The present study was aimed to determine whether Th2 cytokine IL-4 and its soluble receptor IL-4R-alpha take part in the fibrogenic process seen in chronic EAA, which is thought to exhibit a Th2 cytokine milieu. Data obtained from chronic EAA patients were compared to sarcoidosis patients. Sarcoidosis is considered to be a typical Th1 disease, moreover studies of IL-4R gene polymorphisms have showed no association with sarcoidosis, and no differences were found in IL-4 BALF concentrations from sarcoidosis patients compared to healthy controls [10, 11].
In this study the patients groups differed in age, and this is not surprising as the usual age of sarcoidosis manifestation is < 40 years. None of chronic EAA patients were considered professional EAA, and it could be related to lower age .
Initially, BALF concentrations of IL-4 and IL-4R-alpha were compared in chronic EAA and sarcoidosis patients. Even though the difference was not statistically significant, higher IL-4 concentrations were observed in EAA group. Previous studies have documented the role of IL-4 as well as interferon gamma in the pathogenesis of EAA, both cytokines being produced by non-immune cells . The effect of IL-4 on fibrosis development in EAA patients varies form study to study. Some authors even documented a remarkable improvement in murine EAA after IL-4 administration, while others did not find such effect for this cytokine [14, 15]. Even though IL-4 augments the effects of fibroblasts in in vitro studies, its contribution to fibro-proliferation in EAA patients has not been studied. Nevertheless studies performed in IPF patients have shown a clear role for IL-4 in fibrogenesis; distinct polymorphisms of the gene for IL-4 have been associated with more severe radiological involvement [16, 17]. Because skewing of the immune response from Th1 in the reversible stages of the disease to Th2 in the later fibrotic stages has been observed, we might think that the role of IL-4/IL-4R could fluctuate according to the manifestation of the disease. Chronic EAA may share radiologic as well as histological features of usual interstitial pneumonia and perhaps IL-4 can play a similar role in IPF and chronic EAA. According to the studies by Pechkovsky et al., IL-4 is able to induce further expression of IL-4R, which together with the above mentioned findings may suggest a link to higher concentrations of IL-4, resulting in higher concentrations of IL-4R-alpha and more pronounced fibro-proliferation .
More severe impairment of Dlco (p < 0.05), significantly higher BALF total protein concentrations and IL-4R-alpha (p < 0.05) were accompanied by non-significantly higher HRCT interstitial scores in chronic EAA patients. We can speculate that BALF soluble IL-4R-alpha concentrations perhaps could be an even more sensitive marker of fibrosis than the more commonly mentioned HRCT [19, 20].
The second aim of this study was to determine if an association between IL-4/IL-4R-alpha and EAA etiopathogenesis could be supported by a correlation between BALF concentrations and the clinical presentation of the disease (i.e., BALF differential cell count, pulmonary function tests and HRCT scores). Because there were only six patients in the sarcoidosis group, correlations were not performed for this group. A positive correlation between HRCT interstitial scores and IL-4R-alpha BALF concentrations was observed, as well as a negative correlation relative to FVC, FEV1,Dlco and IL-4R-alpha BALF concentrations. The relevance of this finding was supported by the same results of IL-4R-alpha adjusted for BALF total protein concentrations. These observations might suggest that IL-4R-alpha acts like a carrier of biological functions of IL-4 in chronic EAA with HRCT signs of fibrotic lung impairment.
BALF total protein concentration in EAA patients with a history of smoking was significantly higher than in non-smoking patients. This findings agree with the results of Sato et al., even though their study measured albumin concentrations . Others have found lower concentrations of proteins and mRNAs in smoking subjects [22, 23]. This discrepancy may be due to the fact that, while acute exposure to cigarette smoke reduces blood-airway permeability, chronic exposure to smoke may disrupt the endothelial barrier integrity and thus influence endothelial permeability [24, 25]. Thus a history of smoking could be an important factor influencing concentrations of various proteins in BALF.
It can be objected that smoking (or exposure to nicotine) was thought to be a protective factor for granulomatous involvement in EAA patients . However, Furuiye et al. showed that long term smoking may lead to more pronounced fibrogenesis even in EAA subjects, probably because cigarette smoke is composed by many substances, some of which could possibly have pro-fibrotic effects. It also should be kept in mind that smoking is a risk factor for other interstitial lung diseases including IPF and rheumatoid arthritis associated interstitial lung disease .
In our study group BALF IL-4R-alpha concentrations were found to be higher in men with EAA than in women. There is no consistent information concerning gender effect on EAA manifestation, perhaps because most studies found in literature concern special types of EAA (farmer’s lung, pigeon breeder’s lung, etc.). On the other hand, interstitial lung diseases, like IPF, dominate in the male population. Of course, smoking status has to be considered, and smoking is still more common in men than in women in most countries including the Czech Republic. However, the effect of gender on immune reactions has been reported by many authors. Pinzan et al. observed different immunological reactions in men and women infected by Paracoccidioidesbrasiliensis infection . According to results of this study, men who were more often infected presented with a dominant Th2 immune response, while women exhibited a Th1 immune response. Different immune responses are usually explained by the influence of estrogen or testosterone on different T cell populations, resulting in different cytokine milieu [29, 30]. Murine studies have shown that not only gender of infected mice, but also IL-4 alpha status can influence outcomes of some infectious diseases . As mentioned above, higher BALF concentrations were associated with more severe radiological and pulmonary function test deterioration. It is possible that differences in IL-4R-alpha expression between males and females are part of different manifestations of the condition and perhaps outcomes of some interstitial lung diseases.
In our opinion the present study has several limitations. First of all, the number of enrolled patients was small, however the study was performed prospectively and consecutive patients suspected of interstitial lung disease were enrolled, among which we finally chose those with a definite diagnosis of chronic EAA or sarcoidosis. We assessed the concentrations of IL-4R-alpha, which is not only part of IL-4R, but also of the IL-13 receptor. Nevertheless, the soluble form of IL-13R was formed by IL-13R-alpha-2 and IL-4R-alpha may be part of a membrane bound IL-13R.