肺纤维化模型小鼠肺泡巨噬细胞凋亡的动态变化及意义

The dynamic change and significance of apoptosis on lung alveolar macrophages in pulmonary fibrosis model mice

目的探讨博莱霉素诱导的肺纤维化模型小鼠肺泡巨噬细胞(AM)凋亡的动态变化及意义。方法 100只C57BL/6雄性小鼠随机分为对照组和模型组,每组各50只。模型组采用口咽吸入法给予博莱霉素A2(2.5 mg/kg)建立肺纤维化模型,对照组给予等体积生理盐水。建模后第1、3、7、14和21天每组随机处死10只动物,5只采用苏木素-伊红(HE)染色观察肺组织病理变化,计算炎症评分及肺系数;另5只采用碱水解法测定肺组织羟脯氨酸(Hyp)的含量;记录肺泡灌洗液(BALF)中AM数目,流式细胞术检测AM凋亡情况。结果模型组小鼠第3天肺组织出现炎症细胞浸润、肺泡壁水肿增厚,炎症评分明显升高;第7天时炎症程度继续加重,炎症评分进一步升高;第14天和第21天时出现纤维增生和胶原沉积;第21天时炎症评分较第7天和第14天下降。第21天时模型组Hyp含量较第1、3、7天均明显升高,但与第14天相比无明显差异。在整个实验过程中,模型组的肺系数,AM数目及凋亡率呈现先升高后降低的变化,于第7天达到峰值。结论在博莱霉素诱导的小鼠肺纤维化模型中,AM的数目和凋亡水平在前期的炎症反应阶段明显升高,在肺纤维化阶段出现下降。动态变化的AM可能在肺纤维化进展过程中发挥不同的作用。

Objective To observe the dynamic change and significance of apoptosis levels in the alveolar macrophages (AM) of the bleomycin-induced pulmonary fibrosis model mice. Methods A total of 100 mice were randomly divided into control group (n=50) and model group (n=50). The model group was received with bleomycin at 2.5 mg/kg via oropharyngeal instillation to molding. The control group was conducted with same volume of normal saline. Ten mice randomly selected were executed on the day 1, 3, 7, 14, and 21 after modeling in the two groups. Hematoxylin and eosin (HE) staining was used to observe the pathological changes of lung tissue and calculate the inflammation score and pulmonary coefficients in five mice of two groups. Alkali hydrolysis method was used to detect the content of hydroxyproline, and the bronchoalveolar lavage fluid (BALF) was collected to assess the cell counts and apoptotic levels of AM in another five mice. Results Mice of model group showed inflammatory cell infiltration and alveolar wall widening on day 3, along with an aggravate inflammation and increased inflammation score on day 7. The fibroplasia and collagen sediment were increased on day 14 and day 21. The inflammation score was decreased significantly in day 21 compared with those in day 7 and day 14. The hydroxyproline contents were increased significantly on day 21 than those on day 1, day 3 and day 7 in model group, but no significant difference when compared with those of day 14. The cell count and apoptotic rates of AM, and pulmonary coefficients showed changes of increasing first and then decreasing in model group, which reached the peak at day 7.Conclusion The cell counts and apoptotic levels increase in the preliminary stage of inflammatory and decrease in pulmonary fibrosis phase in bleomycin-induced pulmonary fibrosis model mice. These changes may play different roles in the progression of pulmonary fibrosis.