CN115814086B - circPI4KB在诊断和治疗非酒精性脂肪性肝病以及相关药物筛选中的应用 - Google Patents
circPI4KB在诊断和治疗非酒精性脂肪性肝病以及相关药物筛选中的应用 Download PDFInfo
- Publication number
- CN115814086B CN115814086B CN202210837047.8A CN202210837047A CN115814086B CN 115814086 B CN115814086 B CN 115814086B CN 202210837047 A CN202210837047 A CN 202210837047A CN 115814086 B CN115814086 B CN 115814086B
- Authority
- CN
- China
- Prior art keywords
- circpi
- fatty liver
- liver disease
- alcoholic fatty
- nafld
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
- 208000008338 non-alcoholic fatty liver disease Diseases 0.000 title claims abstract description 97
- 239000003814 drug Substances 0.000 title claims abstract description 21
- 238000003745 diagnosis Methods 0.000 title abstract description 10
- 238000012216 screening Methods 0.000 title abstract description 6
- 238000011282 treatment Methods 0.000 title description 9
- 238000002360 preparation method Methods 0.000 claims abstract description 4
- 210000002966 serum Anatomy 0.000 claims description 22
- 238000011529 RT qPCR Methods 0.000 claims description 13
- 108091027967 Small hairpin RNA Proteins 0.000 claims description 11
- 239000004055 small Interfering RNA Substances 0.000 claims description 11
- 210000003494 hepatocyte Anatomy 0.000 claims description 9
- 230000002401 inhibitory effect Effects 0.000 claims description 8
- 210000005228 liver tissue Anatomy 0.000 claims description 5
- 239000012807 PCR reagent Substances 0.000 claims description 4
- 108091028043 Nucleic acid sequence Proteins 0.000 claims description 3
- 230000014509 gene expression Effects 0.000 abstract description 34
- 230000008021 deposition Effects 0.000 abstract description 13
- 150000002632 lipids Chemical class 0.000 abstract description 13
- 229940079593 drug Drugs 0.000 abstract description 12
- 238000000034 method Methods 0.000 abstract description 11
- 108090000623 proteins and genes Proteins 0.000 abstract description 9
- 210000005229 liver cell Anatomy 0.000 abstract description 6
- 238000011160 research Methods 0.000 abstract description 6
- 102000004169 proteins and genes Human genes 0.000 abstract description 5
- 230000009471 action Effects 0.000 abstract description 4
- 239000000090 biomarker Substances 0.000 abstract description 4
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 abstract description 3
- 201000010099 disease Diseases 0.000 abstract description 2
- 238000007877 drug screening Methods 0.000 abstract description 2
- 210000004027 cell Anatomy 0.000 description 35
- 206010053219 non-alcoholic steatohepatitis Diseases 0.000 description 14
- 108020004999 messenger RNA Proteins 0.000 description 12
- HVYWMOMLDIMFJA-DPAQBDIFSA-N cholesterol Chemical compound C1C=C2C[C@@H](O)CC[C@]2(C)[C@@H]2[C@@H]1[C@@H]1CC[C@H]([C@H](C)CCCC(C)C)[C@@]1(C)CC2 HVYWMOMLDIMFJA-DPAQBDIFSA-N 0.000 description 10
- 239000002609 medium Substances 0.000 description 10
- 108091070501 miRNA Proteins 0.000 description 10
- 239000002679 microRNA Substances 0.000 description 10
- 210000004185 liver Anatomy 0.000 description 9
- 230000008859 change Effects 0.000 description 8
- 238000001514 detection method Methods 0.000 description 8
- 206010073071 hepatocellular carcinoma Diseases 0.000 description 7
- 102100031181 Glyceraldehyde-3-phosphate dehydrogenase Human genes 0.000 description 6
- 230000015572 biosynthetic process Effects 0.000 description 6
- 238000006243 chemical reaction Methods 0.000 description 6
- 108020004445 glyceraldehyde-3-phosphate dehydrogenase Proteins 0.000 description 6
- 230000002018 overexpression Effects 0.000 description 6
- 235000012000 cholesterol Nutrition 0.000 description 5
- 238000011161 development Methods 0.000 description 5
- 239000007788 liquid Substances 0.000 description 5
- 238000010839 reverse transcription Methods 0.000 description 5
- 239000000523 sample Substances 0.000 description 5
- 239000006228 supernatant Substances 0.000 description 5
- 238000003786 synthesis reaction Methods 0.000 description 5
- 238000001890 transfection Methods 0.000 description 5
- 102100022089 Acyl-[acyl-carrier-protein] hydrolase Human genes 0.000 description 4
- 108010039731 Fatty Acid Synthases Proteins 0.000 description 4
- 206010016654 Fibrosis Diseases 0.000 description 4
- 238000013232 NAFLD rodent model Methods 0.000 description 4
- 230000000694 effects Effects 0.000 description 4
- 235000021588 free fatty acids Nutrition 0.000 description 4
- 239000001963 growth medium Substances 0.000 description 4
- 208000019423 liver disease Diseases 0.000 description 4
- 238000002156 mixing Methods 0.000 description 4
- 239000000203 mixture Substances 0.000 description 4
- 230000001105 regulatory effect Effects 0.000 description 4
- 239000000243 solution Substances 0.000 description 4
- 108091032973 (ribonucleotides)n+m Proteins 0.000 description 3
- 102100029077 3-hydroxy-3-methylglutaryl-coenzyme A reductase Human genes 0.000 description 3
- 101000988577 Homo sapiens 3-hydroxy-3-methylglutaryl-coenzyme A reductase Proteins 0.000 description 3
- 239000012124 Opti-MEM Substances 0.000 description 3
- 238000003556 assay Methods 0.000 description 3
- 238000004364 calculation method Methods 0.000 description 3
- 239000003153 chemical reaction reagent Substances 0.000 description 3
- 230000007882 cirrhosis Effects 0.000 description 3
- 208000019425 cirrhosis of liver Diseases 0.000 description 3
- 239000002299 complementary DNA Substances 0.000 description 3
- 230000007423 decrease Effects 0.000 description 3
- 239000013604 expression vector Substances 0.000 description 3
- 231100000844 hepatocellular carcinoma Toxicity 0.000 description 3
- 230000005764 inhibitory process Effects 0.000 description 3
- 230000003834 intracellular effect Effects 0.000 description 3
- 230000007246 mechanism Effects 0.000 description 3
- 230000001681 protective effect Effects 0.000 description 3
- 238000003753 real-time PCR Methods 0.000 description 3
- 230000002829 reductive effect Effects 0.000 description 3
- 230000032258 transport Effects 0.000 description 3
- UFTFJSFQGQCHQW-UHFFFAOYSA-N triformin Chemical compound O=COCC(OC=O)COC=O UFTFJSFQGQCHQW-UHFFFAOYSA-N 0.000 description 3
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 3
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 description 2
- 108091028075 Circular RNA Proteins 0.000 description 2
- 101150042222 DGAT1 gene Proteins 0.000 description 2
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 2
- 208000004930 Fatty Liver Diseases 0.000 description 2
- 206010019708 Hepatic steatosis Diseases 0.000 description 2
- KFZMGEQAYNKOFK-UHFFFAOYSA-N Isopropanol Chemical compound CC(C)O KFZMGEQAYNKOFK-UHFFFAOYSA-N 0.000 description 2
- 241001529936 Murinae Species 0.000 description 2
- 206010028980 Neoplasm Diseases 0.000 description 2
- 102000006382 Ribonucleases Human genes 0.000 description 2
- 108010083644 Ribonucleases Proteins 0.000 description 2
- BCKXLBQYZLBQEK-KVVVOXFISA-M Sodium oleate Chemical compound [Na+].CCCCCCCC\C=C/CCCCCCCC([O-])=O BCKXLBQYZLBQEK-KVVVOXFISA-M 0.000 description 2
- 102000009822 Sterol Regulatory Element Binding Proteins Human genes 0.000 description 2
- 108010020396 Sterol Regulatory Element Binding Proteins Proteins 0.000 description 2
- 239000013599 cloning vector Substances 0.000 description 2
- 238000010586 diagram Methods 0.000 description 2
- 235000005911 diet Nutrition 0.000 description 2
- 230000000378 dietary effect Effects 0.000 description 2
- 230000005750 disease progression Effects 0.000 description 2
- 230000029142 excretion Effects 0.000 description 2
- 208000010706 fatty liver disease Diseases 0.000 description 2
- IPCSVZSSVZVIGE-UHFFFAOYSA-M hexadecanoate Chemical compound CCCCCCCCCCCCCCCC([O-])=O IPCSVZSSVZVIGE-UHFFFAOYSA-M 0.000 description 2
- IPCSVZSSVZVIGE-UHFFFAOYSA-N hexadecanoic acid Chemical compound CCCCCCCCCCCCCCCC(O)=O IPCSVZSSVZVIGE-UHFFFAOYSA-N 0.000 description 2
- 230000037041 intracellular level Effects 0.000 description 2
- 230000037356 lipid metabolism Effects 0.000 description 2
- 239000003550 marker Substances 0.000 description 2
- 239000013610 patient sample Substances 0.000 description 2
- 239000013612 plasmid Substances 0.000 description 2
- 239000013600 plasmid vector Substances 0.000 description 2
- 230000008569 process Effects 0.000 description 2
- 230000002441 reversible effect Effects 0.000 description 2
- 230000035945 sensitivity Effects 0.000 description 2
- 231100000240 steatosis hepatitis Toxicity 0.000 description 2
- 230000000638 stimulation Effects 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- 210000001519 tissue Anatomy 0.000 description 2
- 239000012096 transfection reagent Substances 0.000 description 2
- 238000012546 transfer Methods 0.000 description 2
- 238000013519 translation Methods 0.000 description 2
- 239000013598 vector Substances 0.000 description 2
- WRIDQFICGBMAFQ-UHFFFAOYSA-N (E)-8-Octadecenoic acid Natural products CCCCCCCCCC=CCCCCCCC(O)=O WRIDQFICGBMAFQ-UHFFFAOYSA-N 0.000 description 1
- LQJBNNIYVWPHFW-UHFFFAOYSA-N 20:1omega9c fatty acid Natural products CCCCCCCCCCC=CCCCCCCCC(O)=O LQJBNNIYVWPHFW-UHFFFAOYSA-N 0.000 description 1
- QSBYPNXLFMSGKH-UHFFFAOYSA-N 9-Heptadecensaeure Natural products CCCCCCCC=CCCCCCCCC(O)=O QSBYPNXLFMSGKH-UHFFFAOYSA-N 0.000 description 1
- BSYNRYMUTXBXSQ-UHFFFAOYSA-N Aspirin Chemical compound CC(=O)OC1=CC=CC=C1C(O)=O BSYNRYMUTXBXSQ-UHFFFAOYSA-N 0.000 description 1
- 208000024172 Cardiovascular disease Diseases 0.000 description 1
- 108020004414 DNA Proteins 0.000 description 1
- 102000002148 Diacylglycerol O-acyltransferase Human genes 0.000 description 1
- 108010001348 Diacylglycerol O-acyltransferase Proteins 0.000 description 1
- 206010061818 Disease progression Diseases 0.000 description 1
- 208000032928 Dyslipidaemia Diseases 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 108700039887 Essential Genes Proteins 0.000 description 1
- 102000002754 Glycerol-3-Phosphate O-Acyltransferase Human genes 0.000 description 1
- 108010018837 Glycerol-3-Phosphate O-Acyltransferase Proteins 0.000 description 1
- 108010043121 Green Fluorescent Proteins Proteins 0.000 description 1
- 229940121710 HMGCoA reductase inhibitor Drugs 0.000 description 1
- 101100215371 Homo sapiens ACTB gene Proteins 0.000 description 1
- 238000012404 In vitro experiment Methods 0.000 description 1
- 206010061218 Inflammation Diseases 0.000 description 1
- 208000017170 Lipid metabolism disease Diseases 0.000 description 1
- ZQPPMHVWECSIRJ-UHFFFAOYSA-N Oleic acid Natural products CCCCCCCCC=CCCCCCCCC(O)=O ZQPPMHVWECSIRJ-UHFFFAOYSA-N 0.000 description 1
- 239000005642 Oleic acid Substances 0.000 description 1
- 102000004316 Oxidoreductases Human genes 0.000 description 1
- 108090000854 Oxidoreductases Proteins 0.000 description 1
- 235000021314 Palmitic acid Nutrition 0.000 description 1
- 102000008079 Sterol Regulatory Element Binding Protein 2 Human genes 0.000 description 1
- 108010074438 Sterol Regulatory Element Binding Protein 2 Proteins 0.000 description 1
- 108091036066 Three prime untranslated region Proteins 0.000 description 1
- 229960001138 acetylsalicylic acid Drugs 0.000 description 1
- AVKUERGKIZMTKX-NJBDSQKTSA-N ampicillin Chemical compound C1([C@@H](N)C(=O)N[C@H]2[C@H]3SC([C@@H](N3C2=O)C(O)=O)(C)C)=CC=CC=C1 AVKUERGKIZMTKX-NJBDSQKTSA-N 0.000 description 1
- 229960000723 ampicillin Drugs 0.000 description 1
- 230000003321 amplification Effects 0.000 description 1
- 230000003110 anti-inflammatory effect Effects 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 102000023732 binding proteins Human genes 0.000 description 1
- 108091008324 binding proteins Proteins 0.000 description 1
- 230000033228 biological regulation Effects 0.000 description 1
- 238000003759 clinical diagnosis Methods 0.000 description 1
- 238000010367 cloning Methods 0.000 description 1
- 230000002860 competitive effect Effects 0.000 description 1
- 238000010276 construction Methods 0.000 description 1
- 230000001276 controlling effect Effects 0.000 description 1
- 230000003247 decreasing effect Effects 0.000 description 1
- 230000007850 degeneration Effects 0.000 description 1
- 238000013461 design Methods 0.000 description 1
- 230000009266 disease activity Effects 0.000 description 1
- 208000016097 disease of metabolism Diseases 0.000 description 1
- ZQPPMHVWECSIRJ-MDZDMXLPSA-N elaidic acid Chemical compound CCCCCCCC\C=C\CCCCCCCC(O)=O ZQPPMHVWECSIRJ-MDZDMXLPSA-N 0.000 description 1
- 210000001808 exosome Anatomy 0.000 description 1
- 238000010195 expression analysis Methods 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 230000004761 fibrosis Effects 0.000 description 1
- 238000009472 formulation Methods 0.000 description 1
- 208000006454 hepatitis Diseases 0.000 description 1
- 230000004054 inflammatory process Effects 0.000 description 1
- 230000003993 interaction Effects 0.000 description 1
- 238000011835 investigation Methods 0.000 description 1
- QXJSBBXBKPUZAA-UHFFFAOYSA-N isooleic acid Natural products CCCCCCCC=CCCCCCCCCC(O)=O QXJSBBXBKPUZAA-UHFFFAOYSA-N 0.000 description 1
- 208000018191 liver inflammation Diseases 0.000 description 1
- 230000036210 malignancy Effects 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 208000030159 metabolic disease Diseases 0.000 description 1
- 229960003105 metformin Drugs 0.000 description 1
- XZWYZXLIPXDOLR-UHFFFAOYSA-N metformin Chemical compound CN(C)C(=N)NC(N)=N XZWYZXLIPXDOLR-UHFFFAOYSA-N 0.000 description 1
- -1 methylglutaryl Chemical group 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 230000009456 molecular mechanism Effects 0.000 description 1
- 238000000465 moulding Methods 0.000 description 1
- WQEPLUUGTLDZJY-UHFFFAOYSA-N n-Pentadecanoic acid Natural products CCCCCCCCCCCCCCC(O)=O WQEPLUUGTLDZJY-UHFFFAOYSA-N 0.000 description 1
- 239000002547 new drug Substances 0.000 description 1
- 238000003199 nucleic acid amplification method Methods 0.000 description 1
- 230000008506 pathogenesis Effects 0.000 description 1
- 230000002265 prevention Effects 0.000 description 1
- 230000003449 preventive effect Effects 0.000 description 1
- 230000001737 promoting effect Effects 0.000 description 1
- 230000005180 public health Effects 0.000 description 1
- 230000009467 reduction Effects 0.000 description 1
- 230000028327 secretion Effects 0.000 description 1
- 238000012163 sequencing technique Methods 0.000 description 1
- 229940045870 sodium palmitate Drugs 0.000 description 1
- GGXKEBACDBNFAF-UHFFFAOYSA-M sodium;hexadecanoate Chemical compound [Na+].CCCCCCCCCCCCCCCC([O-])=O GGXKEBACDBNFAF-UHFFFAOYSA-M 0.000 description 1
- 238000001228 spectrum Methods 0.000 description 1
- 238000007619 statistical method Methods 0.000 description 1
- 239000011550 stock solution Substances 0.000 description 1
- 239000013589 supplement Substances 0.000 description 1
- 230000009885 systemic effect Effects 0.000 description 1
- 230000007723 transport mechanism Effects 0.000 description 1
- 150000003626 triacylglycerols Chemical class 0.000 description 1
- 230000003827 upregulation Effects 0.000 description 1
- 238000005303 weighing Methods 0.000 description 1
- 230000004580 weight loss Effects 0.000 description 1
- 238000001262 western blot Methods 0.000 description 1
- 238000012049 whole transcriptome sequencing Methods 0.000 description 1
Landscapes
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
本发明涉及非酒精性脂肪性肝病诊断治疗方法以及药物技术领域,具体涉及circPI4KB在诊断和治疗非酒精性脂肪性肝病以及相关药物筛选中的应用。本技术方案提示了circPI4KB与miRNA‑122的表达水平以及其从肝细胞内到外的转运相关,且其还与下游脂质沉积相关蛋白的表达相关。上述研究结果提示了,该分子是一种潜在的非酒精性脂肪性肝病相关药物的作用靶点以及药物筛选靶点,以及一种潜在的诊断非酒精性脂肪性肝病生物标志物。本技术方案解决了现有技术缺乏专门针对非酒精性脂肪性肝病的药物以及生物标志物的技术问题。将本方案应用在非酒精性脂肪性肝病的药物制备、疾病诊断以及药物筛选中,具有极大的应用价值和市场前景。
Description
技术领域
本发明涉及非酒精性脂肪性肝病诊断治疗方法以及药物技术领域,具体涉及circPI4KB在诊断和治疗非酒精性脂肪性肝病以及相关药物筛选中的应用。
背景技术
随着人民生活方式和饮食结构的改变,非酒精性脂肪性肝病(Nonalcoholicfatty liver disease,NAFLD)发病率逐渐增高,全球约25%的人群患有NAFLD。NAFLD的全球发病率高达25%,在中国发病率为29.2%,已成为全球和中国排名第1位的慢性肝病和重要的公共卫生问题,给患者和社会带来巨大的疾病负担和经济影响。NAFLD疾病谱包括非酒精性单纯性脂肪肝(NAFL)和非酒精性脂肪性肝炎(NASH)、肝硬化和肝细胞癌。在NASH发生后,可出现肝脏炎症、纤维化,导致疾病活动、进展,甚至发生肝硬化、肝细胞癌(Hepatocellular carcinoma,HCC)。根据模型研究预测,美国、欧洲和中国与NAFLD相关的HCC的发病率和患病率将在2016年至2030年期间迅速增加至56%。
一旦患者进展到NASH阶段后,NASH相关疾病进展风险加快,发生肝硬化、心血管疾病、HCC和其他系统恶性肿瘤的几率均明显增高。但是,针对NAFLD和NASH人群进展成HCC尚未有行之有效的预防措施。目前,除了通过服用他汀类药物、二甲双胍、阿司匹林等控制血脂血糖异常和预防性的抗炎治疗,或者通过减体重治疗外,临床上有效治疗NASH的药物及方案非常有限,急需研发有效的NASH的防治措施。可以预见,非酒精性脂肪性肝病将会成为今后我国慢性肝病的主要社会经济负担,加强对NASH的基础与临床研究,寻找NASH的治疗新手段与新靶点,具有重大的社会价值和临床实际需求,亟待进一步的研究和探索。
发明内容
本发明意在提供circPI4KB在制备治疗非酒精性脂肪性肝病的药物中的应用,以解决现有技术缺乏专门针对非酒精性脂肪性肝病的药物的技术问题。
为达到上述目的,本发明采用如下技术方案:
circPI4KB在制备治疗非酒精性脂肪性肝病的药物中的应用。
本方案还提供了circPI4KB在制备诊断非酒精性脂肪性肝病的系统中的应用。
本方案还提供了circPI4KB筛选治疗非酒精性脂肪性肝病药物中的应用。
本技术方案的原理以及有益效果在于:
circRNA是一类共价闭合环状RNA,在炎症和肿瘤的发生发展中起重要作用。近年来Nature和Science等国际顶级期刊相继报道,circRNA可作为细胞内的竞争性内源性RNA,通过结合并吸附miRNA,从而阻止miRNA在3′非翻译区与mRNA相互作用,进而干扰miRNA对靶基因的调控作用。也有报道称circRNA可结合miRNA并增加其稳定性,还可实现对miRNA的转运功能,解锁了circRNA研究的新功能和新领域。针对NAFLD关键影响因子miRNA-122,其从肝细胞内到外的转运机制,以及如何维持其在NAFLD患者血清中的稳定性,现有技术中尚未见报道。发明人通过对NAFLD细胞模型进行研究,筛选出了若干表达量产生显著变化的候选circRNA。并发现其中的circPI4KB与miRNA-122的表达以及由细胞内到外的转运相关。并且,circPI4KB表达上调后,miRNA-122的下游5个脂质沉积相关蛋白及其mRNA的表达量上调;而抑制circPI4KB表达后,5个脂质沉积相关蛋白及其mRNA的表达量下调。进一步说明了circPI4KB可以作为治疗NAFLD的药物作用靶点,应用在制备治疗非酒精性脂肪性肝病的药物的实践操作中。除此之外,circPI4KB作为药物作用靶点,还可以针对该靶点进行药物筛选,以找到对circPI4KB形成抑制的适合于治疗NAFLD的新药。
除此之外,发明人还对45例和45例正常人和NAFLD病人的血清样本进行了测试,发现正常人和病人血清中,circPI4KB表达量存在差异,其可以作为诊断NAFLD的生物标志物应用在临床诊断的时间操作中。通过对样本的研究,验证了circPI4KB对NAFLD的诊断能力,结果表明其诊断效能曲线下面积为0.925(0.878-0.972),p<0.001,诊断效力理想。
综上所述,非酒精性脂肪性肝病(NAFLD)目前已经成为我国第一位的慢性肝病。随着人民生活方式和饮食结构的改变,NAFLD的发病率逐渐升高,据文献报道我国2018年预估的脂肪肝发病率高达32.95%,其即将成为今后我国慢性肝病的主要社会经济负担。加强对NAFLD的基础与临床研究,寻找治疗的新手段与新靶点,具有重大的社会价值和临床实际需求。本研究体外实验调节circPI4KB导致对肝内脂质沉积的抑制作用减弱,促使NAFLD的发生。本研究为NAFLD发生的分子机制提供新的线索,并为NAFLD的诊断和治疗提供新的靶点。
进一步,circPI4KB在制备治疗非酒精性脂肪性肝病的药物中的应用,包括抑制circPI4KB表达水平的步骤。通过抑制circPI4KB表达水平来实现对非酒精性脂肪性肝病的治疗。
进一步,使用shRNA抑制circPI4KB表达。shRNA是一种能够有效抑制环状RNA表达水平的手段,且针对于shRNA的设计和操作方法相对成熟,容易操作和掌握。
进一步,所述shRNA的DNA序列为:5’-AGAATGAGGATGAGCTTGGAA-3’。采用上述序列的shRNA可以对circPI4KB的表达形成有效抑制,进而减少miRNA-122从肝细胞内到外的转运,延缓肝脏脂质沉积以及NAFLD的发生发展。
进一步,circPI4KB在制备诊断非酒精性脂肪性肝病的试剂中的应用,circPI4KB的接收号为hsa_circ_0006982;诊断非酒精性脂肪性肝病的系统用于检测circPI4KB在血清、肝细胞或者肝组织中的含量。通过检测circPI4KB在血清、肝细胞或者肝组织中的含量,可以较为准确地反应肝内脂质沉积以及NAFLD的发展趋势的大致情况,对诊断非酒精性脂肪性肝病起到有效的指示作用。
进一步,诊断非酒精性脂肪性肝病的系统包括qRT-PCR试剂。
进一步,所述qRT-PCR试剂包括引物F和引物R;引物F的序列为:3’-CAGCCAGCAACCCTAAAGTG-5’;引物R的序列为:5’-ACTGTATCTCCCATGGCCAC-3’。
进一步,circPI4KB在血清中的诊断参考值为28.16。通过RT-qRCR检测以及2-ΔΔCt法计算,可以获得circPI4KB在血清中的相对拷贝数。经过统计分析发现,circPI4KB在血清中的诊断参考值为28.16。小于该值,可以初步判断患者具有一定的可能性患有非酒精性脂肪性肝病,提示需要采用进一步的诊断手段,对患者是否患有非酒精性脂肪性肝病进行进一步判断。
附图说明
图1为实施例1的miRNA-122在NAFLD的发生过程中的作用机制示意图。
图2为实施例1的NAFLD和健康对照相比circRNA表达量变化情况火山图。
图3为实施例1的NASH和健康对照相比circRNA表达量变化情况火山图。
图4为实施例2的NASH和NAFLD相比circRNA表达量变化情况火山图。
图5为实施例1的qRT-PCR检测NAFLD细胞模型中差异表达的circRNAs(图中数据为三次重复实验的实验结果,图中左侧箭头表示具有显著表达差异的circRNAs,其中,circPI4KB的差异表达水平为0.58±0.11fold change;*p<0.05、**p<0.01)。
图6为实施例1的正常细胞和NAFLD细胞模型中circPI4KB在细胞内外表达水平统计图(图中数据为三次重复实验的实验结果;circPI4KB在细胞内的水平为:0.51±0.06fold change;ircPI4KB在上清液中的水平为:2.27±0.28fold change;*p<0.05、**p<0.01)。
图7为实施例2的ROC曲线。
图8为实施例3的qRT-PCR检测调节circPI4KB水平后miRNA-122在肝细胞内外水平变化(图中数据为三次重复实验的实验结果;miRNA-122在细胞内的水平为:0.41±0.23fold change;miRNA-122在上清液的水平为:3.78±0.78fold change;*p<0.05、**p<0.01、***p<0.001;ns:不存在显著差异)。
图9为实施例3的Western blot检测miRNA-122的下游五个脂质代谢相关蛋白水平变化图片。
图10为实施例3的qRT-PCR检测miRNA-122的下游五个脂质代谢相关mRNA水平变化统计图(图中数据为三次重复实验的实验结果;针对FFA+circPI4KB组,HMGCR为3.53±0.98;FAS,3.12±0.54;SREBP2,2.78±0.54;Agpat1,2.35±0.34;Dgat1,2.14±1.11;*p<0.05、**p<0.01;ns:不存在显著差异)。
具体实施方式
下面结合实施例对本发明做进一步详细的说明,但本发明的实施方式不限于此。若未特别指明,下述实施例以及实验例所用的技术手段为本领域技术人员所熟知的常规手段,且所用的材料、试剂等,均可从商业途径得到。
实施例1:
NAFLD是一种由于甘油三酯和胆固醇等脂质过度沉积而诱发肝细胞变性的代谢性疾病。miRNA-122占肝脏miRNA总量的70%,被定义为肝脏特异性miRNA。miRNA-122可以通过两种途径抑制肝脏脂质沉积,分别是抑制胆固醇的合成和抑制甘油三酯的合成,对肝脏起保护性作用。其一,miRNA-122通过转录后抑制胆固醇调节元件结合蛋白(SREBP)、脂肪酸合成酶(FAS)和甲基戊二酰辅酶A还原酶(HMGCR)的mRNA翻译而减少胆固醇合成;其二,miRNA-122还可通过转录后抑制甘油磷酸酰基转移酶1(Agpat1)和二酰甘油酰基转移酶1(Dgat1)的mRNA翻译而减少甘油三酯合成,从而减少脂质沉积。综上所述,miRNA-122可通过转录后抑制胆固醇和甘油三酯合成的相关mRNA,减少肝脏脂质沉积,在NAFLD的发生中起保护性作用(图1)。miRNA-122如何实现从肝细胞内到外的转运,如何维持其在NAFLD患者血清中的稳定性,进一步深入研究miRNA-122“逃逸”有助于阐明NAFLD发生的机制。
发明人对miRNA-122“逃逸”机制进行了深入研究。其中,通过对健康对照、NAFLD患者和NASH患者肝脏组织的进行全转录组测序(包含circRNA,miRNA和mRNA等),利用差异表达分析共筛选到34个差异表达circRNAs,参见图2-4(circRNA测序火山图)。图2为NAFLD和健康对照相比circRNA表达量变化情况;图3为NASH和健康对照相比circRNA表达量变化情况;图4为NASH和NAFLD相比circRNA表达量变化情况。
发明人对在NAFLD或NASH中有差异表达的若干circRNAs进行了进一步研究。发明人建立了NAFLD细胞模型,然后使用qRT-PCR对这些差异表达的circRNAs进行筛选以及验证。实验结果详见图5,circC7orf44、circSPECC1-2、circRBBP8、circPI4KB和circAFF1-1等circRNAs出现了差异表达的情况。NAFLD细胞模型由如下方式建立:人源肝细胞L02以5-10×105/mL接种,使用含有游离脂肪酸刺激(FFA)的培养基培养人源肝细胞L02 24_h,获得NAFLD细胞模型。含有游离脂肪酸刺激(FFA)的培养基的配方具体参见实施例3。
利用NAFLD细胞模型观察到circRNA-PI4KB(circPI4KB)(hsa_circ_0006982)在肝脏细胞/肝内含量下降,细胞上清/血清中含量上升。即,在NAFLD细胞模型中,circPI4KB在细胞内水平下降,在细胞外水平上升。circPI4KB降低和逃逸出肝细胞是潜在的NAFLD发病的原因,而其分泌增多也是其作为血清标物的潜在靶点,实验结果参见图6。其他前述检测到的在NAFLD细胞模型和正常细胞差异表达的circRNAs,并未呈现上述现象,说明circPI4KB相对于其他circRNAs具有特殊功能。
实施例2:利用circPI4KB作为诊断标志物的效力研究
搜集45例和45例正常人和NAFLD病人的血清样本,患者情况参见表1。
表1:患者情况统计
除了对血清、细胞和组织中的总RNA的提取的具体流程如下:
(1)收集6孔板内细胞或称量50μg研磨后的肝组织或者取200μL血清于无RNA酶的EP管中,加入TRIzol 500μL,振荡混匀,室温孵育5分钟;
(2)加入氯仿100μL,振荡混匀15秒,低温离心机离心15分钟,转速12000×g,此时EP管中的液体分三层;
(3)吸取最上层清亮液体于另一干净的EP管中,加入250μL异丙醇,振荡混匀,室温孵育10分钟,低温离心机离心10分钟,转速12000×g;
(4)轻轻吸弃上清,加入75%乙醇(DEPC水配)500mL,轻微颠倒混匀,低温离心机离心5分钟,转速7500×g;
(5)吸弃上清,室温干燥10-15分钟,每管加入提前60℃预热的DEPC水30-50μL,移液器吹打混匀,保存于-80℃冰箱。
通过qRT-PCR的方法检测血清样本中circPI4KB的表达量。本研究采用SYBR Green荧光定量PCR方法检测目的基因表达水平。对mRNA逆转录后的cDNA进行检测,使用人源管家基因ACTB或者鼠源GAPDH作为内参对照;采用2-ΔΔCt计算组织/细胞目的基因mRNA的相对拷贝数(以人源ACTB或者鼠源GAPDH为参照)。利用Roche罗氏LC96定量PCR仪LightCycler96System对其拷贝值进行测定。
计算公式为:ΔΔCt=ΔCtexp-ΔCtcon=(Ctexp-target-Ctexp-ACTB/GAPDH)-(Ctcon-target-Ctcon-ACTB/GAPDH);
其中,Ctexp-target:NAFLD病人样本的circPI4KB扩增的Ct值;
Ctcon-target:正常样本的circPI4KB扩增的Ct值;
Ctexp-ACTB/GAPDH:NAFLD病人样本的内参扩增的Ct值;
Ctcon-ACTB/GAPDH:正常样本的内参扩增的Ct值。
采用2-ΔCt计算细胞/外泌体/血清中目的miRNA的相对拷贝数(以U6为参照,针对miRNA的一个reverse引物。)。计算公式为:ΔΔCt=ΔCtexp-ΔCtcon=(Ctexp-target-Ctexp-U6)-(Ctcon-target-Ctcon-U6)。其中,circPI4KB的检测是和普通mRNA检测方法一致。
其中,circPI4KB的引物序列为:
3’-CAGCCAGCAACCCTAAAGTG-5’(即:5’-GTGAAATCCCAACGACCGAC-3’,SEQ IDNO.1);
5’-ACTGTATCTCCCATGGCCAC-3’(SEQ ID NO.2)。
ACTB的引物序列为:
3’-CTCCATCCTGGCCTCGCTGT-5’(即:5’-TGTCGCTCCGGTCCTACCTC-3’,SEQ IDNO.3);
5’-GCTGTCACCTTCACCGTTCC-3’(SEQ ID NO.4)。
反应体系以及反应条件参见表2、表3、表4和表5所示。
表2:逆转录反应体系(20μL)(TAKARA RR047A逆转录试剂盒)
试剂 | 用量 |
5×PrimeScript Buffer(for Real Time) | 4μL |
PrimeScript RT Enzyme Mix | 1μL |
Random 6mers(100μM) | 1μL |
Total RNA | 100ng-1μg |
RNase Free dH2O | 补齐至20μL |
Total | 20μL |
表3:逆转录反应条件
温度 | 时间 |
37℃ | 15min |
85℃ | 5s |
4℃ | 1min |
表4:qPCR反应体系(20μL)(Premix Ex TaqTM II(Tli RNaseH Plus),RR820Q/A/B,TAKARA RR047A逆转录试剂盒配套的qPCR试剂盒)
2×concentrated master mix | 10μL |
Former primer(浓度10μM) | 0.5μL |
Reverse primer(浓度10μM) | 0.5μL |
cDNA | 2μL |
dH2O | 7μL |
Total | 20μL |
表5:qPCR反应条件
根据检测结果,45例正常人的检测结果ΔΔCt为:29.99±4.34,45例NAFLD病人ΔΔCt为:27.34±2.13绘制ROC曲线,以判定circPI4KB作为诊断标志物的效力。对上述样本检测的数据进行youden约登指数的最佳指数,诊断临界值(截断值,cut-off)为28.16,用来反映单个指标(circPI4KB含量)对整体的诊断和预测效果。采用该生物标志物的相对拷贝数作为诊断变量,计算在每个诊断变量具体取值下的敏感性和特异性,进而绘制出ROC曲线。ROC曲线是以假阳性率(False positive rate,1-特异性)为横轴,真阳性率(Truepositive rate,敏感性)为纵轴所组成的坐标图,测试样本在不同的判断标准(阈值)得出的不同结果画出的曲线。利用Roche罗氏LC96定量PCR仪LightCycler 96System对血清circPI4KB拷贝值进行测定。通过对上述样本的研究,验证了circPI4KB对NAFLD的诊断能力,结果表明其诊断效能曲线下面积为0.925(0.878-0.972),p<0.001,ROC曲线详见图7。
实施例3:circPI4KB的过表达以及抑制表达的研究
构建circPI4KB的过表达以及抑制表达的细胞模型,具体过程如下:
(1)表达载体构建
将circPI4KB的DNA序列通过本领域常规手段整合到克隆载体pcDNA3.1(+)(吉玛基因;克隆载体pcDNA3.1(+);克隆位点:BamHI/EcoRI;质粒抗性:Ampicillin)上,获得circPI4KB过表达载体。将shRNA的DNA通过本领域常规手段整合到空载体pGPH1/GFP/Neo(现有技术可通过商业途径获取的用于shRNA真核表达载体)上,获得shRNA表达载体。
其中,circPI4KB(circPI4KB)(hsa_circ_0006982)的cDNA全长序列(937bp)如SEQID NO.5所示(5’→3’):
CTTGGAAGCTCGAAGTCTGGCTGTGGCCATGGGAGATACAGTAGTGGAGCCTGCCCCCTTGAAGCCAACTTCTGAGCCCACTTCTGGCCCACCAGGGAATAATGGGGGGTCCCTGCTAAGTGTCATCACGGAGGGGGTCGGGGAACTATCAGTGATTGACCCTGAGGTGGCCCAGAAGGCCTGCCAGGAGGTGTTGGAGAAAGTCAAGCTTTTGCATGGAGGCGTGGCAGTCTCTAGCAGAGGCACCCCACTGGAGTTGGTCAATGGGGATGGTGTGGACAGTGAGATCCGTTGCCTAGATGATCCACCTGCCCAGATCAGGGAGGAGGAAGATGAGATGGGGGCCGCTGTGGCCTCAGGCACAGCCAAAGGAGCAAGAAGACGGCGGCAGAACAACTCAGCTAAACAGTCTTGGCTGCTGAGGCTGTTTGAGTCAAAACTGTTTGACATCTCCATGGCCATTTCATACCTGTATAACTCCAAGGAGCCTGGAGTACAAGCCTACATTGGCAACCGGCTCTTCTGCTTTCGCAACGAGGACGTGGACTTCTATCTGCCCCAGTTGCTTAACATGTACATCCACATGGATGAGGATGTGGGTGATGCCATTAAGCCCTACATAGTCCACCGTTGCCGCCAGAGCATTAACTTTTCCCTCCAGTGTGCCCTGTTGCTTGGGGCCTATTCTTCAGACATGCACATTTCCACTCAACGACACTCCCGTGGGACCAAGCTACGGAAGCTGATCCTCTCAGATGAGCTAAAGCCAGCTCACAGGAAGAGGGAGCTGCCCTCCTTGAGCCCGGCCCCTGACACAGGGCTGTCTCCCTCCAAAAGGACTCACCAGCGCTCTAAGTCAGATGCCACTGCCAGCATAAGTCTCAGCAGCAACCTGAAACGAACA GCCAGCAACCCTAAAGTGGAGAATGAGGATGAG(SEQ ID NO.5,下划线为qRT-PCR引物结合位点);
shRNA序列为:5’-AGAATGAGGATGAGCTTGGAA-3’(SEQ ID NO.6)。
(2)质粒转染
以5-10×105/mL的人源肝细胞L02(包括正常细胞以及游离脂肪酸刺激(FFA)刺激的NAFLD模型细胞)密度接种于6孔板,显微镜下观察细胞形态,待细胞贴壁生长至50%-60%。其中,正常细胞使用的培养基为Opti-MEM。游离脂肪酸刺激(FFA)刺激的NAFLD模型细胞使用的培养基由如下方式配制:1)配制FFA原液:油酸钠(sodium oleate,OA)0.0609g溶于10ml质量分数为25%的BSA(20mM),棕榈酸钠(sodium palmitate,PA)0.0557g溶于20ml质量分数为25%的BSA(10mM),55℃水浴溶解1h,震荡,混匀,超声10分钟(30s×20次),37h水浴过夜。2)细胞1M FFA培养基:将0.333ml(OA)和0.333ml(PA)溶于10mL培养基。
然后,将上一步构建的质粒载体、Opti-MEM培养基、脂质体LipofectamineTM3000,严格按照转染试剂说明书配比成所需转染液(赛默飞,Invitrogen Lipofectamine 3000转染试剂)。将上一段所述的培养基换成转染液,在转染液中,质粒载体的浓度为1μg/ml,LipofectamineTM3000的浓度为125μl/ml。上述混合液体轻轻混匀后置37℃孵育10-20分钟,每孔加入250μL的转染液并充分混匀,置于恒温培养箱培养6小时以后,更换正常普通培养基(Opti-MEM)。
(3)检测
在转染之后24h,对FFA组换取棕榈酸+油酸(1:1,即FFA培养基)继续造模24小时,正常组更换正常的普通培养基。对空白对照、circPI4KB过表达组、sh-circPI4KB组的细胞(liver)中以及培养基(serum)中的miRNA-122的表达水平进行了qRT-PCR检测,实验结果参见图8。可见在正常细胞中,circPI4KB过表达会导致大量miRNA-122外排至细胞外;circPI4KB被抑制后,miRNA-122会在细胞内一定程度地累积,并且外排至胞外的量大大减少。所以,对于正常细胞而言,circPI4KB表达上调会促进miRNA-122从胞内排出至胞外。而miRNA-122从肝细胞内到外的转运,导致细胞内miRNA-122含量减少而保护性作用降低,会促进肝脏脂质沉积以及NAFLD的发生发展。而在NAFLD模型细胞(FFA)中,细胞中的miRNA-122含量进一步减小,circPI4KB过表达组中细胞中的miRNA-122含量最少,其次是空白对照和sh-circPI4KB组。而在NAFLD模型细胞(FFA)中,培养基中,miRNA-122含量进一步增加,circPI4KB过表达组中培养基中的miRNA-122含量最高,其次是空白对照和sh-circPI4KB组。上述实验结果说明,使用sh-circPI4KB可以通过抑制circPI4KB的表达,来抑制miRNA-122的表达以及外排,可以作为治疗由于miRNA-122引起的NAFLD的潜在手段。
发明人进而对miRNA-122的下游5个脂质沉积相关蛋白及其mRNA(SREBP、FAS、HMGCR、Agpat1和Dgat1)的水平变化进行了检测。实验结果参见图9和图10。上述实验结果明确了circPI4KB的水平变化可影响肝细胞内脂质沉积。
以上所述的仅是本发明的实施例,方案中公知的具体技术方案和/或特性等常识在此未作过多描述。应当指出,对于本领域的技术人员来说,在不脱离本发明技术方案的前提下,还可以作出若干变形和改进,这些也应该视为本发明的保护范围,这些都不会影响本发明实施的效果和专利的实用性。本申请要求的保护范围应当以其权利要求的内容为准,说明书中的具体实施方式等记载可以用于解释权利要求的内容。
/>
/>
Claims (3)
1.抑制circPI4KB的shRNA在制备治疗非酒精性脂肪性肝病的药物中的应用,其特征在于:所述shRNA的DNA序列为:5’-AGAATGAGGATGAGCTTGGAA-3’。
2.检测circPI4KB在血清、肝细胞或者肝组织中的含量的系统在制备诊断非酒精性脂肪性肝病的系统中的应用,其特征在于:诊断非酒精性脂肪性肝病的系统包括qRT-PCR试剂;所述qRT-PCR试剂包括引物F和引物R;引物F的序列为:3’-CAGCCAGCAACCCTAAAGTG-5’;引物R的序列为:5’-ACTGTATCTCCCATGGCCAC-3’;circPI4KB在血清中的诊断参考值为28.16。
3.根据权利要求2所述的circPI4KB在制备诊断非酒精性脂肪性肝病的系统中的应用,其特征在于,circPI4KB的接收号为hsa_circ_0006982。
Priority Applications (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN202210837047.8A CN115814086B (zh) | 2022-07-15 | 2022-07-15 | circPI4KB在诊断和治疗非酒精性脂肪性肝病以及相关药物筛选中的应用 |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN202210837047.8A CN115814086B (zh) | 2022-07-15 | 2022-07-15 | circPI4KB在诊断和治疗非酒精性脂肪性肝病以及相关药物筛选中的应用 |
Publications (2)
Publication Number | Publication Date |
---|---|
CN115814086A CN115814086A (zh) | 2023-03-21 |
CN115814086B true CN115814086B (zh) | 2024-05-14 |
Family
ID=85522832
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN202210837047.8A Active CN115814086B (zh) | 2022-07-15 | 2022-07-15 | circPI4KB在诊断和治疗非酒精性脂肪性肝病以及相关药物筛选中的应用 |
Country Status (1)
Country | Link |
---|---|
CN (1) | CN115814086B (zh) |
Citations (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2009028457A1 (ja) * | 2007-08-29 | 2009-03-05 | Shinshu University | 非アルコール性脂肪肝炎治療薬 |
KR102328770B1 (ko) * | 2020-06-01 | 2021-11-19 | 고려대학교 산학협력단 | 비알코올 지방간염 진단용 바이오마커 miRNA-4449 |
WO2022025387A1 (ko) * | 2020-07-28 | 2022-02-03 | 고려대학교 산학협력단 | Microrna조합을 이용한 비알코올 지방간염 진단용 바이오마커 |
Family Cites Families (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2020146639A2 (en) * | 2019-01-11 | 2020-07-16 | Viscient Biosciences, Inc. | Compositions and methods for the diagnosis and treatment of diseases of the liver |
-
2022
- 2022-07-15 CN CN202210837047.8A patent/CN115814086B/zh active Active
Patent Citations (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2009028457A1 (ja) * | 2007-08-29 | 2009-03-05 | Shinshu University | 非アルコール性脂肪肝炎治療薬 |
KR102328770B1 (ko) * | 2020-06-01 | 2021-11-19 | 고려대학교 산학협력단 | 비알코올 지방간염 진단용 바이오마커 miRNA-4449 |
WO2022025387A1 (ko) * | 2020-07-28 | 2022-02-03 | 고려대학교 산학협력단 | Microrna조합을 이용한 비알코올 지방간염 진단용 바이오마커 |
Non-Patent Citations (1)
Title |
---|
微小RNA在非酒精性脂肪肝病中调控作用的研究进展;杨燕;童南伟;;重庆医科大学学报;20191127;44(12);第1537-1541页 * |
Also Published As
Publication number | Publication date |
---|---|
CN115814086A (zh) | 2023-03-21 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
Jiang et al. | Hepatocyte-derived extracellular vesicles promote endothelial inflammation and atherogenesis via microRNA-1 | |
Yuwen et al. | MiR-146a-5p level in serum exosomes predicts therapeutic effect of cisplatin in non-small cell lung cancer. | |
Wu et al. | Hepatic exosome-derived miR-130a-3p attenuates glucose intolerance via suppressing PHLPP2 gene in adipocyte | |
Fang et al. | MicroRNA‐29b suppresses tumor angiogenesis, invasion, and metastasis by regulating matrix metalloproteinase 2 expression | |
Chen et al. | Long non-coding RNA FOXD2-AS1 aggravates nasopharyngeal carcinoma carcinogenesis by modulating miR-363-5p/S100A1 pathway | |
Li et al. | RETRACTED ARTICLE: The lncRNA RHPN1-AS1 downregulation promotes gefitinib resistance by targeting miR-299-3p/TNFSF12 pathway in NSCLC | |
Ye et al. | LncRNA LINC00460 promotes tumor growth of human lung adenocarcinoma by targeting miR-302c-5p/FOXA1 axis | |
Zhang et al. | Circular RNA Circ_0000442 acts as a sponge of MiR-148b-3p to suppress breast cancer via PTEN/PI3K/Akt signaling pathway | |
Song et al. | Long non-coding RNA 319 facilitates nasopharyngeal carcinoma carcinogenesis through regulation of miR-1207-5p/KLF12 axis | |
Sun et al. | Long noncoding RNA SNHG12 facilitates the tumorigenesis of glioma through miR-101-3p/FOXP1 axis | |
Mu et al. | Long noncoding RNA TMPO-AS1 promotes lung adenocarcinoma progression and is negatively regulated by miR-383-5p | |
CN113476618B (zh) | miR-199a-3p在制备治疗鼻咽癌药物中的应用 | |
Xie et al. | Long non-coding RNA 520 is a negative prognostic biomarker and exhibits pro-oncogenic function in nasopharyngeal carcinoma carcinogenesis through regulation of miR-26b-3p/USP39 axis | |
Hu et al. | MicroRNA-21 promotes cell proliferation in human hepatocellular carcinoma partly by targeting HEPN1 | |
Ma et al. | Long non-coding RNA ANRIL promotes chemoresistance in triple-negative breast cancer via enhancing aerobic glycolysis | |
Guo et al. | Downregulation of TNFRSF19 and RAB43 by a novel miRNA, miR-HCC3, promotes proliferation and epithelial–mesenchymal transition in hepatocellular carcinoma cells | |
Lu et al. | Circ_0078710 promotes the development of liver cancer by upregulating TXNDC5 via miR-431-5p | |
TianHao | miR-5100 mediates migration and invasion of melanomatous cells in vitro via targeting SPINK5 | |
CN115814086B (zh) | circPI4KB在诊断和治疗非酒精性脂肪性肝病以及相关药物筛选中的应用 | |
Li et al. | Circular ribonucleic acid nei‐like deoxyribonucleic acid glycosylase 3 governs the microribonucleic acid‐3150b‐3p/laminin subunit gamma 1 network to partially promote the development of hepatocellular carcinoma | |
Wang et al. | MicroRNA-124 expression in Kupffer cells modulates liver injury by targeting IL-6/STAT3 signaling | |
Zheng et al. | Circ_0003159 upregulates LIFR expression through competitively binding to miR-221-3p/miR-222-3p to block gastric cancer development | |
CN114480647A (zh) | 膀胱癌检测试剂盒、核酸检测芯片、信号通道抑制剂及其应用 | |
CN110819718B (zh) | miR-154-5p在前列腺癌骨转移的新应用 | |
Lv et al. | Circ_0003907 modulates sepsis-induced myocardial injury via enhancing MYD88/NLRP3/NF-κB axis by sponging miR-944 |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
PB01 | Publication | ||
PB01 | Publication | ||
SE01 | Entry into force of request for substantive examination | ||
SE01 | Entry into force of request for substantive examination | ||
CB03 | Change of inventor or designer information |
Inventor after: Liu Changhai Inventor after: Zeng Qingmin Inventor after: Wu Dongbo Inventor after: Jiang Wei Inventor after: Tang Hong Inventor before: Liu Changhai |
|
CB03 | Change of inventor or designer information | ||
GR01 | Patent grant | ||
GR01 | Patent grant |