Abstract

ZMIZ1 acts as an oncogene in hepatocellular carcinoma (HCC). circZMIZ1 (hsa_circ_0018964) derives from ZMIZ1; its underlying mechanism in HCC has not been reported. Peripheral blood and peripheral blood mononuclear cells (PBMCs) were obtained from HCC patients and healthy volunteers. CD8⁺ T cells were sorted from PBMCs of HCC patients. Applying flow cytometry, cell apoptosis and the proportion of KCNJ2/CD8⁺ T cells were examined. The cytotoxicity of CD8⁺ T cells against HCC cells was evaluated. The interaction among circZMIZ1, miR-15a-5p, and KCNJ2 was investigated by dual luciferase assay, RNA immunoprecipitation, and RNA pull-down assay. An orthotopic mouse model of HCC was constructed by intrahepatic injection of H22 cells. Upregulation of circZMIZ1 and KCNJ2 and downregulation of miR-15a-5p were observed in peripheral blood and PBMCs of HCC patients. The proportion of KCNJ2/CD8⁺ T cells was also increased in HCC patients. circZMIZ1 knockdown restrained apoptosis of CD8⁺ T cells and elevated cytotoxicity of CD8⁺ T cells. Mechanically speaking, circZMIZ1 elevated KCNJ2 expression by sponging miR-15a-5p. miR-15a-5p inhibitor reversed circZMIZ1 silencing-mediated inhibition of apoptosis and promotion of cytotoxicity in CD8⁺ T cells. In vivo, orthotopic mice of HCC exhibited increased expression of circZMIZ1 and KCNJ2, elevated proportion of KCNJ2/CD8⁺ T cells, and decreased expression of miR-15a-5p. This work demonstrated that circZMIZ1 inhibited the anti-tumor activity of CD8⁺ T cells in HCC by regulating the miR-15a-5p/KCNJ2 axis. This provides a theoretical basis for the development of effective circZMIZ1 in tumor immunotherapy.

中文翻译：

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敲除circZMIZ1可增强CD8+ T细胞的抗肿瘤活性，从而缓解肝细胞癌

摘要

ZMIZ1 在肝细胞癌 (HCC) 中充当癌基因。 circZMIZ1 (hsa_circ_0018964) 源自 ZMIZ1；其在HCC中的作用机制尚未见报道。外周血和外周血单核细胞（PBMC）取自 HCC 患者和健康志愿者。从 HCC 患者的 PBMC 中分选CD8 ^{+ T 细胞。应用流式细胞术检测细胞凋亡及KCNJ2/CD8 +} T细胞比例。评估CD8 ⁺ T细胞对HCC细胞的细胞毒性。通过双荧光素酶测定、RNA 免疫沉淀和 RNA Pull-down 测定研究了 circZMIZ1、miR-15a-5p 和 KCNJ2 之间的相互作用。通过肝内注射H22细胞构建原位小鼠肝癌模型。在 HCC 患者的外周血和 PBMC 中观察到 circZMIZ1 和 KCNJ2 上调以及 miR-15a-5p 下调。 HCC 患者中 KCNJ2/CD8 ⁺ T 细胞的比例也有所增加。 circZMIZ1 敲低抑制了 CD8 ⁺ T 细胞的凋亡并提高了 CD8 ⁺ T 细胞的细胞毒性。从机制上讲，circZMIZ1 通过海绵 miR-15a-5p 来升高 KCNJ2 表达。 miR-15a-5p 抑制剂逆转 circZMIZ1 沉默介导的细胞凋亡抑制并促进 CD8 ⁺ T 细胞的细胞毒性。在体内，HCC原位小鼠表现出circZMIZ1和KCNJ2表达增加、KCNJ2/CD8 ⁺ T细胞比例升高以及miR-15a-5p表达降低。这项工作证明，circZMIZ1通过调节miR-15a-5p/KCNJ2轴抑制HCC中CD8 ⁺ T细胞的抗肿瘤活性。这为开发有效的circZMIZ1在肿瘤免疫治疗中提供了理论基础。