慢性阻塞性肺疾病小鼠模型制备方法的比较研究

Comparative study on establishment of a mouse model of chronic obstructive pulmonary disease

目的:采用香烟烟雾(cigarette smoke,CS)暴露、肺炎克雷伯杆菌(Klebsiella pneumoniae,KP)感染、聚肌胞苷酸(polyinosinic-polycytidylic acid,Poly I:C)滴鼻、CS暴露联合KP感染和CS暴露联合Poly I:C滴鼻5种方法,建立并比较慢性阻塞性肺疾病(chronic obstructive pulmonary disease,COPD)小鼠模型。方法:将288只雄性BALB/c小鼠随机分为正常组(normal组)、CS组、KP组、CS+KP组、Poly I:C组及CS+Poly I:C组,每组48只。第1~8周造模,分别于第4、8、16和24周末取材。观察小鼠肺组织平均肺泡数(mean alveolar number,MAN)、肺泡平均截距(mean linear intercept,MLI)、肺功能呼气峰流速(peak expiratory flow,PEF)和50%潮气量呼气流量(50%tidal volume expiratory flow,EF50)的变化,检测肺组织中肿瘤坏死因子α(tumor necrosis factor-α,TNF-α)、白细胞介素6(interleukin-6,IL-6)和基质金属蛋白酶2(matrix metalloproteinase-2,MMP-2)的表达,采用R值综合评价法分析不同造模方法的COPD小鼠病变程度。结果:与normal组比较,第4周,CS+KP组PEF和MAN显著降低,MLI及TNF-α水平和IL-6显著升高(P<0.05);第8周,各模型组小鼠肺组织出现大量炎症细胞浸润、肺泡腔扩张、肺泡壁断裂融合、气管壁增厚等病理变化,MAN显著降低,MLI显著升高,肺功能PEF和EF50显著降低,TNF-α、IL-6和MMP-2水平显著升高(P<0.05);观察至第24周CS+KP组和CS+Poly I:C组仍然表现出COPD的病理特征;综合所有时点R值综合评价结果显示,CS+KP组和CS+Poly I:C组R综合值较CS组、KP组和Poly I:C组显著升高,KP组和Poly I:C组R综合值较CS组显著降低(P<0.05)。结论:CS暴露联合细菌感染或Poly I:C滴鼻组COPD的病理特征明显、出现早、持续时间长、远后观察稳定。

AIM:To establish the mouse model of chronic obstructive pulmonary disease(COPD),we employed five approaches including cigarette smoke(CS)exposure,Klebsiella pneumoniae(KP)infection,polyinosinicpolycytidylic acid(Poly I:C)nasal drip,the combination of CS exposure and KP infection,and the combination of CS exposure and Poly I:C nasal drip.METHODS:Male BALB/c mice(n=288)were randomly divided into normal group,CS group,KP group,CS+KP group,Poly I:C group and CS+Poly I:C group(n=48 in each group). The mice were exposed to CS,KP and/or Poly I:C from the 1 st to 8 th weeks,and sacrificed at the end of the 4 th,8 th,16 th and 24 th weeks. Mean alveolar number(MAN),mean linear intercept(MLI),peak expiratory flow(PEF)and 50 % tidal volume expirtory flow(EF50)were determined. The expression levels of tumor necrosis factor-α(TNF-α),interleukin-6(IL-6)and matrix metalloproteinase-2(MMP-2)in the mouse lungs were detected. The R value was used to analyze the pathological changes of COPD mice.RESULTS:Compared with normal group,the PEF and MAN were significantly decreased,while the levels of MLI,TNF-α and IL-6 were significantly increased in CS+KP group at week 4(P<0. 05). At week 8,massive inflammatory infiltration,alveolar cavity expansion,alveolar wall destruction,airway wall thickening and other pathological changes were observed in the lung tissues of mice in each model group. Furthermore,MAN,PEF and EF50 were significantly decreased,while the levels of MLI,TNF-α,IL-6 and MMP-2 were significantly increased(P<0. 05). The pathological characteristics of COPD were still observed in CS+KP group and CS+Poly I:C group at week 24. The results of comprehensive evaluation of R values at all time points displayed that the R values of CS+KP group and CS+Poly I:C group were significantly higher than those of CS group,KP group and Poly I:C group,while the R values of KP group and Poly I:C group were significantly lower than that of CS group(P<0. 05).CONCLUSION:The mice in the group of combination of CS and KP or Poly I:C had distinctly pathological characteristics of COPD,which appeared to occur early,last longer and even during the entire observation period.