慢性阻塞性肺疾病肺血管内皮细胞和肺泡上皮细胞凋亡的实验研究

Apoptosis of alveolar epithelial cells and pulmonary vascular endothelial cells in chronic obstructive pulmonary disease

目的研究COPD患者及烟雾暴露COPD大鼠肺血管内皮细胞和肺泡上皮细胞凋亡的相互关系及其与肺功能和肺气肿的相关性。方法采用烟雾暴露法建立COPD大鼠模型。分别采集COPD患者（13例）和对照患者（12例）及烟雾暴露80d大鼠（11只）和对照大鼠（12只）的肺组织标本，用HE染色评估肺部病理改变，用平均内衬间隔（MH）与平均肺泡数（MAN）评估大鼠肺气肿程度；用原位末端标记法对肺血管内皮细胞和肺泡上皮细胞凋亡进行定量检测，分析这两种细胞凋亡的相关性，及其与肺功能和肺气肿指标的相关性。正态分布的计量资料以x±s表示，采用两独立样本的t检验进行分析；非正态分布的计量资料以中位数（四分位间距）表示，采用两个独立样本比较的Wilcoxon秩和检验；采用Spearman秩相关进行相关分析。结果COPD患者和大鼠均出现明显肺气肿病理改变。COPD患者肺血管内皮细胞和肺泡上皮细胞凋亡指数[（13．2±2．6）％和（28．9±3．1）％]明显高于对照患者[（5．6±1．5）％和（5．8±1．2）％]。COPD患者肺泡上皮细胞凋亡指数[（28．9±3．1）％]明显高于肺血管内皮细胞凋亡指数[（13．2±2．6）％]，二者间呈正相关（r=0．60，P〈0．05）；COPD患者肺血管内皮细胞和肺泡上皮细胞的凋亡指数与FEV，占预计值％呈负相关（r值分别为-0．83和-0．69，均P〈0．05），与FEV1／FVC呈负相关（r值分别为-0．95和-0．71，均P〈0．05），与残气容积／肺总量呈正相关（r值分别为0．93和0．70，均P〈0．05）。COPD大鼠肺血管内皮细胞和肺泡上皮细胞凋亡指数[（4．1±0．4）％和（10．0±1．0）％]明显高于对照大鼠[（0．2±0．1）％和（2．1±0．4）％]，COPD大鼠肺组织MH与肺泡上皮细胞凋亡指数呈正相关（r=0．59，P〈0．05），MAN与肺泡上皮细胞凋亡指数呈负相关（r=-0．81，P〈0．05）。结论COPD患者和大鼠肺内存在异常的细胞凋亡现象，其肺泡上皮细胞凋亡较肺血管内皮细胞凋亡更为明显，且二者呈正相关；COPD患者和大鼠肺血管内皮细胞和肺泡上皮细胞的凋亡与其肺功能及肺气肿的改变明显相关，推测肺血管内皮细胞和肺泡上皮细胞凋亡可能参与COPD的发病过程。

Objective To investigate the relationship between apoptosis of alveolar epithelial cells and pulmonary vascular endothelial cells in chronic obstructive pulmonary disease （COPD） and the relationship between apoptosis and lung function and emphysema. Methods Twenty-four male Sprague- Dawley rats were randomly divided into a smoke exposure group （ COPD group） and a normal control group （NC group）. The rat model of COPD was established by exposure to cigarette smoke for 80 days, and lung tissues were obtained. Lung tissue samples were also collected respectively from patients with COPD （ n = 13） and without COPD （n = 12）. Lung sections stained by HE were observed to study the morphological alteration, and mean linear intercept （MLI） and mean alveolar numbers （MAN） were measured to estimate the extent of emphysema in rats. Quantitative analysis of apoptosis of alveolar epithelial cells and pulmonary vascular endothelial cells was undertaken by TdT-mediated dUTP nick end labeling （TUNEL）. The data distributed normally were expressed as x ± s, and the independent-samples t-test was used for comparison of means. Nonparametric data were expressed as median （quartile range）, and Wilcoxon rank sum test was used for comparison. Correlation between apoptosis of the two kinds of ceils in patients with COPD was estimated by Spearman rank correlation coefficients, and the correlations between the specific indexes, such as FEV1% Pre, FEV1/FVC（% ）, residual volume/total lung capacity（% ） [ RV/TLC（% ） ], MLI, MAN, and the cell apoptosis were also investigated. Statistical difference was accepted at P 〈 0. 05. Results Destruction of alveolar walls and enlargement of alveolar space were observed as pathological changes of lung tissues from patients and rats with COPD. The apoptotic index （AI） of alveolar epithelial cells and pulmonary vascular endothelial cells were （28.9 ± 3. 1 ）%, （13. 2 ± 2.6）% in patient COPD group; （10.0±1.0）%, （4. 1 ±0.4）% in rat COPD group; the differences being significant （t = -23.946, - 8. 820, - 2.4. 273, - 36. 422, all P 〈 0. 05 ）, as compared with NC groups [ （5. 8 ± 1.2） %, （5. 6 ± 1.5）%, （2. 1 ± 0.4）%, （0.2 ± 0. 1）% respectively]. In patient COPD group, the AI of alveolar epithelial cells was significantly higher than that of pulmonary vascular endothelial cells （t = - 13. 889, P 〈 0.05 ） and both were positively correlated with each other （ r = 0. 60, P 〈 0.05 ）. In patient COPD group, both the apoptosis of alveolar epithelial cells and pulmonary vascular endothelial cells revealed negative correlations to FEV1 （ % ） pre （ r = - 0. 83, - 0. 69, all P 〈 0. 05 ） and FEV1/FVC （ % ） （ r = - 0. 95, - 0. 71, all P 〈 0. 05 ）, but positive correlations were demonstrated between them and RV/TLC （ % ） （ r = 0. 93, 0. 70, all P 〈0. 05）. In rat COPD group, MLI of lung tissue showed a positive correlation with the AI of alveolar epithelial cells （ r = 0. 59, P 〈 0. 05 ） while MAN a negative correlation with the latter （ r = -0. 81, P 〈 0. 05 ）. Conclusions Abnormal apoptosis of alveolar epithelial cells and pulmonary vascular endothelial cells was present in patients and rats with COPD, which was related to the changes of lung function or pathological changes of lung tissues. The results suggest that the abnormal changes of apoptosis of pulmonary tissue in COPD may be involved in the pathogenesis of the disease.