[摘 要] 目的：探讨干扰素刺激基因家族成员黏病毒抵抗蛋白2（MX2）在调控肿瘤细胞对呼肠孤病毒（Reo）溶瘤敏感性中的 作用及其机制。方法：选取4株具有不同耐药特征的人源肿瘤细胞，通过CCK-8法评估其对Reo的溶瘤敏感性；通过转录组测序 筛选出差异表达基因MX2，qPCR法及WB法验证MX2在4株人源肿瘤细胞中的表达；使用siRNA敲低溶瘤低敏感的COC1/DDP 细胞中的MX2基因。在细胞感染Reo病毒后，通过CCK-8法检测细胞存活率；qPCR法检测细胞中Reo病毒S1基因表达；免疫荧 光法检测细胞内Reo病毒蛋白的积累；半数组织培养感染剂量（TCID50 ）法测定病毒滴度；流式细胞术分别检测细胞内Reo病毒 dsRNA、活性氧（ROS）水平以及细胞凋亡率；透射电镜观察细胞内质网形态变化并采用WB法检测内质网应激相关蛋白（JNK、 p-JNK、eIF2α、p-eIF2α、CHOP、PERK）的表达。结果：在4株肿瘤细胞中，SKOV3细胞对Reo溶瘤作用高度敏感，而COC1/DDP、 HuH-7SRB 及SNU-398细胞均为溶瘤低敏感性。转录组测序结果显示，MX2在溶瘤低敏感肿瘤细胞中的表达水平显著高于溶 瘤高敏感细胞（P < 0.01）；在溶瘤低敏感性的COC1/DDP细胞中，敲低MX2显著促进Reo病毒复制、诱导细胞凋亡增加，并升高 细胞内活性氧水平（均P < 0.001）。透射电镜观察显示，敲低MX2的COC1/DDP细胞感染Reo病毒后出现内质网肿胀、扩张及断 裂等典型内质网应激超微结构改变。WB结果显示，内质网应激关键标志物eIF2α/p-eIF2α、PERK、CHOP及凋亡相关调节蛋白 JNK/p-JNK的表达均显著上调（P < 0.05或P < 0.01）。结论：肿瘤细胞对Reo的溶瘤敏感性与其细胞内MX2表达水平密切相关。 敲低MX2可显著增强Reo在细胞内的复制，进而促进ROS积累，触发内质网应激并促进凋亡。病毒复制增加与细胞凋亡激活的 双重作用，最终协同增强Reo的溶瘤作用。

[Abstract] Objective: To investigate the role and underlying mechanism of myxovirus resistance protein 2 (MX2), a member of the interferon-stimulated gene family, in modulating tumor cell sensitivity to reovirus (Reo)-mediated oncolysis. Methods: Four human tumor cell lines with distinct drug-resistance characteristics were selected, and their sensitivity to Reo-mediated oncolysis was evaluated using the CCK-8 assay. Transcriptome sequencing was performed to identify differentially expressed genes, and MX2 was screened as a candidate gene. The differential expression of MX2 was validated by qPCR and WB assay. In the Reo-low-sensitive COC1/DDP cells, MX2 was knocked down using siRNA, followed by Reo infection, and cell viability was assessed using CCK-8 assay. Viral replication was evaluated by measuring S1 gene expression using qPCR, intracellular viral protein accumulation by immunofluorescence staining, and viral titer by the TCID50 assay. Intracellular Reo dsRNA levels, reactive oxygen species (ROS) production, and apoptosis rates were measured using flow cytometry. Ultrastructural changes in the endoplasmic reticulum (ER) were examined using transmission electron microscopy (TEM). Additionally, WB was used to assess the expression of ER stress-related proteins, including JNK, p-JNK, eIF2α, p-eIF2α, CHOP, and PERK. Results: Among the four tested tumor cell lines, SKOV3 cells exhibited high sensitivity to Reo-mediated oncolysis, whereas COC1/DDP, HuH-7SRB , and SNU-398 cells showed low sensitivity. Transcriptome sequencing revealed that MX2 expression was significantly higher in Reo-low-sensitive cell lines compared with Reo-high-sensitive cells (P < 0.01). In COC1/DDP cells, MX2 knockdown markedly enhanced Reo replication, increased apoptosis, and elevated intracellular ROS levels (all P < 0.001). TEM revealed typical ultrastructural features of ER stress in MX2-knockdown COC1/DDP cells following Reo infection, including ER swelling, dilation, and fragmentation. WB analysis showed significant upregulation of key ER stress markers (p-eIF2α/eIF2α, PERK, and CHOP) and apoptosis-related regulatory proteins JNK/ p-JNK (P < 0.05 or P < 0.01). Conclusion: Tumor cell sensitivity to Reo-mediated oncolysis is closely associated with intracellular MX2 expression levels. Knockdown of MX2 significantly enhanced intracellular Reo replication, leading to ROS accumulation, activation of ER stress, and induction of apoptosis. The synergistic effects of enhanced viral replication and apoptosis ultimately potentiate the oncolytic efficacy of Reo.

[基金项目] 国家自然科学基金（82460604）；贵州省科技计划支撑项目资助[黔科合支撑（2025）一般126号]；贵州医科大学 肿瘤免疫治疗工程研究中心项目（校工程中心，第2024[001]号）