急性心肌梗死患者循环miRNA-21、miRNA-126的表达水平及其临床意义(7)

中文摘要研究背景：急性心肌梗死（acute myocardial infarction，AMI）是在冠状动脉病变的基础上，发生冠状动脉血供急剧减少或者中断，使相应的心肌严重而持久地急性缺血导致心肌坏死。

临床表现有持久的胸骨后剧烈疼痛、发热、白细胞计数和血清心肌坏死标志物增高以及心电图进行性改变；可发生心律失常、休克或心力衰竭，属冠心病的严重类型。

据2012年不完全统计，在我国每年约有54万人死于急性心肌梗死，平均每天约有1480人死于心肌梗死，平均每分钟就达1人。

近年来，心肌梗死人群朝着年轻化的趋势扩散。

对急性心肌梗死诊断、治疗、预后等方面的研究一直是心血管界的热点。

MicroRNA（miRNA）是近年来发现的一类广泛存在于真核生物的细胞内小的非编码RNAs。

目前国内外研究提示循环miRNA的水平在各类疾病中有显著变化，它们可能可以作为疾病的生物标志物，或者可以作为评估疾病严重程度和预后的指标。

本研究所针对的是miRNA-21和miRNA-126,它们与肿瘤疾病的相关研究已经比较成熟。

近年，它们与急性心肌梗死相关的动物、细胞学实验研究也开展起来。

目前实验提示miRNA-21可以在心肌梗死发生后减少心肌梗死的面积，而miRNA-126可以促进心肌梗死后侧枝循环的建立。

所以，研究miRNA-21和miRNA-126在急性心肌梗死中作用和机制并找到相应的作用靶点对心梗患者的诊断、预后评估、靶向治疗等方面有重要的作用。

但目前miRNA-21、miRNA-126这个研究主要集中在动物和细胞实验,在人体中的研究少见报道,而且样本量小,得出的结论不明确。

本研究纳入较大样本量，观察急性心肌梗死患者循环中miRNA-21和miRNA-126的表达是否有显著性的变化，以明确miRNA-21和miRNA-126表达的变化与急性心梗预后的相关性并探讨其可能机制，为心肌梗死的防治提供新的思路和靶点。

分别观察急性心肌梗死患者循环miRNA-21、miRNA-126的表达水平较健康人是否有显著差异。

不同发病时间组之间是否有显著差异。

它们的表达水平与患者发病时间、冠脉Gensini积分分值、LVEF值、MYO、CKMBII、cTnI、WBC 的关系。

方法：资料收集：选择2012年3月-2013年2月在我院进行治疗的急性心肌梗死患者74例为观察组，选择2012年3月-2013年2月在我院体检的健康人群50例为对照组，对这两组患者抽取肘静脉血样本。

血样本收集：如有行冠脉造影的患者血样本均在进行冠状动脉造影术前采集。

所有急性心肌梗死患者血标本抽取的同时记录距发病(胸痛)的时间，并将标本送至本院检验科检测MYO（肌红蛋白）、CK-MBII（肌酸激酶MB同工酶2）、cTnI （肌钙蛋白I）、血常规。

运用逆转录实时荧光定量PCR检测出它们循环miRNA-21、miRNA-126的相对表达量。

用适合的统计学方法作以下对比。

1.用独立样本T检验比较观察组与对照组中miRNA-21、miRNA-126的表达水平是否有显著差异。

2.对观察组进行细分，用独立样本T检验比较观察组中循环miRNA-21、miRNA-126的表达在距发病（胸痛发作）时间<12小时和>=12小时两组间是否有显著差异。

3.挑选出观察组中有相应资料的样本分别观察循环miRNA-21、miRNA-126的表达与MYO、CKMBII、cTnI、冠脉Gensini积分分值、LVEF值、WBC是否有相关性。

以上以P＜0.05标准作为差异有统计学意义。

计量资料用均数±标准差表示（1）观察组循环miRNA-21表达水平为1.8196±1.02, 显著高于对照组的1.1226±0.63(P<0.01)。

观察组循环miRNA-126水平为2.0716±1.21，显著高于对照组的1.253±0.61 (P<0.01)。

（2）观察组中发病时间<12小时组的循环miRNA-21水平为1.5823±1.01，发病时间>=12小时组的miRNA-21水平1.9814±0.93,两组间无显著差异(P>0.05)。

发病时间<12小时组的循环miRNA-126水平为1.8340±1.06，发病时间>=12小时组的miRNA-126水平2.2336±1.01,两组间无显著差异(P>0.05)。

（3）用Spearman相关分析显示急性心肌梗死患者循环中miRNA-21、miRNA-126表达量与患者MYO、CKMBII、cTnI、冠脉Gensini积分分值、LVEF 值、WBC无显著相关性（P>0.05)。

结论：1、急性心肌梗死患者循环miRNA-21及miRNA-126水平显著高于健康人群。

推测其升高可能为机体的一种代偿机制。

2、急性心肌梗死患者循环miRNA-21及miRNA-126水平与急性心梗的发病时间可能无明显相关性，在急性心肌梗死发病<12小时和>=12小时组的表达无显著差异。

3、急性心肌梗死患者循环中miRNA-21、miRNA-126水平与MYO、CKMBII、cTnI、冠脉Gensini积分分值、LVEF值、WBC无显著相关性。

关键词：miRNA-21；miRNA-126；心肌梗死ABSTRACTBackground:Acute myocardial infarction(AMI) is the disease which is characterized by that coronary blood supply is drastically reduced or interrupted on the basis of coronary artery disease,leading to severe acute myocardial ischemia,eventually leading to myocardial necrosis. As a serious type of coronary heart disease, it's clinical manifestations contains severe pain,fever,elevated white blood cell count and serum myocardium necrosis markers, progressive ECG changes, even arrhythmia,shock or heart failure.According to incomplete statistics, in 2012, there are about 540,000 people died of acute myocardial infarction every year in our country. In another word, there are about 1480 people died of myocardial infarction per day on average, which means that one person died of that disease per minute on average. In recent years, the trend of myocardial infarction (mi) develops towards younger. The research of diagnosis, treatment and prognosis for acute myocardial infarction has always been the hot spot in cardiovascular field.MicroRNA (miRNA), which was discovered in recent years, is a class of small non-coding RNAs which widely exists in eukaryotic cells. Overseas and domestic research suggest the expression of circulating MicroRNA in all kinds of disease has significant changes, which may can be served as the biomarker of disease, or the assessment of disease severity and the prognosis index.The relevant research about the miRNA - 21 and miRNA – 126 associated with tumor disease become more mature. Their target genes had also been obtained preliminary prediction and verification. The animal experiment research which discusses the relationship between them and acute myocardial infarction has been carried out. From the target genes in miRNA - 21 and miRNA – 126 which is acknowledged widely, we consider that their function is directly related to the pathological changes of acute myocardial infarction.That means the miRNA - 21 and miRNA – 126 may play an important role in thediagnosis, evaluation, treatment of patients with acute myocardial infarction.The research which discusses that the miRNA - 21 and miRNA - 126 is associated with cardiovascular clinical has been carried out, but there is few sample for research analysis about the expression of circulating miRNA - 21 and miRNA – 126 in patients with acute myocardial infarction (mi).Objective:1. Whether the expression in circulating miRNA –21 and miRNA –126 has significant difference between the patients with acute myocardial infarction (mi) and the healthy people.2. Whether the expression in circulating miRNA –21 and miRNA –126 has significant difference between the groups at different time.3. The relationship between the expression levels of Circulating miRNA -21 and miRNA - 126 in patients who died of acute myocardial infarction (mi) and chest pain time, coronary Gensini score, LVEF value, MYO, CKMBII, cTnI, the WBC.Methods:Data collection: Choose 74 cases of patients with acute myocardial infarction in our hospital for treatment in March 2011 - February 2013 as observation group, choose 50 cases of healthy people in our hospital for check-up in March 2011 - February 2013 as control group, to extract elbow venous blood samples from above two groups of patients.Blood samples collection: The learning sample of patients who have a line of coronary angiography all be collected before undergoing coronary angiography. When we drawn the blood sample from the patients with acute myocardial infarction, we also record the time from the onset of chest pain in patients at the same time, and send parts of them to our clinical laboratory to test MYO (myoglobin), CKMBII (creatine kinase MB isoenzyme 2), cTnI (troponin I). Some patients tested the blood routine at the same time.We use quantitative reverse transcription-PCR (qRT-PCR) to detect the relativeexpression of circulating miRNA -21 and miRNA - 126, and make the following comparison by appropriate statistical methods.1. Using independent sample T test to compare that whether the expression of the miRNA - 21 and miRNA –126 has significant difference between the observation group and control group.2. We subdivided the observation group, and used the independent sample T test to compare whether the expression of circulating miRNA –21 and miRNA - 126 has significant difference between the group which onset time is within 12 hours and the group onset time is more than 12 hours.3. Pick out the corresponding data samples from the observation group to observe whether the expression of circulating miRNA – 21 and miRNA - 126 has correlation with MYO, CKMBII, cTnI, coronary Gensini score, LVEF value and the WBC. Above methods which take P < 0.05 standard as the difference has statistical significance.Results:Measurement data expresses with mean± standard deviation.(1) In the observation group, the expression of circulating miRNA - 21 was 1.8196 ±1.02, which is significantly higher than the control group which shows 1.1226 ± 0.63 (P < 0.01). In the observation group, the expression of circulating miRNA - 126 was 2.0716 ±1.21, which is significantly higher than the control group which shows1.253±0.61 (P < 0.01).(2) In the observation group, the expression of circulating miRNA - 21 in the group which onset time is within 12 hours was 1.5823 ±1.01, and the expression of circulating miRNA - 21 in the group which onset time is more than 12 hours was 1.9814 ± 0.93; therefore, there is no significant difference between above two groups (P > 0.05). The expression of circulating miRNA - 126 in the group which onset time is within 12 hours was 1.8340±1.06, and the expression of circulating miRNA - 126 in the group which onset time is more than 12 hours was 2.2336±1.01; therefore, there is no significant difference between above two groups (P > 0.05).(3) Using Spearman test analysis to show that there is no significant correlation between the expression of circulating miRNA –21 and miRNA –126 and MYO, CKMBII, cTnI, coronary Gensini score, LVEF value, the WBC in patients with acute myocardial infarction (mi) (P > 0.05).Conclusion:1.Circulating miRNA -21 and miRNA - 126 in patients with acute myocardial infarction (mi) were significantly higher than healthy people. We speculated that its rise may be a compensatory mechanism for the body.2. The expression of Circulating miRNA -21 and miRNA - 126 in patients with acute myocardial infarction (mi) has no obvious difference between the group that onset time is within 12 hours and the group that onset time is more than 12 hours.3. Circulating miRNA -21 and miRNA - 126 in patients with acute myocardial infarction (mi) have no significant correlation with MYO, CKMBII, cTnI, coronary Gensini score, LVEF value and the WBC.Key words: miRNA-21; miRNA-126; Myocardial infarction (MI).第一章前言心血管疾病是目前主要的致死性疾病之一，而其中急性心肌梗死为较为常见的心血管疾病，它为猝死的首要病因。