痰热清注射液对慢性阻塞性肺疾病大鼠支气管上皮多药耐药相关蛋白1水平的影响

Effects of Tanreqing injection on expression levels of multidrug resistance-associated protein 1 in bronchial epithelial cells of rats with chronic obstructive pulmonary disease

目的观察痰热清注射液对慢性阻塞性肺疾病（COPD）大鼠支气管上皮多药耐药相关蛋白1（MRP1）表达水平的影响，探讨痰热清治疗COPD的作用机制。方法选择雄性Wistar大鼠30只，按随机数字表法分为正常对照组、COPD模型组和痰热清治疗组，每组10只。采用气管内滴人脂多糖（LPS）加香烟烟熏方法复制COPD模型，正常对照组不予任何处理。制模后痰热清治疗组尾静脉注射痰热清注射液2mUkg，COPD模型组和正常对照组则注射0．9％氯化钠注射液2mUkg；各组均连续给药14d，然后观察大鼠体质量及呼吸功能，用蛋白质免疫印迹试验（Western Blot）测定肺组织MRP1蛋白表达，取肺组织观察大鼠肺组织病理学改变。结果治疗14d后3组大鼠体质量均较治疗前增加，但COPD模型组增加缓慢且低于痰热清治疗组（g：247．8±15．9比265．4±18．7，P〈0．05）。COPD模型组0．3s用力呼气容积占用力肺活量的比值（FEV0．3／FVC）较正常对照组明显降低（0．688±0．213比0．973±0．052），呼气峰流速[PEF（mUs）：3．28±0．74比4．58±1．48]、肺顺应性[Cydn（mL/cmH2O）：317．1±130．3比515．1±140．2]和MRP1蛋白水平[吸光度（A）值：0．698±0．103比1．318±0．169]均较正常对照组明显降低，残气容积／肺总量比值（RVffLC）较正常对照组明显升高（0．502±0．128比0．316±0．153）；痰热清治疗组FEV0．3／FVC（0．801±0．142比0．688±0．213）、PEF（mUs：4．24±0．36比3．28±0．74）、Cydn（421．2±342．0比317．1±130．3）、MRP1蛋白（A值：0．964±0．095比0．698±0．103]均较COPD模型组增高，RVffLC较COPD模型组降低（0．411±0．105比0．502±0．128），差异均有统计学意义（均P〈O．05）。光镜下可见：COPD模型组大鼠肺泡壁变薄、断裂，肺泡融合扩大，支气管黏膜大量上皮纤毛脱落、倒伏、粘连，杯状细胞增生，各级细支气管壁可见大量炎性细胞浸润，管腔内有炎性分泌物潴留；痰热清治疗组上述变化较COPD模型组有一定程度改善。结论痰热清注射液可能通过增加大鼠支气管上皮细胞中MRP1的表达起到有效减轻COPD气道炎症、延缓COPD肺功能恶化和改善症状的作用。

Objective To observe the effects of Tanreqing injection on the expression levels of muhidrug resistance-associated protein 1 （MRP1） mRNA in bronchial epithelial cells of rats with chronic obstructive pulmonary disease （COPD） model and to explore the mechanism of Tanreqing for treating COPD. Methods Thirty male Wistar rats were randomly divided into normal control, COPD model and Tanreqing treatment groups, 10 rats in each group. The COPD rat model was established with the methods of tracheal instillation of lipopolysaccharide （LPS） and quantitative stimulation with tobacco smoke; no treatment was given to the normal control group. After the model was established, the Tanreqing therapy group was injected with Tanreqing injection 2 mL/kg; the COPD model and the normal control groups were injected with 0.9% sodium chloride injection 2 mL/kg through caudal vein; in each group, the respective agent was administered continuously for 14 days, and then the body mass and respiratory function were observed, the MRP1 protein expression of pulmonary tissue was detected by Western Blot and the pathological changes of rat lung tissues were observed in each group. Results The body mass of three groups was increased after 14 days of treatment, but the increase of body mass in the COPD model group was slower than that in the Tanreqing therapy group （g： 247.8 ± 15.9 vs. 265.4 ± 18.7, P 〈 0.05）. Compared with the normal control group, the forced expiratory volume in 0.3 second/forced vital capacity （FEV0.3/FVC） was decreased significantly in the COPD model group （0.688 ± 0.213 vs. 0.973 ± 0.052）; the peak expiratory flow rate [PEF（mL/s）： 3.28 ± 0.74 vs. 4.58 ± 1.48], the lung compliance [Cydn （mL/cmH20）： 317.1 ± 130.3 vs. 515.1 ± 140.2] and the protein expression of the bronchial epithelial MRP1 [absorbance （A） value： 0.698 ± 0.103 vs. 1.318 ±0.169] in the COPD model group were decreased obviously, residual volume/total lung capacity （RV/TLC） was increased significantly （0.502± 0.128 vs. 0.316 ± 0.153）. Compared with the COPD model group, FEV0.3/FVC （0.801-±0.142 vs. 0.688±0.213）, PEF （mL/s： 4.24±0.36 vs. 3.28±0.74）, Cydn （mL/cmH20： 421.2 ± 342.0 vs.317.1 ± 130.3）, the protein expression of MRPI （A value： 0.964± 0.095 vs. 0.698± 0.103） in Tanreqing treatment group were increased significantly, RV/TLC was decrease significantly （0.411±0.105） vs. （0.502±0.128） in the Tanreqing treatment group （all P 〈 0.05）. Under light microscope： in COPD model group, the alveolar wall became thin and broken, the alveoli fused and enlarged, a large quantity of cilia of bronchial mucosal epithelium detached, fallen down and adherent together, goblet cell proliferating, a large amount of inflammatory cell infiltration in various levels of bronchiole mucosal epithelial walls and retention of inflammatory secretion in lumens could be seen; in Tanreqing treatmeat group, the above changes were improved to a certain degree compared with those in COPD model group. Conclusions Tanreqing can effectively alleviate the lung bronchial inflammation, postpone deterioration of lung function and improve the symptoms in rats with COPD; the mechanism is possibly related to the increase of MRP1 expression of bronchial epithelial cells.