雷帕霉素对慢性阻塞性肺疾病大鼠气道重塑中血管内皮生长因子表达作用的研究

Effect of Rapamycin on the Expression of Vascular Endothelial Growth Factor in Airway Remodeling of Rats with Chronic Obstructive Pulmonary Disease

目的探讨雷帕霉素对慢性阻塞性肺疾病(COPD)大鼠气道重塑中血管内皮生长因子(VEGF)表达的作用。方法 2015年6月—2016年3月,采用随机数字表法将30只SPF级健康雄性Wistar大鼠分为正常组、模型组、雷帕霉素组,每组10只。模型组与雷帕霉素组采用气道内滴入脂多糖(LPS)加烟熏双重刺激法建立COPD气道重塑大鼠模型,正常组不予任何干预。雷帕霉素组给予雷帕霉素治疗,模型组、正常组给予等量的0.9%氯化钠溶液。干预治疗14 d后,收集各组大鼠血清、肺泡灌洗液(BALF)、肺组织标本;取部分右侧肺组织行HE染色,剩余右侧肺组织行Masson染色与免疫组化染色,分别计算小气道基底膜单位长度的胶原厚度(WAc/Pbm)、细支气管VEGF阳性表达面积占气管管壁总面积的百分比(Aim/WAt);采用酶联免疫吸附实验(ELISA)法检测血清、BALF及肺组织中VEGF水平。结果 HE染色结果显示,与模型组比较,雷帕霉素组大鼠细支气管、肌性动脉形态及肺组织病理改变有所改善;Masson染色结果显示,模型组、雷帕霉素组大鼠WAc/Pbm大于正常组,雷帕霉素组大鼠WAc/Pbm小于模型组(P<0.05)。免疫组化染色结果显示,模型组、雷帕霉素组大鼠Aim/WAt大于正常组,雷帕霉素组大鼠Aim/WAt小于模型组(P<0.05)。ELISA法检测结果显示,模型组、雷帕霉素组大鼠血清、BALF、肺组织中VEGF水平高于正常组,雷帕霉素组大鼠血清、BALF、肺组织中VEGF水平低于模型组(P<0.05)。结论雷帕霉素可以改善COPD大鼠气道重塑,这可能与雷帕霉素干预治疗减少了COPD大鼠气道重塑中VEGF水平,进而抑制血管新生有关。

Objective To explore the role of rapamycin on the expression of vascular endothelial growth factor（ VEGF） in airway remodeling of rats with chronic obstructive pulmonary disease（ COPD）. Methods From June 2015 to March2016,30 SPF grade male Wistar rats were randomly divided into three groups（ normal group, model group and rapamycin group） by random number table method,and each group had 10 rats. The dual stimulation way of lipopolysaccharide（ LPS）through the airway plus fumigation was conducted to rats in model group and rapamycin group to construct COPD rats＇ model of airway remodeling,while no intervention was given to rats in normal group. Rats in rapamycin group received treatment of rapamycin,while rats in model group and normal group received the same amount of 0. 9% solution of sodium chloride. After 14- day intervention,the serum,bronchial alveolar lavage fluid（ BALF） and lung tissue specimens of rats in each group were collected; part of the right lung tissue was taken to perform HE staining and Masson staining and immunohistochemical staining were given to the remaining tissue,collagen thickness（ WAc / Pbm） per unit length of small airway basement membrame,and the percentage of bronchioles VEGF positive expression area in the total area of the corresponding tracheal tube wall（ Aim / WAt）were calculated respectively; the enzyme- linked immunosorbent assay（ ELISA） was used to detect VEGF expression level in serum,BALF and lung tissue. Results The HE staining results showed that compared with model group,the pathologic change of bronchiole,muscular artery morphology and lung tissue of rats in rapamycin group had improved; Masson staining results showed that the WAc / Pbm of rats in model group and rapamycin group was greater than that in normal group,while WAc / Pbm of rats in rapamycin group was less than that in model group（ P〈 0. 05）. Immunohistochemical staining results showed that Aim/ WAt of rats in model group and rapamycin group was greater than that in normal group,while Aim / WAt of rats in rapamycin group was less than that in model group（ P 〈0. 05）. ELISA testing results showed that the VEGF expression level of serum,BALF and lung tissue in rapamycin group and model group was higher than that in normal group,while the VEGF expression level of serum,BALF and lung tissue in rapamycin group was lower than that in model group（ P 〈0. 05）. Conclusion Rapamycin can improve the airway remodeling of COPD rats,which may be related to the intervention treatment of rapamycin on decreasing of VEGF level in airway remodeling of COPD rats and thus inhibiting angiogenesis in rats.