| 引用本文: | 冯靖洋,申晓宝,李子烨,王百林,吴帆.EGFL9通过EGFR/PI3K/AKT/mTOR通路促进肝癌细胞的增殖、侵袭和迁移能力[J].中国临床新医学,,():-. |
| Feng Jingyang,Shen Xiaobao,Li Ziye,Wang Bailin.EGFL9通过EGFR/PI3K/AKT/mTOR通路促进肝癌细胞的增殖、侵袭和迁移能力[J].中国临床新医学,,():-. |
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| 摘要: |
| 目的:分析表皮生长因子样结构9（epidermal growth factor-like domain，EGFL9）通过EGFR/PI3K/AKT/mTOR通路促进肝癌增殖、侵袭、迁移的具体分子机制。 方法:利用慢病毒介导的siRNA技术，构建针对两个不同靶点敲减EGFL9基因的肝癌细胞系（Huh-7、SNU-449）。运用MTT法、细胞凋亡试验、Transwell试验，评估细胞增殖、凋亡、迁移与侵袭能力。采用蛋白印迹方法检测EGFL9敲减及过表达的肝癌细胞系和裸鼠皮下种植瘤中EGFR/PI3K/AKT/mTOR信号通路蛋白的表达及磷酸化水平。然后再以EGFR及AKT抑制剂分别处理EGFL9过表达的Huh-7细胞系（前期研究已构建），再通过细胞功能实验检测上述细胞增殖、迁移能力的变化，并以蛋白印迹方法检测相应分子通路蛋白含量的变化。 结果:RT-qPCR结果显示双靶点敲减对Huh-7、SNU-449肝癌细胞系的EGFL9基因抑制效率均超过80%。细胞功能试验表明，通过两个不同靶点敲减EGFL9基因均可明显抑制Huh-7、SNU-449肝癌细胞的增殖、侵袭、迁移能力，并促进其凋亡。蛋白印迹实验结果提示，敲减EGFL9可显著抑制肝癌细胞及相应裸鼠皮下种植瘤中EGFR、PI3K、AKT、mTOR蛋白的磷酸化水平，但不影响其总表达水平；而在EGFL9过表达的Huh-7细胞中，这些蛋白的的磷酸化水平则明显高于对照组。经EGFR及AKT抑制剂分别处理后，EGFL9过表达Huh-7肝癌细胞中相关通路蛋白的磷酸化水平明显下降，细胞的增殖和迁移能力也明显减弱。 结论:EGFL9可能通过EGFR-PI3K-AKT-mTOR信号通路促进肝癌细胞的增殖、侵袭及迁移能力。提示其可能是一个潜在的肝癌治疗靶点。 |
| 关键词: EGFL9 肝细胞癌 信号传导通路 增殖 侵袭 |
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| 基金项目:国家自然科学基金（81974442）；广东省自然科学基金（2020A1515010799）；广州市科技计划-市校（院）企联合资助项目（2024A03J0670） |
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| EGFL9 enhances the proliferative, invasive, and migratory capacities of hepatocellular carcinoma (HCC) cells through activation of the EGFR/PI3K/AKT/mTOR signaling pathway |
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Feng Jingyang, Shen Xiaobao, Li Ziye, Wang Bailin
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Department of Hepatobiliary Surgery,Guangzhou Red Cross Hospital of Jinan University,Guangzhou
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| Abstract: |
| Objective: To analyze the specific molecular mechanisms through which epidermal growth factor-like domain 9 (EGFL9) promotes hepatocellular carcinoma (HCC) proliferation, invasion, and migration via the EGFR/PI3K/AKT/mTOR signaling pathway. Methods: Lentivirus-mediated siRNA technology was employed to establish EGFL9-knockdown HCC cell lines (Huh-7 and SNU-449) targeting two distinct genomic sites. Cellular proliferation, apoptosis, migration, and invasion capabilities were evaluated using MTT assays, apoptosis detection, and Transwell assays. Western blotting was performed to examine expression and phosphorylation levels of key EGFR/PI3K/AKT/mTOR pathway components in both EGFL9-knockdown and EGFL9-overexpressing cells , as well as in corresponding subcutaneous xenograft tumor tissues. Pre-established EGFL9-overexpressing Huh-7 cells were treated with EGFR and AKT inhibitors, respectively, followed by reassessment of proliferative and migratory capacities through functional assays and corresponding pathway protein expression changes via Western blotting. Results: RT-qPCR analysis demonstrated over 80% suppression efficiency of EGFL9 expression in both Huh-7 and SNU-449 HCC cell lines with dual-targeting knockdown. Functional assays revealed that EGFL9 knockdown significantly inhibited proliferative, invasive, and migratory capacities while promoting apoptosis in both HCC cell lines. Western blot analysis indicated that EGFL9 knockdown substantially reduced phosphorylation levels of EGFR, PI3K, AKT, and mTOR proteins without affecting their total expression levels. Conversely, EGFL9-overexpressing Huh-7 cells maintained comparable total protein expression but exhibited markedly elevated phosphorylation levels of these pathway components relative to controls. Both EGFR and AKT inhibitor treatments significantly diminished pathway protein phosphorylation and correspondingly attenuated proliferative and migratory capacities in EGFL9-overexpressing Huh-7 cells. Conclusion: EGFL9 may enhance HCC cell proliferation, invasion, and migration capabilities through the EGFR-PI3K-AKT-mTOR signaling pathway, suggesting its potential as a novel therapeutic target for hepatocellular carcinoma. |
| Key words: EGFL9 Hepatocellular carcinoma Signal transduction pathway Proliferation Invasion |
