循环microRNA用于低氧性肺动脉高压早期诊断的实验研究

发布时间：2018-05-30 16:01

本文选题：低氧性肺动脉高压 + 循环miRNAs　；参考：《石河子大学》2015年硕士论文

【摘要】：研究目的的::模拟6,000米海拔低压低氧条件,建立低压低氧诱发肺动脉高压(HPH)的大鼠模型,研究低压低氧暴露条件下HPH模型大鼠循环mi RNA表达谱特征及其与HPH病理变化的相关性,探索循环miRNA作为HPH诊断生物标志物的可行性。研究方法:1.随机将120只雄性6周龄SD大鼠分为低氧4d、7d、10d、14d和21d暴露组及各应时点对照组,置人工气候舱常规饲养,继以模拟6,000m海拔进行低压低氧暴露,按分组时点定时采用直接测压法检测平均肺动脉压(m PAP)并采集血液及组织标本,心肺组织标本经固定、包埋后进行HE染色及病理分析。2.依据低压低氧后m PAP的变化,取低氧14d暴露组及对照组大鼠血清标本,利用基因芯片技术进行循环mi RNA表达谱检测,不同组别间循环mi RNA表达谱差异分析采用聚类分析。3.利用荧光定量PCR技术对候选循环mi RNA进行验证,采用相对定量法进行数据分析。研究结果:1.低压低氧条件暴露10d、14d、21d的SD大鼠m PAP、右心室肥厚指数(RVHI)及肺小动脉血管壁厚占外径比值(MT%)较同时点对照组显著升高(P0.01)。2.基因芯片结果显示,累计从大鼠血清中检测到723种循环mi RNAs,按照FC2、Flag/Call值3及P0.05的标准,从低氧14d组及对照组大鼠间筛选出6种差异化表达的mi RNAs,其中低氧组中表达上调的mi RNA有3种(let-7d、mi R-451、mi R-505),表达下调的mi RNA有3种(mi R-455-3p、mi R-455-5P、mi R-214)。3.荧光定量PCR定量验证结果显示,与对照组相比,低氧14d组大鼠血清中mi R-451、mi R-505、let-7d表达上调(P0.01),mi R-214表达下调(P0.05),mi R-455和mi R-455-5p表达与对照组相比无显著差异;ROC曲线表明,mi R-451、mi R-505及let-7d的用于鉴别低氧性HPH大鼠和对照组大鼠的曲线下面积(AUC)均大于0.8,且与机体脂质过氧化指标丙二醛(MDA)含量均正相关。结论:模拟海拔6000m高原环境低压低氧14d即可成功建立低压低氧诱发肺动脉高压的大鼠模型;基于大鼠模型,获得了低压低氧暴露条件下HPH模型大鼠循环mi RNA表达谱,初步发现4种循环mi RNA在HPH大鼠和对照组大鼠间存在显著性差异化表达,可能与HPH病理变化相关。
[Abstract]:The purpose of this study is to simulate the hypobaric hypoxia conditions of 6000 m, establish a rat model of pulmonary hypertension (HPH) induced by low pressure hypoxia, and study the characteristics of MI RNA expression in HPH model rats under hypobaric hypoxia exposure and the correlation with the pathological changes of HPH, and explore the feasibility of circulating miRNA as a biomarker for HPH diagnosis. Method: 1. the 120 male 6 week old SD rats were randomly divided into hypoxia 4D, 7d, 10d, 14d and 21d exposure group and each time point control group, and the artificial climate cabin was routinely raised. The average pulmonary arterial pressure (m PAP) was detected by direct pressure measurement at the time point of the group at the time point of the group, and the blood and tissue specimens were collected, and the blood and tissue specimens were collected and the heart and lung were collected. The tissue specimens were fixed, embedded after HE staining and pathological analysis,.2. based on the changes of M PAP after hypobaric hypoxia, the serum samples of hypoxia 14d exposure group and control group were taken, the expression of circulating mi RNA expression was detected by gene chip technology. The difference analysis of RNA expression spectrum of circulating mi between different groups was analyzed by cluster analysis.3. using fluorescence quantitative PCR. A relative quantitative method was used to verify the candidate cycle mi RNA. The results were as follows: 1. the low oxygen and hypoxia conditions exposed 10d, 14d, 21d SD rats, m PAP, the right ventricular hypertrophy index (RVHI) and the ratio of the diameter of the pulmonary artery wall thickness (MT%) significantly higher than those in the same group (P0.01).2. gene chip results showed the cumulative from In the rat serum, 723 kinds of circulating mi RNAs were detected. According to the standard of FC2, Flag/Call value 3 and P0.05, 6 kinds of differential expression mi RNAs were screened from the hypoxia 14d group and the control group, of which there were 3 kinds of MI RNA in the hypoxia group. There were 3 kinds of downregulation. Compared with the control group, the serum mi R-451, MI R-505, let-7d expression up regulated (P0.01) and MI R-214 expression decreased (P0.05) compared with the control group, and there was no significant difference in the expression of MI R-214 expression between the 14d and the control group. The area under the curve (AUC) of the control group was more than 0.8, and it was positively correlated with the content of malondialdehyde (MDA) in the lipid peroxidation index of the body. Conclusion: the rat model of pulmonary hypertension induced by hypobaric hypoxia at the altitude of 6000m plateau can be successfully established. Based on the rat model, the HPH model under the condition of low pressure and hypoxia exposure is obtained. The expression of MI RNA in rats circulated, and it was found that there was a significant difference in the expression of 4 kinds of circulating mi RNA between the HPH rats and the control rats, which may be related to the pathological changes of HPH.
【学位授予单位】：石河子大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R544.1;R440

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