线粒体损伤在支气管哮喘发生机制中的作用
发布时间：2018-08-21 08:38
【摘要】：支气管哮喘是一种复杂的炎症疾病，它伴有各种程度的气流受限，气道高反应性和气道炎症。它通常是由环境因素及炎症刺激相互结合引起的。最近人们研究发现线粒体的功能和哮喘的发病机制有着密切的联系，他们发现线粒体的功能紊乱在哮喘的气道炎症中起重要作用。那么在哮喘中线粒体会发生怎样的变化呢？本次研究我们将通过哮喘动物模型及对线粒体超微结构的观察来探讨哮喘中线粒体结构发生了哪些变化及这些变化在哮喘中的作用。线粒体是细胞内的一种重要的细胞器，是细胞内的能量发生器,可以产生机体需要的ATP。同时，线粒体也在其他许多生理过程中都扮演着重要角色,例如在体内葡萄糖的代谢作用以及在细胞水平上影响胞内的钙信号、活性氧(ROS)的生成和细胞凋亡等。那么，线粒体对胞质内钙信号和活性氧会有怎样的调控作用呢？研究已表明生理状态下，线粒体通过释放和摄取的途径对胞浆内钙离子浓度进行调控，而胞浆的钙稳态对维持细胞内的正常生理活动有重要作用，当外界炎症刺激导致线粒体破坏时，细胞内钙稳态会破坏，钙离子浓度会产生波动。本次研究我们通过测定炎症刺激时钙离子浓度的变化来探讨钙离子和炎症的关系。同样，线粒体的破坏也与活性氧的含量呈正相关，外界炎症刺激细胞导致各种自由基的释放，它们会损伤线粒体，导致线粒体超微结构的改变，被破坏的线粒体会通过氧化磷酸化产生过量的活性氧，大量的活性氧会对细胞组织造成损伤，这对气道上皮细胞的纤毛摆动频率会产生怎样的影响呢？本次研究将通过测定不同浓度活性氧模拟物过氧化氢对细胞纤毛摆动频率的影响来反映活性氧与炎症的关系。 目的 本次研究利用卵清蛋白来激发大鼠建立哮喘模型，并对大鼠的气道反应性和各项炎症反应进行检测；通过电镜观察气道线粒体的结构来说明线粒体在支气管哮喘中的变化；通过测定炎症刺激时气道上皮细胞中活性氧及钙离子浓度以及不同浓度活性氧对气道上皮细胞纤毛摆动频率的影响来反映线粒体损伤在支气管哮喘中的机制。 方法 1.用卵清蛋白来激发大鼠建立哮喘模型，利用呼吸机和多导生理信号记录仪来测定大鼠气道反应性，利用共聚焦显微镜测定纤毛摆动频率，用瑞士染色涂片对嗜酸性粒细胞进行计数，，用ELISA对血浆中炎症因子进行检测，用HE染色分析分析大鼠的肺组织形态， 2.用电镜来观察线粒体超微结构， 3.用IL-4刺激气道上皮细胞并用共聚焦显微镜测定细胞内的ROS及钙离子浓度；用不同浓度的活性氧刺激细胞并测定纤毛摆动频率。 结果 1.与正常组大鼠相比，哮喘大鼠气道反应性明显增高（P0.05）；气道结构发生重构，气管管壁明显增厚（P0.05）,管壁周围有大量炎症细胞浸润；气管上皮纤毛摆动频率降低；肺泡灌洗液中嗜酸性粒细胞数量明显增多；血浆中IL-8和TNF-的含量明显增高（p0.05）；这些结果说明哮喘时大鼠机体发生了炎症反应; 2.与正常组大鼠相比，哮喘组大鼠气道内线粒体的超微结构发生改变，出现明显的肿胀与破坏；说明哮喘时气道内线粒体结构发生破坏，其导致的功能损伤在哮喘的发病机制起重要作用; 3.与正常组细胞相比，IL-4刺激的气道上皮细胞其活性氧含量与细胞内钙离子浓度明显增高（P0.05），其纤毛摆动频率明显降低（P0.05）；用不同浓度H2O2刺激细胞时，细胞纤毛摆动频率随浓度的增高而降低，均低于正常组（P0.05）。 结论 上述结果表明哮喘时线粒体的结构会发生明显的破坏，损伤后的线粒体会导致胞浆内钙离子浓度的升高及大量活性氧的释放，活性氧的释放会导致纤毛摆动频率的降低，使其无法维持正常的呼吸道功能，这将进一步加重炎症，导致哮喘迁延不愈。
[Abstract]:Bronchial asthma is a complex inflammatory disease accompanied by various degrees of airflow restriction, airway hyperresponsiveness and airway inflammation. It is usually caused by the combination of environmental factors and inflammatory stimuli. Recent studies have found that the function of mitochondria is closely related to the pathogenesis of asthma. They have found that the function of mitochondria is closely related to the pathogenesis of asthma. Disturbance plays an important role in airway inflammation in asthma. So what happens to mitochondria in asthma? In this study, we will explore the changes of mitochondrial structure in asthma and the role of these changes in asthma through asthma animal models and observation of mitochondrial ultrastructure. An important organelle is an intracellular energy generator that produces the ATP needed by the body. At the same time, mitochondria also play an important role in many other physiological processes, such as glucose metabolism in vivo, calcium signaling at the cellular level, production of reactive oxygen species (ROS) and cell apoptosis. How does mitochondria regulate intracytoplasmic calcium signaling and reactive oxygen species? Studies have shown that under physiological conditions, mitochondria regulate intracytoplasmic calcium concentration by means of release and uptake, and cytoplasmic calcium homeostasis plays an important role in maintaining normal cellular physiological activities, when inflammation stimulates mitochondria. In this study, we investigated the relationship between calcium ion and inflammation by measuring the changes of calcium ion concentration during inflammatory stimulation. Similarly, the destruction of mitochondria is positively correlated with the content of reactive oxygen species (ROS). External inflammation stimulates the release of various free radicals in cells, and they Mitochondria will be damaged, resulting in ultrastructural changes in mitochondria, the damaged mitochondria will produce excessive reactive oxygen species through oxidative phosphorylation, a large number of reactive oxygen species will cause damage to the cell tissue, which will affect the frequency of cilia swing in airway epithelial cells? The effect of hydrogen peroxide on cell cilia oscillation frequency reflects the relationship between reactive oxygen species and inflammation.
objective
In this study, ovalbumin was used to stimulate asthma model in rats, and the airway reactivity and inflammatory reactions were detected; the structure of airway mitochondria was observed by electron microscopy to explain the changes of mitochondria in bronchial asthma; the concentration of reactive oxygen species (ROS) and calcium ions in airway epithelial cells were measured by inflammatory stimulation. And the effect of different concentrations of reactive oxygen species on cilia oscillation frequency of airway epithelial cells to reflect the mechanism of mitochondrial damage in bronchial asthma.
Method
1. Establish asthma model in rats stimulated by ovalbumin, measure airway responsiveness by respirator and multi-channel physiological signal recorder, measure cilia swing frequency by confocal microscope, count eosinophils by Swiss staining smear, detect inflammatory factors in plasma by ELISA, and analyze by HE staining. Lung tissue morphology in rats
2. electron microscopy was used to observe the ultrastructure of mitochondria.
3. The airway epithelial cells were stimulated by IL-4 and the intracellular concentrations of ROS and Ca2+ were measured by confocal microscopy, and the ciliary oscillation frequencies were measured by reactive oxygen species with different concentrations.
Result
1. Compared with the normal group, the airway responsiveness of asthmatic rats was significantly increased (P 0.05); the airway structure was reconstructed, the tracheal tube wall was significantly thickened (P 0.05), there were a large number of inflammatory cells infiltrated around the tracheal wall; the frequency of ciliary oscillation of tracheal epithelium was decreased; the number of eosinophils in alveolar lavage fluid was significantly increased; the contents of IL-8 and TNF-in plasma were significantly increased. The amount was significantly higher (P0.05); these results indicated that inflammation occurred in asthmatic rats.
2. Compared with normal rats, the ultrastructure of airway mitochondria in asthmatic rats was changed, and there was obvious swelling and destruction.
3. Compared with the normal group, the content of reactive oxygen species (ROS) and intracellular calcium concentration of IL-4-stimulated airway epithelial cells were significantly increased (P 0.05), and the frequency of ciliary oscillation was significantly decreased (P 0.05).
conclusion
These results suggest that the structure of mitochondria in asthma may be destroyed obviously, and the damaged mitochondria will lead to the increase of calcium concentration and the release of a large number of reactive oxygen species in the cytoplasm. The release of reactive oxygen species will lead to the decrease of ciliary oscillation frequency, making it unable to maintain normal respiratory function, which will further aggravate inflammation and lead to asthma. Delay is not healed.
【学位授予单位】：华中科技大学
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R562.25
[Abstract]:Bronchial asthma is a complex inflammatory disease accompanied by various degrees of airflow restriction, airway hyperresponsiveness and airway inflammation. It is usually caused by the combination of environmental factors and inflammatory stimuli. Recent studies have found that the function of mitochondria is closely related to the pathogenesis of asthma. They have found that the function of mitochondria is closely related to the pathogenesis of asthma. Disturbance plays an important role in airway inflammation in asthma. So what happens to mitochondria in asthma? In this study, we will explore the changes of mitochondrial structure in asthma and the role of these changes in asthma through asthma animal models and observation of mitochondrial ultrastructure. An important organelle is an intracellular energy generator that produces the ATP needed by the body. At the same time, mitochondria also play an important role in many other physiological processes, such as glucose metabolism in vivo, calcium signaling at the cellular level, production of reactive oxygen species (ROS) and cell apoptosis. How does mitochondria regulate intracytoplasmic calcium signaling and reactive oxygen species? Studies have shown that under physiological conditions, mitochondria regulate intracytoplasmic calcium concentration by means of release and uptake, and cytoplasmic calcium homeostasis plays an important role in maintaining normal cellular physiological activities, when inflammation stimulates mitochondria. In this study, we investigated the relationship between calcium ion and inflammation by measuring the changes of calcium ion concentration during inflammatory stimulation. Similarly, the destruction of mitochondria is positively correlated with the content of reactive oxygen species (ROS). External inflammation stimulates the release of various free radicals in cells, and they Mitochondria will be damaged, resulting in ultrastructural changes in mitochondria, the damaged mitochondria will produce excessive reactive oxygen species through oxidative phosphorylation, a large number of reactive oxygen species will cause damage to the cell tissue, which will affect the frequency of cilia swing in airway epithelial cells? The effect of hydrogen peroxide on cell cilia oscillation frequency reflects the relationship between reactive oxygen species and inflammation.
objective
In this study, ovalbumin was used to stimulate asthma model in rats, and the airway reactivity and inflammatory reactions were detected; the structure of airway mitochondria was observed by electron microscopy to explain the changes of mitochondria in bronchial asthma; the concentration of reactive oxygen species (ROS) and calcium ions in airway epithelial cells were measured by inflammatory stimulation. And the effect of different concentrations of reactive oxygen species on cilia oscillation frequency of airway epithelial cells to reflect the mechanism of mitochondrial damage in bronchial asthma.
Method
1. Establish asthma model in rats stimulated by ovalbumin, measure airway responsiveness by respirator and multi-channel physiological signal recorder, measure cilia swing frequency by confocal microscope, count eosinophils by Swiss staining smear, detect inflammatory factors in plasma by ELISA, and analyze by HE staining. Lung tissue morphology in rats
2. electron microscopy was used to observe the ultrastructure of mitochondria.
3. The airway epithelial cells were stimulated by IL-4 and the intracellular concentrations of ROS and Ca2+ were measured by confocal microscopy, and the ciliary oscillation frequencies were measured by reactive oxygen species with different concentrations.
Result
1. Compared with the normal group, the airway responsiveness of asthmatic rats was significantly increased (P 0.05); the airway structure was reconstructed, the tracheal tube wall was significantly thickened (P 0.05), there were a large number of inflammatory cells infiltrated around the tracheal wall; the frequency of ciliary oscillation of tracheal epithelium was decreased; the number of eosinophils in alveolar lavage fluid was significantly increased; the contents of IL-8 and TNF-in plasma were significantly increased. The amount was significantly higher (P0.05); these results indicated that inflammation occurred in asthmatic rats.
2. Compared with normal rats, the ultrastructure of airway mitochondria in asthmatic rats was changed, and there was obvious swelling and destruction.
3. Compared with the normal group, the content of reactive oxygen species (ROS) and intracellular calcium concentration of IL-4-stimulated airway epithelial cells were significantly increased (P 0.05), and the frequency of ciliary oscillation was significantly decreased (P 0.05).
conclusion
These results suggest that the structure of mitochondria in asthma may be destroyed obviously, and the damaged mitochondria will lead to the increase of calcium concentration and the release of a large number of reactive oxygen species in the cytoplasm. The release of reactive oxygen species will lead to the decrease of ciliary oscillation frequency, making it unable to maintain normal respiratory function, which will further aggravate inflammation and lead to asthma. Delay is not healed.
【学位授予单位】：华中科技大学
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R562.25
【共引文献】
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