| 王 超,陈 炼,张思宇,等.微小RNA-9-5p干扰RhoA表达调控胰腺癌细胞增殖、侵袭及迁移[J].肿瘤学杂志,2023,29(3):220-225. |
| 微小RNA-9-5p干扰RhoA表达调控胰腺癌细胞增殖、侵袭及迁移 |
| MicroRNA-9-5p Affects Proliferation， Invasion and Migration of Pancreatic Ductal Adenocarcinoma Cells Through RhoA |
| 投稿时间：2022-12-21 |
| DOI：10.11735/j.issn.1671-170X.2023.03.B009 |
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| 中文关键词: 胰腺导管腺癌 微小RNA-9-5p RhoA信号通路 |
| 英文关键词:pancreatic ductal adenocarcinoma microRNA-9-5p RhoA signaling pathway |
| 基金项目:河北省卫生厅科研基金项目（1020140005） |
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| 中文摘要: |
| 摘 要：［目的］ 探讨微小RNA（miR）-9-5p在胰腺导管腺癌（pancreatic ductal adenocarcinoma，PDAC）中表达及影响其生物学行为的机制。［方法］ 实时定量反转录聚合酶链反应（qRT-PCR）测定40例PDAC与正常人胰腺细胞系HPDE6-C7中miR-9-5p的表达。miR-9-5p模拟物转染PDAC细胞株PANC-1；甲基噻唑蓝（MTT）比色法检测细胞增殖活性；划痕实验、Transwell侵袭实验测定细胞侵袭、迁移能力。qPCR及蛋白印迹实验检测RhoA、ROCK1、PCNA、CyclinD1、MMP-2、MMP-9及p21表达。双荧光素酶报告基因分析实验检测miR-9-5p对RhoA的调控功能。［结果］ ①临床样本中miR-9-5p在PDAC组织中的表达显著性低于癌旁组织（0.34±0.08 vs 1.01±0.24，t=13.222，P<0.001）。miR-9-5p表达降低与胰腺癌淋巴结转移、肿瘤分期存在相关性。②miR-9-5p在胰腺癌细胞系PANC-1中表达明显低于正常人胰腺细胞系HPDE6-C7（0.28±0.04 vs 0.53±0.05，t=7.204，P=0.019）。③双荧光素酶报告基因分析结果表明，野生型RhoA的荧光素酶活性在转染miR-9-5p模拟物后较对照组降低（0.99±0.08 vs 0.22±0.02，P<0.001）。④转染后，转染组细胞增殖活性降低（0.53±0.07 vs 1.09±0.11、1.13±0.08，P均<0.001），侵袭距离减少（0.28±0.07 μm vs 0.66±0.05 μm、0.64±0.07 μm，P均<0.001），迁移细胞数减少（32.83±2.99 vs 70.00±4.29、65.67±5.79，P均<0.001）。⑤与阴性对照组及Scramble组比较，miR-9-5p模拟物转染组RhoA、ROCK1、PCNA、Cyclin D1、MMP-2、MMP-9表达显著性降低，p21表达升高（P均<0.05）。［结论］ 胰腺导管腺癌细胞中miR-9-5p表达降低；过表达miR-9-5p后能明显抑制PANC-1细胞的体外增殖、侵袭及迁移，其调控机制可能与RhoA信号通路相关。 |
| 英文摘要: |
| Abstract：［Objective］ To explored the effect and mechanisms of microRNA-9-5p(miR-9-5p) on proliferation， invasion and migration of pancreatic ductal adenocarcinoma cells(PDAC) . ［Methods］ The expression of miR-9-5p was detected by qRT-PCR in tumor tissues and pericancerous tissues of 40 patients with pancreatic ductal adenocarcinoma， the correlation between miR-9-5p expression and clinicopathological features of PDAC was analyzed. The levels of miR-9-5p were detected in pancreatic ductal adenocarcinoma cell lines and normal human pancreatic cell line HPDE6-C7. PDAC PANC-1 cells were transfected with miR-9-5p mimics. The cell proliferation activity was detected with MTT assay， cell invasion and migration ability was determined with wound healing assay and Transwell assay， the expression of RhoA， ROCK1， PCNA， Cyclin D1， MMP-2， MMP-9 and p21 was detected with RT-PCR and Western blot， and the regulatory function of miR-9-5p on RhoA was verified with dual luciferase reporter gene analysis. ［Results］ ①The expression of miR-9-5p in PDAC tissues was significantly lower than that in paracancerous tissues(0.34±0.08 vs 1.01±0.24， t=13.222， P<0.001). Low miR-9-5p expression was correlated with lymph node metastasis and higher tumor stage in PDAC. ②The expression of miR-9-5p in PANC-1 cells was significantly lower than that in HPDE6-C7 cells(0.28±0.04 vs 0.53±0.05， t=7.204， P=0.019). ③Dual luciferase reporter gene analysis showed that the luciferase activity of wild-type RhoA was reduced after transfection with miR-9-5p mimics compared to negative controls group(0.99±0.08 vs 0.22±0.02， t=20.976， P<0.001). ④The cell proliferation activity (0.53±0.07 vs 1.09±0.11， 1.13±0.08， P all<0.001)， invading distance(0.28±0.07 μm vs 0.66±0.05 μm， 0.64±0.07 μm， P all <0.001)， and the number of migration cells(32.83±2.99 vs 70.00±4.29， 65.67±5.79， P all <0.001) were all reduced in PANC-1 cells after transfected with miR-9-5p mimics. ⑤Compared with the negative control and Scramble group， miR-9-5p mimics transfected group showed significantly lower expression of RhoA， ROCK1， PCNA， Cyclin D1， MMP-2， MMP-9 and higher expression of p21(P all <0.05). ［Conclusion］ The miR-9-5p is down-regulated in pancreatic ductal adenocarcinoma cells， overexpression of miR-9-5p significantly inhibits the proliferation， invasion and migration of PANC-1 cells in vitro， and its regulatory mechanism may be related to the RhoA signaling pathway. |
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