背景:矽肺是一种由长期接触二氧化硅粉尘引起的不可逆的肺纤维化疾病,表现为炎症细胞的浸润,促炎细胞因子的过度分泌,和肺弥漫性纤维化。随着疾病的进展,肺功能进一步恶化,导致患者生活质量较差。目前,治疗矽肺的有效药物很少。双环醇(BIC)是广泛用于治疗慢性病毒性肝炎和药物诱导的肝损伤的化合物。虽然最近的研究已经证明了BIC对多个器官的抗纤维化作用,包括肝脏,肺,和肾脏,其对矽肺的治疗益处尚不清楚.在这项研究中,我们建立了大鼠矽肺模型,目的是评估BIC的潜在治疗效果。
方法:我们构建了矽肺大鼠模型,并在损伤后给予BIC。使用带有强制振荡系统的FlexibleVent仪器检测大鼠的肺功能。HE和Masson染色用于评估BIC对二氧化硅诱导的大鼠的影响。RAW264.7细胞巨噬细胞炎症模型,NIH-3T3细胞的成纤维细胞-肌成纤维细胞转变(FMT)模型,体外建立TC-1细胞上皮-间质转化(EMT)模型。并通过WesternBlot分析评估BIC治疗后体内和体外炎症介质和纤维化相关蛋白的水平,RT-PCR,ELISA,和流式细胞术实验。
结果:BIC显著改善了二氧化硅诱导大鼠的肺静态顺应性和呼气和吸气量。此外,BIC减少炎症细胞和细胞因子的数量以及肺胶原沉积,导致矽肺大鼠模型纤维化进展延迟。对潜在分子机制的进一步探索表明,BIC抑制了活化,极化,SiO2诱导RAW264.7巨噬细胞凋亡。此外,BIC抑制SiO2介导的炎症细胞因子IL-1β的分泌,IL-6,TNF-α,和巨噬细胞中的TGF-β1。在体外矽肺模型中,BIC抑制NIH-3T3的FMT以及TC-1的EMT,导致NIH-3T3细胞的增殖和迁移能力降低。对巨噬细胞分泌的细胞因子的进一步研究表明,BIC通过靶向TGF-β1抑制FMT和EMT。值得注意的是,BIC阻断了FMT所需的NIH-3T3细胞中JAK2/STAT3的激活,同时阻止了EMT过程所必需的TC-1细胞中SMAD2/3的磷酸化和核易位。
结论:集体数据表明,BIC阻止FMT和EMT过程,反过来,减少异常胶原沉积。我们的发现首次证明BIC能改善炎性细胞因子的分泌,特别是,TGF-β1,并因此通过TGF-β1规范和非规范途径抑制FMT和EMT,最终导致异常胶原沉积的减少和矽肺的进展缓慢,支持其作为新型治疗剂的潜力。
BACKGROUND: Silicosis is an irreversible fibrotic disease of the lung caused by chronic exposure to silica dust, which manifests as infiltration of inflammatory cells, excessive secretion of pro-inflammatory cytokines, and pulmonary diffuse fibrosis. As the disease progresses, lung function further deteriorates, leading to poorer quality of life of patients. Currently, few effective drugs are available for the treatment of silicosis. Bicyclol (BIC) is a compound widely employed to treat chronic viral hepatitis and drug-induced liver injury. While recent studies have demonstrated anti-fibrosis effects of BIC on multiple organs, including liver, lung, and kidney, its therapeutic benefit against silicosis remains unclear. In this study, we established a rat model of silicosis, with the aim of evaluating the potential therapeutic effects of BIC.
METHODS: We constructed a silicotic rat model and administered BIC after injury. The FlexiVent instrument with a forced oscillation system was used to detect the pulmonary function of rats. HE and Masson staining were used to assess the effect of BIC on silica-induced rats. Macrophages-inflammatory model of RAW264.7 cells, fibroblast-myofibroblast transition (FMT) model of NIH-3T3 cells, and epithelial-mesenchymal transition (EMT) model of TC-1 cells were established in vitro. And the levels of inflammatory mediators and fibrosis-related proteins were evaluated in vivo and in vitro after BIC treatment by Western Blot analysis, RT-PCR, ELISA, and flow cytometry experiments.
RESULTS: BIC significantly improved static compliance of lung and expiratory and inspiratory capacity of silica-induced rats. Moreover, BIC reduced number of inflammatory cells and cytokines as well as collagen deposition in lungs, leading to delayed fibrosis progression in the silicosis rat model. Further exploration of the underlying molecular mechanisms revealed that BIC suppressed the activation, polarization, and apoptosis of RAW264.7 macrophages induced by SiO2. Additionally, BIC inhibited SiO2-mediated secretion of the inflammatory cytokines IL-1β, IL-6, TNF-α, and TGF-β1 in macrophages. BIC inhibited FMT of NIH-3T3 as well as EMT of TC-1 in the in vitro silicosis model, resulting in reduced proliferation and migration capability of NIH-3T3 cells. Further investigation of the cytokines secreted by macrophages revealed suppression of both FMT and EMT by BIC through targeting of TGF-β1. Notably, BIC blocked the activation of JAK2/STAT3 in NIH-3T3 cells required for FMT while preventing both phosphorylation and nuclear translocation of SMAD2/3 in TC-1 cells necessary for the EMT process.
CONCLUSIONS: The collective data suggest that BIC prevents both FMT and EMT processes, in turn, reducing aberrant collagen deposition. Our findings demonstrate for the first time that BIC ameliorates inflammatory cytokine secretion, in particular, TGF-β1, and consequently inhibits FMT and EMT via TGF-β1 canonical and non-canonical pathways, ultimately resulting in reduction of aberrant collagen deposition and slower progression of silicosis, supporting its potential as a novel therapeutic agent.