硫化氢对高肺血流性肺动脉高压大鼠内质网应激的调节作用

发布时间：2018-03-11 22:36

  本文选题：硫化氢　切入点：高肺血流性肺动脉高压　出处：《吉林大学》2011年硕士论文　论文类型：学位论文

【摘要】：目的：高肺血流性肺动脉高压是左向右分流型先天性心脏病的常见并发症,其严重程度直接影响患者手术时机、手术成功率及术后长期生活质量,严重威胁人类的健康和生命,但有关其发病机制尚未完全阐明。近来研究发现新型气体信号分子硫化氢(hydrogen sulfide, H2S)对心血管系统具有重要的保护作用,能够减轻高肺血流性肺动脉高压的形成。内质网应激(endoplasmic reticulum stress, ERS)作为众多心血管疾病发生发展的重要病理机制,近年来日益受到人们的广泛关注。既往研究认为,H2S能够通过调节内质网应激减轻组织的损伤程度,但H2S与ERS在高肺血流性肺动脉高压的发生发展过程中有无关系及其具体机制尚不清楚。本实验以高肺血流性肺动脉高压大鼠为研究对象,探讨H2S对高肺血流性肺动脉高压大鼠内质网应激的影响,进而对其机制做进一步研究。 方法：选取体重140g～160g的健康雄性SD大鼠52只,随机分为六组：对照组(n=8)、对照+硫氢化钠(sodium hybrosulfide, NaHS)组(n=8)对照+炔丙基甘氨酸(propargy lglycine, PPG)组(n=8)、分流组(n=10)、分流+NaHS组(n=10)、分流+PPG组(n=8)。对分流组、分流+NaHS组及分流+PPG组大鼠行腹主动脉壁-下腔静脉穿刺术,造成腹主动脉与下腔静脉瘘道建立左向右分流肺动脉高压动物模型。分别在终止实验前以右心导管测定肺动脉平均压(MPAP),并取肺组织,光镜下计算肺血管中肌性动脉(MA)、部分肌性动脉(PMA)和非肌性血管(NMA)百分比；以敏感硫电极法检测肺组织硫化氢(H2S)含量、肺组织硫化氢生成酶(CSE)的活性；原位缺口末端标记(TUNEL)检测平滑肌细胞的凋亡、采用免疫组织化学染色方法检测肺动脉组织中内质网应激标志糖调节蛋白78(GRP78)、内质网相关性凋亡标志蛋白半胱天冬水解酶-12(caspasel2)和磷酸化的真核细胞蛋白质翻译起始复合体(p-eIF2a)的蛋白表达,并用全自动图像分析系统进行半定量分析。 结果： (1)分流8周时各组肺动脉平均压(MPAP)比较有统计学意义(P0.01)。分流组MPAP明显高于对照组(P0.01)；与分流组比较,分流+NaHS组MPAP明显降低(P0.01)；分流+PPG组MPAP明显升高(P0.01)。 (2)与对照组相比,分流组肺小动脉MA、PMA占肺小血管总数的百分比明显升高,NMA占肺小血管总数的百分比降低。与对照组相比,分流组CSE活性均明显高于对照组(P0.05)。 (3)分流8周时,各组大鼠中、小型肺动脉平滑肌细胞中均有凋亡细胞存在。与对照组相比,分流组肺动脉平滑肌细胞凋亡率明显减少(P0.05)；与分流组比较,分流+NaHS组肺动脉平滑肌细胞凋亡率明显增加(P0.01),分流+PPG组肺动脉平滑肌细胞凋亡率明显减少(P0.01)。 (4)各组大鼠肺动脉内质网应激标志蛋白GRP78、caspase12和p-eIF2a表达结果：①与对照组相比,分流组大鼠肺动脉GRP78、caspase12和p-eIF2a表达均明显增高(P0.05)；与分流组相比,分流+NaHS组大鼠肺动脉GRP78、caspase12和p-eIF2a表达明显降低(P0.05)；与分流组相比,分流+PPG组大鼠肺动脉caspase12和p-eIF2a蛋白表达显著升高(P0.05)。②与对照组相比,对照+PPG组大鼠肺动脉GRP78和caspase12表达明显升高(P0.01)；对照+NaHS组大鼠caspase12表达明显降低(P0.05)。 结论：腹主动脉-下腔静脉分流8周所致高肺血流性肺动脉高压大鼠肺动脉中内质网应激显著增强。H2S可通过抑制过度内质网应激反应拮抗高肺血流性肺动脉高压和肺血管结构重建形成。
[Abstract]:Objective: high flow pulmonary hypertension is a common complication of congenital heart disease with left to right shunt and the severity of patients directly affect the timing of surgery, surgical success rate and long-term life quality after the operation, a serious threat to human health and life, but its pathogenesis has not been fully elucidated. A recent study found that the new gas signal molecular hydrogen sulfide (hydrogen sulfide, H2S) has an important protective effect on the cardiovascular system, can reduce the formation of high flow pulmonary hypertension. Endoplasmic reticulum stress (endoplasmic reticulum, stress, ERS) as an important pathological mechanism of the occurrence and development of many cardiovascular diseases, in recent years has been widespread concern. Previous studies suggest that H2S can through the regulation of endoplasmic reticulum stress and tissue injury, but H2S and ERS in the occurrence and development of high flow pulmonary hypertension in No relationship and its specific mechanism are not yet clear. In this experiment, we studied the effects of H2S on endoplasmic reticulum stress in rats with pulmonary hypertension induced by high pulmonary hemodynamics, and further studied the mechanism.
Methods: the weight of 140g to 160g in 52 healthy male SD rats, were randomly divided into six groups: control group (n=8), control + sodium hydrosulfide (sodium hybrosulfide NaHS) control group (n=8) + propargylglycine (propargy, lglycine, PPG) group (n=8), shunt group (n=10) and shunt +NaHS group (n=10), +PPG group (n=8). The shunt of the shunt group and shunt +NaHS group and shunt +PPG group rats underwent abdominal aorta and inferior vena cava puncture caused by abdominal aorta and inferior vena cava by establishing animal pulmonary hypertension with left to right shunt model. Respectively before the end of experiment with right heart catheterization determination of mean pulmonary arterial pressure (MPAP), and the lung tissue, calculation of muscular artery in pulmonary arteries under light microscope (MA), part of the muscular artery (PMA) and non muscular vessels (NMA) to detect the percentage of lung tissue; sulfur sensitive electrode method of hydrogen sulfide (H2S) content in lung tissue generating enzyme (CSE) the in situ nick end activity; End labeling (TUNEL) detection of apoptosis of smooth muscle cells, immunohistochemical staining method to detect the pulmonary tissue in the endoplasmic reticulum stress marker glucose regulated protein 78 (GRP78), endoplasmic reticulum associated protein markers of apoptosis caspase -12 hydrolase (caspasel2) complex eukaryotic translation initiation protein and phosphorylated protein (p-eIF2a) the expression of semi quantitative analysis and automatic image analysis system.
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