腹腔注射博来霉素诱导小鼠肺纤维化模型的长期稳定性  

Intraperitoneal injection of bleomycin induces pulmonary fibrosis in mice: a long-term stability evaluation

作　　者：苏敏红 江宁[2] 李洪涛[3] 王振国[1] 谢玉芬[1] 郑晓滨[1] 涂常力 黄瑾[1] Su Min-hong Jiang Ning Li Hong-tao Wang Zhen-guo Xie Yu-fen Zheng Xiao-bin Tu Chang-li Huang Jin(Department of Respiratory Medicine, the Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai 519000 Guangdong Province, China Department of Urology, the Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou 510000, Guangdong Province, China Department of Respiratory Medicine, the Third Affiliated Hospital of Sun Yat-sen University, Guangzhou 510630, Guangdong Province, China)

机构地区：[1]中山大学附属第五医院呼吸内科,广东省珠海市519000 [2]中山大学孙逸仙纪念医院泌尿外科,广东省广州市510000 [3]中山大学附属第三医院呼吸科,广东省广州市510630

出　　处：《中国组织工程研究》2017年第4期512-519,共8页Chinese Journal of Tissue Engineering Research

基　　金：国家自然科学基金项目(81470220)~~

摘　　要：背景:至今特发性肺间质纤维化尚无特效治疗药物,这与该疾病动物模型尚不能完全模拟特发性肺间质纤维化病理表现有极大关系,因此建立更理想的动物肺间质纤维化模型对研究人类特发性肺间质纤维化的发病机制及研制有效治疗药物显得至关重要。目的:建立一种更符合人类特发性肺间质纤维化病理组织特征的小鼠肺纤维化模型。方法:雄性C57BL/6小鼠70只,随机均分为两组,博来霉素每周2次组(A组),腹腔注射博来霉素35 mg/kg,2次/周,共注射8次;博来霉素每周1次组(B组)腹腔注射博来霉素35 mg/kg,1次/周,共注射8次。分别于注射完第8次博来霉素后2,4,6,8,10周各处死5只小鼠,取肺组织进行羟脯氨酸测定、苏木精-伊红、Masson和免疫组织化学染色。结果与结论:(1)两组在第8次腹腔注射后2,4,6,8,10周均出现不同程度的肺泡炎、肺纤维化。A组肺泡炎、肺纤维化评分于2周开始逐渐升高,于6-8周达高峰,持续至10周仍无明显降低;B组肺泡炎、肺纤维化评分于2周最高,随后逐渐波动式下降,至6周开始明显比A组评分低(P<0.05);(2)兔疫组织化学显示Ⅰ型胶原蛋白主要沉积在胸膜下、血管周围和肺泡间隔,与Masson染色所见的胶原纤维分布一致;转化生长因子β1和α平滑肌动蛋白在肺泡炎和肺纤维化部位表达增多。A组小鼠肺Ⅰ型胶原蛋白、a-SMA、转化生长因子β1、羟脯氨酸含量于2周开始持续升高,于6-8周达峰值;B组于2周最高,随后逐渐降低,于4周时肺a-SMA和转化生长因子β1表达明显低于A组(P<0.05),于6周时肺Ⅰ型胶原蛋白表达和羟脯氨酸含量明显低于A组(P<0.05);(3)结果表明,1周2次腹腔注射博来霉素35 mg/kg诱导小鼠肺纤维化模型有极好的稳定性,且操作十分简单,可重复性好,其反复损伤修复过程、病理变化、细胞因子变化都与人类特发性肺间质纤维化类似,对研究人类肺纤维化,尤其是特发性肺�BACKGROUND： There is no effective drug for idiopathic pulmonary fibrosis（IPF）, because of a lack of the animal model imitating the complete pathogenesis of human IPF. Therefore, it is critical to establish an ideal animal IPF model used for investigating the underlying pathogenesis and developing a kind of effective drug. OBJECTIVE： To establish an animal model that can mimic more characters of human IPF. METHODS： Seventy male C57BL/6 mice were randomly divided into two groups, followed by subjected to the intraperitoneal injection of bleomycin（35 mg/kg） on days 1, 4, 8, 11, 15, 18, 22, and 25, twice（group A） or once（group B） a week. Mice were sacrificed at 2, 4, 6, 8, and 10 weeks after the eighth injection, and the lung tissues were moved used for hematoxylin-eosin, Masson and immunohistochemical stainings. RESULTS AND CONCLUSION： There were various degrees of alveolitis and pulmonary fibrosis in the two groups at different time points after the last injection. The scores of alveolitis and pulmonary fibrosis in the group A began to gradually increase from the 2nd week and reached the highest level at the 6th-8th weeks until the 10 th week. In contrast, the scores of alveolitis and pulmonary fibrosis in the group B peaked at the 2nd week, then fluctuately decreased, and were significantly lower than those in the group A at the 6th week（P〈0.05）. Immunohistochemistry showed that type I collagen deposition was mainly distributed in the subpleural region, peri-vascular region and alveolar septa, which was consistent with Masson staining findings. The expression levels of transforming growth factor β1（TGF-β1） and α-smooth muscle actin（α-SMA） in the regions developing alveolitis and pulmonary fibrosis were significantly increased. In the group A, the expression levels of type I collagen, TGF-β1, α-SMA, and the hydroxyproline content in the lung tissues reached the peak level at 6-8 weeks. However, in the group B, all above indicators reached the highest level at the 2nd week

关 键 词：疾病模型 动物 肺纤维化 博来霉素 组织构建 组织工程 转化生长因子Β1 特发性肺间质纤维化 普通间质性肺炎 腹腔注射 小鼠 动物模型 动物实验 国家自然科学基金 

分 类 号：R318[医药卫生—生物医学工程]