DRP1和OPA1在高体积氧诱导早产大鼠肺损伤的作用

发布时间：2018-05-30 18:00

本文选题：高浓度氧 + 肺损伤　；参考：《泸州医学院》2013年硕士论文

【摘要】：目的研究DRP1（dynamin related protein1）和OPA1（optic atrophy1)在高体积氧诱导早产大鼠肺损伤时细胞凋亡中的作用。方法1.以早产大鼠肺组织细胞为研究对象。随机将早产Wistar大鼠48只分为高氧组和对照组，高氧组吸入95%的氧（建立高氧肺损伤模型）,对照组吸入21%的氧的常压空气。相同常规喂养1d、3d、7d分批离断早产鼠颈放血处死后取肺组织做石蜡切片。2.检测指标：采用苏木精-伊红（hematein eosin HE）染色在光镜下观察早产鼠肺组织的病理形态学变化；免疫组织化学链霉菌抗生物素蛋白-过氧化物酶连结法（streptavidin-peroxidase SP）观察与线粒体形态变化相关的蛋白：动力相关蛋白(dynamin related protein1DRPl)和视神经萎缩症蛋白(optic atrophyOPAl)在各组早产大鼠肺组织的表达和分布及其比值变化；采用原位末端转移酶标记技术（terminal-deoxynucleotidyl transferase mediated nick endlabeling TUNEL）评价各组早产大鼠肺组织细胞凋亡情况。3.统计学处理：应用SPSS16.0软件，所有数据以x±s表示，组间比较采用单因素方差分析，两组比较进行配对t检验和采用LSD法检验（a=0.05），指标间的关系采用直线相关分析进行处理,P0.05为差异有统计学意义。结果1.高氧组早产大鼠肺组织细胞可见典型的肺损伤病理形态学改变。在光镜下观察，对照组大鼠肺组织细胞紧密排列多角扁平状，呈鹅卵石样改变,透明度好，胞质中颗粒稀少。高氧组大鼠肺组织细胞数量随时间（1d、3d、7d）较对照组显著减少，细胞形态改变不规则，肺泡结构破坏明显，透光度减弱，，较多空泡、脂肪小滴、颗粒聚集物出现在胞质中，细胞间隙变大，细胞碎片大量填充细胞间，间隔增粗肺泡腔变小。2.免疫组织化学法测定DRP1表达：高氧组与对照组相比，高氧组细胞胞质中DRPl表达显著升高，差异有统计学意义(p0．05)。DRP1在高氧组平均光密度(Mean±SD)1d(1634.57±108.85),3d(8008.14±530.26),7d(13132.32±594.57);DRP1在对照组平均光密度值(Mean±SD)1d(789.86±58.95),3d(772.31±42.82),7d(770.39±67.8)。3.免疫组织化学法测定OPAl表达：高氧组与对照组相比,高氧组细胞胞质中OPA1表达显著降低，差异有统计学意义(p0.05)。OPA1在高氧组平均光密度值(Mean±SD)1d(3126.91±264.91),3d(1399.81±268.62),7d(579.76±99.20);OPA1在对照组平均光密度值(Mean±SD)1d(3935.42±263.99),3d(3764.15±281.53),7d(3907.80±231.34)。4.与对照组相比,高氧组DRP1/OPA1表达的比值呈时间依赖性增高，差异有统计学意义(p0．05):高氧组DRP1/OPA1的比值表达(Mean±SD)1d(0.52±0.01),3d(5.72±0.05),7d(22.68±0.07);对照组DRP1/OPA1的比值表达(Mean±SD)1d (0.200±0.01),3d(0.205±0.01),7d（0.197±0.01）。5.对照组中可见少量黄色或棕黄色的TUNEL阳性细胞，高氧组可见大量TUNEL阳性细胞出现在支气管上皮细胞和肺泡上皮细胞中。高氧暴露时间越长，TUNEL阳性细胞越多，细胞凋亡指数增高,细胞凋亡随高氧暴露时间有增加趋势:高氧组凋亡指数1d(26.39±1.45),3d(43.87±1.62)，7d(56.34±1.56)；对照组凋亡指数1d(14.54±1.88),3d(14.68±2.01),7d(14.45±1.75)。6.在高氧组，DRP1/OPA1比值表达与细胞凋亡指数相比，呈显著正相关（r=0.725，P0.01）。结论1.高氧暴露导致早产鼠肺组织产生急性肺损伤病理学改变，这一形态学改变表现出与高氧暴露有时间依赖性。2.高氧暴露可致新生大鼠肺组织细胞凋亡，其机制通过提高DRP1和降低OPA1的表达水平致线粒体融合分离失衡使线粒体片段化加强促进细胞凋亡，参与高氧肺损伤。3.DRP1/OPA1蛋白比值表达增高变化与高氧暴露有时间依赖性，细胞凋亡指数随高氧暴露时间有上升趋势，DRP1/OPA1比值与细胞凋亡指数相比具有显著正相关性。
[Abstract]:Objective to study the role of DRP1 (dynamin related protein1) and OPA1 (optic atrophy1) in the apoptosis of lung injury induced by high-volume oxygen in premature rats.
Methods 1. the preterm rat lung tissue cells were studied. 48 Wistar rats were randomly divided into hyperoxia group and control group. The hyperoxia group inhaled 95% oxygen (high oxygen lung injury model), and the control group inhaled 21% oxygen atmospheric pressure. The same routine feeding 1D, 3D, 7d batches of premature rat neck bleeding after the death of the lung tissue to make paraffin cut .2. detection index: the pathological changes of lung tissue in premature rats were observed under the light microscope with hematoxylin and eosin (Hematein eosin HE) staining; the protein of immuno histochemical Streptomyces anti biotin peroxidase (streptavidin-peroxidase SP) was used to observe the protein of mitochondrial morphologic changes: dynamic related protein (dyn Amin related protein1DRPl) and optic dystrophy protein (optic atrophyOPAl) expression and distribution of lung tissue in each group of preterm rats and the changes in their ratio. The apoptosis of lung tissue in each group of preterm rats was evaluated by in situ terminal transferase labeling technique (terminal-deoxynucleotidyl transferase mediated Nick endlabeling TUNEL) State.3. statistics: using SPSS16.0 software, all the data were expressed in X + s, and the single factor variance analysis was used in the group. The two groups were compared with the paired t test and LSD test (a=0.05). The relationship between the indexes was treated with linear correlation analysis, and P0.05 was statistically significant.
Results the lung tissue cells in the 1. hyperoxic group showed typical pulmonary pathological changes. Under the light microscope, the lung tissue cells in the control group were closely arranged and flattened, with cobblestone like changes, the transparency was good, and the particles in the cytoplasm were scarce. The number of lung tissue cells in the hyperoxic group was significantly higher than that of the control group (1D, 3D, 7D). Decreased cell morphology changed irregularly, alveolar structure was destroyed obviously, light transmittance was weakened, more vacuoles, fat droplets, granular aggregates appeared in the cytoplasm, cell space became larger, cell fragments were filled with large numbers of cells, and spaced thickening alveolar cavity changed small.2. immunohistochemical method to determine DRP1 expression: hyperoxia group compared with control group, hyperoxia group was fine. The expression of DRPl in cytoplasm was significantly higher, the difference was statistically significant (P0.05).DRP1 in the average light density (Mean + SD) 1D (1634.57 + 108.85), 3D (8008.14 + 530.26), 7d (13132.32 + 594.57), DRP1 in the control group (Mean + SD) 1D (789.86 + 58.95), (772.31 + 42.82), 770.39 + 67.8) immunohistochemical method Al expression: compared with the control group, the expression of OPA1 in the cytoplasm of hyperoxia group decreased significantly, and the difference was statistically significant (P0.05).OPA1 in the hyperoxic group (Mean + SD) 1D (3126.91 + 264.91), 3D (1399.81 + 268.62), 7d (579.76 + 99.20), OPA1 in the control group (3935.42 + 263.99) (3935.42 + 263.99), 3764.15 + 281 .53), 7d (3907.80 + 231.34).4. compared with the control group, the ratio of DRP1/OPA1 expression in the hyperoxic group increased in time dependent, and the difference was statistically significant (P0.05): the ratio of DRP1/OPA1 in the hyperoxic group (Mean + SD) 1D (0.52 + 0.01), 3D (5.72 + 0.05), 7d (22.68 + 0.07), and the ratio of DRP1/OPA1 in the control group (0.200 + 0.01), 0.205 + 0. 01) a small amount of yellow or brown yellow TUNEL positive cells were found in the 7d (0.197 + 0.01).5. control group. A large number of TUNEL positive cells appeared in the bronchial epithelial cells and alveolar epithelial cells in the hyperoxia group. The longer the exposure time of the hyperoxia, the more TUNEL positive cells, the increase of apoptosis index, and the increase of apoptosis with the time of hyperoxia exposure. Potential: apoptosis index of hyperoxia group (26.39 + 1.45), 3D (43.87 + 1.62), 7d (56.34 + 1.56), apoptosis index 1D (14.54 + 1.88), 3D (14.68 + 2.01), 7d (14.45 + 1.75).6. in hyperoxia group. The expression of DRP1/OPA1 ratio was significantly positively correlated with apoptosis index (r=0.725, P0.01).
Conclusion 1. hyperoxia exposure leads to the pathological changes of acute lung injury in preterm rat lung tissue. This morphological change shows that the time dependent.2. hyperoxia exposure can induce apoptosis in the lung tissue of newborn rats. The mechanism by raising the expression of DRP1 and reducing the expression of OPA1 to the mitochondrial fusion separation disequilibrium causes the mitochondrial fragment. The increase of.3.DRP1/OPA1 protein ratio in hyperoxia lung injury is dependent on hyperoxia exposure, and the apoptotic index increases with the exposure time of hyperoxia, and the DRP1/OPA1 ratio has a significant positive correlation with the apoptosis index.
【学位授予单位】：泸州医学院
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R722.6

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