具核梭杆菌感染的结肠癌细胞分泌的外泌体通过传递 hsa_circ_0004085 来传递对奥沙利铂和 5-FU 的耐药性。
Exosomes secreted by Fusobacterium nucleatum-infected colon cancer cells transmit resistance to oxaliplatin and 5-FU by delivering hsa_circ_0004085.
机构信息
Department of Oncology and Cancer Rehabilitation Center, The First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, China.
The First Clinical Medical College of Nanjing Medical University, Nanjing, Jiangsu, China.
出版信息
J Nanobiotechnology. 2024 Feb 15;22(1):62. doi: 10.1186/s12951-024-02331-9.
BACKGROUND
A large number of Fusobacterium nucleatum (Fn) are present in colorectal cancer (CRC) tissues of patients who relapse after chemotherapy, and Fn has been reported to promote oxaliplatin and 5-FU chemoresistance in CRC. Pathogens such as bacteria and parasites stimulate exosome production in tumor cells, and the regulatory mechanism of exosomal circRNA in the transmission of oxaliplatin and 5-FU chemotherapy resistance in Fn-infected CRC remains unclear.
METHODS
Hsa_circ_0004085 was screened by second-generation sequencing of CRC tissues. The correlation between hsa_circ_0004085 and patient clinical response to oxaliplatin/5-FU was analyzed. Exosome tracing experiments and live imaging systems were used to test the effect of Fn infection in CRC on the distribution of hsa_circ_0004085. Colony formation, ER tracking analysis and immunofluorescence were carried out to verify the regulatory effect of exosomes produced by Fn-infected CRC cells on chemotherapeutic resistance and ER stress. RNA pulldown, LC-MS/MS analysis and RIP were used to explore the regulatory mechanism of downstream target genes by hsa_circ_0004085.
RESULTS
First, we screened out hsa_circ_0004085 with abnormally high expression in CRC clinical samples infected with Fn and found that patients with high expression of hsa_circ_0004085 in plasma had a poor clinical response to oxaliplatin/5-FU. Subsequently, the circular structure of hsa_circ_0004085 was identified. Fn infection promoted hsa_circ_0004085 formation by hnRNP L and packaged hsa_circ_0004085 into exosomes by hnRNP A1. Exosomes produced by Fn-infected CRC cells transferred hsa_circ_0004085 between cells and delivered oxaliplatin/5-FU resistance to recipient cells by relieving ER stress. Hsa_circ_0004085 enhanced the stability of GRP78 mRNA by binding to RRBP1 and promoted the nuclear translocation of ATF6p50 to relieve ER stress.
CONCLUSIONS
Plasma levels of hsa_circ_0004085 are increased in colon cancer patients with intracellular Fn and are associated with a poor response to oxaliplatin/5-FU. Fn infection promoted hsa_circ_0004085 formation by hnRNP L and packaged hsa_circ_0004085 into exosomes by hnRNP A1. Exosomes secreted by Fn-infected CRC cells deliver hsa_circ_0004085 between cells. Hsa_circ_0004085 relieves ER stress in recipient cells by regulating GRP78 and ATF6p50, thereby delivering resistance to oxaliplatin and 5-FU.
背景
大量的具核梭杆菌（Fn）存在于化疗后复发的结直肠癌（CRC）患者的组织中，并且已有报道称 Fn 可促进 CRC 对奥沙利铂和 5-FU 的耐药性。细菌和寄生虫等病原体可刺激肿瘤细胞产生外泌体，而 Fn 感染的 CRC 中外泌体 circRNA 在传递奥沙利铂和 5-FU 化疗耐药性中的调控机制尚不清楚。
方法
通过对 CRC 组织的第二代测序筛选出 hsa_circ_0004085。分析 hsa_circ_0004085 与患者对奥沙利铂/5-FU 临床反应的相关性。通过外泌体示踪实验和活细胞成像系统检测 Fn 感染 CRC 对 hsa_circ_0004085 分布的影响。通过集落形成、ER 追踪分析和免疫荧光实验验证 Fn 感染 CRC 细胞产生的外泌体对化疗耐药性和 ER 应激的调节作用。通过 RNA 下拉、LC-MS/MS 分析和 RIP 实验探索 hsa_circ_0004085 对下游靶基因的调控机制。
结果
首先，我们筛选出 Fn 感染 CRC 临床样本中异常高表达的 hsa_circ_0004085，并发现血浆中 hsa_circ_0004085 高表达的患者对奥沙利铂/5-FU 的临床反应较差。随后，鉴定了 hsa_circ_0004085 的环状结构。Fn 通过 hnRNP L 促进 hsa_circ_0004085 的形成，并通过 hnRNP A1 将 hsa_circ_0004085 包装成外泌体。Fn 感染的 CRC 细胞产生的外泌体将 hsa_circ_0004085 在细胞间传递，并通过减轻 ER 应激将奥沙利铂/5-FU 耐药性传递给受体细胞。hsa_circ_0004085 通过与 RRBP1 结合增强 GRP78 mRNA 的稳定性，并促进 ATF6p50 的核转位以减轻 ER 应激。
结论
结直肠癌患者体内有胞内 Fn 时，血浆中 hsa_circ_0004085 的水平升高，且与奥沙利铂/5-FU 反应不良相关。Fn 通过 hnRNP L 促进 hsa_circ_0004085 的形成，并通过 hnRNP A1 将 hsa_circ_0004085 包装成外泌体。Fn 感染的 CRC 细胞分泌的外泌体在细胞间传递 hsa_circ_0004085。hsa_circ_0004085 通过调节 GRP78 和 ATF6p50 减轻 ER 应激，从而传递奥沙利铂和 5-FU 的耐药性。
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