| 倪文,王曦,王奇胜,等.长链非编码 RNA MIR4435-2HG靶向微小 RNA-1-3p调控信号通路磷脂酰肌醇激酶 /蛋白激酶 B对胰腺癌细胞增殖、侵袭和迁移的机制研究[J].安徽医药,2021,25(3):467-473. |
| 长链非编码 RNA MIR4435-2HG靶向微小 RNA-1-3p调控信号通路磷脂酰肌醇激酶 /蛋白激酶 B对胰腺癌细胞增殖、侵袭和迁移的机制研究 |
| Mechanism of Lnc RNAMIR4435-2HGregulatedsignalingpathwayPI3K/Aktonproliferation, invasion and migration of pancreatic cancer cells by regulating miR-1-3p expression |
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| DOI：10.3969/j.issn.1009-6469.2021.03.009 |
| 中文关键词: 胰腺肿瘤 RNA，长链非编码 RNA MIR4435-2HG 微小 RNA-1-3p 磷脂酰肌醇激酶 /蛋白激酶 B信号通路 增殖 迁移 侵袭 |
| 英文关键词: Pancreatic neoplasms RNA, long noncoding Proliferation Migration Invasion |
| 基金项目: |
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| 中文摘要: |
| 目的探讨长链非编码 RNA MIR4435-2HG(LncRNA MIR4435-2HG)通过调控微小 RNA-1-3p(miR-1-3p)的表达对胰腺癌细胞增殖、迁移及侵袭的影响及其可能作用机制。方法体外培养正常胰腺细胞系 hTERT-HPNE与胰腺癌细胞系 PANC-1、 SW1990、PaTu8988、BxPC-3，将胰腺癌 PaTu8988细胞按照随机数字表法分为 si-MIR4435-2HG组(转染 MIR4435-2HG siRNA)、 si-con组(转染 siRNA Control)、 miR-1-3p组(转染 miR-1-3p mimics)、 miR-con组(转染 miR-1-3p阴性对照)、 si-MIR4435-2HG+an? ti-miR-1-3p组(共转染 MIR4435-2HG siRNA与 miR-1-3p抑制剂)、 si-MIR4435-2HG+anti-miR-con组(共转染 MIR4435-2HG siR? NA与 miR-1-3p抑制剂的阴性对照)，同时将未经任何处理的细胞作为 NC组。采用实时荧光定量逆转录聚合酶链反应( qRTPCR)检测细胞中 MIR4435-2HG与 miR-1-3p的表达；双荧光素酶报告基因检测 MIR4435-2HG与 miR-1-3p的相互作用。四甲基偶氮唑盐微量酶反应比色法( MTT法)分析敲低 MIR4435-2HG及上调 miR-1-3p表达对胰腺癌细胞增殖的影响； Transwell迁移及侵袭实验检测 MIR4435-2HG及上调 miR-1-3p表达对胰腺癌细胞迁移及侵袭能力的影响。蛋白质印迹法( Western blotting)检测细胞周期蛋白 D1(cyclin D1)、基质金属蛋白酶 2(MMP2)、基质金属蛋白酶 9(MMP9)及磷脂酰肌醇激酶( PI3K)/蛋白激酶 B( Akt)信号通路相关蛋白的表达。结果 MIR4435-2HG在胰腺癌细胞中的表达水平明显高于正常胰腺细胞； MIR4435-2HG可特异性结合 miR-1-3p并调控其表达活性；敲低 MIR4435-2HG与上调 miR-1-3p表达后，可抑制胰腺癌 PaTu8988细胞增殖，明显减弱细胞迁移及侵袭能力［其中 NC组、 miR-con组、 si-MIR4435-2HG组增殖活性分别为 48 h:(1.21±0.10)、(1.18±0.12)、(0.86±0.09)，细胞迁移数量分别为( 90.56±9.16)、(88.56±8.91)、(26.49±2.10)，细胞侵袭数量分别为( 160.79±16.12)、(155.09± 15.49)、(69.23±7.10)均 P < 0.05］，下调 PaTu8988细胞中 cyclin D1、MMP2、MMP9、p-Akt、PI3Kp110α、PI3Kp110β的表达；抑制 miR-1-3p表达可促进胰，腺癌 PaTu8988细胞增殖、迁移及侵袭。结论敲低 LncRNA MIR4435-2HG可通过靶向调控 miR-1-3p表达并抑制 PI3K/Akt信号通路活化而降低细胞增殖、迁移及侵袭相关蛋白表达，进而减弱胰腺癌细胞增殖、迁移及侵袭能力。 |
| 英文摘要: |
| Objective To investigate the effects of long-chain non-coding RNA MIR4435-2HG (LncRNA MIR4435-2HG) on the pro? liferation, migration and invasion of pancreatic cancer cells by regulating the expression of microRNA-1-3p (miR-1-3p) and its possible mechanism.Methods The normal pancreatic cell line hTERT-HPNE and pancreatic cancer cell lines PANC-1, SW1990, PaTu8988 and BxPC-3 were cultured in vitro, pancreatic cancer PaTu8988 cells were divided into si-MIR4435-2HG groups (transfection MIR4435-2HG siRNA), si-con group (transfection siRNA Control), miR-1-3p group (transfection miR-1-3p mimics), mir-con group (transfection negative control), si-MIR4435-2HG anti-miR-1-3p group (co-transfection MIR4435-2HG siRNA and miR-1-3), siMIR4435-2HG anti-miR-con group (co-transfection MIR4435-2HG siRNA negative control with miR-1-3p inhibitors) according to ran?dom number table method p inhibitors, while cells without any treatment were taken as the NC group. The expression of MIR4435-2HG and miR-1-3p was detected by real-time fluorescent quantitative PCR. The dual luciferase reporter gene detected the interaction of MIR4435-2HG with miR-1-3p. MTT assay was used to analyze the effect of knockdown of MIR4435-2HG and up-regulation of miR-13p expression on proliferation of pancreatic cancer cells. Transwell migration and invasion assays were performed to detect the effectsof MIR4435-2HG and up-regulation of miR-1-3p expression on migration and invasion of pancreatic cancer cells. Western blotting was used to detect the expression of cyclin D1, MMP2, MMP9 and phosphatidylinositol kinase (PI3K) / protein kinase B (Akt) signalingpathway-associated proteins.Results The expression level of MIR4435-2HG in pancreatic cancer cells was significantly higher than that of normal pancreatic cells. MIR4435-2HG could bind specifically to miR-1-3p to modulate the expression of miR-1-3p. Knock? down of MIR4435-2HG and up-regulation of miR-1-3p expression inhibited the proliferation of pancreatic cancer PaTu8988 cells, sig?nificantly attenuated cell migration and invasion [The proliferative activity of NC group, miR-con group and si-MIR4435-2HG groupwere 48 h:(1.21±0.10), (1.18±0.12), (0.86±0.09), the number of cell migration was (90.56±9.16), (88.56±8.91), (26.49±2.10), the num?ber of cell invasion was (160.79±16.12), (155.09±15.49), (69.23±7.10), all P < 0.05], and down-regulated the expression of cyclin D1, MMP2, MMP9, p-Akt, PI3Kp110α, and PI3Kp110β in PaTu8988 cells. Inhibition of miR-1-3p expression promoted proliferation, mi? gration and invasion of pancreatic cancer PaTu8988 cells.Conclusion Knockdown of LncRNA MIR4435-2HG can reduce the prolif? eration, migration and invasion of pancreatic cancer cells by targeting the regulation of miR-1-3p expression and inhibiting the activa? tion of PI3K/Akt signaling pathway, thereby reducing cell proliferation, migration and invasion-related protein expression. |
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