Division of Gastrointestinal Surgery, Department of General Surgery, Shanghai Eighth People Hospital, Jiangsu University, Shanghai 200232, P.R. China.
Department of Ophthalmology, Shanghai Eighth People Hospital, Jiangsu University, Shanghai 200232, P.R. China.
Int J Oncol. 2022 Feb;60(2). doi: 10.3892/ijo.2022.5304. Epub 2022 Jan 11.
Resistance to 5‑Fluorouracil (5‑FU) is a frequent occurrence in patients with colorectal cancer (CRC). MicroRNAs (miRNAs) from cancer‑associated fibroblasts (CAFs)‑secreted exosomes have been associated with 5‑FU sensitivity. The potential molecular mechanism of CAFs‑exosomal miRNAs in CRC remains unclear. The aim of the present study was to elucidate the role of exosomal miRNAs in 5‑FU sensitivity in CRC. Exosomes derived from CAFs were extracted. Exosomal miR‑181d‑5p was identified as a miRNA associated with 5‑FU sensitivity. The putative function of exosomal miR‑181d‑5p was evaluated by ethynyl‑2‑deoxyuridine staining, flow cytometry, RNA immunoprecipitation, luciferase reporter assay, tumor xenograft formation, reverse transcription‑quantitative PCR and western blot analysis. Modification of miR‑181d‑5p by the RNA N6‑methyladenosine (mA) methyltransferase like (METTL)3 was examined by m6A methylation analysis. The results indicated that mA modification and METTL3 expression were upregulated in CRC patients. METTL3‑dependent mA methylation promoted the miR‑181b‑5p process by DiGeorge Syndrome Critical Region 8 (DGCR8) in CAFs. CAFs‑derived exosomes inhibited 5‑FU sensitivity in CRC cells through the METTL3/miR‑181d‑5p axis. A mechanistic study revealed that miR‑181d‑5p directly targeted neurocalcin δ (NCALD) to inhibit the 5‑FU sensitivity of CRC cells. Patients with higher NCALD levels exhibited a higher survival rate. Taken together, METTL3‑dependent mA methylation was upregulated in CRC to promote the processing of miR‑181d‑5p by DGCR8. This led to increased miR‑181d‑5p expression, which inhibited the 5‑FU sensitivity of CRC cells by targeting NCALD. The results of the present study provided novel insight into exosomal microRNAs in 5‑FU sensitivity in CRC cells. Furthermore, exosomal miR‑181d‑5p may represent a potential prognostic marker for CRC.
对氟尿嘧啶（5-FU）的耐药性是结直肠癌（CRC）患者中经常发生的现象。来自癌症相关成纤维细胞（CAF）分泌的外泌体的 microRNAs（miRNAs）与 5-FU 敏感性相关。CAF-外泌体 miRNAs 在 CRC 中的潜在分子机制尚不清楚。本研究旨在阐明外泌体 miRNAs 在 CRC 中对 5-FU 敏感性的作用。提取源自 CAF 的外泌体。鉴定出 miR-181d-5p 是与 5-FU 敏感性相关的 miRNA。通过 Ethynyl-2-deoxyuridine 染色、流式细胞术、RNA 免疫沉淀、荧光素酶报告基因检测、肿瘤异种移植形成、逆转录-定量 PCR 和 Western blot 分析评估外泌体 miR-181d-5p 的假定功能。通过 m6A 甲基化分析检查 miR-181d-5p 的 RNA N6-甲基腺苷（mA）甲基转移酶样（METTL）3 修饰。结果表明，CRC 患者中 mA 修饰和 METTL3 表达上调。METTL3 依赖性 mA 甲基化通过 CAFs 中的 DiGeorge 综合征关键区域 8（DGCR8）促进 miR-181b-5p 的加工。CAF 衍生的外泌体通过 METTL3/miR-181d-5p 轴抑制 CRC 细胞对 5-FU 的敏感性。一项机制研究表明，miR-181d-5p 直接靶向神经钙蛋白 δ（NCALD）抑制 CRC 细胞对 5-FU 的敏感性。NCALD 水平较高的患者存活率更高。总之，CRC 中 METTL3 依赖性 mA 甲基化上调以促进由 DGCR8 加工的 miR-181d-5p。这导致 miR-181d-5p 表达增加，通过靶向 NCALD 抑制 CRC 细胞对 5-FU 的敏感性。本研究结果为 CRC 细胞中 5-FU 敏感性的外泌体 microRNAs 提供了新的见解。此外，外泌体 miR-181d-5p 可能代表 CRC 的潜在预后标志物。
