小鼠矽肺纤维化造模成功时间点的比较研究

A comparative study of the time points of successful modeling silicotic fibrosis in mice

目的探索昆明小鼠开始形成稳定矽肺纤维化模型的最佳时间,为矽肺纤维化的研究提供一个可靠且成熟的矽肺纤维化模型的方法。方法口咽法造模。麻醉小鼠之前先称重,根据小鼠的体重给予戊巴比妥钠进行腹腔注射麻醉。口咽法滴注50.0mg/ml SiO_(2)混悬液200μl至小鼠支气管中。待小鼠清醒后,每隔1周进行一次取材,均做HE和Masson染色来观察小鼠肺组织纤维化变化的进展。利用实时荧光定量PCR和免疫组化法检测Ⅰ型胶原纤维(CollagenⅠ)和Ⅲ型胶原纤维(CollagenⅢ)的mRNA及蛋白表达水平。结果滴注SiO_(2)悬浊液1~2周时,小鼠肺组织中开始出现不同程度的炎症表现。第3周开始,巨噬细胞开始聚集,形成大小不一的灰色团块。至第4周后,可见较大面积的矽结节,个数多达数十个以上,肺泡结构也出现不同程度的破坏。与对照组相比,模型组的CollagenⅠ和CollagenⅢmRNA及相应的蛋白表达水平上调,差异有统计学意义(P<0.01)。结论昆明小鼠口咽法滴注50.0mg/ml SiO_(2)混悬液200μl至小鼠支气管中,经过4周后开始形成稳定的矽肺纤维化模型。

Objective To explore the best time for Kunming mice to form a stable model of silicotic fibrosis,so as to provide a reliable and proven method for modeling silicotic fibrosis.Methods The model was made by oropharyngeal method.The mice were weighed before anesthesia,and they were given sodium pentobarbital intraperitoneally for anesthesia according to their body weight.200μl of 50.0 mg/ml SiO_(2)suspension was dripped into the bronchus of the mice by oropharyngeal method.After the mice were awake,samples were taken from them every week,and all the samples were stained with HE and Masson to observe the progress of pulmonary fibrosis.The mRNA and protein expressions of the fibers of Collagen I and CollagenⅢwere measured by real-time fluorescence quantitative PCR and immunohistochemistry.Results At 1~2 weeks after dripping SiO_(2)suspension,different degrees of inflammatory manifestations began to appear in the lung tissues of mice.Since the third week,macrophages began to accumulate and form gray clumps of various sizes.By the fourth week,a large area of silicotic nodules of more than dozens could be seen and their alveolar structures were damaged to varying degrees.Compared with the control group,the model group had up-regulated mRNA and up-regulated corresponding protein expressions for CollagenⅠand CollagenⅢ,and the differences were statistically significant(P<0.01).Conclusion A stable model of silicotic fibrosis can be formed in Kunming mice after 4 weeks of oropharyngeal dripping 200μl of 50.0 mg/ml SiO_(2)suspension into the bronchus of mice.