| 冯佳梅,万 华,高晴倩,瞿文超,邵士珺,孙佳晔,吴雪卿.LncRNA MCM3AP-AS1调节miR-424-5p/PSAT1轴对乳腺癌恶性进展的影响[J].现代生物医学进展英文版,2024,(14):2606-2612. |
| LncRNA MCM3AP-AS1调节miR-424-5p/PSAT1轴对乳腺癌恶性进展的影响 |
| Effect of LncRNA MCM3AP-AS1 on Malignant Progression of Breast Cancer by Regulating miR-424-5p/PSAT1 Axis |
| Received:January 28, 2024 Revised:February 23, 2024 |
| DOI：10.13241/j.cnki.pmb.2024.14.002 |
| 中文关键词: MCM3AP-AS1 miR-424-5p PSAT1 乳腺癌 |
| 英文关键词: MCM3AP-AS1 miR-424-5p PSAT1 Breast cancer |
| 基金项目:上海市卫生健康委员会科研课题计划项目（202040254） |
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| 中文摘要: |
| 摘要 目的：探讨长链非编码核糖核酸（lncRNA）微小染色体维持蛋白3相关蛋白-反义链1（MCM3AP-AS1）调节微小核糖核酸（miR）-424-5p/磷酸丝氨酸氨基转移酶1（PSAT1）轴对乳腺癌恶性进展的影响。方法：采用实时荧光定量聚合酶链式反应（RT-qPCR）法检测人正常乳腺上皮细胞MCF-10A和乳腺癌细胞株MDA-MB-231、BT549、BT20中LncRNA MCM3AP-AS1、miR-424-5p和PSAT1 信使核糖核酸（mRNA）的表达水平。构建LncRNA MCM3AP-AS1低表达模型、miR-424-5p敲低与过表达模型，并使用RT-qPCR方法验证转染模型构建的成功。分别采用四氮甲基唑蓝（MTT）法、Transwell实验检测乳腺癌细胞株的增殖、迁移、侵袭能力。免疫印迹法检测各组MDA-MB-231细胞中细胞周期蛋白D1（CyclinD1）、E-钙黏蛋白（E-cadherin）、N-钙黏蛋白（N-cadherin）和PSAT1蛋白的表达。经生物信息学分析后，采用双荧光素酶报告基因实验与RNA免疫共沉淀（RIP）实验分别验证LncRNA MCM3AP-AS1与miR-424-5p、miR-424-5p与PSAT1之间的靶向关系。结果：在乳腺癌细胞株MDA-MB-231、BT549、BT20中MCM3AP-AS1、PSAT1 mRNA的表达水平显著高于MCF-10A（P＜0.05），miR-424-5p表达水平显著低于MCF-10A（P＜0.05）。敲低MCM3AP-AS1或过表达miR-424-5p均可以降低MDA-MB-231细胞的吸光度（OD490）值、细胞迁移数目和细胞侵袭数目、CyclinD1、N-cadherin和PSAT1蛋白表达水平（P＜0.05），提高细胞E-cadherin蛋白表达水平（P＜0.05）。敲低miR-424-5p的表达逆转了下调MCM3AP-AS1对MDA-MB-231细胞增殖、迁移和侵袭以及相关蛋白表达的影响。双荧光素酶报告基因实验与RIP实验证实MCM3AP-AS1靶向负调控miR-424-5p表达，miR-424-5p靶向负调控PSAT1的表达。结论：LncRNA MCM3AP-AS1在乳腺癌中呈高表达，其低表达可通过靶向调节miR-424-5p/PSAT1轴抑制乳腺癌的恶性进展。 |
| 英文摘要: |
| ABSTRACT Objective: To investigate the effect of long non-coding RNA (lncRNA) microchromosome maintenance protein 3-associated protein-antisense chain 1 (MCM3AP-AS1) on malignant progression of breast cancer by regulating microribonucleic acid (miR)-424-5p/phosphoserine aminotransferase 1(PSAT1) axis. Methods: The expression levels of LncRNA MCM3AP-AS1, miR-424-5p and PSAT1 mRNA in human normal breast epithelial cells MCF-10A and breast cancer cell lines MDA-MB-231, BT549 and BT20 were detected by real-time fluorescence quantitative polymerase chain reaction(RT-qPCR). LncRNA MCM3AP-AS1 low expression model, miR-424-5p knockdown and overexpression model were constructed, and the success of transfection model construction was verified by RT-qPCR method. The proliferation, migration and invasion of breast cancer cell lines were detected by methyl thiazolyl tetrazolium (MTT) assay and Transwell assay respectively. The expressions of cyclin d1(CyclinD1), E-cadherin (E-cadherin), N-cadherin (N-cadherin) and PSAT1 proteins in MDA-MB-231 cells were detected by Western blotting. After bioinformatics analysis, the targeting relationship between LncRNA MCM3 AP-AS1 and miR-424-5p, miR-424-5p and PSAT1 was verified by double luciferase reporter gene assay and RNA immunoprecipitation (RIP) assay respectively. Results: The expression levels of MCM3AP-AS1 and PSAT1 mRNA in breast cancer cell lines MDA-MB-231, BT549 and BT20 were significantly higher than those in MCF-10A(P<0.05), and the expression level of miR-424-5p was significantly lower than that in MCF-10A(P<0.05). Knockdown of MCM3AP-AS1 or overexpression of miR-424-5p could reduce the absorbance(OD490) value of MDA-MB-231 cells, the number of cell migration and cell invasion, the expression levels of CyclinD1, N-cadherin and PSAT1 proteins(P<0.05), and increase the expression level of E-cadherin protein (P<0.05). Knockdown of miR-424-5p reversed the effects of down-regulation of MCM3AP-AS1 on the proliferation, migration and invasion of MDA-MB-231 cells and the expression of related proteins. The dual luciferase reporter gene assay and RIP assay confirm that MCM3AP-AS1 targeted and negatively regulated the expression of miR-424-5p, and miR-424-5p targeted and negatively regulated the expression of PSAT1. Conclusion: LncRNA MCM3AP-AS1 is highly express in breast cancer, and its low expression can inhibit the malignant progression of breast cancer by targeting the miR-424-5p/PSAT1 axis. |
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