慢性阻塞性肺疾病大鼠膈肌纤维类型改变和膈肌细胞凋亡的研究

Adaptations of diaphragmatic muscle fibres and apoptosis of diaphragmatic muscle cell in rats with chronic obstructive pulmonary disease

目的:通过对慢性阻塞性肺疾病(chronic obstructive pulmonary disease,COPD)大鼠膈肌纤维分类的改变及膈肌凋亡的研究来探讨膈肌疲劳的发病机制,并以此寻找治疗膈肌疲劳的新途径。方法:40只雄性Wistar大鼠随机分成健康对照组和COPD模型组,每组20只。模型组大鼠置于烟雾浓度为5%的密封熏箱内(每天2次,每次30 min,间隔8 h,共28 d),第1、14天向气管内注射脂多糖200μg。健康对照组大鼠放入密闭箱内但不进行烟熏,第1、14天气管内注入生理盐水200μl。第29天将大鼠处死取膈肌及肺,测量各组膈肌的凋亡率、Fas蛋白的表达和Caspase-3基因表达,电镜以及膈肌纤维分类染色观察形态。结果:第28天后COPD模型组大鼠膈肌细胞的凋亡率、Fas蛋白及Caspase-3基因的表达水平均高于健康对照组(P<0.01),相对于健康对照组,模型组Ⅰ型纤维比例增加而Ⅱ型纤维比例减少(P<0.01)。结论:在COPD的发生过程中存在膈肌细胞过度凋亡及纤维类型的改变,Caspase-3和Fas/FasL途径参与了膈肌细胞凋亡的调控,并且纤维类型的改变导致COPD患者晚期膈肌疲劳治疗困难。

Objective： To investigate the pathogenesis of diaphragmatic fatigue by diaphragm fibers classification changes and di- aphragmatic apoptosis in rats with chronic obstructive pulmonary disease （COPD）,in order to find new treatment of diaphragmatic fatigue. Methods： Forty male Wistar rats were randomly averagely divided into healthy control group and COPD group. The rats in the COPD group were exposed to 5% smoke in a sealed box （twice for each day, 30 minutes for each time,interval 8 hours,lasting for 28 days） ,and 200 μg lipopolysaccharide was injected intratracheally on the 1st and 14th day. On the 29th day,the rats were sacrificed for detection of apoptosis rate,Fas protein and Caspase-3 gene expression of diaphragmatic muscle cell, and observation of the morphologie changes by electron microscope and diaphragm myofibers classification dyeing. Results： On the 28th day,destruction of alveolar walls and enlargement of alveolar space were observed in the COPD group,the diaphragmatic muscle cell apoptosis rate, the Fas protein and Caspase-3 gene expression were higher than those of the control group（P 〈 0.01）. Compared to the healthy controls, the type I fiber proportion in the COPD group was increased,while the II type fiber proportion was reduced （both P 〈 0.01 ）. Con- clusion： Abnormal apoptosis of diaphragmatic muscle cell and fiber type of change were present in rats with COPD. Caspase-3 and Fas/FasL way could be involved in the apoptosis of diaphragmatic muscle cell, the regulation of fiber type of change in the late patients with COPD ,which causes difficult to treat diaphragmatic muscle fatigue.