目的 探讨博来霉素诱导小鼠肺纤维化最佳剂量和方法.方法 126只8周龄雄性ICR小鼠,随机分成一次性大剂量模型和多次小剂量模型.一次性大剂量模型分为200 mg/(kg·bw) BLM组、150 mg/(kg·bw)BLM组、100 mg/(kg· bw) BLM组及阴性对照组(DN组),每组18只,分别经尾静脉一次性注射BLM 200、150、100mg/(kg·bw)及生理盐水10 mL/(kg·bw),各组分别于第7、14、21天各处死6只.多次小剂量模型分为每日10 mg/(kg·bw)BLM组及阴性对照组(N组),分别经尾静脉注射BLM 10 mg/(kg·bw)及生理盐水10 mL/(kg·bw),每天1次,连续注射14 d,两组分别于第14、21、28天各处死6只.留取肺组织,观察肺组织病理改变,检测Ⅲ型胶原的含量,观察小鼠体重及生存率.结果 ①在一次性大剂量模型中,BLM各剂量组肺泡炎症评分及肺纤维化评分与正常组相比,除100 mg/(kg·bw)BLM组和150 mg/(kg·bw)BLM组在第7天的模型差异无显著性外(P＞0.05),其余各组差异均有显著性(P＜0.05)；各个剂量组Ⅲ型胶原的表达面积与正常组相比,除100 mg/(kg·bw) BLM组在第7天的模型差异无显著性外(P＞0.05),其余各组均较正常组高(P＜0.05),各个剂量组分别在第21天达到高峰,以200 mg/(kg·bw)BLM组第21天组Ⅲ型胶原的表达面积最高；该模型小鼠各剂量组死亡率为0.②在多次小剂量模型中,各组的肺泡炎症与肺纤维化程度与正常组相比差异均有显著性(P＜0.05)；各组Ⅲ型胶原的表达也均高于正常组(P＜0.05),且随着时间的延长呈进行性增加,在第28天达到高峰；该模型小鼠共死亡11只,死亡率为30.56％.结论 在本实验中,以尾静脉一次性注射BLM 200 mg/(kg·bw)后第21天诱导建立的ICR小鼠肺纤维化模型成模最好,其小鼠死亡率低,操作简单,有效安全方便的特点使之有希望成为一种复制肺纤维化的理想模型.
Objective To explore the optimal methods and dose of bleomycin to induce pulmonary fibrosis in mice. Methods One hundred and twenty six 8-week old male ICR mice were randomly divided into singule-large-dose (OLD) and multiple-low-dose (MLD) model groups. The OLD model was classified into four subgroups with BLM injection in a dose of 200 mg/(kg·bw) , 150 mg/(kg·bw) , 100 mg/(kg·bw) and negative control (saline 10 ml/(kg·bw) ) , and six mice were sacrificed at 7, 14 and 21 days after injection, respectively. The multiple low-dose models were classified into 10 mg/(kg·bw)/d group BLM injection once per day for consecutive 14 days, and negative control group. Six mice were killed at 14, 21 and 28 days after BLM and saline injection, respectively. The body weight, survival rate, pathological changes and amount of type Ⅲ collagen in the lung tissue were observed. Results ① In the OLD models, the scores of pulmonary alveolitis and fibrosis of the mice in all groups were significantly higher than that of the normal group (P< 0.05) , except the 100 mg/(kg·bw)BLM and 150 mg/(kgo bw)BLM groups (P>0. 05) sacrificed at 7 days. The expressions of type Ⅱ collagen in all groups were significantly higher than that of the normal group ( P < 0. 05 ) except the 100 mg/(kg·bw)BLM group (P>0. 05) killed at 7 days. The expression of type Ⅲ collagen in each group at 21 days reached a peak, and the 200 mg/(kg·bw) group killed at 21 days was the highest. The mortality rate was 0. ②In the MLD models, compared with the normal group, the pulmonary alveolitis and fibrosis of all the groups were significantly higher than that in the control group (P <0. 05) , progressively increasing with time, and peaked at 28 days. Eleven MLD model mice died and the mortality rate was 30. 56% . Conclusions The results of our study demonstrate that the mouse models developed at 21 days after BLM 200 mg/( kg·bw) injection is most successful, with advantages such as low mortality, simple operation, effective, good safety and convenience in use. Therefore it may become an ideal method in establishing animal model of pulmonary fibrosis.
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