Tumor cells switch from an epithelial to a mesenchymal-like phenotype, which represents a key hallmark of human cancer metastasis, including gallbladder cancer (GBC). A large set of microRNAs (miRNAs/miRs) have been studied to elucidate their functions in initiating or inhibiting this phenotypic switching in GBC cells. In this paper, we attempted to identify the expression pattern of the miR-214/−3120 cluster and its mode of action in the context of GBC, with a specific focus being placed on their effects on EMT and autophagy in GBC cells. Human GBC cells GBC-SD were assayed for their migration, invasion, and autophagy using the Transwell chamber system, MDC staining, and transmission electron microscopy. The tumorigenicity and metastatic behavior of GBC-SD cells were tested in nude mice. The expression of EMT- and autophagy-specific markers (E-cadherin, N-cadherin, vimentin, ATG5, LC3II/LC3I, and Beclin1) was analyzed in cultured GBC-SD cells and in human GBC-SD xenografts. The E2F3 luciferase reporter activity in the presence of miR-214/−3120 was evaluated by a dual luciferase assay. The miR-214/−3120 was downregulated in GBC. Exogenous miR-214/−3120 inhibited the phenotypic switching of GBC cells from epithelial to mesenchymal, prevented autophagy, and suppressed the tumorigenicity and metastatic behavior of GBC-SD cells in vitro and in vivo. E2F3 was demonstrated to be the target gene of miR-214/−3120, and its knockdown in part mimicked the effect of miR-214/−3120 on the EMT, autophagy, tumorigenicity, and metastatic behavior of GBC-SD cells. These results demonstrated that the miR-214/−3120 cluster blocks the process of EMT and autophagy to limit GBC metastasis by repressing E2F3 expression.

肿瘤细胞从上皮细胞转变为间充质样表型，这是人类癌症转移的关键标志，包括胆囊癌 (GBC)。已经研究了大量的 microRNA (miRNA/miR)，以阐明它们在启动或抑制 GBC 细胞中这种表型转换中的功能。在本文中，我们试图确定 miR-214/-3120 簇的表达模式及其在 GBC 环境中的作用方式，特别关注它们对 GBC 细胞中 EMT 和自噬的影响。使用 Transwell 室系统、MDC 染色和透射电子显微镜分析人类 GBC 细胞 GBC-SD 的迁移、侵袭和自噬。在裸鼠中测试GBC-SD细胞的致瘤性和转移行为。在培养的 GBC-SD 细胞和人类 GBC-SD 异种移植物中分析了 EMT 和自噬特异性标志物（E-cadherin、N-cadherin、波形蛋白、ATG5、LC3II/LC3I 和 Beclin1）的表达。通过双荧光素酶测定评估在 miR-214/-3120 存在下的 E2F3 荧光素酶报告基因活性。miR-214/-3120 在 GBC 中被下调。外源性 miR-214/-3120 抑制 GBC 细胞从上皮细胞向间充质细胞的表型转换，阻止自噬，并抑制 GBC-SD 细胞的致瘤性和转移行为体外和体内。E2F3 被证明是 miR-214/-3120 的靶基因，其敲低部分模仿了 miR-214/-3120 对 GBC-SD 细胞的 EMT、自噬、致瘤性和转移行为的影响。这些结果表明 miR-214/-3120 簇通过抑制 E2F3 表达来阻断 EMT 和自噬过程以限制 GBC 转移。