MiR-4739 inhibits the malignant behavior of esophageal squamous cell carcinoma cells via the homeobox C10/vascular endothelial growth factor A/phosphatidylinositol 3-kinase/AKT pathway

Esophageal cancer is a lethal disease, and emerging evidence has shown that microRNAs are involved in its development, progression, and clinical outcome. MicroRNAs are potential biomarkers for esophageal squamous cell carcinoma (ESCC), and may be useful in advanced RNA therapy for ESCC. This study was conducted to evaluate the molecular mechanism of miR-4739 in ESCC. Reverse transcription-quantitative polymerase chain reaction and western blotting were performed to measure RNA and protein levels. Transwell assay, Cell Counting Kit-8 assay, cytometry analysis, and human umbilical vein endothelial cell tube formation assay were conducted to determine the molecular function of miR-4739 in ESCC. Potential targets of miR-4739 were predicted using bioinformatics tools and confirmed in ESCC cells using a luciferase reporter and RNA pulldown assay. Finally, we performed immunohistochemistry to evaluate the effects of administering agomir-4739 to a mouse model of ESCC. MiR-4739 expression was downregulated in ESCC tissues and cells. MiR-4739 overexpression inhibited cell proliferation, migration, and invasion, and promoted apoptosis of ESCC cells. Furthermore, vascular endothelial growth factor A expression was downregulated by miR-4739 mimics in ESCC cells. MiR-4739 negatively regulated homeobox C10 expression. Additionally, agomir-4739 inhibited tumor growth and angiogenesis in vivo. We demonstrated that miR-4739 overexpression exerted an inhibitory effect on ESCC cells by preventing the expression of homeobox C10 via the vascular endothelial growth factor A/phosphatidylinositol 3-kinase/AKT pathway, indicating the potential of this microRNA as a treatment target in ESCC.

食管癌是一种致死性疾病，越来越多的研究证据表明，微小RNA（microRNAs）参与了其发生发展、疾病进展及临床预后过程。微小RNA是食管鳞状细胞癌（esophageal squamous cell carcinoma, ESCC）的潜在生物标志物，或可应用于ESCC的先进RNA治疗领域。本研究旨在探讨miR-4739在ESCC中的分子调控机制。研究采用逆转录定量聚合酶链反应（reverse transcription-quantitative polymerase chain reaction, RT-qPCR）和蛋白质印迹法（western blotting, WB）分别检测RNA与蛋白的表达水平。通过Transwell实验、Cell Counting Kit-8（CCK-8）增殖实验、流式细胞术分析以及人脐静脉内皮细胞管形成实验，明确miR-4739在ESCC中的分子生物学功能。利用生物信息学工具预测miR-4739的潜在靶基因，并通过荧光素酶报告基因实验和RNA下拉实验在ESCC细胞中验证靶基因的结合关系。最后，通过免疫组化实验评估miR-4739激动剂（agomir-4739）对ESCC小鼠模型的干预效果。实验结果显示，miR-4739在ESCC组织及细胞系中表达下调。过表达miR-4739可抑制ESCC细胞的增殖、迁移与侵袭能力，并促进细胞凋亡。此外，miR-4739模拟物可下调ESCC细胞中血管内皮生长因子A（vascular endothelial growth factor A, VEGFA）的表达水平。miR-4739可负向调控同源盒C10（homeobox C10, HOXC10）的基因表达。体内实验进一步证实，agomir-4739可抑制肿瘤生长与血管生成。本研究证实，miR-4739过表达可通过靶向抑制同源盒C10的表达，进而调控血管内皮生长因子A/磷脂酰肌醇3-激酶/AKT（VEGFA/PI3K/AKT）信号通路，最终发挥抑制ESCC细胞恶性表型的作用，提示miR-4739可作为ESCC临床治疗的潜在靶点。