Exosomes are microvesicles secreted by body cells for intercellular communication. The circular RNA circ_0000338 was found to be present in extracellular vesicles and improve the chemoresistance of colorectal cancer (CRC) cells. However, the role of exosomal circ_0000338 in 5-fluorouracil (5-FU) resistance in CRC is largely unknown. The levels of circ_0000338, microRNA 217 (miR-217), and miR-485-3p were detected using quantitative real-time PCR (qRT-PCR). The 50% inhibitory concentration (IC₅₀) values of cells for 5-FU, cell proliferation, and apoptosis were evaluated using cell counting kit 8 (CCK-8), colony formation, flow cytometry, and Western blot assays. The interaction between miR-217 or miR-485-3p and circ_0000338 was confirmed by RNA immunoprecipitation (RIP), dual-luciferase reporter, and pulldown assays. Exosomes were isolated by ultracentrifugation and qualified by transmission electron microscopy (TEM), Nanosight tracking analysis (NTA), and Western blotting. Xenograft models were performed to analyze whether circ_0000338-loaded exosomes could increase resistance of CRC cells to 5-FU in vivo. The circ_0000338 level was elevated in 5-FU-resistant CRC tissues and cells, and circ_0000338 knockdown sensitized 5-FU-resistant CRC cells to 5-FU through enhancing apoptosis and decreasing proliferation in vitro. Mechanistically, circ_0000338 directly bound to miR-217 and miR-485-3p, and the inhibition of miR-217 or miR-485-3p reversed the effects of circ_0000338 knockdown on cell 5-FU resistance in CRC. Additionally, extracellular circ_0000338 could be incorporated into secreted exosomes and transmitted to 5-FU-sensitive cells. Treatment-sensitive cells with exosomes containing circ_0000338 reduced the 5-FU response in CRC both in vitro and in vivo. Besides that, the exosomal circ_0000338 concentration was higher in patients exhibiting resistance to 5-FU and showed good diagnostic efficiency in 5-FU-resistant CRC. The delivery of circ_0000338 via exosomes enhanced 5-FU resistance in CRC through negative regulation of miR-217 and miR-485-3p, indicating a promising diagnostic and therapeutic marker for 5-FU-based chemotherapy in CRC patients.

中文翻译：

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circ_0000338 的外泌体介导的转移通过调节 MicroRNA 217 (miR-217) 和 miR-485-3p 增强结直肠癌的 5-氟尿嘧啶耐药性

外泌体是体细胞分泌的用于细胞间通讯的微泡。发现环状 RNA circ_0000338 存在于细胞外囊泡中并提高结直肠癌 (CRC) 细胞的化学抗性。然而，外泌体 circ_0000338 在 CRC 中 5-氟尿嘧啶 (5-FU) 耐药性中的作用在很大程度上是未知的。使用实时定量 PCR (qRT-PCR) 检测 circ_0000338、microRNA 217 (miR-217) 和 miR-485-3p 的水平。50% 抑制浓度 (IC ₅₀) 使用细胞计数试剂盒 8 (CCK-8)、集落形成、流式细胞术和蛋白质印迹分析评估细胞的 5-FU、细胞增殖和凋亡值。miR-217 或 miR-485-3p 与 circ_0000338 之间的相互作用已通过 RNA 免疫沉淀 (RIP)、双荧光素酶报告基因和下拉分析证实。通过超速离心分离外泌体，并通过透射电子显微镜 (TEM)、Nanosight 跟踪分析 (NTA) 和蛋白质印迹鉴定。进行异种移植模型以分析加载circ_0000338的外泌体是否可以在体内增加CRC细胞对5-FU的抗性. circ_0000338 水平在 5-FU 抗性 CRC 组织和细胞中升高，circ_0000338 敲低通过增强体外细胞凋亡和减少增殖使 5-FU 抗性 CRC 细胞对 5-FU 敏感。从机制上讲，circ_0000338 直接与 miR-217 和 miR-485-3p 结合，抑制 miR-217 或 miR-485-3p 逆转了 circ_0000338 敲低对 CRC 细胞 5-FU 抗性的影响。此外，细胞外 circ_0000338 可以整合到分泌的外泌体中并传输到 5-FU 敏感细胞。具有包含 circ_0000338 的外泌体的治疗敏感细胞在体外和体内都降低了 CRC 中的 5-FU 反应. 除此之外，外泌体 circ_0000338 浓度在对 5-FU 耐药的患者中更高，并且在 5-FU 耐药的 CRC 中显示出良好的诊断效率。通过外泌体递送circ_0000338通过miR-217和miR-485-3p的负调节增强了CRC中的5-FU抗性，表明CRC患者中基于5-FU的化疗的有希望的诊断和治疗标志物。