CN108610409A - Etv5在制备预防或治疗肥胖症及相关代谢性疾病药物中的应用 - Google Patents
Etv5在制备预防或治疗肥胖症及相关代谢性疾病药物中的应用 Download PDFInfo
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- CN108610409A CN108610409A CN201810312927.7A CN201810312927A CN108610409A CN 108610409 A CN108610409 A CN 108610409A CN 201810312927 A CN201810312927 A CN 201810312927A CN 108610409 A CN108610409 A CN 108610409A
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Abstract
本发明公开了一种与脂肪细胞分化成熟相关的标志物，所述标志物为ETV5基因或其编码的蛋白；本发明还公开了一种ETV5基因或蛋白抑制剂，所述抑制剂为抑制ETV5基因表达的shRNA，所述shRNA核酸序列为SEQ ID NO.1；进一步本发明公开了所述标志物和抑制剂在制备预防或治疗肥胖症及相关代谢性疾病治疗药物中的应用。本发明首次公开了ETV5可以作为药物分子靶向治疗靶点，在治疗肥胖及相关代谢性疾病如II型糖尿病领域具有潜在，良好的应用前景。
Description
技术领域
本发明涉及生物医学技术领域，具体涉及ETV5在制备预防或治疗肥胖症及相关代谢性疾病药物中的应用。
背景技术
现代社会中，肥胖症的发病率日益增加。世界卫生组织(World HealthOrganization,WHO)数据显示，截止到2015年，全球超过13亿成年人超重(30kg/m2>BMI>25kg/m2)，且超过6亿人被认定为肥胖(BMI≥30kg/m2)。随着我国经济的快速发展，人民生活水平的提高以及饮食结构的西方化，我国肥胖人数急剧增加，目前已成为世界第二大肥胖国，肥胖人数仅次于美国。肥胖可能引起糖尿病、高血压、高脂血症、非酒精性脂肪肝等多种代谢综合征。例如，肥胖女性患2型糖尿病的几率相较于非肥胖女性高出60倍。由此可见，肥胖给个人和社会带来了沉重的精神和经济负担，严重影响我国人民健康及社会经济的发展。尽管肥胖发病率高，危害巨大，但由于其病因复杂，发病机制和病理生理过程不清楚，目前尚缺乏有效的治疗方法。
脂肪组织是通过脂肪的间充质干细胞(MSCs)分化而产生的。脂肪细胞数量增多，体积增大都会引起脂肪组织增加，从而导致肥胖的产生。脂肪细胞数量增多是由前脂肪细胞增殖和分化过度引起的，而脂肪细胞体积增大是由细胞内甘油三酯累积程度增加引起的。因为肥胖是由于脂肪生成过多和脂肪细胞过度增大引起的，所以抑制脂肪生成被认为是预防和治疗肥胖的一个有效策略。
脂肪细胞分化过程是由体内的多种激素或生长因子作用于脂肪细胞膜表面的特异受体后，经不同的信号通路转导，改变多个脂肪细胞分化的转录因子活性，从而调控对脂肪细胞分化。目前，关于脂肪分化的研究以及药物设计主要集中在几个重要转录因子上，包括过氧化物酶体增殖物激活受体γ(peroxisome proliferator-activated receptors，PPARγ)和CCAAT/结合子蛋白(CCAAT/enhancer-binding proteins，C/EBPs)家族等。近年来，人们进行大规模基因组学分析(Genome-wide association study)，希望利用肥胖相关性的基因多态性分析，发现新的与肥胖紧密相关的基因。
发明内容
为了解决上述技术问题，本发明主要目的在于提供ETV5基因及其编码的蛋白质在脂肪组织分化成熟中起重要作用，可作为治疗肥胖及相关代谢性疾病的的新靶点；所述ETV5基因在制备治疗肥胖症及相关代谢性疾病药物中应用。
本发明首先提供一种与脂肪细胞分化成熟相关的标志物，所述标志物为ETV5基因或其编码的蛋白。
优选的，所述ETV5基因或其蛋白随着脂肪细胞分化过程表达先升高后下降，但仍显著高于分化前水平。
优选的，所述ETV5蛋白在脂肪组织中表达量高，所述ETV5定位于细胞核中。
优选的，所述ETV5能与PPRE结合，上调PPRE的转录活性，激活PPARγ通路相关的成脂基因，促进脂肪合成。
进一步地，本发明提供所述标志物在制备肥胖及相关代谢性疾病治疗产品中的应用。
优选的，所述产品包括试剂、药物及保健品。
进一步地，本发明还提供一种ETV5基因或蛋白抑制剂，所述抑制剂为抑制ETV5基因表达的shRNA，所述shRNA核酸序列为SEQ ID NO.1。
优选的，所述抑制剂在制备预防或治疗肥胖症及相关代谢性疾病药物中的应用。
优选的，所述相关代谢性疾病为糖脂代谢紊乱相关疾病，所述糖脂代谢紊乱相关疾病包括II型糖尿病、高血压、高脂血症、非酒精性脂肪肝。
优选的，所述抑制剂通过抑制ETV5基因或蛋白表达，进而抑制PPARγ通路相关的成脂基因表达，抑制脂肪细胞分化成熟，最终抑制脂肪生成。
优选的，所述成脂基因包括Pparg，Cebpa、Cebpd、Lipe和Adipoq。所述成脂基因是与脂肪细胞分化的相关基因。
所述Pparg(Pparγ)(peroxisome proliferator activated receptor gamma，过氧化物酶体增殖物激活受体γ)是核受体超家族成员，主要在脂肪组织内表达，是对脂肪细胞终末分化具有决定性作用的转录因子，是前脂肪细胞分化重要标志物。多种参与脂肪酸转运和代谢的基因在转录水平受Pparγ调控。
所述Cebpa(CCAAT/增强子结合蛋白α)和Cebpd(CCAAT/增强子结合蛋白δ)是C/EBPs家族成员，在调控前脂肪细胞分化过程中起着重要作用。
所述Lipe是激素敏感性脂肪酶(HSL)的基因，HSL是随着分化进程的增加而逐渐出现的分化中期标志物；所述Adipoq(脂联素AipoQ)只在成熟脂肪细胞中表达，可促进脂肪细胞分化，是含量最多的脂肪细胞因子。
更进一步地，本发明提供一种减肥药物，所述药物能够抑制PPARγ通路相关的成脂基因表达，抑制脂肪细胞分化成熟；所述药物包括ETV5基因或蛋白抑制剂，所述ETV5抑制剂为抑制ETV5基因表达的shRNA，所述shRNA核酸序列为SEQ ID NO.1；所述药物还包括药学上可接受的载体或其他减肥药物。
本发明所述的载体为药学上可以接受的载体，其指的是：一种或多种相容性固体或液体填料或凝胶物质。它们适合于人使用，而且必须有足够的纯度和足够低的毒性。“相容性”在此指的是组合物中各组分能和本发明的活性成分以及它们之间相互掺和，而不明显降低活性成分的药效。
优选的，所述载体包括但不限于：稀释剂、缓冲剂、混悬剂、乳剂、颗粒剂、包囊剂、赋形剂、填充剂、粘合剂、喷雾剂、透皮吸收剂、湿润剂、崩解剂、吸收促进剂、表面活性剂、着色剂、矫味剂或吸附载体。
优选的，所述药物可以制成包括但不限于显微注射剂、适于转染的剂型、注射液、片剂、粉剂、粒剂、胶囊剂。上述各种剂型的药物均可以按照药学领域的常规方法制备。
本发明有益效果：
1、本发明首次发现了ETV5基因及其编码的蛋白质在脂肪组织分化成熟中起重要作用；ETV5基因在制备治疗肥胖及糖脂代谢紊乱相关代谢性疾病药物的应用中，包括通过ETV5基因上下游靶点制备肥胖相关的药物。
2、本发明公开了一种抑制剂，所述抑制剂为抑制ETV5基因表达的shRNA，所述shRNA核酸序列为SEQ ID NO.1，具有抑制脂肪生成的新功能；所述抑制剂在制备治疗肥胖及相关代谢性疾病药物中的应用。
本发明首次公开了ETV5可以作为减肥药物分子靶向治疗药物的靶点，在治疗肥胖及相关代谢性疾病如II型糖尿病领域具有潜在，良好的应用前景。
附图说明
图1 ETV5蛋白在脂肪组织的定位和表达情况；
图2 ETV5在附睾脂肪组织、皮下脂肪组织、棕色脂肪组织中的表达情况；
图3 Western blot法测定ETV5蛋白在细胞分化过程中的表达情况；
图4通过实时荧光定量PCR和Western blot检测ETV5 shRNA#1、#2慢病毒感染小鼠3T3-L1前脂肪细胞后ETV5基因和蛋白的表达水平；
图5油红O染色测定观察ETV5对3T3-L1细胞分化功能的影响；
图6通过实时荧光定量PCR检测ETV5表达下调后脂肪细胞重要的成脂基因表达的情况；
图7敲低ETV5基因后对BMMSC细胞的成脂分化的影响；其中，图7A为Western Blot检测ETV5-shRNA#1慢病毒感染BMMSC细胞后ETV5蛋白表达水平；图7B为实时荧光定量PCR检测敲低ETV5基因表达后，成脂因子Pparg和Cebpa的mRNA表达水平；
图8为双荧光素酶活性检测结果；
图9为在小鼠体内，用ETV5 shRNA慢病毒干扰ETV5基因表达后，通过实时荧光定量PCR检测ETV5基因表达情况；
图10为敲低ETV5基因表达后，检测相关成脂基因Cebpa表达情况；
图11为敲低ETV5基因表达后，检测相关成脂基因Cebpd表达情况。
具体实施方式
以下实施例用于说明本发明，但不用来限制本发明的范围。若未特别指明，实施例中所用的技术手段为本领域技术人员所熟知的常规手段。
实施例中未注明具体条件的实验方法，通常为本领域常规方法，如按照常规条件如Sambrook等人，分子克隆，实验手册(第三版)(科学出版社，2002)中的所述条件，或按照试剂制造厂家所建议的条件。
本发明所述ETV5基因是在本发明之前的已知基因，所述小鼠ETV5基因的Genebank登录号：104156，所述人ETV5基因的Genebank登录号：2119；所述小鼠Pparg基因的Genebank登录号：19016；所述小鼠Cebpa基因的Genebank登录号：12606；所述小鼠Cebpd基因的Genebank登录号：12609；所述小鼠Lipe基因的Genebank登录号：16890；所述小鼠Adipoq基因的Genebank登录号：11450。
本发明中涉及到PPARγ指过氧化物酶体增殖物激活受体γ的蛋白；Pparg(Pparγ)指过氧化物酶体增殖物激活受体γ的基因。
多个基因组关联学研究发现，ETV5基因的变异与肥胖发生具有紧密相关性。ETV5在调节葡萄糖代谢和能量平衡中起重要作用。ETV5基因敲除小鼠体重降低，脂肪量减少，且高脂饮食喂养后出现肥胖抵抗，这表明ETV5会促进体脂和体重的增加，同时说明了ETV5与能量代谢的调控有直接关系，提示了ETV5可能在脂肪发生及成熟中有重要作用。
相关研究已经表明脂肪合成过程受一系列复杂的转录因子调节，如PPARγ、C/EBP及ADD/SREBP-1c等的调节。本发明主要是在细胞水平采用小鼠前脂肪细胞3T3-L1和小鼠骨髓间充质干细胞(BMMSC)为模型，设计2种携带有ETV5shRNA的慢病毒，感染3T3-L1前脂肪细胞及BMMSC后，经实时荧光定量PCR和Westernblot证实ETV5基因的mRNA水平和蛋白水平成功被抑制。ETV5基因被敲减后，3T3-L1前脂肪细胞和BMMSC细胞的分化及脂滴生成显著被抑制。通过基因分析，发现PPARγ通路相关基因的表达被显著抑制，继而通过双荧光素酶实验发现ETV5可以显著上调了PPAR反应元件(PPRE)的转录活性，提示ETV5是一个新颖的脂肪合成促进转录因子。本发明同时向野生型小鼠皮下脂肪中注射ETV5干扰慢病毒，7天后，脂肪生成相关基因的表达明显被抑制。本发明的研究结果说明ETV5是一个新颖的脂肪合成促进转录因子，通过激活PPARγ通路相关成脂基因，发挥促进脂肪合成的作用；而ETV5干扰慢病毒有抑制脂肪合成的作用，证明ETV5具有治疗肥胖症及相关代谢性疾病的应用价值。
本发明提供ETV5shRNA#1只敲低ETV5基因表达，对PEA6转录因子家族的其他成员ETV1、ETV4基因表达没有影响，说明ETV5 shRNA#1具有敲除ETV5基因的特异性。
本发明所用的主要实验材料如下：
细胞材料：小鼠前脂肪细胞3T3-L1购自ATCC公司；HEK293T细胞、BMMSC细胞购自广州赛业公司；
BCA蛋白定量试剂盒购自Thermo公司；4x Laemmli Sample Buffer，鼠二抗，兔二抗，PVDF膜，ECL化学发光底物购自BIO-RAD公司；PPARγ抗体购自Cell SignalingTechnology公司；ETV5，α-tublin,，β-actin和LaminB1抗体购自Proteintech公司；DMEM培养基、0.25％Trypsin-EDTA、Pen Strep、Opti-MEM培养基购自Gibco公司；FBS购自PANBiotech公司；3-isobutyl-1-methyxan-thine，地塞米松，胰岛素，油红O粉末购自Sigma公司；RNA小量提取试剂盒；Trizol试剂，3000转染试剂购自Invitrogen公司；逆转录试剂RT Master Mix，实时荧光定量PCR SYBR Premix Ex Taq试剂盒购自Takara公司；引物由上海立菲生物技术有限公司合成，含有shRNA的慢病毒由广州赛业公司设计及包装完成；ETV5质粒(pCMV-ETV5)购自Origene公司；pCMV-Entry、pGL4.74[hRluc2/TK]质粒，双荧光素酶检测系统购自Promega公司；异丙醇购自天津市科密欧化学试剂有限公司；0.22μm无菌针头式过滤器购自Millipore公司；脱脂奶粉购自伊利公司。
RIPA裂解缓冲液：150mM NaCl,1％Triton X-100,1％sodium deoxycholate,0.1％SDS,50mM Tris-HCl(PH 7.5),and 2mM EDTA(pH 8.0)。
实施例1分析ETV5蛋白在脂肪组织中表达情况
本发明通过免疫组化学确定ETV5蛋白在脂肪组织的定位和表达情况。从WT雄性小鼠中取出附睾脂肪组织，进行石蜡包埋和切片，DAB染色，结果如图1所示，ETV5在附睾脂肪细胞中有较高表达，并定位于细胞核中。
另外，从WT雄性小鼠中分别取出附睾脂肪组织(eWAT)、皮下脂肪组织(scWAT)和棕色脂肪组织(BAT)，对各组织蛋白进行核质分离。
蛋白的核质分离方法按照细胞核蛋白与细胞浆蛋白抽提试剂盒(碧云天，P00028)说明书指示进行。
随后进行常规Western blotting检测，所述Western blotting方法：RIPA细胞裂解液提取细胞蛋白，BCA法测定蛋白浓度，等量蛋白样品经10％SDS-PAGE电泳后转膜印迹，封闭，一抗4℃过夜孵育，洗膜加二抗，室温孵育1h，洗膜后用ECL显色。结果如图2所示，ETV5在皮下脂肪组织中的表达量较高，并进一步验证了ETV5主要分布于细胞核。Lamin B1作为胞核内参，α-tubulin作为胞浆内参。
实施例2采用小鼠前脂肪细胞3T3-L1证实ETV5脂肪细胞发生中的作用。
1、研究ETV5对3T3-L1前脂肪细胞分化功能的影响
3T3-L1前脂肪细胞于37℃，5％CO2培养箱中，常规培养于含有10％FBS+1％双抗的DMEM培养基中。待细胞生长接触融合(day 0)后继续培养48h，然后，通过加入0.5mM 3-isobutyl-1-methyxan-thine(MIX)，1μg/mL胰岛素及1μM地塞米松进行诱导分化，诱导分化48h后(day 2)换用仅含1μg/mL胰岛素的完全培养基，以后每两天换液直到第八天(day 8)细胞完全分化成熟。
3T3-L1前脂肪细胞诱导分化后0，1，2，3，4，7天收集蛋白，进行核质分离后，采用Western blot方法测定ETV5蛋白在细胞分化过程中的表达。
结果如图3所示，ETV5蛋白随着细胞分化过程表达升高，在细胞诱导分化到第二天(day 2)，ETV5的蛋白表达最高，随后蛋白水平稍有下降，但仍显著高于分化前水平。说明ETV5在分化关键期促进3T3-L1前脂肪细胞的分化。LaminB1作为胞核内参，α-tubulin作为胞浆内参。
2、构建ETV5RNA干扰慢病毒质粒转染3T3-L1前脂肪细胞
本发明设计了并制备了携带有ETV5shRNA或scramble shRNA的慢病毒，感染小鼠3T3-L1前脂肪细胞(方法参照《Pi,J.,et al.,Deficiency in the nuclear factor E2-related factor-2transcription factor results in impaired adipogenesis andprotects against diet-induced obesity.J Biol Chem,2010.285(12):p.9292-300.》)，诱导分化培养至第八天，收集细胞，通过实时荧光定量PCR和Western-Blot实验技术检测ETV5的mRNA和蛋白表达水平。
所述Western-Blot实验法同实施例1所述；所述实时荧光定量PCR检测法：使用RNA小量提取试剂盒提取细胞中的总RNA；用RT MasterMix进行逆转录；采用PCRSYBR Premix Ex Taq进行实时定量PCR，具体步骤参照说明书；其中，所述ETV5引物由上海立菲生物技术有限公司合成，所述ETV5引物序列为：
5’-TCAGTCTGATAACTTGGTGCTTC-3’(SEQ ID NO:4)，
5’-GGCTTCCTATCGTAGGCACAA-3’(SEQ ID NO:5)。
所述shRNA#1、shRNA#2和scramble shRNA序列分别为：
shRNA#1：TACATGAGAGGCGGGTATTTC(SEQ ID NO:1)；
shRNA#2：CACCTCCCACCAAGATCAAAC(SEQ ID NO:2)；
scramble shRNA：CCTAAGGTTAAGTCGCCCTCG(SEQ ID NO:3)。
实验结果如图4显示，在两个敲低实验组的样品中，只有ETV5 shRNA#1可以显著性下调ETV5的mRNA水平，ETV5的表达量下调了62％(P<0.05)(图4A)；Western blotting进一步证明ETV5 shRNA#1敲低实验组ETV5蛋白表达水平显著降低(图4B)，Western blotting的结果与实时荧光定量PCR的结果一致。说明在3T3-L1前脂肪细胞中，ETV5 shRNA#1慢病毒shRNA有敲低ETV5的作用，而ETV5 shRNA#2没有此作用。
3、采用油红O染色测定观察ETV5对3T3-L1前脂肪细胞分化功能的影响
在3T3-L1前脂肪细胞中敲低ETV5基因后，再诱导其分化成熟，在细胞诱导分化的第16天，弃去旧培养基后用PBS轻柔冲洗3次，加入4％的多聚甲醛，在室温下固定细胞10min；小心吸去多聚甲醛，并用PBS冲洗3次，然后用60％的异丙醇浸洗；随后加入配置好的油红O工作液，室温下染色30min，避光于摇床上；小心弃去油红O染液，每孔用60％的异丙醇分化至间质清晰，弃去异丙醇；用PBS冲洗3次，倒置显微镜下观察脂滴形成情况。
通过Image J软件对脂肪细胞的油红O染色结果进行定量计算，结果如图5所示，与ETV5-scramble阴性对照组相比，ETV5 shRNA#1实验组其脂滴的数量、总面积和平均大小都显著下降和减小；而ETV5 shRNA#2实验组其脂滴的数量和大小与阴性对照组相比变化不大。
结果表明敲低ETV5基因的表达可明显抑制3T3-L1前脂肪细胞分化，脂滴明显变小，脂滴数量明显减少。
4、通过实时荧光定量PCR法检测ETV5表达下调后脂肪细胞重要的成脂基因表达情况。
通过实时荧光定量PCR法检测ETV5表达下调后脂肪细胞重要的成脂基因(adipogenic genes)表达情况，所述成脂基因是与脂肪细胞分化相关的基因，其包括Pparg(Pparγ)，Cebpa、Cebpd、Lipe和Adipoq(adiponectin)。
所述引物如表1所示：
表1引物序列
结果如图6所示，3T3-L1前脂肪细胞ETV5表达下调后伴随有成脂基因Pparg、Cebpa、Lipe和Adipoq的表达下降。
上述实验结果表明ETV5shRNA#1运用慢病毒shRNA干扰技术有效地降低了ETV5基因的表达水平，而ETV5基因敲减后可以显著降低3T3-L1前脂肪细胞的分化能力。
结果提示ETV5参与前体脂肪细胞向成熟脂肪细胞分化的调控。
实施例3 ETV5敲低后抑制BMMSC细胞的成脂分化
小鼠骨髓间充质干细胞(BMMSC)可定向分化为脂肪细胞，因此也常被作为研究脂肪细胞分化的细胞模型。
运用ETV5 shRNA#1慢病毒感染小鼠BMMSC，通过Western Blot技术检测敲低实验组中ETV5蛋白表达水平。结果如图7A所示，敲低实验组(shETV5)中ETV5蛋白表达水平较阴性对照组scramble shRNA显著降低，说明在小鼠BMMSC细胞中，ETV5 shRNA#1有敲低ETV5的作用。
通过实时荧光定量PCR实验证实其经典成脂基因Pparg和Cebpa的mRNA水平均显著降低，如图7B所示。进一步验证了ETV5基因在脂肪细胞分化的前期具有重要作用，ETV5参与了脂肪细胞分化的调控。
实施例4 ETV5调控PPARγ信号通路
大多数成脂相关基因在启动子区域都含有PPRE(peroxisome proliferatorresponse elements，过氧化物酶体增殖物反应元件)，为了检测转录因子ETV5对PPRE活性的影响，本发明通过脂质体转染法将pGL3-PPRE，ETV5质粒(pCMV-ETV5)和pGL4.74[hRluc2/TK]质粒瞬时共转染至HEK293T细胞，并设置对照组转染相应的空载质粒(pCMV-Entry)，24h后采用双荧光素酶报告基因系统检测其启动子活性。所述脂质体转染法参照3000转染试剂说明进行。pGL4.74[hRluc2/TK]作为内参可消除由于细胞数目及转染效率等不同所带来的组间差异。结果如图8显示，pCMV-ETV5转染HEK293T细胞24h后，pGL3-PPRE的荧光素酶活性显著增强，是pCMV-Entry对照组的9.19倍，两者差异有显著性。双荧光素酶检测结果表明转录因子ETV5能与PPRE结合，从而调控基因在转录水平上的表达。该结果提示ETV5参与调控PPARγ信号通路。
所述pGL3-PPRE质粒构建方法参照《Li,Z.,et al.,Ghrelin promotes hepaticlipogenesis by activation of mTOR-PPARgamma signaling pathway.Proc Natl AcadSci U S A,2014.111(36):p.13163-8.》文献。
实施例5小鼠体内敲低ETV5后抑制成脂分化
本发明向小鼠皮下脂肪注射含有ETV5 shRNA的慢病毒10μL(1X109IU/mL)，注射病毒后7天，处死小鼠，取出皮下脂肪，提取RNA，通过实时荧光定量PCR测定ETV5及关键成脂基因的mRNA变化情况。结果如图9显示，相比于scramble shRNA及shRNA#2，shRNA#1可显著下调ETV5基因水平，同时成脂基因Cebpa及Cebpd也被显著抑制，如图10-11所示，提示成脂过程被抑制。
虽然，上文中已经用一般性说明及具体实施方案对本发明作了详尽的描述，但在本发明基础上，可以对之作一些修改或改进，这对本领域技术人员而言是显而易见的。因此，在不偏离本发明精神的基础上所做的这些修改或改进，均属于本发明要求保护的范围。
序列表
<110> 深圳大学
<120> ETV5在制备预防或治疗肥胖症及相关代谢性疾病药物中的应用
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Claims (10)
1.一种与脂肪细胞分化成熟相关的标志物，其特征在于，所述标志物为ETV5基因或其编码的蛋白。
2.如权利要求1所述标志物，其特征在于，所述ETV5基因或其蛋白随着脂肪细胞分化过程表达先升高后下降，但仍显著高于分化前水平。
3.如权利要求1所述标志物，其特征在于，所述ETV5能与PPRE结合，上调PPRE的转录活性，激活PPARγ通路相关的成脂基因，促进脂肪合成。
4.权利要求1～3所述标志物在制备预防或治疗肥胖症及相关代谢性疾病治疗产品中的应用。
5.一种ETV5基因或蛋白抑制剂，其特征在于，所述抑制剂为抑制ETV5基因表达的shRNA，所述shRNA核酸序列为SEQ ID NO.1。
6.权利要求5所述抑制剂在制备预防或治疗肥胖症及相关代谢性疾病药物中的应用。
7.如权利要求6所述应用，其特征在于，所述相关代谢性疾病为糖脂代谢紊乱相关的代谢性疾病；所述糖脂代谢紊乱相关代谢性疾病包括II型糖尿病、高血压、高脂血症、非酒精性脂肪肝。
8.如权利要求6所述应用，其特征在于，所述抑制剂通过抑制ETV5基因或蛋白表达，进而抑制PPARγ通路相关的成脂基因表达，抑制脂肪细胞分化成熟，最终抑制脂肪生成。
9.如权利要求8所述应用，其特征在于，所述成脂基因包括Pparg，Cebpa、Cebpd、Lipe和Adipoq。
10.一种减肥药物，其特征在于，所述药物能够抑制PPARγ通路相关的成脂基因表达，抑制脂肪细胞分化成熟；所述药物包括ETV5基因或蛋白抑制剂，所述ETV5抑制剂为抑制ETV5基因表达的shRNA，所述shRNA核酸序列为SEQ ID NO.1；所述药物还包括药学上可接受的载体或其他减肥药物。
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