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基于芯片筛选肝内胆管细胞癌差异表达的环状RNA_000585并探讨其潜在的作用机制

易烽明 辛龙祥 冯龙

王传敏, 孟忠吉, 陈悦, 刘莉. 血清降钙素原检测对肝衰竭合并感染的早期诊断价值[J]. 临床肝胆病杂志, 2017, 33(6): 1137-1140. DOI: 10.3969/j.issn.1001-5256.2017.06.024.
引用本文: 王传敏, 孟忠吉, 陈悦, 刘莉. 血清降钙素原检测对肝衰竭合并感染的早期诊断价值[J]. 临床肝胆病杂志, 2017, 33(6): 1137-1140. DOI: 10.3969/j.issn.1001-5256.2017.06.024.
Wang ChuanMin, Meng ZhongJi, Chen Yue, Liu Li. Value of serum PCT in early diagnosis of bacterial infection in patients with liver failure[J]. J Clin Hepatol, 2017, 33(6): 1137-1140. DOI: 10.3969/j.issn.1001-5256.2017.06.024.
Citation: Wang ChuanMin, Meng ZhongJi, Chen Yue, Liu Li. Value of serum PCT in early diagnosis of bacterial infection in patients with liver failure[J]. J Clin Hepatol, 2017, 33(6): 1137-1140. DOI: 10.3969/j.issn.1001-5256.2017.06.024.

基于芯片筛选肝内胆管细胞癌差异表达的环状RNA_000585并探讨其潜在的作用机制

DOI: 10.3969/j.issn.1001-5256.2021.10.017
基金项目: 

江西省科技厅重点研发计划——一般项目 (20203BBGL73144)

详细信息
    通信作者:

    冯龙,longfengefy@163.com

  • 中图分类号: R735.7

Screening of differentially expressed circular RNAs in intrahepatic cholangiocarcinoma based on microarray technique and potential mechanism of circRNA_000585

Research funding: 

Jiangxi Provincial Department of Science and Technology (20203BBGL73144)

  • 摘要:   目的  通过对肝内胆管细胞癌(iCCA)和配对的癌旁组织的circRNA芯片比较分析,发现并验证iCCA中异常表达的circRNA,进而揭示其潜在的机制。  方法  收集2019年7月—12月收治的3例iCCA患者肿瘤组织标本,采用微阵列芯片杂交的方法检测iCCA和癌旁组织中circRNA的差异表达,并收集同一时期15例iCCA患者样本,采用实时定量PCR(RT-PCR)验证差异表达的circRNA。通过生物信息学分析寻找差异表达的circRNA的下游分子,并进一步采用RT-PCR对潜在分子进行验证。计量资料组间比较采用配对样本t检验;计数资料组间比较采用χ2检验或Fisher精确检验。  结果  采用1.5倍作为差异表达界值,结果发现iCCA相比癌旁组织中117个circRNAs出现上调,104个circRNAs出现下调。采用3倍作为界值,发现iCCA中10个circRNAs出现上调(circRNA_002172、circRNA_002144、circRNA_001588、circRNA_000166、circRNA_000585、circRNA_000167、circRNA_402608、circRNA_006853、circRNA_001589、circRNA_008882),3个circRNAs出现下调(circRNA_406083、circRNA_104940、circRNA_006349)。收集15例病理证实的iCCA及癌旁组织,采用RT-PCR进行验证差异表达的circRNA,结果发现circRNA_000585在iCCA中表达显著升高(t=3.607,P=0.003)。进一步通过生物信息学分析发现circRNA_000585/miR-615-5p/AMOT/YAP可能是iCCA发生的潜在通路,RT-PCR验证发现该通路中miR-615-5p在iCCA中显著下调(t=5.724,P<0.001),而AMOT和YAP分子在iCCA中显著上调(t值分别为2.664、2.986,P值分别为0.019、0.009 8)。  结论  多种circRNAs在iCCA中出现异常表达,其中circRNA_000585在iCCA中显著上调,其可能通过circRNA_000585/miR-615-5p/AMOT/YAP信号通路在iCCA的发生中发挥重要作用。

     

  • 图  1  iCCA与癌旁组织circRNAs表达的热图

    注:a,散点图;b,火山图。

    图  2  circRNAs表达的散点图和火山图

    图  3  与癌旁组织相比iCCA中上调的circRNAs

    图  4  与癌旁相比iCCA中下调的circRNAs

    图  5  iCCA与癌旁中circRNA_000585的表达情况

    图  6  iCCA与癌旁组织中mir-615-5p的表达情况

    图  7  iCCA与癌旁组织中AMOT的表达情况

    图  8  iCCA与癌旁组织中YAP的表达情况

    表  1  circRNA_000585和内参PCR信息

    基因名称 引物(5′-3′) 退火温度(℃) 扩增产物长度(bp)
    18S rRNA F: CAGCCACCCGAGATTGAGCA
    R: TAGTAGCGACGGGCGGTGTG 60 252
    hsa_circRNA_000585 F: GAGGTCAGACTGGGCAGGAGAT
    R: ACAGGACGCACTCAGTTCGCT 60 117
    下载: 导出CSV

    表  2  miR-615-5p和内参PCR信息

    基因名称 引物(5′-3′) 退火温度(℃) 扩增产物长度(bp)
    U6 F: GCTTCGGCAGCACATATACTAAAAT
    R: CGCTTCACGAATTTGCGTGTCAT 60 89
    hsa-miR-615-5p GSP: AAGGGGGTCCCCGGT 60 62
    R: GTGCGTGTCGTGGAGTCG
    下载: 导出CSV

    表  3  AMOT/YAP和内参PCR信息

    基因名称 引物(5′-3′) 退火温度(℃) 扩增产物长度(bp)
    AMOT F: TTGGAGGAGAATGTGATGAGAC
    R: TGGTGTTAGGAGAGTGACTGATG 60 97
    YAP F: GCCAGCAGGTTGGGAGAT
    R: TGTGATTTAAGAAGTATCTCTGACC 60 59
    18S rRNA F: CAGCCACCCGAGATTGAGCA
    R: TAGTAGCGACGGGCGGTGTG 60 252
    下载: 导出CSV
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