihood of remission. However, an appreciable number of patients with TDI-induced asthma do not recover completely, even after avoidance and treatment.3,4,7
Therefore, developing an early diagnostic marker based on serologic tests is essential in TDI-induced asthma. Although monocyte chemoattractant protein-1 (MCP-1) could be a sensitive test to identify diisocyanate (TDI, MDI, or HDI) induced asthma,8
this is not easy to repeat in a larger sample. Several investigators have detected anti-TDI-HSA IgE and IgG antibodies in serum samples from patients with TDI-induced asthma, using ELISA or RAST. The prevalence of anti-TDI-HSA IgE antibodies has proven quite variable between laboratories, and sensitivity to IgG was not high enough.4,9
This study is the first to evaluate whether autoantibodies to major bronchial epithelial CKs can be applied to the identification of asthma patients exposed to a single type of isocyanate (patients with TDI-induced asthma vs. TDI-exposed, but asymptomatic workers), and sensitivity, specificity, positive and negative predictive value were compared with those of anti-TDI-HSA IgE and IgG antibodies. The sensitivities and positive predictive values of CK18 and CK19 autoantibodies were higher than those of anti-TDI-HSA IgE and IgG antibodies, while the specificities, negative predictive values, and test efficiencies were similar. Significant associations were not found between the prevalence of specific IgE and IgG antibodies. These findings suggest that autoantibodies to CK18 and CK19 may be used as a supplementary serologic markers to increase the diagnostic sensitivities of serum-specific antibody tests.
Several airway epithelial cell proteins can conjugate with diisocynates in vitro and in vivo
CKs, major intracytoplasmic cytoskeleton proteins, are potential self-antigens, with a total of 20 human epithelial keratins, based on molecular weight and isoelectric point. Pairs of CKs seem to be consistently coexpressed in different types of epithelial cells.13
The three CKs applied in this study have been found in both bronchial and lung alveolar epithelial cells, which are the major target tissues of bronchial asthma. The prevalence of specific CK18 and CK19 autoantibodies were the same with similar inhibition potencies, according to the ELISA inhibition test.
CKs are normally located in the intracellular space and could gain access to the immune system after epithelial damage or cell death. The precise mechanism of their generation is still unknown, although several studies suggest that CKs are proteolyzed during cell apoptosis and can leak into the circulation as a soluble form, where they may serve as new epitopes for antibody generation.14-17
In this study, high levels of specific CK autoantibodies were detected only in patients with TDI-induced asthma, not in those with allergic asthma. Therefore, these findings may not be derived from simple epithelial damage found in asthmatic airways. Among the patients with TDI-induced asthma, those who are more susceptible to epithelial damage by TDI exposure may develop serum-specific CK autoantibodies. In this study, because we found no association between these antibodies and the clinical characteristics of TDI-induced asthma, including asthma symptoms and duration of exposure, we could not speculate as to which patients would be susceptible. However, airway hyper-responsiveness to methacholine was more severe in patients with autoantibodies to any of the three CKs, despite marginal statistical significance.
Further studies are needed to investigate genetically susceptible markers for developing autoantibodies to specific CKs among workers exposed to TDI. How diisocyantes and human airway epithelial cells are involved in airway inflammation has not been studied fully.18
HDI, conjugated with bronchial CK18, can induce T-cell proliferation and cytokine production in patients with diisocyanate asthma.9,18
Further studies are needed to confirm whether the development of autoantibodies to specific CKs is an epiphenomenon of chronic airway damage occurring in patients with TDI-induced asthma or whether pathogenic or autoantibody-mediated mechanisms are involved.
In conclusion, we demonstrated circulating autoantibodies to three bronchial epithelial CKs (8, 18, and 19) in patients with TDI-induced asthma. Among these, autoantibodies to CK18 and 19 may be used as serological markers for the detection of TDI-induced asthma among workers with high-risk exposure. Further studies will be needed to investigate how these specific autoantibodies may be involved in the chronic, persistent airway inflammation found in TDI-induced asthma.