常规脑电图联合血清miR-146a、miR-129-5p水平检测对药物难治性癫痫的临床诊断价值

doi:10.11958/20240666

摘要/Abstract

摘要：

目的探究常规脑电图联合血清miR-146a、miR-129-5p水平检测对药物难治性癫痫的临床诊断价值。方法将2021年6月—2023年6月收治的60例难治性癫痫患者纳入难治性癫痫组，40例健康志愿者纳入对照组。体外培养连续暴露于苯妥英钠的人脑微血管内皮耐药细胞作为耐药组，分别转染miR-NC、miR-146a模拟物和miR-129-5p模拟物，记作miR-NC组、miR-129-5p模拟物组和miR-146a模拟物组。采用蛋白质印迹检测各组细胞和血液中高迁移率族蛋白B1（HMGB1）蛋白表达。采用荧光定量聚合酶链式反应检测血清miR-146a和miR-129-5p水平，WB检测血清HMGB1蛋白表达。以多名专家医师的综合问诊结果为金标准，评估常规脑电图和血清miR-146a、miR-129-5p水平检测以及联合检测对难治性癫痫患者的诊断价值，并绘制受试者工作特征（ROC）曲线。结果与对照组相比，耐药组细胞的HMGB1表达下降；与miR-NC组相比，miR-129-5p模拟物组和miR-146a模拟物组细胞的HMGB1表达下降（P<0.05）。与对照组相比，难治性癫痫组血清HMGB1蛋白表达下调，miR-146a和miR-129-5p表达上调。ROC曲线分析显示，常规脑电图联合血清miR-146a和miR-129-5p水平诊断难治性癫痫的曲线下面积、敏感度、特异度和准确度高于单一检测。结论常规脑电图和血清miR-129-5p、miR-146a水平联合检测可为临床上药物难治性癫痫的诊断提供帮助。

关键词: 耐药性癫痫, 脑电描记术, 微RNAs, ROC曲线, HMGB1蛋白质, miR-146a, miR-129-5p, 药物难治性癫痫

Abstract:

Objective To investigate the clinical diagnostic value of routine electroencephalogram (EEG) combined with serum miR-146a and miR-129-5p levels in drug-resistant epilepsy. Methods Sixty patients with refractory epilepsy admitted to our hospital from June 2021 to June 2023 were included in the refractory epilepsy group, and 40 healthy volunteers were included in the control group. Human microvascular endothelial drug-resistant cells (HBMECs) continuously exposed to PHT2 were cultured in vitro and transfected with miR-NC (the miR-NC group), miR-146a mimics (the miR-146a mimics group) and miR-129-5p mimics (the miR-129-5p mimics group), respectively. Western blod assay was used to detect the expression of high-mobility group protein B1 (HMGB1) in each group. Serum miR-146a and miR-129-5p levels were detected by fluorescent quantitative PCR (polymerase chain reaction), and serum HMGB1 protein expression was detected by Western blod assay. The sensitivity, specificity and accuracy of routine EEG, serum miR-146a and miR-129-5p levels and combined diagnosis of drug-resistant epilepsy were evaluated using the comprehensive consultation results of several expert physicians as the gold standard, and ROC curves were drawn. Results Compared with the control group, the expression of HMGB1 was significantly decreased in the drug-resistant group. Compared with the miR-NC group, HMGB1 expression was significantly decreased in the miR-129-5p mimics group and the miR-146a mimics group (P<0.05). Compared with the healthy group, the serum HMGB1 protein expression was down-regulated and miR-146a and miR-129-5p expression levels were significantly up-regulated in the refractory epilepsy group. ROC curve analysis indicated that the area under the curve, sensitivity, specificity and accuracy of conventional EEG combined with serum miR-146a and miR-129-5p levels were higher in the diagnosis of refractory epilepsy than that of single diagnostic method. Conclusion The combination of routine EEG and serum miR-129-5p and miR-146a levels can provide help for the diagnosis of drug-resistant patients.

Key words: drug resistant epilepsy, electroencephalography, microRNAs, ROC curve, HMGB1 protein, miR-146a, miR-129p, drug refractory epilepsy

中图分类号:

R742.1

图/表 7

表1 miR-129-5p、miR-146a和U6引物序列

Tab.1 Primer sequences for miR-129-5p, miR-146a and U6

基因名称	引物序列（5′→3′）	产物大小/bp
miR-129-5p	上游：CTTTTTGCGGTCTGGGCTTGC	65
	下游：AGTGCAGGGTCCGAGGTATT	65
miR-146a	上游：TGTTGCTGAAGAGCTCTGGTAC	70
	下游：TCGTCTGATGATGTCGAATGTAC	70
U6	上游：CTCGCAACATGGTCGCACAT	66
	下游：GAACGGATCAGATATTGCGT	66

图1 5组细胞中HMGB1蛋白表达水平比较

Fig.1 Comparison of HMGB1 protein expression between five groups of cells

图2 2组受试者血清中HMGB1蛋白表达水平比较

Fig.2 Comparison of serum HMGB1 protein expression between the two groups

表2 2组受试者血清中miR-129-5p和miR-146a表达水平比较（$\bar{x} \pm s$）

Tab.2 Comparison of serum levels of miR-129-5p and miR-146a between the two groups of patients

组别	n	miR-129-5p	miR-146a
对照组	40	0.98±0.19	1.13±0.40
难治性癫痫组	60	1.59±0.32	1.96±0.40
t		10.950^＊＊	10.160^＊＊

图3 难治性癫痫患者血清miR-129-5p和miR-146a水平相关性分析

Fig. 3 Correlation analysis of serum miR-129-5p and miR-146a levels in patients with drug-resistant epilepsy

表3 血清miR-129-5p、miR-146a水平检测、常规脑电图以及三者联合检测对难治性癫痫的诊断价值

Tab.3 The diagnostic value of serum miR-129-5p, miR-146a levels, routine EEG and their combined detection in drug-resistant epilepsy

诊断方法	临界值	AUC	95%CI	敏感度/%	特异度/ %	准确度/ %
常规脑电图	-	0.900	0.830~0.970	91.67	82.50	88.00
血清miR-146a	1.54	0.920	0.869~0.971	88.33	85.00	87.00
血清miR-129-5p	1.12	0.930	0.879~0.981	91.67	87.50	90.00
联合诊断	-	0.946	0.891~1.000	96.67	90.00	94.00

图4 血清miR-129-5p、miR-146a水平、常规脑电图以及三者联合诊断药物难治性癫痫的ROC曲线

Fig.4 ROC curve analysis of serum miR-129-5p, miR-146a levels, routine EEG and combined detection for clinical diagnosis of drug-resistant epilepsy

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