This study aimed to screen osteosarcoma (OS) prognosis relevant genes for methylation dysregulation, and the functional mechanisms of FES overexpression in OS cells were investigated. The OS prognosis relevant genes with differentially methylated positions (DMPs) identified from the GSE36001 and GSE36002 datasets, and the UCSC database, were used as a training set to construct a risk model, while the GSE21257 dataset was used as validation set. The expression levels of several key genes in OS cells after 5-Aza-2′-deoxycytidine treatment were detected by qPCR. The effects of FES overexpression on cell proliferation, cell cycle, migration, and invasion of MNNG/HOS were analyzed by CCK8, flow cytometry, and Transwell assays. A total of 31 candidate genes, corresponding to 36 DMPs, were identified as OS prognosis relevant genes; from these, the top 10 genes were used to construct a risk model. Following validation of the risk model, FES, LYL1, MAP4K1, RIPK3, SLC15A3, and STAT3 showed expression changes between the OS and control samples. qPCR results showed that the expression of FES was significantly downregulated in three OS cell lines and increased after 5-Aza-DC treatment. The proliferation, cell cycle progression, migration, and invasion of MNNG/HOS cells were significantly inhibited after transfection with FES overexpression plasmid, and the protein expression of FYN and β catenin were decreased in MNNG/HOS cells by FES overexpression. The decrease in FES by hypermethylation was associated with OS prognosis, and might contribute to the proliferation, migration, and invasion of OS cells. FES, and its upstream FYN and β catenin, might coordinately exert a tumor suppressor effect in OS cells.

中文翻译：

---

FES的过表达可能抑制骨肉瘤细胞的增殖、迁移和侵袭

本研究旨在筛选骨肉瘤（OS）预后相关基因甲基化失调，并研究OS细胞中FES过表达的功能机制。从 GSE36001 和 GSE36002 数据集和 UCSC 数据库中鉴定的具有差异甲基化位置 (DMP) 的 OS 预后相关基因用作构建风险模型的训练集，而 GSE21257 数据集用作验证集。通过qPCR检测5-Aza-2'-脱氧胞苷处理后OS细胞中几个关键基因的表达水平。通过CCK8、流式细胞术和Transwell检测分析FES过表达对MNNG/HOS的细胞增殖、细胞周期、迁移和侵袭的影响。共有31个候选基因，对应36个DMP，被鉴定为OS预后相关基因；从这些，前 10 个基因用于构建风险模型。在验证风险模型后，FES、LYL1、MAP4K1、RIPK3、SLC15A3 和 STAT3 显示 OS 和对照样本之间的表达变化。qPCR结果显示FES的表达在三种OS细胞系中显着下调，并且在5-Aza-DC处理后增加。转染FES过表达质粒后MNNG/HOS细胞的增殖、细胞周期进程、迁移和侵袭能力受到明显抑制，FES过表达后MNNG/HOS细胞中FYN和β-catenin蛋白表达降低。高甲基化导致的 FES 降低与 OS 预后相关，并可能有助于 OS 细胞的增殖、迁移和侵袭。FES 及其上游 FYN 和 β 连环蛋白，