microRNA-3129 promotes cell proliferation in gastric cancer cell line SGC7901 via positive regulation of pRb

Several microRNAs (miRNAs) have been reported as oncogenes or tumor suppressors in many cancers, including gastric cancer (GC). However, the role and molecular mechanism of miR-3129 in GC is largely unknown. We aimed to explore the function and the underlying molecular mechanism of miR-3129 in GC. Cancer tissues and corresponding adjacent tissues were collected from 50 patients with GC, and the expression of miR-3129 was detected by RT-qPCR. The expression of miR-3129 and pRb in human GC cell line SCG7091 was altered by transient transfection. Thereafter, MTT and flow cytometry assays were used to analyze cell viability and cell cycle. The expression of cyclin E, CDK2, CDK2 inhibitors (p16 and 21), and pRb were detected by RT-qPCR and western blot. A significant up-regulation of miR-3129 was observed in GC tissues compared to adjacent tissues. Overexpression of miR-3129 significantly improved cell viability after 4 days of post-transfection. Flow cytometry assay results showed that the miR-3129 overexpression arrested more SGC7901 cells at S phase. Moreover, overexpression of miR-3129 down-regulated the expression of CDK2 inhibitors while it up-regulated the expression levels of cyclin E, CDK2, and pRb. Interestingly, we found that pRb inhibition reversed the effect of miR-3129 inhibitor on cell proliferation in SGC7901 cells, increased cell viability, reduced cells at G0/1 phase, and modulated the expression of proliferation-related factors. Our results revealed that miR-3129 functioned as an oncogene through positive regulation of pRb and may prove to be a promising option for molecular therapy of GC.

已有研究表明，多种微小RNA（microRNAs, miRNAs）可作为癌基因或抑癌基因参与多种癌症的发生发展，其中包括胃癌（gastric cancer, GC）。然而，miR-3129在胃癌中的具体作用及分子机制仍未明确。本研究旨在探讨miR-3129在胃癌中的功能及潜在分子机制。本研究收集了50例胃癌患者的癌组织及配对癌旁组织，并通过逆转录实时定量聚合酶链反应（reverse transcription quantitative polymerase chain reaction, RT-qPCR）检测miR-3129的表达水平。通过瞬时转染技术调控人胃癌细胞系SCG7091中miR-3129与视网膜母细胞瘤蛋白（retinoblastoma protein, pRb）的表达水平。随后采用噻唑蓝比色法（MTT）与流式细胞术（flow cytometry）分别检测细胞活力与细胞周期分布。采用逆转录实时定量聚合酶链反应与蛋白质印迹法（western blot）检测细胞周期蛋白E（cyclin E）、细胞周期蛋白依赖性激酶2（CDK2）、CDK2抑制剂（p16与p21）及pRb的表达水平。结果显示，与癌旁组织相比，胃癌组织中miR-3129的表达水平显著上调。转染4天后，miR-3129过表达可显著提升细胞活力。流式细胞术结果显示，miR-3129过表达可使更多SGC7901细胞阻滞于S期。此外，miR-3129过表达可下调CDK2抑制剂的表达，同时上调细胞周期蛋白E、CDK2及pRb的表达水平。值得注意的是，本研究发现，抑制pRb可逆转miR-3129抑制剂对SGC7901细胞增殖的影响，具体表现为细胞活力升高、G0/G1期细胞占比减少，并可调控增殖相关因子的表达。本研究结果表明，miR-3129通过正向调控pRb发挥癌基因功能，有望成为胃癌分子治疗的潜在靶点。