Silicosis is a well-known fibrotic lung disease caused by prolonged inhalation of crystalline silica. Crystalline silica entering the airways is engulfed by macrophages, causing necrosis of the phagocytes, and then, the internalized silica is released again and engulfed by other macrophages. The repeated process of phagocytosis, necrosis and rephagocytosis of the cells induces inflammation and activation of the reactive oxygen species system, which is associated with pulmonary interstitial fibrosis [19, 20]. Some metabolites in the pathogenesis of silicosis may play a predictive role in the diagnosis and severity of the disease. In the present study, we evaluated the potential biomarkers in the plasma of patients with silicosis compared with dust-exposed workers without silicosis and healthy controls through pilot and validation metabolomics studies.
In the present study, we found that 20 DMFs, including lipids and carnitines, could distinguish the DEWs from the healthy controls. Lipids are essential nutrients in humans and are the main components of cell membranes and cellular energy storage. Lipids are related to signal transduction, enzyme activation, growth factors and antioxidants, signal recognition and immunity [21–23]. Recently, the role of lipids has attracted increased attention in lung and respiratory diseases, including cystic fibrosis, asthma and chronic obstructive pulmonary disease, which are all associated with abnormal metabolism [24–26]. Researchers have revealed vital information regarding lipid metabolism in IPF patients, and more importantly, a few potentially promising biomarkers were first identified and may have a predictive role in monitoring and diagnosing IPF [17, 27, 28]. Lysophosphatidylcholine (LysoPC), an intermediate metabolite of sphingolipids, was shown to be a potential biomarker in the serum of patients with IPF by pilot and further validation studies using UHPLC-MS [29]. Carnitine is related to the metabolism of fat into energy in the body and can promote the transport and oxidation of fatty acids and the utilization of carbohydrates and amino acids, improve body tolerance, prevent lactic acid accumulation, and delay ageing and is involved in antioxidant activities [30]. Carnitine was significantly decreased in the lung tissue and reduced mitochondrial beta-oxidation in IPF [16]. In the early pathogenesis of silicosis, the enhancement of macrophage phagocytosis, apoptosis, oxidative stress and inflammation is caused by inhaled crystalline silica [31, 32] and is closely related to the abnormal metabolism of lipids and carnitines. This study showed that sphingolipid metabolism was the major metabolic pathway in the DEWs and that LysoPC and lysophosphatidylethanolamine (LysoPE), the intermediate metabolites of sphingolipid metabolism, were upregulated.
Amino acids are the basic components of proteins and the raw materials for protein synthesis, and they participate in the complex metabolic process of the body. Increased levels of some amino acids, including creatine, putrescine, spermidine, 4-hydroxyproline and proline-hydroxyproline dipeptide, were found in the fibrotic lung tissue of patients with IPF compared with normal lung tissue [16]. Our results showed that 3 amino acid DMFs, L-arginine, kynurenine and creatine, could distinguish patients with silicosis from the healthy controls in the validation study. Collagen fibrils are the most abundant protein in the extracellular matrix (ECM), and excess collagen deposition in the ECM is associated with the key pathogenic mechanism of IPF [33]. Ornithine can also be converted to proline and hydroxyproline for collagen formation in fibrosis [17]. Silicosis has similar changes in amino acid metabolism as IPF, which may be related to the fibrotic process [34]. In the late pathogenesis of silicosis, the formation of silicotic nodules is the pathological manifestation of the production of collagen fibres and pulmonary interstitial fibrosis mediated by amino acid metabolism, which is consistent with the finding in the present study that arginine and proline metabolism was the major metabolic pathway in silicosis.
A growing number of studies have suggested that arginine methylation and asymmetric dimethylarginine (ADMA) metabolism may be associated with the progression of IPF [35]. One animal experiment featuring lung fibrosis showed that the direct infusion of ADMA resulted in elevated collagen deposition in mouse lungs and enhanced arginase activity [36]. L-arginine can produce L-ornithine under the action of arginase, and then, through ornithine aminotransferase, L-ornithine converts into proline, which is the main component of collagen [37]. Our study has shown that the plasma level of L-arginine in silicosis was significantly higher than that in the controls and was related to the decline of pulmonary function; thus, this molecule may be a potential biomarker for diagnosing and monitoring the disease. Kynurenine is an intermediate metabolite of tryptophan, an essential amino acid. Kynurenine has immune regulatory functions and can regulate vascular tone, which might be relevant in pulmonary hypertension [38]. We found that compared with that of the controls, the level of kynurenine significantly increased in silicosis. Moreover, the level of kynurenine was higher in Stage III silicosis than in Stage I or Stage II silicosis and was negatively correlated with FVC % predicted. Our unpublished data showed that 30.4% of a cohort of 92 patients with Stage III silicosis had a high or intermediate probability of pulmonary hypertension, which is related to pulmonary artery stenosis resulting from mechanical compression by the lesions of central type progressive massive fibrosis (data not shown). This finding may partially explain the overexpression of kynurenine in severe silicosis. Therefore, kynurenine might be a potential biomarker for diagnosing and monitoring silicosis.
Some limitations of the present study should be mentioned. First, potential enrolment bias existed in the present study and may affect the validity of the results. The study population from a single medical centre may not be fully representative of all patients with silicosis. In addition, more males were enrolled than females because they were at risk of silica dust exposure through engagement in manual labour, such as excavation and digging, polishing and buffing. Second, although the plasma metabolic features of silicosis are accessible and noninvasive, they may not fully represent the metabolic process of the lungs. Further research is warranted to explore the distinct metabolism of sputum as well as bronchoalveolar lavage in patients with silicosis. Third, despite our analysis of distinct metabolic features in silicosis, a subset of progressive fibrotic interstitial lung disease [39], it is still unknown whether plasma metabolic biomarkers in silicosis are observed in various other fibrotic diseases, such as IPF, connective tissue disease-related interstitial lung diseases, chronic hypersensitivity pneumonitis and sarcoidosis. Finally, given the cross-sectional design, the study did not have the power to explore the potential metabolic biomarkers for disease progression and survival, which are clinically significant.