Objective: To explore the changes in the autophagy marker microtubule-associated protein 1 light chain 3 (LC3) and yeast autophagy-related gene 6 (Beclin1) in rat lungs exposed to free silica (SiO(2)) dust for different periods. Methods: A total of 72 male specific pathogen-free Wistar rats were randomly divided into solvent control group and SiO(2) model group. The SiO(2) model group received one-time non-exposed intratracheal instillation of suspension of SiO(2) particles to establish a model of silicosis. The solvent control group received an equal amount of saline. Six rats each were sacrificed at 1, 7, 14, 21, 28, and 60 days after model establishment. The pathological changes and fibrosis of rat lungs at different time points were evaluated by H&E staining and Masson staining, respectively. Enzyme-linked immunosorbent assay (ELISA) was used to determine the levels of transforming growth factor-β (TGF-β) , interleukin-1 (IL-1) , and tumor necrosis factor-α (TNF-α) in lung tissue homogenate. Western blot was used to determine the relative expression levels of LC3 and Beclin1 in the lung tissue. Results: The results of H&E staining showed that the model group had continuous inflammation in the lung tissue from day 1 to day 60, and the inflammatory scores were significantly higher in the model group than in the control group (P<0.05) . The results of Masson staining showed that rats in the model group had a small amount of collagen fibers in the lung tissue on day 14 and a large amount of collagen fibers on day 60. The fibrosis score was significantly higher in the model group than in the control group (P<0.05) . No collagen fibrosis was observed in the lung tissue in the control group. The results of ELISA showed that the model group had significantly higher levels of IL-1, TNF-α, and TGF-β in lung tissue homogenate than the control group at each time point after exposure (P<0.05) . The results of Western blot showed that the model group had decreased expression of Beclin1 protein in the lung tissue on days 7, 14, 21, 28, and 60, which was significantly higher than that of the control group (P<0.05) . The model group also had a decreased ratio of LC3II/LC3I on days 1, 7, 14, 21, 28, and 60, which were significantly higher than that of the control group (P<0.05) . Conclusion: In the rat model of silicosis induced by free SiO(2) dust, the expression levels of autophagy-related proteins, LC3 and Beclin1, are correlated with different stages of silicosis. In the early stage of silicosis, the lung tissue has inflammation, substantially increased ratio of LC3II/LC3I and expression of Beclin1, and active autophagy. With the progression of silicosis, the ratio of LC3II/LC3I and expression level of Beclin1 gradually decrease and autophagy becomes weak.
目的： 探讨游离二氧化硅（SiO(2)）粉尘暴露大鼠肺组织不同时间点自噬标志物微管相关蛋白1轻链3（Microtubules associated protein 1 light chain 3，LC3）和酵母自噬相关基因6（Beclin1）的变化。 方法： 将72只无特定病原体级（SPF）Wistar雄性大鼠随机分为溶剂对照组和SiO(2)模型组，模型组大鼠采用非暴露式气管滴注法一次性给予SiO(2)悬浮液建立矽肺模型，溶剂对照组大鼠给予等量生理盐水。造模后两组分别于1、7、14、21、28、60 d处死6只大鼠，用苏木精-伊红（HE）染色和Masson染色观察两组大鼠不同时间点肺组织常规病理变化和纤维化程度；用酶联免疫吸附（ELISA）法检测肺组织匀浆中转化生长因子（TGF）-β、白细胞介素（IL）-1和肿瘤坏死因子（TNF）-α的水平；用Western blot法检测肺组织中自噬相关蛋白LC3和Beclin1的相对表达水平。 结果： HE染色结果显示，模型组大鼠肺组织在第1天至第60天持续存在炎性反应，炎性评分均明显高于对照组，差异有统计学意义（P<0.05）；Masson染色观察结果显示，模型组大鼠肺组织从第14天开始出现少量胶原纤维增生，至第60天可见大量胶原纤维，纤维化评分明显高于对照组，差异有统计学意义（P<0.05）；对照组大鼠肺组织未见胶原纤维增生。ELISA法测定结果显示，模型组大鼠在染尘后各时间点肺组织匀浆中IL-1、TNF-α和TGF-β的含量均明显高于对照组，差异有统计学意义（P<0.05）。Western blot结果显示，模型组大鼠肺组织Beclin1蛋白表达水平在第7、14、21、28和60天时呈下降趋势，均明显高于对照组，差异有统计学意义（P<0.05）；模型组大鼠肺组织LC3II/LC3I的比值在第1、7、14、21、28和60天时呈下降趋势，均明显高于对照组，差异有统计学意义（P<0.05）。 结论： 游离SiO(2)粉尘致大鼠矽肺发病过程中，自噬相关蛋白LC3和Beclin1的表达水平与矽肺的不同发展阶段有关。在大鼠染矽尘早期，肺组织以炎性变化为主；随着染矽尘大鼠纤维化程度的加重。.
Keywords: Autophagy; Fibrosis; Inflammatory response; Silicosis.