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栏目介绍 彤心飞传是由天津医科大学第二医院心脏科刘彤教授和郑州大学附属洛阳中心医院心内科谷云飞共同推出的一档心血管前沿文献速读栏目。栏目的理念是“专注 坚持 积累”，每期精选国际顶级期刊心血管领域的最新文献进行深度呈现，以期让国内相关专业人员可以同步跟踪学科进展，了解前沿动态资讯，获取最新研究结论。 
心房颤动（AF）是临床最常见的房性心律失常，明显增加栓塞和心力衰竭发生的风险[1,2]。指南推荐症状性房颤首选导管射频消融治疗[3]。然而，虽然阵发性房颤疗效较好，但持续性房颤术后长期复发率仍然较高。心房重构是房颤发生的主要病理机制，心房纤维化是心房重构的重要标志，心房纤维化的严重程度与房颤的预后有关[4]。改善左房纤维化基质可改善房颤的预后[5]。因此、寻找心房纤维化的早期生物标志物并探讨房颤的发病机制对改善预后至关重要。miRNA是一种长度在18-22碱基的非编码RNA分子，目前研究证实其参与调控多种心血管疾病的发生发展。且循环中的miRNAs可作为各种疾病的诊断生物学标志物[6,7]。 最近，云南省第一人民医院范洁课题组在Journal of Cardiology发表原创研究，题目为miR-425-5p is negatively associated with atrial fibrosis and promotes atrial remodeling by targeting CREB1 in atrial fibrillation，通过使用高通量测序来筛选房颤患者及非房颤患者血液中差异表达的miRNAs，并分析其与左房纤维化的关系及射频消融术后的表达变化。同时在细胞和动物模型中验证miR-425-5p在房颤心房重构中的作用及作用分子机制。 首先通过高通量测序发现mIR-425-5p在房颤患者外周血中表达下降，并进一步通过qRT-PCR在80例房颤患者外周血及外科手术房颤患者心房组织标本验证其表达下降，更为重要的是，房颤患者左房血液标本中miR-425-5p表达低于外周血，进一步提示miR-425-5p的异常表达可能与房颤发生发展密切相关。 其次分析miR-425-5p在不同类型房颤的表达变化，结果发现miR-425-5p在持续性心房颤动患者中表达低于阵发性房颤，同时对接受房颤导管射频消融术的持续性房颤术中标测左房低电压区，分析miR-425-5p表达与左房低电压的关系，结果显示miR-425-5p与持续性房颤患者左房低电压区呈负相关，提示miR-425-5p与左房纤维化相关。更为重要的是，导管消融可逆转血浆中下降的miR-425-5p表达。 最后通过构建房颤细胞模型与动物模型，利用分子生物学技术，证实下调miR-425-5p表达可促进心房成纤维细胞增殖，而过表达miR-425-5p表达可改善房颤心房重构。进一步利用靶基因预测软件，荧光素酶报告基因实验，免疫印迹等实验，证实miR-425-5p通过直接靶向调控CREB1参与调控上述生物学功能。 Figure1: The expression level of miRNAs in plasma in two dependent cohorts by miRNA sequencing and qRT-PCR. A: Heatmap of miRNA expression data from plasma samples of patients with AF(n=3) and healthy controls (n=3). Relative miRNA expression was depicted according to the color scale shown on the botton. Red indicated upregulation; green indicated downregulation, Ctl indicated healthy control samples. B-C: Validation of miRNA sequencing data by qRT-PCR in patients with AF(n=3) and healthy controls (n=3), **P<0.01 vs control.D-F: Validation of 2 downregulation miRNAs and 1 upregulation miRNAexpression in an independent cohort with 80 AF patients and 80 healthy control by qRT-PCR, ***P<0.001. The vertical axis was ΔCt. whereby a lower ΔCt value mean a higher expression of miRNA.
Figure2: miR-425-5p was associated with left atrial fibrosis. A: The miR-425-5p expression level in persistent AF and paroxysmal AF, paro AF= paroxysmal AF, pers AF= persistent AF, The vertical axis was ΔCt. whereby a lower ΔCt value mean a higher expression of miRNA. B: The correlation between plasma miR-425-5p and the extent of left atrial low voltage area, all valve were log-transformed to normalize their distribution before statistical analysis, the extent of left atrial low voltage was used to evaluate left fibrosis. C-E: The representative image of the left atrial low voltage area(LVA), the red color represented the left atrial low area which reflect the severity of left atrial fibrosis. F: Relative miR-425-5p expression according to LVA group(group 1: LVAs <10%; group 2: LVAs 10% to 20%; group 3: LVAs>20%, *P<0.05, ***P<0.001).
Figure3: The miR-425-5p expression level in atrial tissue in AF patients and mouse AF model. A: The expression level of miR-425-5p in AF patients(n=6); B: The expression level of miR-425-5p in mouse AF model(n=12); C: The expression level of miR-425-5p in left and right atrium tissue in AF model and sham, **P<0.01.
Figure4: The plasma miR-425-5P expression level in the pre-RFCA and post-RFCA. A: The plasma level of miR-425-5p in the control group (n=20) and pre-RFCA (n=20), post-RFCA (n=20), **P<0.01 vs control group, ##P<0.01 vs pre-RFCA. B: The expression change of miR-425-5p in each AF patients in pre-RFCA and post-RFCA.
Figure5: Effect of miR-425-5p on atrial remodeling. A: The miR-425-5p expression level in fibroblast cell transfected with miR-425-5p inhibitor. B: Fibroblast cell proliferation was detected with CCK-8. C: The proliferation markers of Ki67 and Cyclin D. D: The expression level of fibrosis biomarkers. E: The miR-425-5p expression level in mouse atrial tissue injected with miR-425-5p mimics. F: The AF incidence rate. G: Typical representative Masson’s trichrome staining. H: The statistical histogram, ***P<0.001, **P<0.01
Figure6: CREB1 was the direct target gene of miR-425-5p. A: The CREB1 expression was upregulated in atrial tissue in AF patients and mouse AF model. B: The CREB1 expression was negatively with miR-425-5p in AF patients by Pearson’s correlation coefficients. C-D: Inhibition of CREB1 expression could reverse the phenotype of downregulation of miR-425-5p to fibroblast cell proliferation and fibrosis. cell proliferation was detected with CCK-8. E: The potential binding site of miR-425-5p with CREB1 3’-URT was indicated with TargetScan 7.2 database. F-G: The CREB1 protein expression level in atrial fibroblast cell transfected with miR-NC and miR-425-5p inhibitor, GAPDH was used as loading control. H: Luciferase assay confirmed that CREB1 was a target of miR-425-5p, **P<0.01,*P<0.05.综述结果表明，miR-425-5p在AF患者血浆和心房组织中表达下调，miR-425-5p下调可作为心房纤维化的无创生物标志物，进而通过靶向调控CREB1表达参与调控房颤心房重构。进一步揭示了miRNAs在房颤发生发展中的作用，miR-425-5p有望成为未来房颤干预的分子靶点及预后的分子标志物。1. Odutayo A, et al. Atrial fibrillation and risks of cardiovascular disease, renal disease, and death: systematic review and meta-analysis. BMJ 2016;354:i4482.2. Bekwelem W, et al. Extracranial systemic embolic events in patients with nonvalvular atrial fibrillation. Circulation 2015;132:796-803.3. Tilz RR, et al. Ten-year clinical outcome after circumferential pulmonary vein isolation utilizing the Hamburg approach in patients with symptomatic drug-refractory paroxysmal atrial fibrillation. Circ Arrhythm Electrophysiol 2018;11:e005250.4. Thanigaimani S, et al. Molecular mechanisms of atrial fibrosis: implications for the clinic. Expert Rev Cardiovasc Ther 2017;15:247-56.5 Yang B, et al. STABLE-SR (Electrophysiological Substrate Ablation in the Left Atrium During Sinus Rhythm) for the Treatment of Nonparoxysmal Atrial Fibrillation: A prospective, multicenter randomized clinical trial. Circ Arrhythm Electrophysiol 2017;10:e005405.6. Fan PC, et al. A circulating miRNA signature for early diagnosis of acute kidney injury following acute myocardial infarction. J Transl Med 2019;17:139.7. Aghabozorgi AS, et al. Circulating exosomal miRNAs in cardiovascular disease pathogenesis: New emerging hopes. J Cell Physiol 2019;234:21796-809.范洁，云南省第一人民医院心内科副主任、国家卫计委心律失常介入培训基地负责人，博士生导师、二级教授， 现任中华医学会心电生理和起搏分会委员，女医师联盟的常委，中国医师协会心律失常学专业委员会常委，云南省医学会心电生理和起搏学会的副主任委员。先后发表学术论文40余篇，主编专著一本。获云南省科技进步一等奖，获得国家基金3项、省基金面上项目资助3项。
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