Molecular Toxicology Key Laboratory of Sichuan Provincial Education Office, West China School of Public Health and West China Fourth Hospital, Sichuan University, Chengdu, Sichuan 610041, China; Department of Toxicology and Pathology, West China School of Public Health and West China Fourth Hospital, Sichuan University, Chengdu, Sichuan 610041, China.
Department of Toxicology and Pathology, West China School of Public Health and West China Fourth Hospital, Sichuan University, Chengdu, Sichuan 610041, China.
Toxicology. 2023 Dec;500:153673. doi: 10.1016/j.tox.2023.153673. Epub 2023 Nov 16.
Silicosis is a severe worldwide occupational hazard, characterized with lung tissue inflammation and irreversible fibrosis caused by crystalline silicon dioxide. As the most common and abundant internal modification of messenger RNAs or noncoding RNAs, N6-methyladenosine (m6A) methylation is dysregulated in the chromic period of silicosis. However, whether m6A modification is involved in the early phase of silica-induced pulmonary inflammation and fibrosis and its specific effector cells remains unknown. In this study, we established a pulmonary inflammation and fibrosis mouse model by silica particles on day 7 and day 28. Then, we examined the global m6A modification level by m6A dot blot and m6A RNA methylation quantification kits. The key m6A regulatory factors were analyzed by RTqPCR, Western blot, and immunohistochemistry (IHC) in normal and silicosis mice. The results showed that the global m6A modification level was upregulated in silicosis lung tissues with the demethylase FTO suppression after silica exposure for 7 days and 28 days. METTL3, METTL14, ALKBH5, and other m6A readers had no obvious differences between the control and silicosis groups. Then, single-cell sequencing analysis revealed that thirteen kinds of cells were recognized in silicosis lung tissues, and the mRNA expression of FTO was downregulated in epithelial cells, endothelial cells, fibroblasts, and monocytes. These results were further confirmed in mouse lung epithelial cells (MLE-12) exposed to silica and in the peripheral blood mononuclear cells of silicosis patients. In conclusion, the high level of global m6A modification in the early stage of silicosis is induced by the downregulation of the demethylase FTO, which may provide a novel target for the diagnosis and treatment of silicosis.
矽肺是一种严重的全球性职业危害，其特征是肺部组织炎症和由结晶二氧化硅引起的不可逆转纤维化。作为信使 RNA 或非编码 RNA 最常见和丰富的内部修饰物，N6-甲基腺苷（m6A）甲基化在矽肺的慢性期失调。然而，m6A 修饰是否参与二氧化硅诱导的肺炎症和纤维化的早期阶段及其特定效应细胞尚不清楚。在这项研究中，我们通过第 7 天和第 28 天的二氧化硅颗粒在小鼠中建立了肺炎症和纤维化模型。然后，我们通过 m6A 点印迹和 m6A RNA 甲基化定量试剂盒检测了全局 m6A 修饰水平。通过 RTqPCR、Western blot 和免疫组织化学（IHC）在正常和矽肺小鼠中分析了关键的 m6A 调节因子。结果表明，在暴露于二氧化硅后 7 天和 28 天，矽肺肺组织中的全局 m6A 修饰水平升高，去甲基酶 FTO 受到抑制。METTL3、METTL14、ALKBH5 和其他 m6A 阅读器在对照组和矽肺组之间没有明显差异。然后，单细胞测序分析显示，在矽肺肺组织中识别出十三种细胞，上皮细胞、内皮细胞、成纤维细胞和单核细胞中的 FTO mRNA 表达下调。这些结果在暴露于二氧化硅的小鼠肺上皮细胞（MLE-12）和矽肺患者的外周血单核细胞中得到进一步证实。总之，矽肺早期高水平的全局 m6A 修饰是由去甲基酶 FTO 的下调诱导的，这可能为矽肺的诊断和治疗提供一个新的靶点。
