颈动脉粥样硬化性狭窄患者外周血miRNA-210-5P表达及其靶基因生物信息学分析
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摘要
目的通过检测颈动脉粥样硬化性狭窄患者外周血miRNA-210-5P相对表达量,分析miRNA-210-5P及其靶基因功能。方法选择2015年7月至2018年9月诊断明确的颈动脉粥样硬化性狭窄患者(CAS组,146例)作为观察对象,采用逆转录-聚合酶链反应(RT-PCR)检测不同处理组受试者外周血miRNA-210-5P相对表达量,并以TargetScan和CoMeTa数据库进行靶基因预测、DAVID数据库进行靶基因功能富集分析(GO分析)和KEGG信号通路分析。结果与无颈动脉粥样硬化性狭窄患者或正常受试者(对照组,60例)相比,CAS组患者血清miRNA-210-5P相对表达量升高(t=14.759,P=0.000)。其中,重度狭窄组(31例;q=23.028,P=0.000)、中度狭窄组(53例;q=6.657,P=0.000)、轻度狭窄组(62例;q=42.612,P=0.000)患者外周血miRNA-210-5P相对表达量均高于对照组;而中度狭窄组(q=34.538,P=0.000)和重度狭窄组(q=11.914,P=0.000)miRNA-210-5P相对表达量高于轻度狭窄组;重度狭窄组亦高于中度狭窄组(q=16.983,P=0.000)。ROC曲线显示,miRNA-210-5P预测颈动脉粥样硬化中至重度狭窄的曲线下面积为0.943,当最佳临界值为1.495时,其预测灵敏度90.33%、特异度92.54%;生物信息学分析提示,miRNA-210-5P潜在靶基因包括VEGFA、KCMF1、HMGCS1、KLF12、EFNA3、GIT2等54个基因;GO分析显示,miRNA-210-5P靶基因功能主要富集于血管生成、神经元发育、DNA转录因子活性的正性调控、内皮细胞趋化性、细胞迁移与分化黏附等;对KEGG信号通路的检测显示,miRNA-210-5P靶基因主要富集于突触导向信号转导通路。结论颈动脉粥样硬化性狭窄患者血清miRNA-210-5P表达上调,且可能通过调控多种靶基因而作用于突触导向信号转导通路中,参与颈动脉粥样硬化性狭窄的发生与发展。
Objective To investigate the relative expression of miRNA-210-5 P in serum of patients with carotid atherosclerotic stenosis(CAS)and to explore the function of miRNA-210-5 P and its target genes using bioinformatics methods.Methods We selected 146 patients with CAS from July 2015 to September 2018 in our hospital.Reverse transcriptase-polymerase chain reaction(RT-PCR)was used to detect the relative expression of miRN A-210-5 P in peripheral blood of all enrolled patients.The target genes were predicted by using TargetScan and CoMeTa databases.The target genes of miRNA-210-5 P were enriched by Gene Ontology(GO)using DAVID data and were performed with KEGG Pathway analysis.Results Compared with non-CAS patients or normal subjects(control group,N = 60),the relative expression of miRNA-210-5 P in the serum of CAS group was significantly increased(t= 14.759,P=0.000).The relative expressions of serum miRNA-210-5 P in severe stenosis group(N=31; q=23.028,P=0.000),moderate stenosis group(N =53; q =6.657,P =0.000)and mild stenosis group(N =62; q = 42.612,P =0.000)were higher than that in control group.The relative expressions of serum miRN A-210-5 P in moderate stenosis group(q = 34.538,P = 0.000)and severe stenosis group(q = 11.914,P = 0.000)were significantly higher than that in mild stenosis group.The relative expressions of serum miRNA-210-5 P in severe stenosis group was significantly higher than that in moderate stenosis group(q= 16.983,P=0.000).Receiver operating characteristic(ROC)curve showed that the miRNA-210-5 P predicted the area under the curve(AUC)of moderates to severe stenosis in CAS to be 0.943,the sensitivity was 90.33% and the specificity was 92.54% at the best cutoff value of 1.495.Bioinformatics analysis showed there were 54 potential target genes of miRNA-210-5 P,such as VEGFA,KCMF1,HMGCS1,KLF12,EFNA3,GIT2,etc.GO analysis showed that the target genes of miRN A-210-5 P were involved in angiogenesis,neuronal development,positive regulation of DNA transcription factor activity,endothelial cell c he mo taxis,cell migration and differentiation and adhesion processes.KEGG Pathway analysis displayed miRNA-210-5 P target genes were mainly enriched in synaptic-directed factor signal transduction pathways.Conclusions The expression of miRNA-210-5 P in peripheral blood of CAS patients is upregulated,and it may participate in the process of CAS occurrence and development by regulating multiple target genes and acting on synaptic-directed signaling pathways.
Objective To investigate the relative expression of miRNA-210-5 P in serum of patients with carotid atherosclerotic stenosis(CAS)and to explore the function of miRNA-210-5 P and its target genes using bioinformatics methods.Methods We selected 146 patients with CAS from July 2015 to September 2018 in our hospital.Reverse transcriptase-polymerase chain reaction(RT-PCR)was used to detect the relative expression of miRN A-210-5 P in peripheral blood of all enrolled patients.The target genes were predicted by using TargetScan and CoMeTa databases.The target genes of miRNA-210-5 P were enriched by Gene Ontology(GO)using DAVID data and were performed with KEGG Pathway analysis.Results Compared with non-CAS patients or normal subjects(control group,N = 60),the relative expression of miRNA-210-5 P in the serum of CAS group was significantly increased(t= 14.759,P=0.000).The relative expressions of serum miRNA-210-5 P in severe stenosis group(N=31; q=23.028,P=0.000),moderate stenosis group(N =53; q =6.657,P =0.000)and mild stenosis group(N =62; q = 42.612,P =0.000)were higher than that in control group.The relative expressions of serum miRN A-210-5 P in moderate stenosis group(q = 34.538,P = 0.000)and severe stenosis group(q = 11.914,P = 0.000)were significantly higher than that in mild stenosis group.The relative expressions of serum miRNA-210-5 P in severe stenosis group was significantly higher than that in moderate stenosis group(q= 16.983,P=0.000).Receiver operating characteristic(ROC)curve showed that the miRNA-210-5 P predicted the area under the curve(AUC)of moderates to severe stenosis in CAS to be 0.943,the sensitivity was 90.33% and the specificity was 92.54% at the best cutoff value of 1.495.Bioinformatics analysis showed there were 54 potential target genes of miRNA-210-5 P,such as VEGFA,KCMF1,HMGCS1,KLF12,EFNA3,GIT2,etc.GO analysis showed that the target genes of miRN A-210-5 P were involved in angiogenesis,neuronal development,positive regulation of DNA transcription factor activity,endothelial cell c he mo taxis,cell migration and differentiation and adhesion processes.KEGG Pathway analysis displayed miRNA-210-5 P target genes were mainly enriched in synaptic-directed factor signal transduction pathways.Conclusions The expression of miRNA-210-5 P in peripheral blood of CAS patients is upregulated,and it may participate in the process of CAS occurrence and development by regulating multiple target genes and acting on synaptic-directed signaling pathways.
引文
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[21]Bulysheva AA,Burcus N,Lundberg CG,Francis MP,Heller R.VEGF-B electrotransfer mediated gene therapy induces cardiomyogenesis in a rat model of cardiac ischemia[J].Bioelectrochemistry,2018,124:105-111.
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[2]Gao ZY, Wang W, Chen M,Qin HN,Shi J, Gao XG.Relationship between carotid artery color ultrasonography indexes and brain white matter lesions in elderly patients[J].Zhonghua Lao Nian Xin Nao Xue Guan Bing Za Zhi,2019,21:270-273.[高智玉,王雯,陈敏,秦海宁,石净,高旭光.老年患者颈动脉彩色超声检查指标与脑白质病变相关性分析[J].中华老年心脑血管病杂志,2019,21:270-273.]
[3]Zhang XH.Expression profiles of six circulating micrornas critical atherosclerosis in patients with subclinical hypothyroidism:a clinical study[D].Ji'nan:Shandong University,2014.[张新焕.动脉硬化关键信号分子相关的血清microRNA在亚临床甲减病人中表达谱初探[D].济南:山东大学,2014.]
[4]Li JZ,Xia X, Huan SY, Ke XW, Jian Z, Ding HL,Yu LY.Identification of microRNA transcriptome reveals that miR-100is involved in the renewal of porcine intestinal epithelial cells[J].Sci China Life Sci,2019,62:816-828.
[5]Pan YB,Sun ZL,Feng DF.The role of microRNA in traumatic brain injury[J].Neuroscience,2017,367:189-199.
[6]Chen YJ,Chang WA,Huang MS,Chen CH,Wang KY,Hsu YL,Kuo PL.Identification of novel genes in aging osteoblastsusing next-generation sequencing and bioinformatics[J].Oncotarget,2017,8:1 13598-113613.
[7]Martinez SR,Ma Q,Dasgupta C,Meng X,Zhang L.MicroRNA-210 suppresses glucoc-orticoid receptor expression in response to hypoxia in fetal rat cardiomyocytes[J].Oncotarget,2017,8:80249-80264.
[8]Kuang YQ,Liu L,Xu FF,Chen Y,Su YW.Analysis of microRNA-122 as a biomarker for early evaluation of drug hepatotoxicity[J].Nanjing Yi Ke Da Xue Xue Bao(Zi Ran Ke Xue Ban),2018,38:322-327.[匡雨琼,刘亮,许飞飞,陈芸,苏钰文.MicroRNA-122作为新药肝毒性早期评价标志物的相关分析[J].南京医科大学学报(自然科学版),2018,38:322-327.]
[9]Hu H,Ding Y,Wang Y,Geng S,Liu J,He J,Lu Y,Li X,Yuan M,Zhu S,Zhao S.MitoKATP channels promote theproliferation of hypoxic human pulmonary artery smooth muscle cells via the ROS/HIF/miR 210/ISCU signaling pathway[J].Exp Ther Med,2017,14:6105-61 12.
[10]Zhang WH,Song X.The role of hypoxia-relaterd microRNA-210in malignant process of tumor[J].Zhongguo Zhong Liu Sheng Wu Zhi Liao Za Zhi,2016,23:733-738.[张文会,宋鑫.缺氧相关microRNA-210在肿瘤恶性进程中的作用[J].中国肿瘤生物治疗杂志,2016,23:733-738.]
[11]Duan YY,Zhang J,Ye P.Recent advances in microRNA and atherosclerosis[J].Zhonghua Lao Nian Xin Nao Xue Guan Bing Za Zhi,2016,18:1220-1223.[段媛媛,张蛟,叶平.微小RNA与动脉粥样硬化最新研究进展[J].中华老年心脑血管病杂志,2016,18:1220-1223.]
[12]Karlsson L,Kangefjard E,Hermansson S,Stromberg S,Osterberg K,Nordanstig A,Ryndel M,Gellerman K,Fred,nLindqvist J,Bergstrom GM.Risk of recurrent stroke in patients with symptomatic mild(20%-49%NASCET)carotid artery stenosis[J].Eur J Vasc Endovasc Surg,2016,52:287-294.
[13]Xu XT,Ma N.Application of transcranial Doppler in interventional department[J].Zhongguo Zu Zhong Za Zhi,2018,13:1237-1242.[徐晓彤,马宁.经颅多普勒超声在神经介入临床中的应用[J].中国卒中杂志,2018,13:1237-1242.]
[14]Shariat A,Niknam L,Izadi S,Salehi A.Prevalence of intracranial artery stenosis in Iranian patients with ac ute ischemic stroke using transcranial Doppler ultrasonography[J].Iran J Neurol,2016,15:133-139.
[15]Mei YY,Huang DS.Research progress of microRN A-210[J].Yi Xue Zong Shu,2016,22:2913-2917.[梅妍妍,黄东生.微RN A-210研究进展[J].医学综述,2016,22:2913-2917.]
[16]Fang H,Wu D,An Y,Li D.Recent advances in relationship between interstitial microenvironment of atherosclerosis plaque and formation of new angiogenesis[J].Xin Xue Guan Bing Xue Jin Zhan,2018,39:75-79.[房昊,吴丹,安毅,李丹.动脉粥样硬化斑块内间质微环境与新生血管生成关系的最新研究进展[J].心血管病学进展,2018,39:75-79.]
[17]Jiang H,Toscano JF,Schiraldi M,Song SS,Schlick KH.Dumitrascu OM,Liou R,Lyden PD,Pan J,Zhan R,Saver JL.Gonzalez NR.Differential expression of vascular endothelial growth factor-A165 isoforms between intracranial atherosc:lerosis and moyamoya disease[J].J Stroke Cerebrovasc Dis,2019,28:360-368.
[18]Tian X,Xiong H,Wu D,Zhang R,Lu M,Zhang YT.Age and sex-specific relationships between blood pressure variability and carotid intima-media thickness[J]. Australas Phys Eng Sci Med,2016,39:967-976.
[19]Vera Janavel G,Crottogini A,Cabeza Meckert P,Cuniberti L.Mele A,Papouchado M,Fernandez N,Bercovich A,Criscuolo M,Melo C,Laguens R.Plasmid-mediated VEGF gene transfer induces cardiomyogenesis and reduces myocardial infarct size in sheep[J].Gene Ther,2006,13:1133-1142.
[20]Li JJ, Li S,Zhu CG,Wu CC,Zhang NQ,Guo Y,Gao LY,Li Y,Qing XL,Cui P,Xu CJ,Jiang RX,Sun ZW,Liu J, Dong G,Li Q.Familiar hypercholesterolemia phenotype in Chinese patients under-going coronary angiography[J].Arterioscler Thromb Vase Biol,2017,37:570-579.
[21]Bulysheva AA,Burcus N,Lundberg CG,Francis MP,Heller R.VEGF-B electrotransfer mediated gene therapy induces cardiomyogenesis in a rat model of cardiac ischemia[J].Bioelectrochemistry,2018,124:105-111.
[22]Godin-Heymann N,Brabetz S,Murillo MM,Saponaro M,Santos CR,Lobley A,East P,Chakravarty P,Matthews N,Kelly G.Jordan S,Castellano E,Downward J.Tumour-suppression function of KLF12 through regulation of anoikis[J].Oncogene,2016,35:3324-3334.
[23]Li J,Zhou Y,Hao YE.Advances in the study of G proteincoupled receptor kinase interacting protein 2(G1T2)involved in immune regulation[J].Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi.2016,32:1416-1418.[李佳,周艺,郝玉娥.G蛋白偶联受体激酶相互作用蛋白2(GIT 2)参与免疫调控的研究进展[J].细胞与分子免疫学杂志,2016,32:1416-1418.]
[24]Kang SG,Kim C,Cortez LM,Carmen Garza M,Yang J,WilleH, Sim VL, Westaway D,McKenzie D,Aiken J.Toll-like receptor-mediated immune response inhibits prion propagation[J].Glia,2016,64:937-951.
[25]Jones DP,True HD,Patel J.Leukocyte trafficking in cardiovascular disease:insights from experimental models[J].Mediators Inflamm,2017:ID9746169.