Wang Zijin, Liu Xing Chen, Gao Zhen Gya, Shi Wo Da, Wang Wen Cai
1Department of Cardiothoracic Surgery, Affiliated Hospital 6 of Nantong University The Yancheng School of Clinical Medicine of Nanjing Medical University, Yancheng Third People's Hospital, Yancheng Jiangsu, 224000 China.
Acta Pharm. 2025 Apr 10;75(1):69-86. doi: 10.2478/acph-2025-0009. Print 2025 Mar 1.
This study aims to investigate the regulatory mechanisms of METTL3, YTHDF1, and the long non-coding RNA FOXD2-AS1 in the proliferation and apoptosis of esophageal cancer, with the goal of providing a basis for molecular diagnosis and targeted therapies. Gene expression was evaluated using qRT-PCR (METTL3/14) and Western blot analysis. The Cell Counting Kit-8 (CCK-8) assay, flow cytometry, and Transwell assay were employed to assess cell proliferation and apoptosis. The EpiQuik m6A RNA Methylation Quantification Kit was utilized to quantify total m6A levels. The interaction between YTHDF1, FOXD2-AS1, and METTL3 was confirmed using RNA Binding Protein Immunoprecipitation (RIP), Co-Immunoprecipitation (CO-IP), and RNA pull-down assays. Methylated RNA Immuno preci pitation (MeRIP) was employed to assess the m6A modification levels of FOXD2-AS1. Tissue samples from animal models were analyzed via Hematoxylin-eosin staining (HE) staining and immunohisto-chemistry to assess METTL3 expression. The expression of was up-regulated in esophageal cancer tissues and cells. Flow cytometry and CCK-8 detection showed that silencing could inhibit the proliferation of esophageal cancer cells but accelerate their apoptosis. MeRIP-qPCR and Prediction of m6A-modified sites indicated that METTL3 regulated the m6A modification of FOXD2-AS1. and experiments showed that YTHDF1 binds to METTL3 and regulates the m6A modification of FOXD2-AS1 to affect esophageal cancer. Our results indicate that METTL3 regulates FOXD2-AS1 in an m6A-dependent manner through its interaction with YTHDF1, thereby influencing EC proliferation and apoptosis. This suggests a potential therapeutic target for the treatment of esophageal cancer.
本研究旨在探讨METTL3、YTHDF1和长链非编码RNA FOXD2-AS1在食管癌增殖和凋亡中的调控机制，为分子诊断和靶向治疗提供依据。采用qRT-PCR（METTL3/14）和蛋白质免疫印迹分析评估基因表达。使用细胞计数试剂盒-8（CCK-8）检测、流式细胞术和Transwell检测评估细胞增殖和凋亡。利用EpiQuik m6A RNA甲基化定量试剂盒定量总m6A水平。通过RNA结合蛋白免疫沉淀（RIP）、免疫共沉淀（CO-IP）和RNA下拉实验证实YTHDF1、FOXD2-AS1和METTL3之间的相互作用。采用甲基化RNA免疫沉淀（MeRIP）评估FOXD2-AS1的m6A修饰水平。通过苏木精-伊红染色（HE）和免疫组化分析动物模型的组织样本，以评估METTL3的表达。 在食管癌组织和细胞中表达上调。流式细胞术和CCK-8检测表明，沉默 可抑制食管癌细胞的增殖，但加速其凋亡。MeRIP-qPCR和m6A修饰位点预测表明，METTL3调节FOXD2-AS1的m6A修饰。 和 实验表明，YTHDF1与METTL3结合并调节FOXD2-AS1的m6A修饰，从而影响食管癌。我们的结果表明，METTL3通过与YTHDF1相互作用以m6A依赖的方式调节FOXD2-AS1，从而影响食管癌的增殖和凋亡。这提示了一种治疗食管癌的潜在治疗靶点。
