CN114262705B - 防治糖酯代谢疾病的miRNA、药物组合物及其应用 - Google Patents
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Abstract
本发明公开了防治糖酯代谢疾病的miRNA、药物组合物及其应用,所述miRNA包括no‑miRNA‑18、no‑miRNA‑6中的至少一种,所述miRNA具有降脂、保肝、抗氧化应激和抗炎的作用;所述miRNA可以单独使用,也可以与miRNA‑592、miRNA‑1247‑3P、miRNA‑3072等10种miRNA进行组合,用于防治糖酯代谢疾病,降脂保肝具有显著的疗效。
Description
技术领域
本发明涉及外泌体技术领域,尤其涉及一种防治糖酯代谢疾病的miRNA、药物组合物及其应用。
背景技术
外泌体(Exosome)自首次于绵羊血液内被发现后,经过学者对其结构和功能研究后正式命名为“Exosomes”。外泌体存在广泛,来源丰富,可在植物和动物中的体液及体外培育的动物细胞内获得。现今,其特指直径在40-200nm左右的微小纳米样颗粒或囊泡物质,显微形态为囊状或茶托状。
糖和脂的代谢作为机体能量供给的主要来源,在维持正常的生命活动中起着关键作用,但糖脂代谢紊乱会产生一系列的慢性疾病,诸如脂肪肝、糖尿病、高脂血症、动脉硬化性心脑血管病等,且长期的糖脂代谢紊乱会损害全身脏器的技能,最终致残致死。目前,非酒精性脂肪肝患者被认为是糖酯代谢紊乱中最普遍的患病人群,可直接诱发2型糖尿病(T2DM)及其他慢性疾病,对全球医疗保健造成越来越大的负担,严重影响中老年人群的身心健康。
中医药具有“多组分,多途径,多靶点,整体调节”的特点,在糖尿病及其并发症防治方面具有独特优势。熊胆是四大名贵动物药之一,为熊科动物的干燥胆囊(胆汁),被誉为“药中黄金”。其始载于《药性论》记载其“主小儿五疳、杀虫、治恶疮”;历代医书记载了其凉肝血、泻肝火、清热解毒、清肝明目等功效,迄今应用千年疗效确切。熊胆作为药用始载于唐代的《新修本草》,距今已有一千三百多年历史。明代李时珍的《本草纲目》也载有:熊胆,气味苦寒、无毒,入肝胆心经。现代医药学研究证实,熊胆具有清热解毒、疏肝利胆,去翳明目,解痉镇痛作用。熊胆的抗炎、保肝、镇痛等多种药理活性也被发现。外泌体作为细胞质粒,在细胞与细胞交流传递中起着重要作用。目前发现外泌体具有较强的药理活性,且较一般化学成分具有更优的治疗效果。
熊胆(汁/粉)作为“血肉有情之品”类动物药,其主要药效成分除胆汁酸类化学成分外,其中含有的外泌体类物质有可能参与了其调节糖脂代谢类疾病(高脂血症、糖尿病、脂肪肝、肥胖、动脉硬化疾病等)的药效作用发挥,因此有必要研究熊胆外泌体,通过提取和分析熊胆(汁/粉)外泌体中含有的miRNA类有效作用成分并为制备防治糖酯代谢疾病提供新的研究方向,进而为临床从根本上治疗糖脂代谢疾病提供科学有效的药物。
发明内容
鉴于此,本发明的目的在于提供防治糖脂代谢类疾病(高脂血症、糖尿病、脂肪肝、肥胖、动脉硬化疾病等)的miRNA、药物组合物及其应用,为开发防治糖酯代谢疾病的药物提供新的方向。
一种防治糖酯代谢疾病的miRNA,所述miRNA为no-miRNA-18、no-miRNA-6中的至少一种;所述no-miRNA-18的核苷酸序列如SEQ ID NO:1所示,所述no-miRNA-6的核苷酸序列如SEQ ID NO:2所示。
进一步地,所述miRNA还包含miRNA-592、miRNA-1247-3P、miRNA-3072中的一种或多种;所述miRNA-592的核苷酸序列如SEQ ID NO:3所示,所述miRNA-1247-3P的核苷酸序列如SEQ ID NO:4所示,所述miRNA-3072的核苷酸序列如SEQ ID NO:5所示。
进一步地,所述miRNA还包含miRNA-3541、miRNA-343、miRNA-673-3p、miRNA-6324、miRNA-1247-5p、miRNA-370-3p、miRNA-675-5p中的一种或多种,核苷酸序列依次如SEQ IDNO:6-12所示。
本发明提供上述miRNA在制备防治糖脂代谢类疾病药物中的应用,所述糖脂代谢类疾病包括高脂血症、糖尿病、脂肪肝、肥胖、动脉硬化等疾病。
本发明提供一种防治糖酯代谢疾病的药物组合物,包括上述防治糖酯代谢疾病的miRNA,还包括可药用的载体,所述可药用载体包括但不限于:软磷脂、硬脂酸铝、氧化铝、离子交换材料、自乳化药物传递系统、吐温或其他表面活化剂、血清蛋白、缓冲物质如磷酸盐、氨基乙酸、山梨酸、水、盐、电解质如硫酸盐精蛋白、磷酸氢二钠、磷酸氢钾、氯化钠、锌盐、硅酸镁、饱和脂肪酸部分甘油酯混合物等。
进一步地,所述药物组合物包括可药用的辅料。常用的药物辅料如粘合剂(如微晶纤维素)、填充剂(如淀粉、葡萄糖、无水乳糖和乳糖珠粒)、崩解剂(如交联PVP、交联羧甲基淀粉钠、交联羧甲基纤维素钠、低取代羟丙基纤维素)、润滑剂(如硬脂酸镁)以及吸收促进剂、吸附载体、香味剂、甜味剂、赋形剂、稀释剂、润湿剂等。
进一步地,本发明的miRNA成分及熊胆以及其药物组合物可按本领域常规方法制备并可以通过肠道或非肠道或局部途径给药。口服制剂包括胶囊剂、片剂、口服液、颗粒剂、丸剂、散剂、丹剂、膏剂等;非肠道给药制剂包括注射液等;局部给药制剂包括霜剂、贴剂、软膏剂、喷雾剂等。优选单一成分溶解后经脉给药。
本发明的miRNA成分及熊胆以及其药物组合物的给药途径可以为经脉、口服、舌下、经皮、经肌肉或皮下、皮肤粘膜、尿道、阴道等。
与现有技术相比,本发明的有益效果为:
本发明首次提出了no-miRNA-18和no-miRNA-6具有降脂、保肝、抗氧化应激和抗炎的作用,可作为防治糖酯代谢疾病的药物,为制备防治糖酯代谢疾病的药物提供新的方向。
本发明提出的miRNA-592、miRNA-1247-3P和miRNA-3072具有较其他miRNA更优的降脂、保肝等药效,可作为防治糖酯代谢疾病的潜在有效药物。
本发明所述miRNA简单易得,可以单独使用或以药物组合物的形式使用,范围广泛,经济可行。
附图说明
图1 为本发明提供的熊胆汁中12种miRNA含量示意图。
图2 为本发明提供的12种miRNA对NAFLD细胞活力的影响对比图。
图3为本发明提供的12种miRNA的保肝药效对比图。
图4 为本发明提供的12种miRNA的降脂药效对比图。
图5 为本发明提供的12种miRNA的抗炎药效对比图。
图6 为本发明提供的12种miRNA的抗氧化药效对比图。
图7 为本发明提供的12种miRNA的综合药效及曲线下总面积对比图。
图8 为本发明提供的miRNA-592、miRNA-1027-3p和miRNA-3072的降脂功效对比图。
图9为本发明提供的miRNA-592、miRNA-1027-3p和miRNA-3072对相关蛋白的影响对比图。
具体实施方式
实施例1 熊胆汁miRNA的制备及对糖脂代谢细胞模型的药物活性研究
材料:本发明所述新鲜的熊胆汁样品来源:资溪康仁堂生物开发有限公司,批号:20191211。
1 . 熊胆汁中miRNA药物制备
取适量新鲜的熊胆汁,按照相关文献方法(差速离心法)进行分离。取适量蒸馏水溶解熊胆粉,先于3000g/min离心20min,再于5000g/min离心15min,取溶液再于10000g/min离心40min,取溶液于100000g/min离心60min,收集沉淀,使用PBS清洗三次,再根据制造商的说明,按TRIzol试剂操作要求从沉淀中提取总RNA。总RNA进行反转录,以使用SYBR互补DNA (cDNA)® PrimeScript® RT-PCR试剂盒和基因组DNA (gDNA) Eraser (TaKaRa公司,日本)。主混合物(10 μL)包括:PrimeScript RT酶混合物I (1.0 μL),RT Primer Mix *4(1.0 μL),5×PrimeScript Buffer 2 (4.0 μL,实时)和RNase Free dH 2O (4.0 μL)。PCR扩增按以下步骤进行:第1阶段,在95 °C下预热变性40 s,第2阶段,循环系统在95 °C下循环40次15 s,在58 °C下循环1 min。表1中提供了用于该反应的引物的序列(10种已知的miRNA和2种本文新发现的no-miRNA)。每个样品测量三次。将上述基因的表达水平标准化为甘油醛3-磷酸脱氢酶(GAPDH)的表达水平,并通过比较2- ΔΔCt方法进行测量。
表1: 10种miRNA引物序列
基因名称 | RT引物序列(5’-3’) |
miRNA-592 | ATTGTGTCAATATGCGATGATGT |
miRNA-675-5p | TGGTGCGGAAAGGGCCCACAGT |
miRNA-1247-3p | CGGGAACGTCGAGACTGGAGC |
miRNA-3072 | TGCCCCCTCCAGGAAGCCTTCTT |
miRNA-3541 | TCCCTCCCCCTCACTGCA |
miRNA-343 | TCTCCCTCCGTGTGCCCAGA |
miRNA-673-3p | TCCGGGACTGAGTTCTGTGCAC |
miRNA-6324 | TCAGTAGGCCAGACAGCAAGCAC |
miRNA-1247-5p | ACCCGTCCCGTTCGTCCCCGGA |
miRNA-370-3p | GCCTGCTGGGGTGGAACCTGGT |
no-miRNA-18 | TCTTTATCCCTGTAAACAGA |
no-miRNA-6 | TCCCTTTCGCAAGCTTCGAC |
GAPDH | ACGGATTTGGTCGTATTGGG |
2. 糖酯代谢细胞模型的建立及给药
相关药物的配置:参考相关文献配置FFA溶液(OA:PA=2:1)配置成1.0 mmol/mL的溶液,需要时按相关要求稀释成相应浓度使用。所有miRNA使用DMEM培养基稀释成20 mmol/L溶液使用,避光,现用现配。miRNA转染混合液配制。A液:按说明书操作要求用适量的无血清培养基稀释转染试剂,室温孵育5 min。B液:miRNA用适量的无血清培养基稀释至10pmol,室温孵育5 min。再将A和B液体充分混匀,室温孵育20 mim后,即得转染混合液,现配现用,所有操作均避光。
通过使用适量FFA混合溶液诱导NAFLD细胞模型,分别加入FFA(OA:PA=2:1,1.0mmol/L)诱导HepG2细胞,作用24 h以诱导NAFLD细胞模型。本章实验共分14个组别,分别为空白对照(NC)组,非酒精性脂肪肝模型(NAFLD)组,12种miRNA治疗组(10pmol/L),每组重复8 孔。在FFA成功诱导NALFD细胞模型后,分别按要求加入指定药物,共培育24 h。最后分别按相关试剂盒要求检测相关指标。
3. 细胞活力测定
除去上清液后,每孔加入的MTT(20 μL,5 mg/mL用PBS配置),共培育4 h后,去除MTT溶液,加入二甲基亚砜(150 μL/孔,DMSO),使用酶标仪在490 nm处测量吸光度,可反映活细胞的数量,并计算细胞存活率。根据以下公式计算细胞活力:
细胞相对存活率=(实验组OD值-空白组OD值)/(阴性组OD值-空白组OD值)×100%(注:实验中应设无细胞的空白调零孔;MTT试剂配制完成后应避光储存,避免反复冻融)。
4. 细胞生化指标的测定
取出细胞,小心移去所有培养基,加入适量PBS缓冲溶液(2 mL)缓慢冲洗3次,用细胞刮刀将细胞刮下,移至1.5 mL离心管中,12000 g离心10 min,收集沉淀。加入适量PBS,轻轻振摇重悬细胞,用超声波破碎仪破碎细胞,获得细胞匀浆液,最后按照试剂盒相关要求检测细胞内TG、TC、AST、ALT、NO、SOD、MDA和GSH蛋白含量。
5.综合药理学评价
分析所有生化指标使用归一化处理后绘制药效雷达图,并计算综合药效面积(三角函数计算),筛选出综合药效排行前三的miRNA被认为是熊胆汁中具有主要药效作用的miRNA,并进行以下实验。
6. 细胞病理学观察
取出细胞6孔板,小心弃去所有上清液,并按照以下细胞油红O染色方法评估3种主要药效miRNA对NAFLD细胞模型的降脂功效。
(1)按试剂盒要求配置相应的苏木精和油红O染液,避光,现配现用;
(2)将六孔板中生长的细胞用磷酸盐缓冲盐水(PBS)洗涤3 次,加入适量多聚甲醛(4%)静置20 min;
(3)将附着在盖玻片上的细胞用油红O(ORO)固定剂染色15 min,缓慢振摇使染色充分;
(4)加入适量60%异丙醇(1 mL)浸泡5 min,缓慢振摇使浸泡充分;
(5)加苏木精染色剂1 min,用蒸馏水洗涤3 次;
(6)用显微镜观察被油红O染色的脂质滴。
7. Western blot分析
所有细胞样本按照标准程序用所指示的抗体进行免疫印迹,将细胞悬液在裂解液中溶解,并在4℃,以12000 rpm离心15 min,弃去沉淀。上清液用SDS-PAGE分离蛋白质样品,并用BCA蛋白分析试剂盒(thermofisher)测定组织匀浆的蛋白质浓度。通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(50μg)分离总蛋白,然后将其转移至PVDF膜(Millipore Corp,Billerica),5%脱脂奶粉中密封,37 ℃环境静置1 h。随后,将针对PPARα、NF-κB、CYP7A1和GAPDH的兔抗小鼠单克隆抗体(1:1,000; Abcam,Cambridge,MA)添加到膜中,并在4 ℃环境下振摇过夜。将膜用磷酸盐缓冲盐水和吐温(PBST)洗涤3 次,每次5 min。然后,用辣根过氧化物酶标记的山羊抗兔二抗(1:4,000;Cell Signaling Technology,Danvers,MA)添加到膜中,并将其在室温下孵育2 h。将膜用带Tween的tris-缓冲盐水(TBST)洗涤两次,每次10min,然后用电化学发光(ECL)光致发光溶液处理以进行成像,最后使用ImageJ软件分析所有蛋白的测试结果。
8. 统计分析
数据表示为平均值(mean)±标准差(S.D.),n=8。使用SPSS 18.0软件(SPSS公司,美国芝加哥)对组间所有数据进行统计分析,平均比较检验采用T检验或方差分析。当*P<0.05时差异具有统计学意义,认为**P<0.01时具有显著差异,而***P<0.001时认为是极其显著性差异。
研究结果表明:结果如图1所示,本专利首次于熊胆汁中发现了10种miRNA及其表达量。另外,本发明基于基因测序技术还发现了两种新miRNA,且在新鲜的熊胆汁中均有较高的表达水平,可通过相应技术提取,并将其定义其为“no-miRNA-18”和“no-miRNA-6”。上述10种miRNA及新的miRNA信息如表2所示。
表2 :12种主要miRNA信息
序号 | 基因名称 | 序列 |
SEQ ID NO:1 | no-miR-18 | AGGAGAAACGAAGUAGAACCCU |
SEQ ID NO:2 | no-miR-6 | CAACACUGCACUGGAAGAUGGA |
SEQ ID NO:3 | miR-592 | AUUGUGUCAAUAUGCGAUGAUGU |
SEQ ID NO:4 | miR-1247-3p | CGGGAACGUCGAGACUGGAGC |
SEQ ID NO:5 | miR-3072 | UGCCCCCUCCAGGAAGCCUUCUU |
SEQ ID NO:6 | miR-3541 | UCCCUCCCCCUCACUGCA |
SEQ ID NO:7 | miR-343 | UCUCCCUCCGUGCCCAGA |
SEQ ID NO:8 | miR-673-3p | UCCGGGACUGAGUUCUGUGCAC |
SEQ ID NO:9 | miR-6324 | UCAGUAGGCCAGACAGCAAGCAC |
SEQ ID NO:10 | miR-1247-5p | ACCCGUCCGUUCGUCCCCGGA |
SEQ ID NO:11 | miR-370-3p | GCCUGCUGGGGUGGAACCUGGU |
SEQ ID NO:12 | miR-675-5p | UGGUGCGGAAAGGGCCCACAGU |
如图2所示,12种miRNA和UDCA能显著增加NAFLD细胞模型的细胞活力。较空白对照组,NAFLD模型组的细胞活力显著性降低(P<0.05)。较NAFLD模型组,12种miRNA作用于NAFLD细胞模型后细胞活力均有不同的显著升高(P<0.05),其中miRNA592、miRNA1247-3p、miRNA3072最为显著,且本研究首次发现的no-miRNA-18和no-miRNA-6也具有明显的保肝药理活性。
实施例2 测定miRNA对NAFLD细胞模型肝功能影响
ALT和AST可以直接反应肝功能异常的严重程度。为了研究熊胆汁中12种miRNA对肝功能的影响,测定了NAFLD细胞模型中的ALT和AST水平。如图3A、3B所示,与NC组相比,NAFLD组的ALT和AST水平显着升高(p<0.01)。经治疗后,治疗组的大鼠ALT和AST水平均显着降低(p<0.05)。以上结果提示着12种miRNA可通过调节ALT和AST水平起到降血糖作用。从图3A、3B中明显看出,no-miRNA-18和no-miRNA-6对ALT和AST具有较强调节作用,可作为未来保肝的潜在基因类药物。
实施例3 测定miRNA对NAFLD细胞模型中血脂指标的影响
血脂四项是临床诊断血脂水平的常规指标,它反映糖脂代谢异常疾病发展的严重程度。为了研究熊胆汁中12种miRNA对脂质代谢能力的改善作用,测定了NAFLD细胞模型中的TG和TC水平。如图4所示,与NC组相比,NAFLD组的TG(图4A)和TC(图4B)水平显着升高(p<0.01)。经治疗后,治疗组的大鼠TG和TC水平均显着降低(p<0.05)。以上结果提示着12种miRNA可通过调节TG和TC水平起到降血糖作用。值得一提的是,no-miRNA-18和no-miRNA-6对血脂指标具有较强调节作用,可作为未来治疗糖酯代谢异常疾病的潜在基因类药物。
实施例4 测定miRNA对NAFLD细胞模型的炎症改善作用
细胞出现炎症时常会释放出大量NO,因此其常用来作为细胞模型炎症程度的评价指标。为了研究熊胆汁中12种miRNA对细胞炎症的改善作用,本实施例测定了NAFLD细胞模型中的的NO水平。如图5所示,与NC组相比,NAFLD组的NO水平显着升高(p<0.01)。经治疗后,治疗组的大鼠NO水平均显着降低(p<0.05)。以上结果提示着12种miRNA可通过调节NO水平起到抗炎作用。炎症是许多慢性疾病的重要影响因素,本发明首次发现no-miRNA-18和no-miRNA-6对炎症指标具有较强调节作用,可作为未来治疗多种肝脏慢性疾病的潜在基因类药物。
实施例5 测定miRNA对NAFLD细胞模型氧化应激的作用
氧化应激作为肝脏细胞损伤最重要的致病机制,SOD、MDA和GSH作为重要氧化应激评价指标,相关研究常用来评价药物的抗氧化药理活性。为了研究熊胆汁中12种miRNA对细胞氧化应激水平的改善作用,本实施例测定了NAFLD细胞模型中的的SOD、GSH和MDA水平。如图6所示,与NC组相比,NAFLD组的SOD(图6A)、GSH(图6B)和MDA(图6C)水平显着降低(p<0.01),而MDA水平显著升高(p<0.05)。经治疗后,治疗组的大鼠SOD和GSH水平均显着升高(p<0.05),而MDA水平显著降低(p<0.05)。以上结果提示着12种miRNA可通过调节SOD、GSH和MDA水平起到调节氧化应激作用。另外,no-miRNA-18和no-miRNA-6对氧化应激指标具有较强调节作用,这意味着对于许多慢急型疾病具有潜在的治疗作用。
实施例6 miRNA对NAFLD细胞模型的药理活性差异分析
现代学者发现miRNA具有突出药理活性,如抗炎、调节氧化应激和抗肿瘤等。然而,并非所有miRNA均有显著的药理活性,因此本发明基于NAFLD细胞模型对于熊胆汁中12种miRNA的降脂、保肝、抗炎、抗氧化应激进行综合药效比较分析,以求筛选出BES中的主要功效miRNA。图7中,12种miRNA药效指数图如图7A所示,12种miRNA的AUC如图7B所示。较NC组,NAFLD模型组的AUC显著降低(P<0.05),几乎为0。较NAFLD组,所有治疗组的AUC显著上升(P<0.05),其中miRNA592、1247-3p、3072的AUC排在前三,且均强于UDCA。基于以上所有结果,认定 miRNA592、miRNA1247-3p、miRNA3072可能是BES中起主要药效的miRNA。另外,本专利首次发现no-miRNA-18和no-miRNA-6是可通过多途径、多靶点治疗相关疾病。
实施例7 miRNA对NAFLD细胞模型肝组织和细胞油红O染色的影响
肝组织和细胞油红O染色常被用于观察NAFLD疾病的严重程度,可更为直观的观察细胞内脂质代谢异常的程度。如图8所示,空白组中细胞结构正常,正常分裂,无红色的油脂存在。NAFLD组中的细胞出现显著的细胞结构异常,并伴有大量的红色油脂浸润。所有药物治疗组的细胞结构得到恢复,红色油脂面积显著减少,其中miRNA-592组仅具有少量红色油脂。
实施例8 miRNA对NAFLD细胞模型中蛋白表达的影响
氧化应激、炎症和糖脂代谢紊乱是NALFD的重要致病因素。为了验证miRNA-592、miRNA-1247-3p、miRNA-3072在NAFLD细胞模型中的保护作用机制,本实施例测定了NAFLD细胞模型中PPARα、NF-κBp65和CYP7A1的蛋白表达水平。如图9所示,NAFLD-T2DM组PPARα(图9A)和NF-κB(图9B)蛋白的表达水平与NC组相比较显著降低(P<0.05),而CYP7A1(图9C)蛋白的表达水平显著降低(P<0.05)。四个治疗组PPARα和NF-κB的表达与NAFLD组比较均显著降低(P<0.05),而CYP7A1的表达显著升高(P<0.05)。总的来说,这些结果表明miRNA-592、miRNA-1247-3p、miRNA-3072对NALFD细胞模型中PPARα、NF-κB蛋白的活化和CYP7A1蛋白的抑制有显著的调节作用,且功效均优于UDCA。
现代学者认为胆汁酸转运可通过调节体内脂质代谢治疗NAFLD,为胆汁酸类成分的重要保肝作用机制。为了探讨miRNA-592、miRNA-1247-3p、miRNA-3072在NAFLD细胞模型中的作用机制,本实施例测定了NAFLD细胞模型中FXR1和TGR5的蛋白表达水平。如图9所示,与NC组相比较,NAFLD-T2DM组的FXR1(图9D)蛋白表达水平显著升高(P<0.05),而TGR5(图9E)蛋白表达水平显著降低(P<0.05)。与NAFLD-T2DM组比较,四个治疗组FXR1和TGR5蛋白表达均显著降低(P<0.05),而TGR5蛋白表达水平显著上升(P<0.05),表明其均可调节胆汁转运途径。这些结果表明miRNA-592、miRNA-1247-3p、miRNA-3072对NAFLD细胞模型中FXR1的抑制和TGR5蛋白的活化有明显的调节作用。综上所述,miRNA-592、miRNA-1247-3p、miRNA-3072是可通过调节FXR1和TGR5蛋白表达水平治疗NAFLD。
综上所述,本发明提供的熊胆汁中12种miRNA可以糖酯代谢疾病及相关症状,故而其可以作为防治糖酯代谢疾病的潜在药物。特别地,本发明在熊胆汁中新发现了两种miRNA:no-miRNA-18和no-miRNA-6,并通过相关药理学实验探究了二者的相关药理活性。结果表明,no-miRNA-18和no-miRNA-6具有降脂、保肝、抗氧化应激和抗炎的作用,为防治糖酯代谢疾病的治疗提供新的潜在有效药物。本申请特别发现miRNA-592、miRNA-1247-3P和miRNA-3072具有较其他miRNA更优的降脂、保肝等药效,因此本文特此强调miRNA-592、miRNA-1247-3P和miRNA-3072可作为防治糖酯代谢疾病的潜在有效药物。
序列表
<110> 武汉萃绿科技有限公司
<120> 防治糖酯代谢疾病的miRNA、药物组合物及其应用
<160> 25
<170> SIPOSequenceListing 1.0
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<211> 22
<212> RNA
<213> no-miR-18(no-miR-18)
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<213> miR-592(miR-592)
<400> 3
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<212> RNA
<213> miR-1247-3p(miR-1247-3p)
<400> 4
cgggaacguc gagacuggag c 21
<210> 5
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<212> RNA
<213> miR-3072(miR-3072)
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<400> 9
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<210> 10
<211> 21
<212> RNA
<213> miR-1247-5p(miR-1247-5p)
<400> 10
acccguccgu ucguccccgg a 21
<210> 11
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<212> RNA
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gccugcuggg guggaaccug gu 22
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<213> 人工序列(Artificial Sequence)
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<213> 人工序列(Artificial Sequence)
<400> 14
tggtgcggaa agggcccaca gt 22
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cgggaacgtc gagactggag c 21
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tgccccctcc aggaagcctt ctt 23
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<213> 人工序列(Artificial Sequence)
<400> 19
tccgggactg agttctgtgc ac 22
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<213> 人工序列(Artificial Sequence)
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<210> 21
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<212> DNA
<213> 人工序列(Artificial Sequence)
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tctttatccc tgtaaacaga 20
<210> 24
<211> 20
<212> DNA
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tccctttcgc aagcttcgac 20
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acggatttgg tcgtattggg 20
Claims (5)
1.一种防治非酒精性脂肪肝的miRNA,其特征在于,所述miRNA为no-miRNA-18,核苷酸序列如SEQ ID NO:1所示。
2.权利要求1所述miRNA在制备防治非酒精性脂肪肝药物中的应用。
3.一种防治非酒精性脂肪肝的药物组合物,其特征在于,包含权利要求1所述miRNA及可药用的载体。
4.根据权利要求3所述的药物组合物,其特征在于,还包括可药用的辅料。
5.根据权利要求3所述的药物组合物,其特征在于,所述药物组合物剂型包括口服制剂、非肠道给药制剂的任意一种。
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