【作者】 王文静；

【导师】 孙武装；

【作者基本信息】 河北医科大学 ， 内科学， 2018， 硕士

【摘要】 目的:研究氢气对烟雾所致慢性阻塞性肺疾病（chronic obstructive pulmonary disease,COPD）模型大鼠肺组织病理变化的影响,及对氧化抗氧化相关指标（SOD、MDA）、线粒体自噬相关指标（PINK1、Parkin）表达变化的影响,探讨氢气对COPD发展的治疗作用及其机制。方法:1 SD大鼠（160-200 g）70只随机分为7组:正常对照组（Control group）,COPD组（COPD group）,低浓度氢气组（Low Hydrogen group,Hl group）,中浓度氢气组（Middle Hydrogen group,Hm group）,高浓度氢气组（High Hydrogen group,Hh group）,极高浓度氢氧组（Very high Hydrogen and Oxygen groups,Hvh group）,氧气组（Oxygen group,O₂ group）,采用烟熏法建立COPD大鼠模型。观察大鼠日常生活状态。2黄嘌呤氧化酶法测定各组大鼠肺组织匀浆SOD水平;应用硫代巴比妥酸比色分析法测定各组大鼠肺组织匀浆MDA水平。HE染色观察各组大鼠肺组织病理变化。免疫组化及WB法检测肺组织中PINK1、Parkin蛋白的表达。结果:1各组大鼠肺组织匀浆SOD、MDA比较与Control组比较,COPD组SOD活力显著降低（P<0.01）;与COPD组比较,Hvh、Hh组显著增高（P<0.01）,Hm、Hl、O₂组增高（P<0.05）;Hvh组显著高于Hh、Hm、Hl、O₂组（P<0.01）。与Control组比较,COPD组MDA含量显著增高（P<0.01）;与COPD组比较,Hvh、Hh组显著降低（P<0.01）,Hm、Hl组降低（P<0.05）。2 HE染色肺组织病理学变化Control组大鼠肺组织中气管黏膜假复层纤毛柱状上皮完整,肺泡大小及结构正常。COPD组大鼠肺组织较Control组大,灰白色、凹凸不平、弹性差。显微镜下观察肺泡管、肺泡囊、肺泡显著扩张,肺泡结构破坏,肺泡壁变薄,断裂,融合为肺大疱,纤毛细胞变性和坏死,肺小动脉管壁增厚、管腔狭窄,可以观察到大量的炎性细胞浸润在周围。Hvh、Hl、Hm、Hh及O₂组大鼠肺脏较COPD组肺脏体积略有缩小,外观情况也有所改善。肺动脉管壁增厚较局限,出现不同程度的炎性细胞浸润,但较COPD组呈不同程度的减轻。肺泡壁较薄,出现不同程度的断裂,但较COPD组有改善。3采用免疫组化比较PINK1、Parkin蛋白的表达与Control组比较,COPD组Parkin蛋白表达升高;与COPD组比较,各Hvh、Hh、Hm、Hl组Parkin蛋白表达降低,O₂组无显著差异。各组间PINK1蛋白表达无显著差异。4采用Western blot比较PINK1、Parkin的蛋白表达与Control组比较,COPD组Parkin蛋白表达升高;与COPD组比较,各Hvh、Hh、Hm、Hl组Parkin蛋白表达降低,O₂组无显著差异。各组间PINK1蛋白表达无显著差异。结论:1吸入氢气能够减轻COPD大鼠肺病理损伤,不同浓度的氢气效果不同,中、高、极高浓度优于低浓度氢气。2吸入氢气通过抗氧化对COPD的发展有延缓作用。3吸入氢气可能通过线粒体自噬延缓COPD进展。更多还原

【Abstract】 Objective:Through research the effects of hydrogen in pathology to a cigarette-smoke induced chronic obstructive pulmonary disease rat model,observe the expression of the related indicators of oxidation and antioxidation（SOD,MDA）and mitochondrial autophagy（PINK1,Parkin）in COPD rat lung tissue,to discuss the effects of hydrogen in delaying the development of COPD and its mechanisms.Methods:1 70 healthy male Sprague-Dawley rats（body weight 160-200 g）were randomized into the following seven groups（10 rats in each group）:control group,COPD group,low-hydrogen group（Hl group）,intermediate Hydrogen group（Hm group）,high-Hydrogen group（Hh group）,very high Hydrogen（Hvh group）,and oxygen group（O₂ group）.The rats with COPD-like lung disease were established by the smoking method.The daily condition of rats were observed.2 Xanthine oxidase method was used to measure the level of SOD in lung homogenate of rats in each group,and thiobarbituric acid colorimetric assay was used to measure the level of MDA in lung tissue of each group.Hematoxylin and eosin（HE）staining was used to observe the pathological changes of lung tissue in each group.Immunohistochemistry and WB technique were used to detect the expression of PINK1 and Parkin in lung tissue.Results:1 Comparison of SOD and MDA in lung homogenate of each groupSOD activity:Compared with the control group,the COPD group decreased significantly（P<0.01）;compared with the COPD group,Hvh,Hh group was significantly increased（P<0.01）,Hm,Hl,O₂ group was increased（P<0.05）;Hvh group significantly higher than Hh,Hm,Hl,O₂ group（P<0.01）.MDA content:Compared with the control group,the COPD group was significantly increased（P<0.01）;compared with the COPD group,Hvh,Hh group was significantly decreased（P<0.01）,Hm,Hl group was decreased（P<0.05）.2 pathologic changes of lung in cigarette smoke-induced rat modelIn the control group,the ciliated columnar epithelium of the tracheal mucosa was intact in the rat lung tissue.There was no inflammatory cell infiltration around the airways,no expansion and congestion of the capillaries,and normal alveolar size and structure.Compared with the control group,the lung volume in the COPD group was increased;on gross morphology,it was white and uneven,rather than smooth,and had poor flexibility.Under the microscope,the alveolar ducts,alveolar sacs and pulmonary alveoli were significantly expanded,the alveoli were structurally disordered,the alveolar wall showed signs of thinning and breaking,and some had fused into bullae,the ciliated cells degenerated and necrosis,the pulmonary arterial wall thickened,the lumen narrowed,and a large number of inflammatory cell infiltrates around.The volume of lungs in Hvh,Hl,Hm,Hh and O₂ groups was slightly smaller than that in COPD group,and the appearance was also improved.Pulmonary arterial wall thickening is more limited,with varying degrees of inflammatory cell infiltration,but compared with the COPD group to varying degrees of reduction.The alveolar walls were thin,with varying degrees of fracture,but improved compared with the COPD group3 Protein of PINK1 and Parkin of the lung tissue was detected by immunohistochemistry.Compared with control group,Parkin protein expression in COPD group increased.Compared with COPD group,the expression of Parkin protein in Hvh,Hh,Hm,Hl group decreased,but there was no significant difference in O₂ group.There was no significant difference in the expression of PINK1protein among the groups.4 Protein of PINK1 and Parkin of the lung tissue was detected by Western blot.Compared with control group,Parkin protein expression in COPD group increased.Compared with COPD group,the expression of Parkin protein in Hvh,Hh,Hm,Hl group decreased,but there was no significant difference in O₂ group.There was no significant difference in the expression of PINK1protein among the groups.Conclusions:1 Inhalation of hydrogen reduces lung pathological damage.Different concentrations of hydrogen have different effects.Medium,high,and extremely high concentrations are superior to low concentrations of hydrogen.2 Hydrogen can delay the development of COPD through antioxidation.3 Hydrogen may delay COPD progression through mitochondrial autophagy.更多还原