Silica dust alters fecal microbiota that contributes silicosis through the lung-gut axis in rats

Abstract

Abstract Background：Silicosis is a lung disease with diffuse nodular pulmonary fibrosis because of long-term inhalation of a large number of free silica dust. Research has been reported that dysbiosis of fecal microbiota was existed in silicosis patients. However, few studies have examined the effects of silica on the intestinal tract. Objective: In this study, we aimed to investigate the change of fecal microbiota, lung and ileum tissues of rats exposed to silica dust and explore the regulatory role of fecal microbiota in rats after silica exposure. Methods: The Wistar male rats were intratracheally instilled with 50 mg/mL silicon dioxide (1 mL per rat). Hematoxylin and eosin (HE), Masson staining, enzyme-linked immunosorbent assay (ELISA) and Western blot were used to exam the pulmonary inflammation and fibrosis in rats. HE, Western blot and Transmission Electron Microscopy (TEM) were used to exam the ileac injury. The 16s rRNA gene sequences, non-targeted metabolomics, transcriptome analysis were used to exam the fecal microbiota, metabolites and ileac mRNAs respectively. Then, we employed fecal microbiota transplantation (FMT) experiment to examine whether fecal microbiota play the important roles in the change of silica-induced pulmonary inflammation, fibrosis and ileum injury. Meanwhile, Pearson correlation tests were used to detect the differential microbiota and metabolites of feces, and mRNAs of ileum on day 56. Results: The results showed that silica exposure resulted in dynamic change of pulmonary inflammation, fibrosis, fecal microbiota dysbiosis and ileum epithelial injury. FMT up-regulated the level of Bifidobacterium, restored the level of tight junction proteins of ileum. Then we found the level of Bifidobacterium was significantly down-regulated on day 56 in silica-exposed rats. Further we mainly predicted 3 potential mechanisms through conjoint analysis and KEGG analysis: (ⅰ) the change of Bifidobacterium may be related to the production of oleoyl 3-carba cyclic phosphatidic acid (3-CCPA) and the expression of Cldn8, which involved in silica-induced pulmonary inflammatory response and ileac barrier function injury; (ⅱ) Silica-induced fecal microecological dysbiosis and inflammatory respond may affect the arginine biosynthesis and utilization of arginine by regulating the level of N-Acetyl-L-Glutamic Acid and Nos2, which effect ileac architectural integrity; and (ⅲ) Silica-induced fecal microecology disorder may induce ileac injury by regulating Ido1, Kynu and Indole-3-ethanol mediated “Tryptophan metabolism” pathway. Discussion: This study provided evidence that silica could alter fecal microbiota which may in turn play an important role in silica-induced pulmonary fibrosis and ileac barrier injury in rats. Three predicted mechanistic pathways deserved further study. The findings may provide a starting roadmap to intervene in the development of silica-induced pulmonary fibrosis.