两种矽肺大鼠模型肺组织炎性因子和氧化应激指标变化分析

Analysis of changes in inflammatory factors and oxidative stress indicators in lung tissue of two silicosis rat models

关键词: 矽肺  一次性气管灌注  口鼻吸入  炎性反应  氧化应激

Keywords: silicosis  single tracheal perfusion  oronasal inhalation  inflammatory response  oxidative stress

摘 要: 目的 比较口鼻吸入法和一次性气管灌注法建立的矽肺大鼠模型肺组织病理改变、炎性因子和氧化应激指标的差异,为矽肺模型选择提供依据。方法 60 只雄性 Wistar 大鼠随机分为对照组、一次性气管灌注组(100 mg/ml SiO₂ ,1 ml/只)和口鼻吸入组(1 000 mg/m³ SiO₂,6 h/d, 连续吸入 28 d,饲养至 90 d);分别于第 14、28、56 和 90 天取左上肺与右上肺组织, 采用 HE 和 Masson 染色方法评估肺组织病理改变和纤维化程度;通过酶联免疫吸附(ELISA)方法检测左上肺肺组织匀浆中炎性因子白细胞介素(IL)-8、IL-18 和肿瘤坏死因子(TNF)-α 及氧化应激指标过氧化氢酶(CAT)、超氧化物歧化酶(SOD)、丙二醛(MDA)、还原型谷胱甘肽(GSH)和谷胱甘肽过氧化物酶(GSH-Px)表达水平。结果 吸入组和灌注组大鼠左上肺炎症出现时间相近,均随时间延长而加重,灌注组矽肺大鼠纤维化出现早、程度高,但仅吸入组右上肺出现炎症和纤维化, 黏液分泌增加。在 28 d 时吸入组 IL-8(232 pg/ml)、TNF-α(41 pg/ml)、CAT(64 U/ mg prot)、MDA(2.0 nmol mg prot) 和 GSH-Px (2 655 μmol/L) 均分别显著高于灌注组(47 pg/ml,31 pg/ml,53 U/ mg prot,1.6 nmol/mg prot,1 730 μmol/L);56 d 和 90 d 时,吸入组炎性因子 IL-8、IL-18、TNF-α,以及氧化应激指标 CAT、MDA 和 GSH-Px 水平也均显著高于灌注组(P<0.05),而在上述时间吸入组 SOD 水平均显著低于灌注组(P<0.001)。提示灌注组早期氧化应激反应更剧烈,但吸入组整体激活程度更高,累积效应更显著。结论 一次性气管灌注法适用于矽肺早期急性氧化应激反应、快速炎症激活及对应药物研究;口鼻吸入法更适用于炎性因子长期累积效应以及慢性药物干预研究。

Abstract: Objective To compare the differences in lung histopathology, inflammatory factors, and oxidative stress indicators between the silicosis rat models established by oronasal inhalation and one-time tracheal perfusion methods, thereby provide a basis for the selection of silicosis rat models. Methods Sixty male Wistar rats were randomly divided into control group, single intratracheal instillation group (100 mg/ml SiO₂, 1 ml per rat), and oronasal inhalation group (1 000 mg/m³ SiO₂, 6 hours per day, continuous inhalation for 28 days, and feeding until 90 days); on the 14th, 28th, 56th and 90th days, the left upper lung and right upper lung were taken for HE and Masson staining to evaluate the pathological changes and degree of pulmonary fibrosis in the lung tissue, and the expression levels of inflammatory factors (IL-8, IL-18, and TNF-α) and oxidative stress markers (CAT, SOD, MDA, GSH, and GSH-Px ) in the left upper lung tissue homogenates were measured by enzyme-linked immunosorbent assay (ELISA). Results The results showed that the onset time of pulmonary inflammation in the left upper lobe was similar between the inhalation and perfusion groups, and the condition worsened over time in both. However, the fibrosis of silicosis rats in the perfusion group appeared earlier and to a higher degree, but only inflammation and fibrosis occurred in the right upper lung of the inhalation group, with increased mucus secretion. At 28 days, the levels of IL-8 (232 pg/ml), TNF-α (41 pg/ml), CAT (64 U/mg prot), MDA (2.0 nmol/mg prot), and GSH-Px (2 655 μmol/L) in the inhalation group were significantly higher than those in the perfusion group (47 pg/ml, 31 pg/ml, 53 U/mg prot, 1.6 nmol/mg prot, 1 730 μmol/L); At 56 and 90 days, the levels of IL-8, IL-18, TNF-α, CAT, MDA, and GSH-Px in the inhalation group were significantly higher than those in the perfusion group (P<0.05). At above-mentioned time, the SOD levels in the inhalation group were significantly lower than those in the perfusion group (P<0.001), indicating that the early oxidative stress in the perfusion group was more severe, but the overall activation level in the inhalation group was higher, and the cumulative effect was more significant. Conclusion The single intratracheal perfusion method is suitable for the study for early acute oxidative stress responses, rapid inflammatory activation, and corresponding rapid effect drug, while the oronasal inhalation method is more suitable for long-term cumulation of inflammation effects and chronic drug intervention studies.