长链非编码RNA PCAT19对胰腺癌细胞增殖与侵袭的影响及其作用机制
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湖北医药学院附属随州医院/随州市中心医院 普通外科

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赵银峰,湖北医药学院附属随州医院/随州市中心医院主治医师,主要从事肝胆及甲状腺乳腺肿瘤基础与临床方面的研究。

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Effect of long non-coding RNA PCAT19 on proliferation and invasion in pancreatic cancer cells and its action mechanism
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Department of General Surgery, Suizhou Hospital of Hubei University of Medicine/Suizhou Central Hospital, Suizhou, Hubei 441300, China

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    摘要:

    背景与目的 长链非编码RNA PCAT19(lncRNA PCAT19,简称PCAT19)在多种肿瘤中表达上调,且与恶性进展和不良预后密切相关。然而,PCAT19在胰腺癌中的表达、功能及作用机制尚无研究报道。笔者前期通过starBase数据库预测PCAT19可与miR-195-5p互补结合,因此,本研究探讨PCAT19在胰腺癌细胞中的表达与作用,及其与靶基因和相应miRNA的调控关系。方法 用qRT-PCR检测人胰腺导管上皮细胞系(HPNE)和胰腺癌细胞系(PANC-1、SW1990、HS766T、CFPAC-1)中PCAT19及miR-195-5p的表达。用双萤光素酶报告基因分析PCAT19与miR-195-5p的靶向结合作用。将PANC-1细胞分别转染PCAT19沉默序列si-PCAT19(si-PCAT19组)、阴性对照序列(si-NC组)、miR-195-5p抑制序列+si-PCAT19(共转染组)后,用MTT测定细胞增殖能力,Transwell测定细胞侵袭能力,Western blot检测Wnt/β-Catenin信号通路相关蛋白的表达。结果 各胰腺癌细胞系中PCAT19表达水平均明显高于HPNE细胞,而miR-195-5p的表达水平均明显低于HPNE细胞(均P<0.05)。双萤光素酶实验显示miR-195-5p是PCAT19的靶miRNA。与si-NC组PANC-1细胞比较,si-PCAT19组PANC-1中miR-195-5p表达水平明显升高,细胞增殖能力与侵袭能力明显降低,β-catenin、c-Myc和cyclin D1表达水平明显下调(均P<0.05),而共转染组以上各项指标与si-NC组差异均无统计学意义(均P>0.05)。结论 PCAT19在胰腺癌细胞中表达升高,其可促进胰腺癌细胞增殖和侵袭,机制可能与调控miR-195-5p并影响Wnt/β-Catenin信号通路有关。

    Abstract:

    Background and Aims Long non-coding RNA PCAT19 (lncRNA PCAT19, hereafter referred to as PCAT19) is upregulated in various tumors and closely associated with malignant progression and poor prognosis. However, no studies have reported the expression, function, and mechanisms of PCAT19 in pancreatic cancer. The authors predicted that PCAT19 could interact with miR-195-5p in a complementary manner through the starBase database. Therefore, this study was conducted to investigate the expression and function of PCAT19 in pancreatic cancer cells, as well as its regulatory relationship with target genes and corresponding miRNA.Methods The expressions of PCAT19 and miR-195-5p in human pancreatic ductal epithelial cells (HPNE) and pancreatic cancer cell lines (PANC-1, SW1990, HS766T, CFPAC-1) were detected by qRT-PCR method. Dual-luciferase reporter gene analysis was conducted to assess the targeted binding interaction between PCAT19 and miR-195-5p. PANC-1 cells were transfected with PCAT19 silencing sequence si-PCAT19 (si-PCAT19 group), negative control sequence (si-NC group), and miR-195-5p inhibitor plus si-PCAT19 (co-transfection group). Then, cell proliferation ability was determined using MTT assay, cell invasion ability was assessed using Transwell assay, and the expressions of proteins related to the Wnt/β-Catenin signaling pathway were determined by Western blot analysis.Results In all studied pancreatic cancer cell lines, the expression levels of PCAT19 were significantly higher than that in HPNE cells. In contrast, the expression levels of miR-195-5p were significantly lower than that in HPNE cells (all P<0.05). Dual-luciferase experiments showed that miR-195-5p was a target miRNA of PCAT19. In the si-PCAT19 group compared with the si-NC group, the expression of miR-195-5p was significantly increased, cell proliferation and invasion abilities were significantly decreased, and the expression levels of β-catenin, c-Myc, and cyclin D1 were significantly downregulated (all P<0.05). However, in the co-transfection group, there were no statistically significant differences in these indexes compared with the si-NC group (all P>0.05).Conclusion PCAT19 is upregulated in pancreatic cancer cells and can promote the proliferation and invasion of pancreatic cancer cells. The mechanism may be related to regulating miR-195-5p and affecting the Wnt/β-Catenin signaling pathway.

    图1 PCAT19与miR-195-5p在胰腺癌细胞及正常胰腺导管上皮细胞中的表达水平Fig.1 The expression levels of PCAT19 and miR-195-5p in pancreatic cancer cells and normal pancreatic ductal epithelial cells
    图2 PCAT19的靶点预测与验证 A:PCAT19与miR-195-5p存在靶向结合位点;B:萤光素酶实验示miR-195-5p为PCAT19的靶标分子Fig.2 Target prediction and validation of PCAT19 A: PCAT19 has binding sites for miR-195-5p; B: Dual-luciferase experiments demonstrate that miR-195-5p is a target molecule of PCAT19
    图3 PCAT19与miR-195-5p对PANC-1细胞增殖的影响 A:各组细胞miR-195-5p的表达比较;B:各组细胞的增殖曲线比较Fig.3 The impact of PCAT19 and miR-195-5p on the proliferation of PANC-1 cells A: Comparison of miR-195-5p expression among groups of cells; B: Comparison of the proliferation curves of cells in each group
    图4 PCAT19与miR-195-5p对PANC-1细胞侵袭能力的影响Fig.4 The influence of PCAT19 and miR-195-5p on the invasion capability of PANC-1 cells
    图5 PCAT19与miR-195-5p对Wnt/β-Catenin信号通路蛋白表达的影响Fig.5 The effect of PCAT19 and miR-195-5p on the expression of proteins in the Wnt/β-Catenin signaling pathway
    图1 PCAT19与miR-195-5p在胰腺癌细胞及正常胰腺导管上皮细胞中的表达水平Fig.1 The expression levels of PCAT19 and miR-195-5p in pancreatic cancer cells and normal pancreatic ductal epithelial cells
    图2 PCAT19的靶点预测与验证 A:PCAT19与miR-195-5p存在靶向结合位点;B:萤光素酶实验示miR-195-5p为PCAT19的靶标分子Fig.2 Target prediction and validation of PCAT19 A: PCAT19 has binding sites for miR-195-5p; B: Dual-luciferase experiments demonstrate that miR-195-5p is a target molecule of PCAT19
    图3 PCAT19与miR-195-5p对PANC-1细胞增殖的影响 A:各组细胞miR-195-5p的表达比较;B:各组细胞的增殖曲线比较Fig.3 The impact of PCAT19 and miR-195-5p on the proliferation of PANC-1 cells A: Comparison of miR-195-5p expression among groups of cells; B: Comparison of the proliferation curves of cells in each group
    图4 PCAT19与miR-195-5p对PANC-1细胞侵袭能力的影响Fig.4 The influence of PCAT19 and miR-195-5p on the invasion capability of PANC-1 cells
    图5 PCAT19与miR-195-5p对Wnt/β-Catenin信号通路蛋白表达的影响Fig.5 The effect of PCAT19 and miR-195-5p on the expression of proteins in the Wnt/β-Catenin signaling pathway
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赵银峰,梁云,魏天天,嘉玉成.长链非编码RNA PCAT19对胰腺癌细胞增殖与侵袭的影响及其作用机制[J].中国普通外科杂志,2023,32(9):1333-1340.
DOI:10.7659/j. issn.1005-6947.2023.09.006

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  • 收稿日期:2022-05-23
  • 最后修改日期:2023-03-06
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  • 在线发布日期: 2023-11-03