Metal-Regulatory Transcription Factor-1 Targeted by miR-148a-3p Is Implicated in Human Hepatocellular Carcinoma Progression.
Metal-Regulatory Transcription Factor-1 Targeted by miR-148a-3p Is Implicated in Human Hepatocellular Carcinoma Progression.
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DOI：
10.3389/fonc.2021.700649
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发表时间：
2021
影响因子：
4.7
通讯作者：
Yang Z
中科院分区：
文献类型：
--
作者：
Lyu Z;Yang M;Yang T;Ma M;Yang Z
金属调节转录因子-1（MTF-1）在维持体内金属平衡中起重要作用。铜暴露显著刺激具有增强的MTF-1表达的肝细胞癌（HCC）细胞的增殖。然而，潜在的分子机制尚未完全阐明。在这项研究中，我们利用不同的方法来研究MTF-1在HCC进展中的潜在作用。使用实时聚合酶链反应（PCR）、蛋白质印迹和免疫组织化学测定MTF-1和miR-148 a-3 p的表达水平。分别采用免疫沉淀法和双荧光素酶报告基因法检测MTF-1与脱嘌呤脱嘧啶核酸内切酶/氧化还原效应因子1（APE/Ref-1）或miR-148 a-3 p的相互作用。采用集落形成法、MTT法、划痕法和Transwell法检测细胞存活率和转移能力，流式细胞仪检测凋亡细胞。还使用异种移植小鼠模型确定MTF-1和miR-148 a-3 p的生物学功能。MTF-1在HCC细胞中表达上调，并与生存率和复发相关。MTF-1过表达可增强肝癌细胞的增殖和转移能力。进一步的机制分析表明，MTF-1与APE/Ref-1结合，并且MTF-1是miR-148- 3 p的直接靶点，miR-148- 3 p反向调节MTF-1的转录活性。miR-148 a-3 p过表达可有效抑制MTF-1刺激的HCC细胞增殖和转移，并增加细胞凋亡。从HCC患者血浆和HCC细胞培养上清液中分离的外泌体中miR-148 a-3 p表达降低。HCC细胞与来自肝细胞条件培养基的外来体共孵育抑制细胞迁移并引起凋亡。体内研究显示，与对照组相比，MTF-1敲低和miR-148 a-3 p过表达的Hep 3B来源的异种移植物生长缓慢，肿瘤体积和重量减少。总的来说，这些发现暗示MTF-1作为HCC肿瘤发生和进展的调节剂。选择性靶向外泌体miR-148 a-3 p，这可能有助于至少部分地在HCC中负调节MTF-1，证明了对HCC患者的治疗益处。
Metal-regulatory transcription factor-1 (MTF-1) is of importance in maintaining metal homeostasis. Copper exposure considerably stimulates the proliferation of hepatocellular carcinoma (HCC) cells with enhanced MTF-1 expression. However, the underlying molecular mechanisms have not been completely elucidated. In this study, we utilized different approaches to investigate the potential role of MTF-1 involved in HCC progression. The expression levels of MTF-1 and miR-148a-3p were determined using real-time polymerase chain reaction (PCR), Western blotting, and immunohistochemistry. The interaction of MTF-1 with apurinic apyrimidinic endonuclease/redox effector factor 1 (APE/Ref-1) or miR-148a-3p was determined using immunoprecipitation or dual-luciferase reporter assay, respectively. Cell viability and metastatic ability were evaluated using colony formation, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), wound scratch, and Transwell assays, and apoptotic cells were detected by flow cytometry. The biological functions of MTF-1 and miR-148a-3p were also determined using a xenograft mouse model. MTF-1 expression was upregulated in HCC cells and was associated with poor survival and recurrence. MTF-1 overexpression enhanced the proliferation and metastatic potential of HCC cells. Further mechanistic analyses demonstrated that MTF-1 bound to APE/Ref-1 and that MTF-1 is a direct target of miR-148-3p, which inversely regulated MTF-1 transcription activity. MiR-148a-3p overexpression effectively inhibited HCC cell proliferation and metastasis stimulated by MTF-1, with increased apoptosis. There was a decrease in miR-148a-3p expression in exosomes isolated from the plasma of patients with HCC and HCC cell culture supernatants. Co-incubation of HCC cells with exosomes from hepatocyte-conditioned media inhibited cell migration and caused apoptosis. The in vivo study revealed slow growth of MTF-1-knockdown and miR-148a-3p-overexpressing Hep3B-derived xenografts, with reduced tumor volume and weight compared with the control group. Collectively, these findings implicate MTF-1 as a modulator of HCC tumorigenesis and progression. Selective targeting towards exosomal miR-148a-3p, which might contribute to the negative regulation of MTF-1 at least partially in HCC, demonstrates therapeutic benefits for patients with HCC.
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