Qin Yue-Chao, Yan Xin, Yuan Xiao-Lin, Yu Wei-Wei, Qu Fan-Jie
Department of Oncology, Affiliated Dalian Third People's Hospital of Dalian Medical University, Dalian 116033, Liaoning Province, China.
Research Center, Affiliated Zhongshan Hospital of Dalian University, Dalian 116001, Liaoning Province, China.
World J Gastrointest Oncol. 2023 Sep 15;15(9):1544-1555. doi: 10.4251/wjgo.v15.i9.1544.
Gastric cancer (GC) is one of the most common malignant tumors. Osteopontin (OPN) is thought to be closely related to the occurrence, metastasis and prognosis of many types of tumors.
To investigate the effects of OPN on the proliferation, invasion and migration of GC cells and its possible mechanism.
The mRNA and protein expression of OPN in the GC cells were analyzed by real-time quantitative-reverse transcription polymerase chain reaction and western blotting, and observe the effect of varying degree expression OPN on the proliferation and other behaviors of GC. Next, the effects of OPN knockdown on GC cells migration and invasion were examined. The short hairpin RNA (shRNA) and negative control shRNA targeting OPN-shRNA were transfected into the cells according to the manufacturer's instructions. Non transfected cells were classified as control in the identical transfecting process. 24 h after RNA transfection cell proliferation activity was detected by 3-(4,5)-dimethylthiahiazo (-z-y1)-3,5-di-phenytetrazoliumromide assay, and cell invasiveness and migration were detected by Trans well assay. Meanwhile, the expression of protein kinase B (AKT), matrix metalloproteinase 2 (MMP-2) and vascular endothelial growth factor (VEGF) in the human GC cell lines was detected by reverse transcription polymerase chain reaction and western blotting.
The results of this study revealed that OPN mRNA and protein expression levels were highly expressed in SGC-7901 cells. OPN knockdown by specific shRNA noticeably reduced the capabilities of proliferation, invasion and migration of SGC-7901 cells. Moreover, in the experiments of investigating the underlying mechanism, results showed that OPN knockdown could down-regulated the expression of MMP-2 and VEGF, it also decreased the phosphorylation of AKT. Meanwhile, the protein expression levels of MMP-2, VEGF and phosphorylated AKT was noticeable lower than that in control group in the GC cells after they were added to phosphatidylinositol-3-kinase (PI3K) inhibitor (LY294002).
These results suggested that OPN though PI3K/AKT/mammalian target of rapamycin signal pathway to up-regulate MMP-2 and VEGF expression, which contribute SGC-7901 cells to proliferation, invasion and migration. Thus, our results demonstrate that OPN may serve as a novel prognostic biomarkers as well as a potential therapeutic targets for GC.
胃癌（GC）是最常见的恶性肿瘤之一。骨桥蛋白（OPN）被认为与多种肿瘤的发生、转移及预后密切相关。
探讨OPN对GC细胞增殖、侵袭和迁移的影响及其可能机制。
采用实时定量逆转录聚合酶链反应和蛋白质印迹法分析GC细胞中OPN的mRNA和蛋白表达，并观察不同程度表达OPN对GC增殖及其他行为的影响。接下来，检测OPN敲低对GC细胞迁移和侵袭的影响。按照制造商的说明，将靶向OPN-shRNA的短发夹RNA（shRNA）和阴性对照shRNA转染到细胞中。在相同的转染过程中，未转染的细胞作为对照。RNA转染24小时后，通过3-（4,5）-二甲基噻唑（-z-y1）-3,5-二苯基四氮唑溴盐法检测细胞增殖活性，通过Transwell法检测细胞侵袭和迁移能力。同时，通过逆转录聚合酶链反应和蛋白质印迹法检测人GC细胞系中蛋白激酶B（AKT）、基质金属蛋白酶2（MMP-2）和血管内皮生长因子（VEGF）的表达。
本研究结果显示，OPN mRNA和蛋白表达水平在SGC-7901细胞中高表达。特异性shRNA敲低OPN可显著降低SGC-7901细胞的增殖、侵袭和迁移能力。此外，在探究潜在机制的实验中，结果表明OPN敲低可下调MMP-2和VEGF的表达，还可降低AKT的磷酸化水平。同时，在GC细胞中加入磷脂酰肌醇-3-激酶（PI3K）抑制剂（LY294002）后，MMP-2、VEGF和磷酸化AKT的蛋白表达水平明显低于对照组。
这些结果表明，OPN通过PI3K/AKT/雷帕霉素哺乳动物靶点信号通路上调MMP-2和VEGF表达，促进SGC-7901细胞增殖、侵袭和迁移。因此，我们的结果表明，OPN可能作为一种新的预后生物标志物以及GC的潜在治疗靶点。
