清肺口服液缓解放射性肺纤维化肺泡上皮细胞焦亡的作用及机制研究

The Role and Underlying Mechanisms of Qingfei Oral Liquid in Alleviating Pyroptosis of Alveolar Epithelial Cells in Radiopulmonary Fibrosis

关键词:  清肺口服液  放射性肺纤维  肺泡上皮细胞焦亡  肺成纤维细胞活化

Key words:  Qingfei oral liquid  radiation-induced pulmonary fibrosis  alveolar epithelial cell pyroptosis  pulmonary fibroblast activation

摘要:

目的 明确清肺口服液对放射性肺纤维化(radiation-induced pulmonary fibrosis, RIPF)的治疗作用,并探究其潜在作用机制。方法 SD大鼠随机均分成对照组、模型组、地塞米松组和清肺口服液组。大鼠接受胸部放射构建RIPF模型。地塞米松组和清肺口服液组大鼠分别每日灌胃给予地塞米松和清肺口服液。8周后,核磁共振检测大鼠肺纤维化,苏木素-伊红、马森染色观察肺组织形态和胶原沉积,Elisa检测肺泡灌洗液中TGF-β1、血清中丙二醛和超氧化物歧化酶量,试剂盒检测肺组织羟脯氨酸含量。放射处理人肺泡二型上皮细胞(AT2)后给予清肺口服液,CCK-8检测细胞活力,Western blotting和Elisa检测细胞焦亡标志物表达。放射处理AT2后与人胚肺成纤维细胞(MRC-5)和人支气管上皮细胞(Beas-2b)共培养,Western blotting检测成纤维细胞活化和上皮细胞间质化标志物表达。分析清肺口服液入肺组织的有效成分,并结合网络药理学分析清肺口服液治疗RIPF的潜在机制。结果 清肺口服液可显著缓解放射诱导的肺组织炎症、胶原沉积和纤维化,且与地塞米松效果类似；清肺口服液可显著抑制放射诱导的AT2细胞焦亡,减少炎症因子释放；放射处理的AT2细胞可诱导MRC-5活化和Beas-2b间质化；网络药理学分析提示清肺口服液通过TNF和NOD-like receptor等通路抑制放射诱导的细胞焦亡和肺纤维化。结论 清肺口服液通过TNF和NOD-like receptor等通路抑制AT2细胞焦亡进而缓解RIPF。

Abstract:

OBJECTIVE To clarify the therapeutic effect of Qingfei oral liquid on radiation-induced pulmonary fibrosis (RIPF), and explore its potential mechanism of action. METHODS SD rats were randomly divided into control group, model group, dexamethasone group and Qingfei oral liquid group. RIPF model was constructed by radiation exposure. The rats in dexamethasone group and Qingfei oral liquid group were given dexamethasone and Qingfei oral liquid daily by intragastric administration, respectively. After 8 weeks, pulmonary fibrosis of rats was detected by MRI, lung tissue morphology and collagen deposition were observed by Hematoxylin-eosin staining and Masson staining, TGF-β1 in alveolar lavage fluid, malondialdehyde and superoxide dismutase in serum were detected by Elisa, and hydroxyproline content in lung tissue was detected by kit. Human alveolar type II epithelial cells (AT2) were treated with radiation and given Qingfei oral liquid. CCK-8 was used to detect cell viability, Western blotting and Elisa were used to detect pyroptosis markers. After radiation exposure, AT2 was co-cultured with human embryonic lung (MRC-5) and human epithelial cells (Beas-2b), and the activation of fibroblasts and the expression of mesenchymal transition markers were detected by Western blotting. The active components of Qingfei oral liquid in lung tissue were analyzed, and the underlying mechanisms of Qingfei oral liquid in treating RIPF was analyzed by network pharmacology. RESULTS Qingfei oral liquid significantly alleviated radiation-induced inflammation, collagen deposition, and fibrosis in lungs, and the effect is similar to dexamethasone. Qingfei oral liquid significantly inhibited radiation-induced pyroptosis in AT2 and reduced the release of inflammatory factors. Co-culture with radiation-treated AT2 resulted in MRC-5 activation and Beas-2b mesenchymal transition. Network pharmacology analysis suggested that Qingfei oral liquid inhibited radiation-induced pyroptosis and pulmonary fibrosis through TNF and NOD-like receptor pathways. CONCLUSION Qingfei oral liquid inhibited pyroptosis of AT2 cells through TNF and NOD-like receptor pathways and thereby alleviating RIPF.