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中华危重症医学杂志(电子版)

中华危重症医学杂志(电子版) ›› 2021, Vol. 14 ›› Issue (02) : 113 -119. doi: 10.3877/cma.j.issn.1674-6880.2021.02.004

所属专题： 文献；

论著

非编码RNA在脓毒症心肌细胞中表达及分析
黄爱蓉1,†(), 梁冬施1, 金益梅1, 林苗苗2, 夏小娇2, 陈晓丽2   
  1. 1. 325000　浙江温州，温州医科大学附属第二医院、育英儿童医院儿童普通内科
    2. 325000　浙江温州，温州医科大学附属第二医院、育英儿童医院中西医结合科
  • 收稿日期:2020-12-25 出版日期:2021-04-30
  • 通信作者: 黄爱蓉
  • 基金资助:
    浙江省医药卫生科技计划项目(2019311934); 温州市公益性科技计划项目(Y20180009)

Differential expression of non-coding RNA in septic cardiomyocytes

Airong Huang1,†(), Dongshi Liang1, Yimei Jin1, Miaomiao Lin2, Xiaojiao Xia2, Xiaoli Chen2   

  1. 1. Department of Pediatrics, the Second Affiliated Hospital and Yuying Children's Hospital of Wenzhou Medical University, Wenzhou 325000, China
    2. Department of Integrated Chinese and Western Medicine, the Second Affiliated Hospital and Yuying Children's Hospital of Wenzhou Medical University, Wenzhou 325000, China
  • Received:2020-12-25 Published:2021-04-30
  • Corresponding author: Airong Huang
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黄爱蓉, 梁冬施, 金益梅, 林苗苗, 夏小娇, 陈晓丽. 非编码RNA在脓毒症心肌细胞中表达及分析[J]. 中华危重症医学杂志(电子版), 2021, 14(02): 113-119.

Airong Huang, Dongshi Liang, Yimei Jin, Miaomiao Lin, Xiaojiao Xia, Xiaoli Chen. Differential expression of non-coding RNA in septic cardiomyocytes[J]. Chinese Journal of Critical Care Medicine(Electronic Edition), 2021, 14(02): 113-119.

目的

探讨长链非编码RNA（lncRNA）及微RNA（miRNA）在脓毒症心肌细胞中的表达情况。

方法

采用基因芯片筛选对照组与脓毒症组小鼠心肌细胞中lncRNA与miRNA的差异基因，通过基因本体和京都基因与基因组百科全书对差异基因进行分析以判定差异基因主要参与的生物学功能及相关通路。通过starbase数据库及NCBI数据库中的基因数据库进一步筛选与心肌细胞及线粒体损伤相关的差异lncRNA和miRNA。采用实时定量PCR方法验证上述lncRNA与miRNA基因在脓毒症心肌细胞中的表达情况。

结果

对照组与脓毒症组小鼠心肌细胞中有显著差异的lncRNA共5 520个，上调的lncRNA有3 186个，下调的lncRNA有2 334个；有显著差异的miRNA共有90个，其中上调的miRNA共59个，下调的miRNA共有31个。生物学功能筛选发现，lncRNA中的xist、1700020I14Rik及miRNA中的miR-7a-5p、miR-30c-1-3p与心肌细胞及线粒体损害密切相关。脓毒症组细胞中xist（t = 5.640，P = 0.002）与1700020I14Rik （t = 6.044，P = 0.002）表达水平较对照组均显著升高，miR-7a-5p（t = 27.277，P < 0.001）与miR-30c-1-3p（t = 12.910，P < 0.001）表达水平较对照组均显著下降。

结论

lncRNA及miRNA均参与了脓毒症心肌细胞损伤的过程，且差异基因xist、1700020I14Rik、miR-7a-5p、miR-30c-1-3p可能在其中起到关键作用。

关键词: 脓毒症, 肌细胞, 心脏, 非编码RNA
Objective

To explore the differential expression levels of long non-coding RNA (lncRNA) and microRNA (miRNA) in septic cardiomyocytes.

Methods

The gene microarray was used to detect the differential lncRNA and miRNA in the cardiomyocytes of mice in the sepsis group and control group. The gene ontology and Kyoto Encyclopedia of Genes and Genomes were used to predict the major biological functions and signal pathways involved in the differentially expressed lncRNA and miRNA. Then the differential lncRNA and miRNA related to myocardial cell and mitochondrial injury were further screened through starbase and NCBI databases. The above related genes were verified and detected by real-time quantitative PCR.

Results

A total of 5 520 distinctly differentially expressed lncRNAs and 90 distinctly differentially expressed miRNAs were found in the sepsis group as compared with the control group, among which 3 186 lncRNAs and 59 miRNAs were up-regulated, and 2 334 lncRNAs and 31 miRNAs down-regulated. Biological function analysis showed that xist, 1700020I14Rik in lncRNA and miR-7a-5p, miR-30c-1-3p in miRNA were closely related to damage of cardiomyocytes and mitochondria. Compared with the control group, the expression levels of xist (t = 5.640, P = 0.002) and 1700020I14Rik (t = 6.044, P = 0.002) significantly increased, and the expression levels of miR-7a-5p (t = 27.277, P < 0.001) and miR-30c-1-3p (t = 12.910, P < 0.001) markedly decreased in the sepsis group.

Conclusion

Both lncRNA and miRNA are involved in the pathophysiological process of sepsis in cardiomyocytes, and the xist, 1700020I14Rik, miR-7a-5p and miR-30c-1-3p may play a key role in the sepsis-induced cardiac dysfunction.

Key words: Sepsis, Myocytes, cardiac, Non-coding RNA
表1 lncRNA与miRNA的引物序列
引物名称 序列
miR-7a-5p上游引物 GCCGAGGGAAGACTAGTGAT
miR-7a-5p下游引物 CTCAACTGGTGTCGTGGAGT
miR-30c-1-3p上游引物 CGAGTGGGAGAGGGTTGT
miR-30c-1-3p下游引物 CTCAACTGGTGTCGTGGAGT
U6上游引物 CTCGCTTCGGCAGCACA
U6下游引物 AACGCTTCACGAATTTGCGT
1700020I14Rik上游引物 TCCCGTGAGAAGCGGAGTA
1700020I14Rik下游引物 CGTGCCAAGTCCTGAGTTCA
xist上游引物 GGCTCAAAGAATGCCACCCT
xist下游引物 GATCACGCTGAAGACCCAGTT
GAPDH上游引物 AGGTCGGTGTGAACGGATTTG
GAPDH下游引物 TGTAGACCATGTAGTTGAGGTCA
表2 两组心肌细胞RNA样本浓度和纯度（ ± s）
组别 浓度（ng/μL） A260/A280 A260/A230
对照组 777.7 ± 1.6 1.89 ± 0.07 1.931 ± 0.019
脓毒症组 865.3 ± 3.7 1.85 ± 0.05 1.975 ± 0.043
图1 lncRNA中显著富集的差异表达基因GO分析及pathway分析
图2 miRNA中显著富集的差异表达基因GO分析及pathway分析
图3 两组心肌细胞lncRNA及miRNA中显著富集的差异表达基因比较（n = 4.5 × 105）
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