短暂性脑缺血发作患者血清microRNA的水平变化及其临床意义

发布时间：2018-08-17 16:11

【摘要】：短暂性脑缺血性发作(transient ischemic attacks,TIA)是指由多种原因引发局灶性脑缺血,进而导致的突发性、短暂性、可逆性神经功能障碍。TIA易反复发作,既往有一过性TIA病史的患者再次发生脑卒中的风险明显增加,致残率、病死率更高。血清微小核糖核酸(microRNAs,miRNAs)是近年来的研究热点,与缺血性脑血管疾病的发生、发展密切相关,但有关TIA患者血清miRNAs表达水平及其变化尚缺乏系统性的研究,血清miRNAs对TIA预测及TIA后再发卒中风险评估的临床价值仍有待进一步研究探讨。目的:通过分析TIA患者血清miRNAs水平变化,筛选TIA患者血清特异变化的miRNAs,比较并探讨血清miRNAs及现有脑损伤相关血清学标志物,对TIA预测及TIA后再发卒中风险评估的临床价值,以期为临床TIA患者的病情评估与监测提供新的途径与理论依据。方法:选取126例TIA患者和44例对照者,依据TIA患者再发卒中风险评估体系ABCD3-I评分,将其分为低危(0～3分,46例)、中危(4～7分,45例)、高危(8～13 分,35 例)三组。采用实时荧光定量 PCR(quantitative reverse-transcription PCR,qRT-PCR)技术,检测候选的13种与缺血性脑血管疾病相关的miRNAs,在TIA患者和对照者血清中的相对表达水平(包括miR-15b、miR-30a、miR-21、miR-29b、miR-181 a、miR-124、miR-335、miR-146a、miR-499、miR-144、miR-208b、miR-215和miR-23b),以筛选TIA患者血清特异变化的miRNAs。同时,检测TIA患者脑损伤相关血清学标志物的水平,包括血清脂蛋白相关磷脂酶A2(Lipoprotein-associated phospholipase A2,Lp-PLA2)、氧化低密度脂蛋白(Oxidized low-density lipoprotein,ox-LDL)、神经元特异性烯醇化酶(Neuron-Specific Enolase,NSE)和脑源性神经营养因子(Brain Derived Neurotrophic Factor,BDNF)。采用 Spearman 相关性分析,探讨血清 miRNAs 和脑损伤相关血清学标志物之间的相关性。采用受试者工作曲线(receiver operating characteristic curves,ROC)分析和多元 Logistic 回归分析,比较并探讨血清miRNAs及脑损伤相关血清学标志物应用于TIA预测及TIA后再发卒中风险评估的临床价值。结果:一、qRT-PCR 结果显示,TIA 患者组血清 miR-23b-3p、miR-208b、miR-215、miR-181a-5p水平较对照组均显著升高(P均0.05);TIA患者中,高危组、中危组、低危组的血清 miR-23b-3p、miR-208b、miR-215、miR-181a-5p 水平也较对照组均显著升高(P均0.05)。相关性分析显示,上述四种miRNAs水平两两之间均呈显著正相关(P均0.05)。二、TIA患者组血清Lp-PLA2、ox-LDL及NSE水平较对照组均显著升高(P均0.05);TIA患者中,仅高危组患者血清Lp-PLA2、ox-LDL及NSE水平较对照组显著升高(P均0.05)。相关性分析显示,血清Lp-PLA2与ox-LDL水平呈显著正相关(r= 0.264,P=0.002)、NSE与BDNF水平呈显著正相关(r=0.197,P=0.041)。三、ROC曲线分析结果显示,血清miR-23b-3p、miR-208b、miR-215和miR-181a-5p区分TIA患者与对照者的曲线下面积(area under the curve,AUC)分别为 0.784、0.795、0.768 和 0.779(P 均0.01),四种 miRNAs 的组合区分 TIA患者与对照者的AUC为0.812(P=0.001);血清Lp-PLA2和NSE区分TIA患者的AUC分别为0.725和0.743(P均0.05),两指标的组合区分TIA患者与对照者的AUC为0.817(P=0.001);四种miRNAs联合Lp-PLA2和NSE的组合区分TIA患者与对照者的AUC为0.925(P0.001)。四、以TIA患者不同危险分层组及对照组作为独立的四分类变量,以对照组作为参照类别,分别对血清miRNAs和脑损伤相关指标进行多元Logistic回归分析。单因素Logistic回归分析结果显示,血清高miR-23b-3p、miR-208b、miR-215水平均与TIA患者发生卒中的各层风险相关;血清高Lp-PLA2、ox-LDL、NSE及BDNF水平均与TIA患者发生卒中的高风险相关,高NSE水平还与TIA患者发生卒中的中、低风险相关。多因素Logistic回归分析结果显示,在调整年龄、性别、血压和血脂参数的影响后,血清miR-23b-3p水平的升高仍与TIA患者发生卒中的高、中风险密切相关,miR-208b水平的升高仍与TIA患者发生卒中的高风险密切相关;血清Lp-PLA2、ox-LDL及NSE水平的升高仍与TIA患者发生卒中的高风险密切相关,Lp-PLA2、NSE水平的升高仍与TIA患者发生卒中的中、低风险密切相关。结论:TIA 患者血清 miR-23b-3p、miR-181a-5p、miR-208b 及 miR-215 水平显著增加,上述miRNAs有望成为TIA预测及TIA后卒中风险评估的新型生物标志物,为临床TIA患者的病情评估与监测提供新的途径与理论依据。
[Abstract]:Transient ischemic attacks (TIA) refers to sudden, transient, reversible neurological dysfunction caused by focal cerebral ischemia caused by a variety of reasons. TIA is prone to recurrence. Patients with a history of transient TIA have a significantly increased risk of stroke recurrence, disability rate, and higher mortality. MicroRNAs (microRNAs) are the hotspots of research in recent years. They are closely related to the occurrence and development of ischemic cerebrovascular diseases. However, there is no systematic study on the expression and changes of serum microRNAs in TIA patients. The clinical value of serum microRNAs in predicting TIA and assessing the risk of recurrent stroke after TIA remains to be further studied. Objective: To analyze the changes of serum microRNAs levels in patients with TIA, screen serum microRNAs with specific changes, compare and explore the clinical value of serum microRNAs and existing serum markers related to brain injury in predicting TIA and assessing the risk of recurrent stroke after TIA, so as to provide a new way for clinical evaluation and monitoring of TIA patients. Methods: 126 TIA patients and 44 controls were divided into three groups according to the ABCD3-I score of TIA patients with recurrent stroke: low-risk group (0-3 points, 46 cases), medium-risk group (4-7 points, 45 cases) and high-risk group (8-13 points, 35 cases). The relative expression levels of 13 candidate microRNAs associated with ischemic cerebrovascular diseases (including microRNAs-15b, microRNAs-30a, microRNAs-21, microRNAs-29b, microRNAs-181a, microRNAs-124, microRNAs-335, microRNAs-146a, microRNAs-499, microRNAs-144, microRNAs-208b, microRNAs-215 and microRNAs-23b) in TIA patients and controls were measured to screen for serum-specific changes in microRNAs. Levels of serum biomarkers associated with brain injury, including Lipoprotein-associated phospholipase A2 (Lp-PLA2), oxidized low-density lipoprotein (ox-LDL), neuron-specific enolase (NSE) and brain-derived neurotrophic factor (Brain Der) Ived Neurotrophic Factor, BDNF. Spearman correlation analysis was used to explore the correlation between serum microRNAs and serum markers associated with brain injury. receiver operating characteristic curves (ROC) analysis and multiple logistic regression analysis were used to compare and explore serum microRNAs and brain injury related blood. Results: QRT-PCR results showed that serum levels of microRNAs-23b-3p, microRNAs-208 b, microRNAs-215, microRNAs-181a-5p in TIA patients were significantly higher than those in control group (all P 0.05); serum levels of microRNAs-23b-3p, microRNAs-208 b, microRNAs-215, and microRNAs-181a-5p in high-risk group, middle-risk group and low-risk group were significantly higher than those in TIA patients (all P 0.05). The levels of serum Lp-PLA2, ox-LDL and NSE in TIA patients were significantly higher than those in control group (all P 0.05). In TIA patients, the levels of serum Lp-PLA2, ox-LDL and NSE in high-risk group were higher than those in control group (all P 0.05). Correlation analysis showed that serum Lp-PLA2 and ox-LDL levels were positively correlated (r = 0.264, P = 0.002), NSE and BDNF levels were positively correlated (r = 0.197, P = 0.041). 3. ROC curve analysis showed that serum microRNA23b-3p, microRNA208b, microRNATI-215 and microRNA181a-5p could distinguish the area under the curve between A patients and controls (are A under the curve, AUC were 0.784, 0.795, 0.768 and 0.779 respectively (P 0.01). The combination of four microRNAs distinguished AUC between TIA patients and controls was 0.812 (P = 0.001); serum Lp-PLA2 and NSE distinguished AUC between TIA patients and controls was 0.725 and 0.743 (P 0.05), and the combination of the two indicators distinguished AUC between TIA patients and controls was 0.817 (P = 0.001). The AUC of TIA patients and controls was 0.925 (P 0.001). Fourth, the risk stratification group and control group of TIA patients were used as independent four classified variables. The control group was used as control group. Multiple logistic regression analysis was performed for serum microRNAs and brain injury related indicators. The results showed that high levels of serum microRNAs-23b-3p, microRNAs-208b, and microRNAs-215 were associated with the risk of stroke in TIA patients; high levels of serum Lp-PLA2, ox-LDL, NSE and BDNF were associated with high risk of stroke in TIA patients; high levels of NSE were also associated with moderate and low risk of stroke in TIA patients. After adjusting for the effects of age, sex, blood pressure and lipid parameters, the increase of serum microRNAs-23b-3p levels was still closely related to the high and moderate risk of stroke in TIA patients, the increase of microRNAs-208b levels was still closely related to the high risk of stroke in TIA patients, and the elevation of serum Lp-PLA2, ox-LDL and NSE levels was still closely related to the high risk of stroke in TIA patients. Conclusion: Serum levels of microRNAs - 23B - 3p, microRNAs - 181a - 5p, microRNAs - 208b and microRNAs - 215 in TIA patients are significantly increased. These microRNAs may be new biomarkers for predicting TIA and assessing the risk of stroke after TIA. And provide new ways and theoretical basis for monitoring.
【学位授予单位】：南方医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R743.31

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