CN110093412A - 用于诊断动脉粥样硬化的分子标志物ugp2及应用 - Google Patents

用于诊断动脉粥样硬化的分子标志物ugp2及应用 Download PDF

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CN110093412A
CN110093412A CN201910250626.0A CN201910250626A CN110093412A CN 110093412 A CN110093412 A CN 110093412A CN 201910250626 A CN201910250626 A CN 201910250626A CN 110093412 A CN110093412 A CN 110093412A
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ugp2
atherosclerosis
disease
protein
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郑磊
王前
胡炎伟
丁立
何鑫
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Southern Hospital Southern Medical University
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Abstract

本发明提供一种用于诊断动脉粥样硬化的分子标志物UGP2及其应用，包括检测UGP2蛋白质的存在的试剂盒或诊断装置在制备用于诊断和/或预防和/或治疗动脉粥样硬化引起的疾病的药剂中的应用，所述试剂盒或诊断装置用于动脉粥样硬化引起的疾病的体外诊断和/或危险分层。本发明发现通过检测患者组织样品中的UGP2蛋白质浓度，可以预测动脉粥样硬化引起的疾病的发生，或对上述疾病进行诊断、危险分层等。

Description

用于诊断动脉粥样硬化的分子标志物UGP2及应用
技术领域
本发明涉及生物检测技术领域，特别是涉及一种用于诊断动脉粥样硬化的分子标志物UGP2及应用。
背景技术
心血管疾病是当前威胁全球人类健康的最重要的疾病之一，是发达国家的疾病和死亡的主要原因。根据世界心脏联盟的最新统计结果显示，每年全球死于心血管疾病的患者超过1700万人，已成为危害人类健康的全球第一杀手。中国的心血管疾病(CVD)现状同样令人堪忧。《中国心血管病报告2015》调查结果表明，2014年中国心血管病死亡率仍居疾病死亡构成的首位，高于肿瘤及其他疾病，CVD占居民疾病死亡构成在农村为44.60％，在城市为42.51％，每5例死亡者中就有2例死于CVD。中国心血管病危险因素流行趋势明显，心血管病的发病人数也在持续增加。
高脂血症与动脉粥样硬化(atherosclerosis，AS)、冠心病、脑卒中等心血管病关系密切已有共识，因此，寻找高血脂人群心血管疾病新型诊断标志物，对于早期干预、延缓疾病进展，减少死亡率与并发症发生率起着至关重要的作用。
目前本领域还没有一种成熟的心血管靶向治疗和检测方法，因此，急需研究开发新的特异性靶点和检测试剂盒，用于心血管疾病的预防和治疗。
发明内容
鉴于以上所述现有技术的缺点，本发明的目的在于提供一种用于诊断动脉粥样硬化的分子标志物UGP2及应用。
为实现上述目的及其他相关目的，本发明提供检测UGP2蛋白质的存在的试剂盒或诊断装置在制备用于诊断和/或预防和/或治疗动脉粥样硬化引起的疾病的药剂中的应用，所述试剂盒或诊断装置用于动脉粥样硬化引起的疾病的体外诊断和/或危险分层。
可选地，所述试剂盒用于检测体液样品中UGP2蛋白质的表达量。
可选地，所述试剂盒通过蛋白质印迹法、酶联免疫吸附测定、放射免疫分析、放射免疫扩散法、奥克特洛尼免疫扩散法、火箭免疫电泳、免疫组化染色、免疫沉淀分析法、补体结合分析法、流式细胞荧光分边技术和蛋白质芯片中的至少一种方法来检测UGP2蛋白质的表达量。
可选地，所述体液样品中UGP2蛋白质的表达量与动脉粥样硬化引起的疾病严重程度正相关。
可选地，所述体液样品选自血清。
可选地，所述药剂用于诊断或监测动脉粥样硬化引起的疾病的存在和/或过程和/或严重程度和/或预后。
本发明还提供UGP2(即尿苷二磷酸葡萄糖焦磷酸化酶2，UDP-glucosepyrophosphorylASe2)或其肽段作为生物标志物在制备或筛选动脉粥样硬化引起的疾病诊断试剂中的应用，所述诊断试剂用于动脉粥样硬化引起的疾病的体外诊断和/或危险分层。
可选地，所述动脉粥样硬化引起的疾病包括心血管疾病。
可选地，所述心血管疾病包括慢性心血管疾病。
可选地，所述慢性心血管疾病包括冠心病。
可选地，所述冠心病包括慢性冠心病、不稳定心绞痛、心肌梗死、稳定心绞痛。
可选地，动脉粥样硬化引起的疾病患者的UGP2蛋白质水平呈高表达。
本发明还提供用于表达UGP2蛋白质的DNA或其mRNA或保有其功能的同源物作为生物标志物在制备或筛选动脉粥样硬化引起的疾病的诊断试剂中的应用。
如上所述，本发明的用于诊断动脉粥样硬化的分子标志物UGP2及应用，具有以下有益效果：本发明发现通过检测患者组织样品中的UGP2蛋白质浓度，可以预测动脉粥样硬化引起的疾病的发生，或对上述疾病进行诊断、危险分层等。
附图说明
图1显示为本发明实施例1的基因芯片热图。
图2显示为本发明实施例1的浓度梯度Western Blot结果图。
图3显示为本发明实施例1的时间梯度Western Blot结果图。
具体实施方式
以下通过特定的具体实例说明本发明的实施方式，本领域技术人员可由本说明书所揭露的内容轻易地了解本发明的其他优点与功效。本发明还可以通过另外不同的具体实施方式加以实施或应用，本说明书中的各项细节也可以基于不同观点与应用，在没有背离本发明的精神下进行各种修饰或改变。
尿苷二磷酸葡萄糖焦磷酸化酶2(UDP-glucose pyrophosphorylASe 2，UGP2)基因定位于人类2号染色体的短臂上，全长49853bp，编码UDPG焦磷酸化酶，其氨基酸序列如序列1所示，其核苷酸序列(其mRNA对应的cDNA序列，不包含内含子和启动子序列)如序列2所示，NCBI上查询到的Gene ID如下：Gene ID:7360。在生理条件下，UDPG焦磷酸化酶可催化1-磷酸葡萄糖与尿苷三磷酸反应生成尿苷二磷酸葡萄糖(UDPG)及焦磷酸，UDPG可看作“活性葡萄糖”，在糖原合酶的作用下参与糖原的合成。由此可见，UGP2作为糖原合成起始途径的关键酶，通过催化葡萄糖转化生成UDPG并进一步生成糖原，在维持体内糖代谢平衡中发挥重要作用。研究表明，糖代谢紊乱与AS关系密切，高血糖可通过诱导血管内皮细胞和血管平滑肌细胞炎症反应，促进单核巨噬细胞向泡沫细胞转化和粥样斑块形成，因此，在临床表现为糖尿病患者出现AS的时间早、程度重和预后差。
在前期实验结果中，我们发现，与正常内膜组织相比，动脉斑块组织中UGP2表达明显上调。此外，在人脐静脉内皮细胞中，使用不同浓度、时间梯度Ox-LDL处理构建内皮细胞氧化应激受损模型，UGP2呈现递增的高表达量趋势。这些实验结果提示UGP2可能参与内皮细胞受损和AS进展。
目前本领域还没有一种成熟的针对UGP2的心血管靶向治疗和检测方法，因此，急需研究开发新的特异性靶点和检测试剂盒，用于心血管疾病的预防和治疗。
本发明涉及UGP2作为靶点用于预防和治疗心血管疾病，芯片结果分析显示UGP2在动脉粥样硬化病人动脉内膜组织和正常组织中差异表达明显，在内皮细胞氧化应激受损模型中，UGP2呈现明显的高表达量趋势。因此，UGP2可作为潜在的心血管疾病治疗靶点。
以下实施例中，图1涉及的实验方法参照文献：Xiao G,Martinez-Vaz B,Pan W,Khodursky AB.Operon information improves gene expression estimation for cDNAmicroarrays.BMC Genomics.2006；7:87.Published 2006Apr 21.doi:10.1186/1471-2164-7-87。
图2、图3涉及的实验方法参照文献：Gilda JE,Ghosh R,Cheah JX,West TM,Bodine SC,Gomes AV.Western Blotting Inaccuracies with Unverified Antibodies:Need for a Western Blotting Minimal Reporting Standard(WBMRS).PLoS One.2015；10(8):e0135392.Published 2015Aug 19.doi:10.1371/journal.pone.0135392。
实施例1
1、采用基因芯片分析AS病人动脉斑块组织和正常组织差异表达的mRNA水平，实验结果如图1所示，发现UGP2上调。
2、用0μg/ml、25μg/ml、50μg/ml、100μg/ml ox-LDL处理内皮细胞24h后经WesternBlot检测UGP2蛋白表达情况，实验结果如图2所示，我们从图中可以发现，UGP2蛋白水平随ox-LDL处理浓度增加，表达水平呈现递增的趋势，差异表达明显。
3、用100μg/ml ox-LDL处理内皮细胞0、6h、12h、24h后经Western Blot检测UGP2蛋白表达情况，实验结果如图3所示，我们从图中可以发现，UGP2蛋白水平随ox-LDL处理时间的增加，表达水平呈现递增的趋势，差异表达明显。
综上所述，本发明发现通过检测患者组织样品中的UGP2蛋白质浓度，可以预测动脉粥样硬化引起的疾病的发生，或对上述疾病进行诊断、危险分层等。
上述实施例仅例示性说明本发明的原理及其功效，而非用于限制本发明。任何熟悉此技术的人士皆可在不违背本发明的精神及范畴下，对上述实施例进行修饰或改变。因此，举凡所属技术领域中具有通常知识者在未脱离本发明所揭示的精神与技术思想下所完成的一切等效修饰或改变，仍应由本发明的权利要求所涵盖。
SEQUENCE LISTING
<110> 南方医科大学南方医院
<120> 用于诊断动脉粥样硬化的分子标志物UGP2及应用
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aatggaaaac gctgtgaatt tgtcatggaa gtcacaaata aaacacgtgc agatgtaaag 840
ggcgggacac tcactcaata tgaaggcaaa ctgagactgg tggaaattgc tcaagtgcca 900
aaagcacatg tagacgagtt caagtctgta tcaaagttca aaatatttaa tacaaacaac 960
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tttctgcctg tcaaaaccac atcagatctc ttgctggtga tgtcaaacct ctatagtctt 1200
aatgcaggat ctctgacaat gagtgaaaag cgggaatttc ctacagtgcc cttggttaaa 1260
ttaggcagtt cttttacgaa ggttcaagat tatctaagaa gatttgaaag tataccagat 1320
atgcttgaat tggatcacct cacagtttca ggagatgtga catttggaaa aaatgtttca 1380
ttaaagggaa cggttatcat cattgcaaat catggtgaca gaattgatat cccacctgga 1440
gcagtattag agaacaagat tgtgtctgga aaccttcgca tcttggacca ctga 1494

Claims (10)

1.检测UGP2蛋白质的存在的试剂盒或诊断装置在制备用于诊断和/或预防和/或治疗动脉粥样硬化引起的疾病的药剂中的应用，所述试剂盒或诊断装置用于动脉粥样硬化引起的疾病的体外诊断和/或危险分层。
2.根据权利要求1所述的应用，其特征在于：所述试剂盒用于检测体液样品中UGP2蛋白质的表达量。
3.根据权利要求1所述的应用，其特征在于：所述试剂盒通过蛋白质印迹法、酶联免疫吸附测定、放射免疫分析、放射免疫扩散法、奥克特洛尼免疫扩散法、火箭免疫电泳、免疫组化染色、免疫沉淀分析法、补体结合分析法、流式细胞荧光分边技术和蛋白质芯片中的至少一种方法来检测UGP2蛋白质的表达量。
4.根据权利要求3所述的应用，其特征在于：所述体液样品中UGP2蛋白质的表达量与动脉粥样硬化引起的疾病严重程度正相关。
5.根据权利要求1所述的应用，其特征在于：所述药剂用于诊断或监测动脉粥样硬化引起的疾病的存在和/或过程和/或严重程度和/或预后。
6.根据权利要求2所述的应用，其特征在于：所述体液样品选自血清。
7.UGP2或其肽段作为生物标志物在制备或筛选动脉粥样硬化引起的疾病诊断试剂中的应用，所述诊断试剂用于动脉粥样硬化引起的疾病的体外诊断和/或危险分层。
8.根据权利要求7所述的应用，其特征在于：所述脉粥样硬化引起的疾病包括心血管疾病。
9.根据权利要求7所述的应用，其特征在于：动脉粥样硬化引起的疾病患者的UGP2蛋白质水平呈高表达。
10.用于表达UGP2蛋白质的DNA或其mRNA或保有其功能的同源物作为生物标志物在制备或筛选动脉粥样硬化引起的疾病的诊断试剂中的应用。
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